Research ArticleAstroglial Plasticity Is Implicated in Hippocampal Remodellingin Adult Rats Exposed to Antenatal Dexamethasone

Vishvesh H Shende1 Simon McArthur23 Glenda E Gillies2 and Jolanta Opacka-Juffry1

1Department of Life Sciences University of Roehampton London SW15 4JD UK2Division of Experimental Medicine Imperial College London W12 0NN UK3Department of Biomedical Sciences Faculty of Science amp Technology University of Westminster London W1W 6UW UK

Correspondence should be addressed to Jolanta Opacka-Juffry jopacka-juffryroehamptonacuk

Received 5 December 2014 Accepted 4 February 2015

Academic Editor Anna Dunaevsky

Copyright copy 2015 Vishvesh H Shende et al This is an open access article distributed under the Creative Commons AttributionLicense which permits unrestricted use distribution and reproduction in any medium provided the original work is properlycited

The long-term effects of antenatal dexamethasone treatment on brain remodelling in 3-month-old male Sprague Dawley ratswhose mothers had been treated with dexamethasone were investigated in the present study Dorsal hippocampus basolateralamygdala and nucleus accumbens volume cell numbers and GFAP-immunoreactive astroglial cell morphology were analysedusing stereology Total brain volume as assessed by micro-CT was not affected by the treatment The relative volume of the dorsalhippocampus ( of total brain volume) showed a moderate by 8 but significant reduction in dexamethasone-treated versuscontrol animals Dexamethasone had no effect on the total and GFAP-positive cell numbers in the hippocampal subregionsbasolateral amygdala and nucleus accumbens Morphological analysis indicated that numbers of astroglial primary processes werenot affected in any of the hippocampal subregions analysed but significant reductions in the total primary process length wereobserved in CA1 by 32 CA3 by 50 and DG by 25 Mean primary process length values were also significantly decreased inCA1 by 25 CA3 by 45 and DG by 25 No significant astroglial morphological changes were found in basolateral amygdalaand nucleus accumbens We propose that the dexamethasone-dependent impoverishment of hippocampal astroglial morphologyis the case of maladaptive glial plasticity induced prenatally

1 Introduction

Astroglia have been acknowledged to play a role in brainresponses to stress and glucocorticoids as its chemical medi-ators [1 2] Such effects implicate brain plasticity and canlead to regional brain remodelling with volume changesobserved within the limbic system in cases of long-termldquotoxicrdquo stress and depression and depression-like conditionsin humans and animals respectively [1 3] Thus increasedamygdala volumes have been observed in teenage adopteeswho experienced early life deprivation [4] On the otherhand hippocampal volume reductions have been reportedin patients with PTSD and major depressive disorder withhistory of early life deprivation [5] Stress-related reductionsin hippocampal volume have also been observed in experi-mental animals both rodents [6 7] and nonhuman primates[8] Hippocampal volume losses albeit usually of modestdegree indicate changes in brain tissue architecture with

most studies reporting on neuroplastic rearrangements [9]However there is also evidence of astroglial involvement inhippocampal remodelling observed in the rat model of earlylife deprivation [7]

Also prenatal stress can result in a reduced hippocampalvolume associated with suppressed neurogenesis in rhesusmonkeys a phenomenon found to be mediated by corticos-teroids [10] Antenatal treatment with synthetic steroids suchas dexamethasone which cross the placenta [11] is often usedin pregnant women at risk for preterm birth [12] It can how-ever affect neurobehavioural development of children whohave lower IQ scores and poormotor and visual coordinationskills during their school age [13] About 85of neonateswithantenatal corticosteroid therapy receivemultiple courses [14]and dexamethasone is commonly administered to ventilator-dependent premature infants with chronic lung insufficiencyto improve lung function [15 16]

Hindawi Publishing CorporationNeural PlasticityVolume 2015 Article ID 694347 8 pageshttpdxdoiorg1011552015694347

2 Neural Plasticity

The first human study on postmortem hippocampi ofneonates who had been antenatally treated with dexametha-sone or betamethasone has shown a glucocorticoid-relatedreduced density of neurons no differences have been found ingliosis or myelination [17] Experimental studies on animalsconfirmnegative effects of antenatal glucocorticoid treatment(AGT) on hippocampal neurogenesis [18] however thereare no reports about AGT effects on brain glia The presentstudy addressed this gap in knowledge and hypothesised thatrat hippocampal astroglia respond to AGT in a maladaptivemanner in the long term

2 Experimental Procedures

21 Animals All animal procedures were carried out inaccordance with the United Kingdom Animals ScientificProcedures Act of 1986 at Imperial College London SpragueDawley rats (Harlan Olac Blackthorn Bicester OxfordshireUK) were kept under controlled lighting (on 0800ndash2000 h)temperature (21ndash23∘C) and humidity (63) with standardrat chow and drinking water (except as described below) pro-vided ad libitum Male and female rats were caged separatelyand allowed to acclimatize to their new environment for oneweek after which groups of one male and two female ratswere housed together overnight and the presence of vaginalplugs the following morning was taken to confirm matingpregnancy was confirmed approximately 6 days later bypalpationThe timed pregnant rats were housed five per cageuntil gestational day (GD) 15 when they were caged singlyin preparation for giving birth From GD 1920 pregnant ratswere monitored several times a day and the day of birthwas defined as day 0 No more than two progenies per litter(one male one female) were included in each experimentalgroup in order to minimize potential effects of litter-of-origin Offspring were weaned at three weeks male andfemale progenywere housed separately and theywere allowedto grow to adulthood with no further interventions otherthan normal husbandry At 68 plusmn 2 days of age animals weredecapitated between 0900 and 1000 h Male rats were used inthe present study

22 Dexamethasone Treatment Dexamethasone was admin-istered noninvasively as dexamethasone sodium phosphate(Faulding Pharmaceuticals Plc Royal Leamington Spa UK)in the drinking water given to pregnant rats at GD 16ndash19 atthe dose of 05 120583gmL [19] with an estimated daily intake ofapproximately 75120583gkg [20]Withdrawing dexamethasone atGD 19 allowed for clearance of the steroid from the maternalcirculation prior to birth with no observable effects on theoutcomes of pregnancy maternal behaviour or adult bodyweight of the offspring [19ndash21]

23 pQCT Scanning Total frozen brain volume (119899 = 6 pergroup) was analysed by means of peripheral quantitativecomputed tomography (pQCT) on a Stratec Research SA+scanner (Stratec Medizintechnik Pforzheim Germany) asdescribed elsewhere [7] Serial coronal CT scans were per-formed covering entire brain region Further analysis tomeasure total brain volume was carried out by using software

Avizo (version 5 Mercury Computer Systems ChelmsfordMA USA)

24 Brain Tissue Preparation Frozen coronal sections(25 120583m) were cut in an anterior-posterior direction Thesectioning procedure was designed to yield the followingregions of interest (ROIs) (in brackets AP coordinates frombregma in mm) nucleus accumbens core (300 to 216)basolateral amygdala (minus172 to minus228) and dorsal hippo-campus (minus204 to minus468) according to the rat brain atlas byPaxinos and Watson [22]

25 Total Cell Count Brain sections were stained with hema-toxylin (HX) for total cell count and then gradually dehy-drated air-dried and protected with a histological mountingmedium and covered with cover slips

26 Immunohistochemistry To visualise astrocytes glial fib-rillary acidic protein (GFAP) immunohistochemistry wasperformed [23] Briefly peroxidase inactivation was carriedout in PBS with 20 methanol 15 H

2O2 and 03

Triton X-100 blocking medium contained 01 Triton X-100 and 10 normal horse serum and the anti-GFAP pri-mary antibody (Sigma UK) was diluted 1 500 and appliedovernight at 4∘C Controls (blanks) without the primaryantibody were processed in parallel All sections were incu-bated with the secondary antibody and streptavidin-biotin-peroxidase complex according to manufacturerrsquos instruc-tion (ABC kit Vector Laboratories UK) Sections weretreated with 331015840-diaminobenzidine (5mgmL 001 H

2O2)

rinsed dehydrated and mounted in xylene-based Histo-mount (BDHMerck Poole UK) prior to stereology

27 Stereological Procedures

271 Volume Analysis Volume estimation using the Cava-lieri principle was performed with Stereo-Investigator soft-ware (Version 9 MicroBrightField Inc Germany) andOlympus BX51 microscope fitted with a motorized stage andvideo camera on a Zeiss UPlan FLN with a 4x objective(NA = 025) Starting at a random position every tenthsection was analyzed leading to an average of 12 sections fordorsal hippocampus 6 sections for basolateral amygdala and5 sections for nucleus accumbens per brain These sets ofconsecutive sections were used for the estimation of volumesfor both control and test groups The investigator was blindto the experimental groups A point counting grid (PCG)(ie 119889 = 50 120583m) was used for volume estimation to obtainmaximum efficiency Representative area per point (119886119901)was2500 120583m2 After applying the PCG on the sampled sectionsin a systematic randommanner the number of points hittingregion of interest was counted (Figure 1(a)) The efficiency ofsampling and volume estimation were checked by estimationof coefficient of error and coefficient of variation [24]

272 Cell Counts Cells in the dorsal hippocampal subre-gions (CA1 CA2 CA3 and DG) basolateral amygdala andnucleus accumbens were counted using the optical fraction-ator (Stereo-Investigator Version 9 MicroBrightField Inc

Neural Plasticity 3

(a) (b)

Control

(c)

Dex-treated

(d)

Figure 1 Dorsal hippocampal volume HX stain (a) and GFAP-positive astroglial cell count (b) estimated by means of stereology Changesin astroglial morphology in response to AGT GFAP-positive astroglial cells in control (c) and Dex-treated (d) male rats (dorsal hippocampaldentate gyrus area) GFAP-positive primary process length is reduced in Dex-treated male rats when compared with control (d) versus (c)Astroglial processes observed at 100x magnification

Germany) An unbiased estimation of total number of cellswas achieved by choosing every 10th section according to thesystematic random sampling procedure [24] yielding 8 sec-tions per animal A pilot study was undertaken to determinethe following parameters A sampling area of 1602500120583m2was found to be optimal for this study Dissector height was8 120583m and a 1 120583m guard zone at the top and bottom part of thesection was excluded from the analysis at every step A fixedcounting frame of 50 times 50 and a variable sampling grid size(119909-119910-axis) of 150 times 150 CA1 150 times 150 CA2 300 times 300 CA3and 300 times 300 DG were used for the dorsal hippocampus(Figure 1(b)) and 150 times 150 was used for both basolateralamygdala and nucleus accumbens These sampling gridsresulted in 100ndash200 counting sites per brain HX and GFAP-positive cells were counted using a 100x Nicon UPlan FLNobjective (NA = 130) which allowed accurate recognitionEach cell was counted according to the unbiased countingrules The efficiency of convenient number of sampled cellsand parameters was checked by estimation of coefficient oferror (CE) and coefficient of variation (CV) [24]

273 AstroglialMorphology Morphological analysiswas per-formed on 80 astrocytes in each brain (10 astrocytes persection) usingNeurolucida software (Version 9MicroBright-Field Inc Germany) for tracing boundaries of the ROIsand astrocytes Astrocytes within the ROI were traced in

a systematic random manner taking care to avoid thosewhich were superimposed upon other astrocytes or bloodvessels The traced astrocytes were then analysed usingNeurolucida explorer software (Version 9 MicroBrightFieldInc Germany) Morphological analysis was performed byobserver who was blind to the experimental groups Primaryprocess length was measured using 100x objective (NiconUPlan FLN NA = 130) and by counting the primary pro-cesses extending directly from the soma in both the lateraland central quadrants of astrocytes in the same sections

28 Statistical Analysis Using SPSS 170 statistical software(SPSS Inc USA) ANOVA with a post hoc Tukey test wasperformed comparing between group differences Indepen-dent 119905-test was performed tomeasure difference between twogroups Statistical significance was set at 119875 lt 005The resultsare expressed as mean plusmn SEM

3 Results

31 Whole Brain Volume and Brain Weight No significanteffects of AGT were observed in the whole brain volume(119875 = 0376) and brain weight (119875 = 0442) (Table 1)

32 VolumeAnalyses AGT rats showed a significant decreasein the absolute volume of dorsal hippocampus by 65

4 Neural Plasticity

Table 1 Effects of AGT exposure on brain weight and volume ofmale adult Sprague Dawley rats

Control Dex-treatedBrain weight (g) 174 plusmn 002 177 plusmn 003Brain volume (mm3) 165018 plusmn 1775 168124 plusmn 2843The brain weight and volume are expressed as mean plusmn SEM (119899 = 6 pergroup) No significant differences between Dex-treated (AGT) and controlwere observed

Table 2 Effects of AGT on the regional brain volume of male adultSprague Dawley rats

VolumeControl Dex-treated

Absolute values (mm3)Dorsal hippocampus 328 plusmn 046 308 plusmn 050lowast

Basolateral amygdala 081 plusmn 002 080 plusmn 002Nucleus accumbens 228 plusmn 006 226 plusmn 007

Relative values (as brain volume)Dorsal hippocampus 199 plusmn 004 183 plusmn 004lowast

Basolateral amygdala 0046 plusmn 0001 0048 plusmn 0001Nucleus accumbens 0138 plusmn 0004 0135 plusmn 0005

The volume of the dorsal hippocampus basolateral amygdala and nucleusin control and dexamethasone-treated groups (119899 = 6 per group) Data areexpressed as mean plusmn SEM lowast119875 lt 005 versus control independent sample 119905-test

versus control (119875 lt 005) When expressed as percentageof the whole brain volume the volume reduction was 8(119875 lt 005) versus control No significant differences wereobserved between the treatment groups in the basolateralamygdala (119875 = 0834) and nucleus accumbens (119875 = 0905)(Table 2)

33 HX Stained Total Cell Numbers AGT did not affect thetotal cell count (HX stain) in the dorsal hippocampal subre-gions CA1 (119875 = 0821) CA2 (119875 = 0975) CA3 (119875 = 0991)and DG (119875 = 0959) basolateral amygdala (119875 = 0180) andthe nucleus accumbens (119875 = 0474) (Table 3)

34 GFAP-Positive Astroglial Cell Numbers There were nosignificant AGT effects on GFAP-positive cell numbers in thedorsal hippocampal subregions CA1 (119875 = 0607) CA2 (119875 =0517) CA3 (119875 = 0500) and DG (119875 = 0424) basolateralamygdala (119875 = 0495) and the nucleus accumbens (119875 =0384) (Table 4)

35 Astroglia (GFAP-Positive) Cell Morphology AGT didnot affect the numbers of primary processes in the regionanalysed (Tables 5 and 6) and it had no significant effectson the astroglial morphology in the nonhippocampal regions(Table 5 all group differences at 119875 lt 005)

However AGT affected the length of primary processesin GFAP-positive cells within the hippocampus (Figures 1(a)and 1(b)) it led to a significant reduction in the total length ofastrocytic primary processes in the hippocampal subregionsby 32 in CA1 (119875 lt 005) 50 in CA3 (119875 lt 0001) and by

Table 3 Effects of AGT on the number of total (hematoxylin-stained) and GFAP-positive cellsin dorsal hippocampus of maleadult Sprague Dawley rats

Subregion Control Dex-treatedHematoxylin stained cell numbers estimated per region (times103)

CA1 7068 plusmn 88 7141 plusmn 297CA2 959 plusmn 34 958 plusmn 2CA3 4913 plusmn 372 4907 plusmn 326DG 7914 plusmn 316 7891 plusmn 306

GFAP +ve cell numbers estimated per region (times103)CA1 2194 plusmn 6 2136 plusmn 91CA2 268 plusmn 11 258 plusmn 1CA3 1109 plusmn 61 1044 plusmn 69DG 201 plusmn 47 1953 plusmn 51The numbers of total and GFAP-positive cells are shown for control anddexamethasone-treated groups (119899 = 6 per group) Data are expressed asmean plusmn SEM No significant differences between Dex-treated (AGT) andcontrol were observed

Table 4 Effects of AGT on the number of total and GFAP-positivecells in basolateral amygdala and nucleus accumbens region of cellsin male adult Sprague Dawley rats

Subregion Control Dex-treatedHematoxylin stained cell numbers estimated per region (times103)

Basolateral amygdala 5286 plusmn 34 5879 plusmn 24Nucleus accumbens 14658 plusmn 68 13961 plusmn 64

GFAP +ve cell numbers estimated per region (times103)Basolateral amygdala 2368 plusmn 19 2571 plusmn 21Nucleus accumbens 2073 plusmn 18 1828 plusmn 20Thenumbers of total andGFAP-positive cells in control and dexamethasone-treated groups (119899 = 6 per group) Data are expressed as mean plusmn SEMNo significant differences between Dex-treated (AGT) and control wereobserved

Table 5 Effects of AGT on themorphology of GFAP-positive astro-glia in the basolateral amygdala and nucleus accumbens region ofmale adult Sprague Dawley rats

GFAP-astroglia morphologyControl Dex-treated

Total primary process length (120583m)Basolateral amygdala 57618 plusmn 5826 61235 plusmn 5989Nucleus accumbens 40334 plusmn 4039 38901 plusmn 4892

Primary process mean lengthBasolateral amygdala 268 plusmn 16 275 plusmn 21Nucleus accumbens 227 plusmn 18 208 plusmn 17

Number of primary processesBasolateral amygdala 213 plusmn 114 221 plusmn 91Nucleus accumbens 178 plusmn 85 185 plusmn 106

The total primary process length mean primary process length and num-bers of primary processes of GFAP-positive cells in control and dexametha-sone-treated groups (119899 = 6 per group) Data are expressed as mean plusmn SEMNo significant differences between Dex-treated (AGT) and control wereobserved

Neural Plasticity 5

Table 6 Effects of AGT on GFAP-positive astroglial primaryprocess numbers in male adult Sprague Dawley rats

GFAP-astroglia morphologyControl Dex-treated

Number of primary processCA1 362 plusmn 361 3233 plusmn 44CA2 301 plusmn 262 311 plusmn 264CA3 339 plusmn 169 3057 plusmn 81DG 3217 plusmn 143 3148 plusmn 47

The numbers of primary processes of GFAP-positive cells in control anddexamethasone-treated groups (119899= 6 per group) Data are expressed asmeanplusmn SEM No significant differences between Dex-treated (AGT) and controlwere observed

25 in DG (119875 lt 001) (Figure 2) AGT-dependent changeswere also observed in the mean length of astrocytic primaryprocesses reductions by 25 in CA1 (119875 lt 001) 45 in CA3(119875 lt 0001) and 25 in DG (119875 lt 001) (Figure 3)

4 Discussion

The present study was focused on the effects of AGT oncentral astroglia and regional volume of the hippocampusamygdala and nucleus accumbens as the areas of the limbicsystem implicated in response to stress and corticosteroids [23 9] AGTwas administered to pregnant dams noninvasivelythrough drinkingwater to avoid the stress of animal handlingand injections Of the brain areas tested in adult male rats thehippocampus responded to AGT with a long-term reductionin its volume and changes in astroglial morphology It is plau-sible that the observed impoverishment of astrocytic primarybranches underpins the hippocampal volume reduction aswe found no evidence of cell loss To our knowledge suchAGT-dependent long-term hippocampal remodelling withimplications for astroglia has not been reported before

Astroglia were visualised by means of the commonlyused GFAP immunohistochemistry GFAP is an intermediatefilament protein present in cell bodies and primary branchesalthough fine distal processes show little or no GFAP [25]Although GFAP staining is selective to astrocytes it does notlabel all astrocytes in the brain Accepting these limitationsof GFAP immunohistochemistry the present study focusedon the length and numbers of primary astrocytic processeswhere GFAP is reliably detectable [25] Stereology was ourmethod of choice as it offers an unbiased approach to regionalbrain volumetry and morphometry [24]

Previous volumetric studies have considered specificbrain regions expressing their volumes in absolute termswith no reference to the total brain volume which itself isa variable Thus the present application of pQCT to measuretotal brain volume adds the much needed rigour to volu-metric assessment where regional volumes need to be scaledto the total brain volume This is particularly important asperinatal corticosteroid exposure can lead to a decrease inbrain weightvolume For example a significant brain weightreduction has been reported in rhesus monkeys treatedprenatally with corticosteroids [26] and repeated doses of

lowast

lowastlowast

lowastlowastlowast

1200000

1100000

1000000

900000

800000

700000

600000

500000

400000

300000

200000

100000

000

CA1 CA2 CA3 DG

TreatmentControlDEX

Mea

n pr

imar

y pr

oces

s tot

al le

ngth

(120583m

)

Dorsal hippocampal subregions

Figure 2 Effect of prenatal dexamethasone treatment on the totallength of primary astrocytic processes in hippocampal subregionsof male adult Sprague Dawley rats The total primary process lengthofGFAP-positive cells in control and dexamethasone-treated groups(119899 = 6 per group) Data are expressed as mean plusmn SEM lowast119875 lt 005lowastlowast

119875 lt 001 and lowastlowastlowast119875 lt 0001 versus control independent sample119905-test

lowastlowast

lowastlowast

lowastlowastlowast

500

000

1000

1500

2000

2500

3000

CA1 CA2 CA3 DG

TreatmentControlDEX

Mea

n pr

imar

y pr

oces

s mea

n le

ngth

(120583m

)

Dorsal hippocampal subregions

Figure 3 Effect of prenatal dexamethasone treatment on the meanlength of primary astrocytic processes in hippocampal subregions ofmale adult Sprague Dawley rats The mean primary process lengthofGFAP-positive cells in control and dexamethasone-treated groups(119899 = 6 per group) Data are expressed as mean plusmn SEM lowastlowast119875 lt 001and lowastlowastlowast119875 lt 0001 versus control independent sample 119905-test

6 Neural Plasticity

dexamethasone at E17ndash19 have resulted in a significant brainweight reduction in newborn rats [27] The present studyfound no AGT effects on the whole brain weight and volumethe latter measured by pQCT One of the factors which affectthe brain weight and volume is the amount of corticosteroidadministered In the present study a relatively small dosewas given which as discussed previously [19] is in the rangeused in perinatal medicine and mimics the glucocorticoidactivity that is required for normal lung maturation duringlate gestation

Within the developing brain the limbic system richlyexpressing glucocorticoid and mineralocorticoid receptors[28] is sensitive to the endogenous and exogenous corti-costeroids which can affect these brain areas with long-term consequences for their structure and function [29]Reductions in the regional brain volume of the hippocampusand amygdala have been reported in preterm infants [30ndash32] and a cortical brain volume reduction of 35 has beenobserved in premature infants treated with dexamethasone[33] On the other hand there have been reports of no effectof single betamethasone antenatal dose on regional brainvolumes particularly within the limbic system [34]

In the present study the hippocampus was significantlyaffected by AGT in line with the theory that this regiondevelops primarily during the foetal period [35] althoughhippocampal volume losses have also been observed fol-lowing administration of corticosteroids in adult monkeysand rats [36 37] No changes were seen in the basolateralamygdala and nucleus accumbens volume which suggests aregion-selective treatment effect consistent with the densityof glucocorticoid receptor distribution [28]

A transient reduction in hippocampal volume has beenobserved in adolescent but not adult mice exposed to a clin-ically relevant dose of dexamethasone at E15 [18] The extentof hippocampal volume loss is normally mildmoderate Thepresent reduction of the hippocampal volume by 8 fits inthat range while being statistically significant It is compara-ble with some previous reports on stresssteroid-dependentlosses in the hippocampal volume across several speciesThusCoe et al [10] have found hippocampal volume reductions by12 and 10 in the offspring of rhesus monkeys exposed toearly life stress In tree shrews four weeksrsquo cortisol treatmentor psychosocial stress has resulted in a hippocampal volumeloss by 5ndash10 [38] Chronic social defeat stress has led toa similarly mild yet significant hippocampal volume lossin adult rats [6] Human adolescents who were born verypreterm have been found to have a hippocampal volume lossin the range of 12 to 156 [39]

What causes the hippocampal volume losses in responsesto stressglucocorticoids There is a range of factors such asneuronal andor glial cell death reduction of cell size includ-ing atrophy of neuronal dendrites andor glial branchesreduced water content or impaired neurogenesis [1] andorreduced microvasculature [6] The present study did not findcell losses in the brain areas tested and focused on astrogliabecause of their major role in maintaining brain plasticityat the level of synapses and cells [25] Currently published

studies on the mechanisms underlying glucocorticoid-induced brain volume reductions have focused on neuronsAs a result neuronal cell loss has been well researched anddocumented and various dexamethasone treatments havebeen found to lead to neuronal loss in rats [40 41] and pri-mates [26 37] mostly in the hippocampal CA3 and dentategyrus Less attention has been given to astroglial cells despitetheir role in the maintenance of brain homeostasis andplasticity [25]

The impoverishment in astroglial morphology reportedin the present study is consistent with the known glia-inhibitory effects of exogenous glucocorticoids or stress invitro [42 43] and in vivo [44] Treatment with corticosteroneor synthetic glucocorticoids such as dexamethasone has beenknown to inhibit GFAP expression in the neonatal andadult rat brain [45ndash48] thus suggesting that astrocytes canbe affected by glucocorticoid overload The very fact thatdexamethasone can affect GFAP expression raises a validmethodological consideration as to what extent the presentobservations of changes in the morphology of GFAP-positiveastroglia are affected bymethodological biaswhen comparingbetween the groups Central astroglia are heterogeneous[25 49] and using GFAP as the only immunohistochemicalmarker brings in an intrinsic limitation shared by many cur-rent studies as critically reviewed elsewhere [49] Howeverthe present GFAP responses to AGT appear to be selective tothe hippocampus where astroglial primary branches reducedin length but no GFAP-positive cell losses were found Sucha discrete response of GFAP immunoreactivity implies thatdexamethasone administered at the present lowdose asAGTdid not induce a global effect in GFAP expression Thisnot only highlights the unique susceptibility of hippocampalregions but also suggests that the changes observed here aretrue manifestations of impoverished hippocampal astroglialmorphology in response to AGT

It is increasingly evident that nongeneticenvironmentalfactors acting in early life result in long-term alterations ofthe physiological systems affected Perinatal programmingandor plasticity of the physiological system under extremeconditions such as ldquotoxicrdquo stress or AGT can result in long-term neurobehavioural abnormalities [3 29 50] and there isgrowing evidence that prenatal or early postnatal exposure tostressors affects the long-term brain functioning [51]

To conclude this study reports significant maladaptivechanges in astroglia in the hippocampal subregions CA1 CA3and dentate gyrus as a long-term effect of AGT It is plausibleto assume that the reductions in astrocytic primary branchescontribute to the losses in the hippocampal volume Theyare also likely to affect astroglial function and neuron-gliainteractions as retraction of primary astroglial processes candisadvantage local networks formed with glia and neuronsThe observed dexamethasone-induced impoverishment ofastroglial morphology can be treated as a demonstration ofglial maladaptive plasticity a phenomenon that deservesmore research in the context of effects of corticosteroidoverload and stress-related pathologies Epigenetics of thisphenomenon should be of interest considering the clinicaluse of dexamethasone

Neural Plasticity 7

Abbreviations

AGT Antenatal glucocorticoid treatmentDex DexamethasoneGD Gestational dayGFAP Glial fibrillary acidic proteinHX HematoxylinMDD Major depressive disorderPBS Phosphate-buffered salinePTSD Posttraumatic stress disorderROI Region of interest

Conflict of Interests

The authors declare that there is no conflict of interestsregarding the publication of this paper

Acknowledgments

This work was supported by the Wellcome Trust Grant (no086871Z08Z) to G E Gillies Vishvesh H Shende was aPhD student sponsored by the University of RoehamptonThe authors thank Dr Todd Rae (Roehampton) for trainingVishvesh H Shende in the pQCT technique

References

[1] B Czeh and P J Lucassen ldquoWhat causes the hippocampalvolume decrease in depression Are neurogenesis glial changesand apoptosis implicatedrdquo European Archives of Psychiatry andClinical Neuroscience vol 257 no 5 pp 250ndash260 2007

[2] P J Lucassen J Pruessner N Sousa et al ldquoNeuropathology ofstressrdquo Acta Neuropathologica vol 127 no 1 pp 109ndash135 2014

[3] B S McEwen L Eiland R G Hunter andMMMiller ldquoStressand anxiety structural plasticity and epigenetic regulation as aconsequence of stressrdquo Neuropharmacology vol 62 no 1 pp3ndash12 2012

[4] M A Mehta N I Golembo C Nosarti et al ldquoAmygdala hip-pocampal and corpus callosum size following severe earlyinstitutional deprivation the English and Romanian Adopteesstudy pilotrdquo Journal of Child Psychology and Psychiatry andAllied Disciplines vol 50 no 8 pp 943ndash951 2009

[5] T Frodl and V OrsquoKeane ldquoHow does the brain deal with cumu-lative stress A review with focus on developmental stress HPAaxis function and hippocampal structure in humansrdquo Neurobi-ology of Disease vol 52 pp 24ndash37 2013

[6] B Czeh N Abumaria R Rygula and E Fuchs ldquoQuantita-tive changes in hippocampal microvasculature of chronicallystressed rats no effect of fluoxetine treatmentrdquo Hippocampusvol 20 no 1 pp 174ndash185 2010

[7] V H Shende C R Pryce D Ruedi-Bettschen T C Rae andJ Opacka-Juffry ldquoEvidence for hippocampal remodeling as along term effect of early life stress in the rat model of earlydeprivationrdquo Recent Developments in Brain Research vol 1 pp1ndash19 2012

[8] A Jackowski T D Perera C G Abdallah et al ldquoEarly-lifestress corpus callosum development hippocampal volumet-rics and anxious behavior in male nonhuman primatesrdquo Psy-chiatry ResearchmdashNeuroimaging vol 192 no 1 pp 37ndash44 2011

[9] C Pittenger and R S Duman ldquoStress depression and neuro-plasticity a convergence of mechanismsrdquoNeuropsychopharma-cology vol 33 no 1 pp 88ndash109 2008

[10] C L Coe M Kramer B Czeh et al ldquoPrenatal stress diminishesneurogenesis in the dentate gyrus of juvenile Rhesus monkeysrdquoBiological Psychiatry vol 54 no 10 pp 1025ndash1034 2003

[11] J D Funkhouser K J Peevey P B Mockridge and E RHughes ldquoDistribution of dexamethasone between mother andfetus after maternal administrationrdquo Pediatric Research vol 12no 11 pp 1053ndash1056 1978

[12] D Roberts and S Dalziel ldquoAntenatal corticosteroids for accel-erating fetal lung maturation for women at risk of pretermbirthrdquo Cochrane Database of Systematic Reviews vol 3 ArticleID CD004454 2006

[13] T F Yeh Y J Lin H C Lin et al ldquoOutcomes at school ageafter postnatal dexamethasone therapy for lung disease of pre-maturityrdquoTheNew England Journal of Medicine vol 350 no 13pp 1304ndash1313 2004

[14] J P Empana M M Anceschi I Szabo E V Cosmi G Breartand P Truffert ldquoAntenatal corticosteroids policies in 14 Euro-pean countries factors associated with multiple courses TheEURAIL surveyrdquo Acta Paediatrica International Journal of Pae-diatrics vol 93 no 10 pp 1318ndash1322 2004

[15] C Romagnoli E Zecca R Luciano G Torrioli and G Tor-torolo ldquoA three year follow up of preterm infants after moder-ately early treatment with dexamethasonerdquo Archives of Diseasein Childhood Fetal and Neonatal Edition vol 87 no 1 pp F55ndashF58 2002

[16] A Charil D P Laplante C Vaillancourt and S King ldquoPrenatalstress and brain developmentrdquo Brain Research Reviews vol 65no 1 pp 56ndash79 2010

[17] D Tijsseling L D E Wijnberger J B Derks et al ldquoEffects ofantenatal glucocorticoid therapy on hippocampal histology ofpreterm infantsrdquoPLoSONE vol 7 no 3 Article ID e33369 2012

[18] C W Noorlander D Tijsseling E V S Hessel et al ldquoAntenatalglucocorticoid treatment affects hippocampal development inmicerdquo PLoS ONE vol 9 no 1 Article ID e85671 2014

[19] S McArthur E McHale and G E Gillies ldquoThe size and distri-bution of midbrain dopaminergic populations are permanentlyaltered by perinatal glucocorticoid exposure in a sex- region-and time-specific mannerrdquo Neuropsychopharmacology vol 32no 7 pp 1462ndash1476 2007

[20] S McArthur E McHale J W Dalley J C Buckingham andG E Gillies ldquoAlteredmesencephalic dopaminergic populationsin adulthood as a consequence of brief perinatal glucocorticoidexposurerdquo Journal of Neuroendocrinology vol 17 no 8 pp 475ndash482 2005

[21] ETheogaraj CD JohnH C Christian J FMorris S F Smithand J C Buckingham ldquoPerinatal glucocorticoid treatmentproduces molecular functional and morphological changes inthe anterior pituitary gland of the adultmale ratrdquoEndocrinologyvol 146 no 11 pp 4804ndash4813 2005

[22] G Paxinos and C WatsonThe Rat Brain in Stereotaxic Coordi-nates Elsevier Academic Press 5th edition 2005

[23] M Leventopoulos D Ruedi-Bettschen I Knuesel J Feldon CR Pryce and J Opacka-Juffry ldquoLong-term effects of early lifedeprivation on brain glia in Fischer ratsrdquo Brain Research vol1142 no 1 pp 119ndash126 2007

[24] M J West L Slomianka and H J G Gundersen ldquoUnbiasedstereological estimation of the total number of neurons in

8 Neural Plasticity

the subdivisions of the rat hippocampus using the optical frac-tionatorrdquo Anatomical Record vol 231 no 4 pp 482ndash497 1991

[25] D T Theodosis D A Poulain and S H R Oliet ldquoActivity-dependent structural and functional plasticity of astrocyte-neuron interactionsrdquo Physiological Reviews vol 88 no 3 pp983ndash1008 2008

[26] H Uno L Lohmiller C Thieme et al ldquoBrain damage inducedby prenatal exposure to dexamethasone in fetal rhesus maca-ques I Hippocampusrdquo Developmental Brain Research vol 53no 2 pp 157ndash167 1990

[27] R Q Carlos F J Seidler and T A Slotkin ldquoFetal dexametha-sone exposure altersmacromolecular characteristics of rat braindevelopment a critical period for regionally selective altera-tionsrdquo Teratology vol 46 no 1 pp 45ndash59 1992

[28] E R de Kloet ldquoStress in the brainrdquo European Journal of Phar-macology vol 405 pp 187ndash198 2000

[29] S G Matthews ldquoAntenatal glucocorticoids and the developingbrain mechanisms of actionrdquo Seminars in Neonatology vol 6no 4 pp 309ndash317 2001

[30] L J Abernethy M Palaniappan and R W I Cooke ldquoQuanti-tative magnetic resonance imaging of the brain in survivors ofvery low birth weightrdquo Archives of Disease in Childhood vol 87no 4 pp 279ndash283 2002

[31] E B Isaacs A LucasW K Chong et al ldquoHippocampal volumeand everyday memory in children of very low birth weightrdquoPediatric Research vol 47 no 6 pp 713ndash720 2000

[32] B S Peterson B Vohr L H Staib et al ldquoRegional brain volumeabnormalities and long-term cognitive outcome in preterminfantsrdquo Journal of the American Medical Association vol 284no 15 pp 1939ndash1947 2000

[33] B P Murphy T E Inder P S Huppi et al ldquoImpaired cerebralcortical graymatter growth after treatmentwith dexamethasonefor neonatal chronic lung diseaserdquo Pediatrics vol 107 no 2 pp217ndash221 2001

[34] P Yossuck M Kraszpulski and A K Salm ldquoPerinatal corti-costeroid effect on amygdala and hippocampus volume duringbrain development in the rat modelrdquo Early Human Develop-ment vol 82 no 4 pp 267ndash272 2006

[35] D Rice and S Barone Jr ldquoCritical periods of vulnerability forthe developing nervous system evidence from humans andanimal modelsrdquo Environmental Health Perspectives vol 108 no3 pp 511ndash533 2000

[36] C S Woolley E Gould and B S McEwen ldquoExposure toexcess glucocorticoids alters dendritic morphology of adulthippocampal pyramidal neuronsrdquo Brain Research vol 531 no1-2 pp 225ndash231 1990

[37] R M Sapolsky H Uno C S Rebert and C E Finch ldquoHippo-campal damage associatedwith prolonged glucocorticoid expo-sure in primatesrdquoThe Journal of Neuroscience vol 10 no 9 pp2897ndash2902 1990

[38] F Ohl T Michaelis G K Vollmann-Honsdorf C Kirschbaumand E Fuchs ldquoEffect of chronic psychosocial stress and long-term cortisol treatment on hippocampus-mediated memoryand hippocampal volume a pilot-study in tree shrewsrdquo Psy-choneuroendocrinology vol 25 no 4 pp 357ndash363 2000

[39] C Nosarti M H S Al-Asady S Frangou A L Stewart L Rif-kin and R M Murray ldquoAdolescents who were born very pre-term have decreased brain volumesrdquo Brain vol 125 no 7 pp1616ndash1623 2002

[40] L E Haynes M R Griffiths R E Hyde D J Barber and I JMitchell ldquoDexamethasone induces limited apoptosis and exten-sive sublethal damage to specific subregions of the striatum andhippocampus implications for mood disordersrdquo Neurosciencevol 104 no 1 pp 57ndash69 2001

[41] N Sousa M M Paula-Barbosa and O F X Almeida ldquoLigandand subfield specificity of corticoid-induced neuronal loss inthe rat hippocampal formationrdquoNeuroscience vol 89 no 4 pp1079ndash1087 1999

[42] K L Crossin M-H Tai L A Krushel V P Mauro and G MEdelman ldquoGlucocorticoid receptor pathways are involved in theinhibition of astrocyte proliferationrdquo Proceedings of the NationalAcademy of Sciences of the United States of America vol 94 no6 pp 2687ndash2692 1997

[43] M Sabolek A Herborg J Schwarz and A Storch ldquoDexam-ethasone blocks astroglial differentiation from neural precursorcellsrdquo NeuroReport vol 17 no 16 pp 1719ndash1723 2006

[44] N R Nichols J N Masters and C E Finch ldquoChanges in geneexpression in hippocampus in response to glucocorticoids andstressrdquo Brain Research Bulletin vol 24 no 5 pp 659ndash662 1990

[45] N R Nichols H H Osterburg J N Masters S L Millarand C E Finch ldquoMessenger RNA for glial fibrillary acidicprotein is decreased in rat brain following acute and chroniccorticosterone treatmentrdquo Molecular Brain Research vol 7 no1 pp 1ndash7 1990

[46] N J Laping N R Nichols J R Day and C E Finch ldquoCortico-sterone differentially regulates the bilateral response of astro-cyte mRNAs in the hippocampus to entorhinal cortex lesions inmale ratsrdquoMolecular Brain Research vol 10 no 4 pp 291ndash2971991

[47] S Tsuneishi S Takada T Motoike T Ohashi K Sano andH Nakamura ldquoEffects of dexamethasone on the expression ofmyelin basic protein proteolipid protein and glial fibrillaryacidic protein genes in developing rat brainrdquo DevelopmentalBrain Research vol 61 no 1 pp 117ndash123 1991

[48] J P OrsquoCallaghan R E Brinton and B S McEwen ldquoGlucocor-ticoids regulate the synthesis of glial fibrillary acidic protein inintact and adrenalectomized rats but do not affect its expressionfollowing brain injuryrdquo Journal of Neurochemistry vol 57 no 3pp 860ndash869 1991

[49] M V Sofroniew and H V Vinters ldquoAstrocytes biology andpathologyrdquoActa Neuropathologica vol 119 no 1 pp 7ndash35 2010

[50] S Maccari M Darnaudery S Morley-Fletcher A R Zuena CCinque and O van Reeth ldquoPrenatal stress and long-termconsequences implications of glucocorticoid hormonesrdquo Neu-roscience and Biobehavioral Reviews vol 27 no 1-2 pp 119ndash1272003

[51] C R Pryce Y Aubert CMaier P C Pearce and E Fuchs ldquoThedevelopmental impact of prenatal stress prenatal dexametha-sone and postnatal social stress on physiology behaviour andneuroanatomy of primate offspring Studies in rhesus macaqueand common marmosetrdquo Psychopharmacology vol 214 no 1pp 33ndash53 2011

Submit your manuscripts athttpwwwhindawicom

Neurology Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

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BioMed Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Research and TreatmentSchizophrenia

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Neural Plasticity

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Parkinsonrsquos Disease

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Research and TreatmentAutism

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Cardiovascular Psychiatry and NeurologyHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

2 Neural Plasticity

The first human study on postmortem hippocampi ofneonates who had been antenatally treated with dexametha-sone or betamethasone has shown a glucocorticoid-relatedreduced density of neurons no differences have been found ingliosis or myelination [17] Experimental studies on animalsconfirmnegative effects of antenatal glucocorticoid treatment(AGT) on hippocampal neurogenesis [18] however thereare no reports about AGT effects on brain glia The presentstudy addressed this gap in knowledge and hypothesised thatrat hippocampal astroglia respond to AGT in a maladaptivemanner in the long term

2 Experimental Procedures

21 Animals All animal procedures were carried out inaccordance with the United Kingdom Animals ScientificProcedures Act of 1986 at Imperial College London SpragueDawley rats (Harlan Olac Blackthorn Bicester OxfordshireUK) were kept under controlled lighting (on 0800ndash2000 h)temperature (21ndash23∘C) and humidity (63) with standardrat chow and drinking water (except as described below) pro-vided ad libitum Male and female rats were caged separatelyand allowed to acclimatize to their new environment for oneweek after which groups of one male and two female ratswere housed together overnight and the presence of vaginalplugs the following morning was taken to confirm matingpregnancy was confirmed approximately 6 days later bypalpationThe timed pregnant rats were housed five per cageuntil gestational day (GD) 15 when they were caged singlyin preparation for giving birth From GD 1920 pregnant ratswere monitored several times a day and the day of birthwas defined as day 0 No more than two progenies per litter(one male one female) were included in each experimentalgroup in order to minimize potential effects of litter-of-origin Offspring were weaned at three weeks male andfemale progenywere housed separately and theywere allowedto grow to adulthood with no further interventions otherthan normal husbandry At 68 plusmn 2 days of age animals weredecapitated between 0900 and 1000 h Male rats were used inthe present study

22 Dexamethasone Treatment Dexamethasone was admin-istered noninvasively as dexamethasone sodium phosphate(Faulding Pharmaceuticals Plc Royal Leamington Spa UK)in the drinking water given to pregnant rats at GD 16ndash19 atthe dose of 05 120583gmL [19] with an estimated daily intake ofapproximately 75120583gkg [20]Withdrawing dexamethasone atGD 19 allowed for clearance of the steroid from the maternalcirculation prior to birth with no observable effects on theoutcomes of pregnancy maternal behaviour or adult bodyweight of the offspring [19ndash21]

23 pQCT Scanning Total frozen brain volume (119899 = 6 pergroup) was analysed by means of peripheral quantitativecomputed tomography (pQCT) on a Stratec Research SA+scanner (Stratec Medizintechnik Pforzheim Germany) asdescribed elsewhere [7] Serial coronal CT scans were per-formed covering entire brain region Further analysis tomeasure total brain volume was carried out by using software

Avizo (version 5 Mercury Computer Systems ChelmsfordMA USA)

24 Brain Tissue Preparation Frozen coronal sections(25 120583m) were cut in an anterior-posterior direction Thesectioning procedure was designed to yield the followingregions of interest (ROIs) (in brackets AP coordinates frombregma in mm) nucleus accumbens core (300 to 216)basolateral amygdala (minus172 to minus228) and dorsal hippo-campus (minus204 to minus468) according to the rat brain atlas byPaxinos and Watson [22]

25 Total Cell Count Brain sections were stained with hema-toxylin (HX) for total cell count and then gradually dehy-drated air-dried and protected with a histological mountingmedium and covered with cover slips

26 Immunohistochemistry To visualise astrocytes glial fib-rillary acidic protein (GFAP) immunohistochemistry wasperformed [23] Briefly peroxidase inactivation was carriedout in PBS with 20 methanol 15 H

2O2 and 03

Triton X-100 blocking medium contained 01 Triton X-100 and 10 normal horse serum and the anti-GFAP pri-mary antibody (Sigma UK) was diluted 1 500 and appliedovernight at 4∘C Controls (blanks) without the primaryantibody were processed in parallel All sections were incu-bated with the secondary antibody and streptavidin-biotin-peroxidase complex according to manufacturerrsquos instruc-tion (ABC kit Vector Laboratories UK) Sections weretreated with 331015840-diaminobenzidine (5mgmL 001 H

2O2)

rinsed dehydrated and mounted in xylene-based Histo-mount (BDHMerck Poole UK) prior to stereology

27 Stereological Procedures

271 Volume Analysis Volume estimation using the Cava-lieri principle was performed with Stereo-Investigator soft-ware (Version 9 MicroBrightField Inc Germany) andOlympus BX51 microscope fitted with a motorized stage andvideo camera on a Zeiss UPlan FLN with a 4x objective(NA = 025) Starting at a random position every tenthsection was analyzed leading to an average of 12 sections fordorsal hippocampus 6 sections for basolateral amygdala and5 sections for nucleus accumbens per brain These sets ofconsecutive sections were used for the estimation of volumesfor both control and test groups The investigator was blindto the experimental groups A point counting grid (PCG)(ie 119889 = 50 120583m) was used for volume estimation to obtainmaximum efficiency Representative area per point (119886119901)was2500 120583m2 After applying the PCG on the sampled sectionsin a systematic randommanner the number of points hittingregion of interest was counted (Figure 1(a)) The efficiency ofsampling and volume estimation were checked by estimationof coefficient of error and coefficient of variation [24]

272 Cell Counts Cells in the dorsal hippocampal subre-gions (CA1 CA2 CA3 and DG) basolateral amygdala andnucleus accumbens were counted using the optical fraction-ator (Stereo-Investigator Version 9 MicroBrightField Inc

Neural Plasticity 3

(a) (b)

Control

(c)

Dex-treated

(d)

Figure 1 Dorsal hippocampal volume HX stain (a) and GFAP-positive astroglial cell count (b) estimated by means of stereology Changesin astroglial morphology in response to AGT GFAP-positive astroglial cells in control (c) and Dex-treated (d) male rats (dorsal hippocampaldentate gyrus area) GFAP-positive primary process length is reduced in Dex-treated male rats when compared with control (d) versus (c)Astroglial processes observed at 100x magnification

Germany) An unbiased estimation of total number of cellswas achieved by choosing every 10th section according to thesystematic random sampling procedure [24] yielding 8 sec-tions per animal A pilot study was undertaken to determinethe following parameters A sampling area of 1602500120583m2was found to be optimal for this study Dissector height was8 120583m and a 1 120583m guard zone at the top and bottom part of thesection was excluded from the analysis at every step A fixedcounting frame of 50 times 50 and a variable sampling grid size(119909-119910-axis) of 150 times 150 CA1 150 times 150 CA2 300 times 300 CA3and 300 times 300 DG were used for the dorsal hippocampus(Figure 1(b)) and 150 times 150 was used for both basolateralamygdala and nucleus accumbens These sampling gridsresulted in 100ndash200 counting sites per brain HX and GFAP-positive cells were counted using a 100x Nicon UPlan FLNobjective (NA = 130) which allowed accurate recognitionEach cell was counted according to the unbiased countingrules The efficiency of convenient number of sampled cellsand parameters was checked by estimation of coefficient oferror (CE) and coefficient of variation (CV) [24]

273 AstroglialMorphology Morphological analysiswas per-formed on 80 astrocytes in each brain (10 astrocytes persection) usingNeurolucida software (Version 9MicroBright-Field Inc Germany) for tracing boundaries of the ROIsand astrocytes Astrocytes within the ROI were traced in

a systematic random manner taking care to avoid thosewhich were superimposed upon other astrocytes or bloodvessels The traced astrocytes were then analysed usingNeurolucida explorer software (Version 9 MicroBrightFieldInc Germany) Morphological analysis was performed byobserver who was blind to the experimental groups Primaryprocess length was measured using 100x objective (NiconUPlan FLN NA = 130) and by counting the primary pro-cesses extending directly from the soma in both the lateraland central quadrants of astrocytes in the same sections

28 Statistical Analysis Using SPSS 170 statistical software(SPSS Inc USA) ANOVA with a post hoc Tukey test wasperformed comparing between group differences Indepen-dent 119905-test was performed tomeasure difference between twogroups Statistical significance was set at 119875 lt 005The resultsare expressed as mean plusmn SEM

3 Results

31 Whole Brain Volume and Brain Weight No significanteffects of AGT were observed in the whole brain volume(119875 = 0376) and brain weight (119875 = 0442) (Table 1)

32 VolumeAnalyses AGT rats showed a significant decreasein the absolute volume of dorsal hippocampus by 65

4 Neural Plasticity

Table 1 Effects of AGT exposure on brain weight and volume ofmale adult Sprague Dawley rats

Control Dex-treatedBrain weight (g) 174 plusmn 002 177 plusmn 003Brain volume (mm3) 165018 plusmn 1775 168124 plusmn 2843The brain weight and volume are expressed as mean plusmn SEM (119899 = 6 pergroup) No significant differences between Dex-treated (AGT) and controlwere observed

Table 2 Effects of AGT on the regional brain volume of male adultSprague Dawley rats

VolumeControl Dex-treated

Absolute values (mm3)Dorsal hippocampus 328 plusmn 046 308 plusmn 050lowast

Basolateral amygdala 081 plusmn 002 080 plusmn 002Nucleus accumbens 228 plusmn 006 226 plusmn 007

Relative values (as brain volume)Dorsal hippocampus 199 plusmn 004 183 plusmn 004lowast

Basolateral amygdala 0046 plusmn 0001 0048 plusmn 0001Nucleus accumbens 0138 plusmn 0004 0135 plusmn 0005

The volume of the dorsal hippocampus basolateral amygdala and nucleusin control and dexamethasone-treated groups (119899 = 6 per group) Data areexpressed as mean plusmn SEM lowast119875 lt 005 versus control independent sample 119905-test

versus control (119875 lt 005) When expressed as percentageof the whole brain volume the volume reduction was 8(119875 lt 005) versus control No significant differences wereobserved between the treatment groups in the basolateralamygdala (119875 = 0834) and nucleus accumbens (119875 = 0905)(Table 2)

33 HX Stained Total Cell Numbers AGT did not affect thetotal cell count (HX stain) in the dorsal hippocampal subre-gions CA1 (119875 = 0821) CA2 (119875 = 0975) CA3 (119875 = 0991)and DG (119875 = 0959) basolateral amygdala (119875 = 0180) andthe nucleus accumbens (119875 = 0474) (Table 3)

34 GFAP-Positive Astroglial Cell Numbers There were nosignificant AGT effects on GFAP-positive cell numbers in thedorsal hippocampal subregions CA1 (119875 = 0607) CA2 (119875 =0517) CA3 (119875 = 0500) and DG (119875 = 0424) basolateralamygdala (119875 = 0495) and the nucleus accumbens (119875 =0384) (Table 4)

35 Astroglia (GFAP-Positive) Cell Morphology AGT didnot affect the numbers of primary processes in the regionanalysed (Tables 5 and 6) and it had no significant effectson the astroglial morphology in the nonhippocampal regions(Table 5 all group differences at 119875 lt 005)

However AGT affected the length of primary processesin GFAP-positive cells within the hippocampus (Figures 1(a)and 1(b)) it led to a significant reduction in the total length ofastrocytic primary processes in the hippocampal subregionsby 32 in CA1 (119875 lt 005) 50 in CA3 (119875 lt 0001) and by

Table 3 Effects of AGT on the number of total (hematoxylin-stained) and GFAP-positive cellsin dorsal hippocampus of maleadult Sprague Dawley rats

Subregion Control Dex-treatedHematoxylin stained cell numbers estimated per region (times103)

CA1 7068 plusmn 88 7141 plusmn 297CA2 959 plusmn 34 958 plusmn 2CA3 4913 plusmn 372 4907 plusmn 326DG 7914 plusmn 316 7891 plusmn 306

GFAP +ve cell numbers estimated per region (times103)CA1 2194 plusmn 6 2136 plusmn 91CA2 268 plusmn 11 258 plusmn 1CA3 1109 plusmn 61 1044 plusmn 69DG 201 plusmn 47 1953 plusmn 51The numbers of total and GFAP-positive cells are shown for control anddexamethasone-treated groups (119899 = 6 per group) Data are expressed asmean plusmn SEM No significant differences between Dex-treated (AGT) andcontrol were observed

Table 4 Effects of AGT on the number of total and GFAP-positivecells in basolateral amygdala and nucleus accumbens region of cellsin male adult Sprague Dawley rats

Subregion Control Dex-treatedHematoxylin stained cell numbers estimated per region (times103)

Basolateral amygdala 5286 plusmn 34 5879 plusmn 24Nucleus accumbens 14658 plusmn 68 13961 plusmn 64

GFAP +ve cell numbers estimated per region (times103)Basolateral amygdala 2368 plusmn 19 2571 plusmn 21Nucleus accumbens 2073 plusmn 18 1828 plusmn 20Thenumbers of total andGFAP-positive cells in control and dexamethasone-treated groups (119899 = 6 per group) Data are expressed as mean plusmn SEMNo significant differences between Dex-treated (AGT) and control wereobserved

Table 5 Effects of AGT on themorphology of GFAP-positive astro-glia in the basolateral amygdala and nucleus accumbens region ofmale adult Sprague Dawley rats

GFAP-astroglia morphologyControl Dex-treated

Total primary process length (120583m)Basolateral amygdala 57618 plusmn 5826 61235 plusmn 5989Nucleus accumbens 40334 plusmn 4039 38901 plusmn 4892

Primary process mean lengthBasolateral amygdala 268 plusmn 16 275 plusmn 21Nucleus accumbens 227 plusmn 18 208 plusmn 17

Number of primary processesBasolateral amygdala 213 plusmn 114 221 plusmn 91Nucleus accumbens 178 plusmn 85 185 plusmn 106

The total primary process length mean primary process length and num-bers of primary processes of GFAP-positive cells in control and dexametha-sone-treated groups (119899 = 6 per group) Data are expressed as mean plusmn SEMNo significant differences between Dex-treated (AGT) and control wereobserved

Neural Plasticity 5

Table 6 Effects of AGT on GFAP-positive astroglial primaryprocess numbers in male adult Sprague Dawley rats

GFAP-astroglia morphologyControl Dex-treated

Number of primary processCA1 362 plusmn 361 3233 plusmn 44CA2 301 plusmn 262 311 plusmn 264CA3 339 plusmn 169 3057 plusmn 81DG 3217 plusmn 143 3148 plusmn 47

The numbers of primary processes of GFAP-positive cells in control anddexamethasone-treated groups (119899= 6 per group) Data are expressed asmeanplusmn SEM No significant differences between Dex-treated (AGT) and controlwere observed

25 in DG (119875 lt 001) (Figure 2) AGT-dependent changeswere also observed in the mean length of astrocytic primaryprocesses reductions by 25 in CA1 (119875 lt 001) 45 in CA3(119875 lt 0001) and 25 in DG (119875 lt 001) (Figure 3)

4 Discussion

The present study was focused on the effects of AGT oncentral astroglia and regional volume of the hippocampusamygdala and nucleus accumbens as the areas of the limbicsystem implicated in response to stress and corticosteroids [23 9] AGTwas administered to pregnant dams noninvasivelythrough drinkingwater to avoid the stress of animal handlingand injections Of the brain areas tested in adult male rats thehippocampus responded to AGT with a long-term reductionin its volume and changes in astroglial morphology It is plau-sible that the observed impoverishment of astrocytic primarybranches underpins the hippocampal volume reduction aswe found no evidence of cell loss To our knowledge suchAGT-dependent long-term hippocampal remodelling withimplications for astroglia has not been reported before

Astroglia were visualised by means of the commonlyused GFAP immunohistochemistry GFAP is an intermediatefilament protein present in cell bodies and primary branchesalthough fine distal processes show little or no GFAP [25]Although GFAP staining is selective to astrocytes it does notlabel all astrocytes in the brain Accepting these limitationsof GFAP immunohistochemistry the present study focusedon the length and numbers of primary astrocytic processeswhere GFAP is reliably detectable [25] Stereology was ourmethod of choice as it offers an unbiased approach to regionalbrain volumetry and morphometry [24]

Previous volumetric studies have considered specificbrain regions expressing their volumes in absolute termswith no reference to the total brain volume which itself isa variable Thus the present application of pQCT to measuretotal brain volume adds the much needed rigour to volu-metric assessment where regional volumes need to be scaledto the total brain volume This is particularly important asperinatal corticosteroid exposure can lead to a decrease inbrain weightvolume For example a significant brain weightreduction has been reported in rhesus monkeys treatedprenatally with corticosteroids [26] and repeated doses of

lowast

lowastlowast

lowastlowastlowast

1200000

1100000

1000000

900000

800000

700000

600000

500000

400000

300000

200000

100000

000

CA1 CA2 CA3 DG

TreatmentControlDEX

Mea

n pr

imar

y pr

oces

s tot

al le

ngth

(120583m

)

Dorsal hippocampal subregions

Figure 2 Effect of prenatal dexamethasone treatment on the totallength of primary astrocytic processes in hippocampal subregionsof male adult Sprague Dawley rats The total primary process lengthofGFAP-positive cells in control and dexamethasone-treated groups(119899 = 6 per group) Data are expressed as mean plusmn SEM lowast119875 lt 005lowastlowast

119875 lt 001 and lowastlowastlowast119875 lt 0001 versus control independent sample119905-test

lowastlowast

lowastlowast

lowastlowastlowast

500

000

1000

1500

2000

2500

3000

CA1 CA2 CA3 DG

TreatmentControlDEX

Mea

n pr

imar

y pr

oces

s mea

n le

ngth

(120583m

)

Dorsal hippocampal subregions

Figure 3 Effect of prenatal dexamethasone treatment on the meanlength of primary astrocytic processes in hippocampal subregions ofmale adult Sprague Dawley rats The mean primary process lengthofGFAP-positive cells in control and dexamethasone-treated groups(119899 = 6 per group) Data are expressed as mean plusmn SEM lowastlowast119875 lt 001and lowastlowastlowast119875 lt 0001 versus control independent sample 119905-test

6 Neural Plasticity

dexamethasone at E17ndash19 have resulted in a significant brainweight reduction in newborn rats [27] The present studyfound no AGT effects on the whole brain weight and volumethe latter measured by pQCT One of the factors which affectthe brain weight and volume is the amount of corticosteroidadministered In the present study a relatively small dosewas given which as discussed previously [19] is in the rangeused in perinatal medicine and mimics the glucocorticoidactivity that is required for normal lung maturation duringlate gestation

Within the developing brain the limbic system richlyexpressing glucocorticoid and mineralocorticoid receptors[28] is sensitive to the endogenous and exogenous corti-costeroids which can affect these brain areas with long-term consequences for their structure and function [29]Reductions in the regional brain volume of the hippocampusand amygdala have been reported in preterm infants [30ndash32] and a cortical brain volume reduction of 35 has beenobserved in premature infants treated with dexamethasone[33] On the other hand there have been reports of no effectof single betamethasone antenatal dose on regional brainvolumes particularly within the limbic system [34]

In the present study the hippocampus was significantlyaffected by AGT in line with the theory that this regiondevelops primarily during the foetal period [35] althoughhippocampal volume losses have also been observed fol-lowing administration of corticosteroids in adult monkeysand rats [36 37] No changes were seen in the basolateralamygdala and nucleus accumbens volume which suggests aregion-selective treatment effect consistent with the densityof glucocorticoid receptor distribution [28]

A transient reduction in hippocampal volume has beenobserved in adolescent but not adult mice exposed to a clin-ically relevant dose of dexamethasone at E15 [18] The extentof hippocampal volume loss is normally mildmoderate Thepresent reduction of the hippocampal volume by 8 fits inthat range while being statistically significant It is compara-ble with some previous reports on stresssteroid-dependentlosses in the hippocampal volume across several speciesThusCoe et al [10] have found hippocampal volume reductions by12 and 10 in the offspring of rhesus monkeys exposed toearly life stress In tree shrews four weeksrsquo cortisol treatmentor psychosocial stress has resulted in a hippocampal volumeloss by 5ndash10 [38] Chronic social defeat stress has led toa similarly mild yet significant hippocampal volume lossin adult rats [6] Human adolescents who were born verypreterm have been found to have a hippocampal volume lossin the range of 12 to 156 [39]

What causes the hippocampal volume losses in responsesto stressglucocorticoids There is a range of factors such asneuronal andor glial cell death reduction of cell size includ-ing atrophy of neuronal dendrites andor glial branchesreduced water content or impaired neurogenesis [1] andorreduced microvasculature [6] The present study did not findcell losses in the brain areas tested and focused on astrogliabecause of their major role in maintaining brain plasticityat the level of synapses and cells [25] Currently published

studies on the mechanisms underlying glucocorticoid-induced brain volume reductions have focused on neuronsAs a result neuronal cell loss has been well researched anddocumented and various dexamethasone treatments havebeen found to lead to neuronal loss in rats [40 41] and pri-mates [26 37] mostly in the hippocampal CA3 and dentategyrus Less attention has been given to astroglial cells despitetheir role in the maintenance of brain homeostasis andplasticity [25]

The impoverishment in astroglial morphology reportedin the present study is consistent with the known glia-inhibitory effects of exogenous glucocorticoids or stress invitro [42 43] and in vivo [44] Treatment with corticosteroneor synthetic glucocorticoids such as dexamethasone has beenknown to inhibit GFAP expression in the neonatal andadult rat brain [45ndash48] thus suggesting that astrocytes canbe affected by glucocorticoid overload The very fact thatdexamethasone can affect GFAP expression raises a validmethodological consideration as to what extent the presentobservations of changes in the morphology of GFAP-positiveastroglia are affected bymethodological biaswhen comparingbetween the groups Central astroglia are heterogeneous[25 49] and using GFAP as the only immunohistochemicalmarker brings in an intrinsic limitation shared by many cur-rent studies as critically reviewed elsewhere [49] Howeverthe present GFAP responses to AGT appear to be selective tothe hippocampus where astroglial primary branches reducedin length but no GFAP-positive cell losses were found Sucha discrete response of GFAP immunoreactivity implies thatdexamethasone administered at the present lowdose asAGTdid not induce a global effect in GFAP expression Thisnot only highlights the unique susceptibility of hippocampalregions but also suggests that the changes observed here aretrue manifestations of impoverished hippocampal astroglialmorphology in response to AGT

It is increasingly evident that nongeneticenvironmentalfactors acting in early life result in long-term alterations ofthe physiological systems affected Perinatal programmingandor plasticity of the physiological system under extremeconditions such as ldquotoxicrdquo stress or AGT can result in long-term neurobehavioural abnormalities [3 29 50] and there isgrowing evidence that prenatal or early postnatal exposure tostressors affects the long-term brain functioning [51]

To conclude this study reports significant maladaptivechanges in astroglia in the hippocampal subregions CA1 CA3and dentate gyrus as a long-term effect of AGT It is plausibleto assume that the reductions in astrocytic primary branchescontribute to the losses in the hippocampal volume Theyare also likely to affect astroglial function and neuron-gliainteractions as retraction of primary astroglial processes candisadvantage local networks formed with glia and neuronsThe observed dexamethasone-induced impoverishment ofastroglial morphology can be treated as a demonstration ofglial maladaptive plasticity a phenomenon that deservesmore research in the context of effects of corticosteroidoverload and stress-related pathologies Epigenetics of thisphenomenon should be of interest considering the clinicaluse of dexamethasone

Neural Plasticity 7

Abbreviations

AGT Antenatal glucocorticoid treatmentDex DexamethasoneGD Gestational dayGFAP Glial fibrillary acidic proteinHX HematoxylinMDD Major depressive disorderPBS Phosphate-buffered salinePTSD Posttraumatic stress disorderROI Region of interest

Conflict of Interests

The authors declare that there is no conflict of interestsregarding the publication of this paper

Acknowledgments

This work was supported by the Wellcome Trust Grant (no086871Z08Z) to G E Gillies Vishvesh H Shende was aPhD student sponsored by the University of RoehamptonThe authors thank Dr Todd Rae (Roehampton) for trainingVishvesh H Shende in the pQCT technique

References

[1] B Czeh and P J Lucassen ldquoWhat causes the hippocampalvolume decrease in depression Are neurogenesis glial changesand apoptosis implicatedrdquo European Archives of Psychiatry andClinical Neuroscience vol 257 no 5 pp 250ndash260 2007

[2] P J Lucassen J Pruessner N Sousa et al ldquoNeuropathology ofstressrdquo Acta Neuropathologica vol 127 no 1 pp 109ndash135 2014

[3] B S McEwen L Eiland R G Hunter andMMMiller ldquoStressand anxiety structural plasticity and epigenetic regulation as aconsequence of stressrdquo Neuropharmacology vol 62 no 1 pp3ndash12 2012

[4] M A Mehta N I Golembo C Nosarti et al ldquoAmygdala hip-pocampal and corpus callosum size following severe earlyinstitutional deprivation the English and Romanian Adopteesstudy pilotrdquo Journal of Child Psychology and Psychiatry andAllied Disciplines vol 50 no 8 pp 943ndash951 2009

[5] T Frodl and V OrsquoKeane ldquoHow does the brain deal with cumu-lative stress A review with focus on developmental stress HPAaxis function and hippocampal structure in humansrdquo Neurobi-ology of Disease vol 52 pp 24ndash37 2013

[6] B Czeh N Abumaria R Rygula and E Fuchs ldquoQuantita-tive changes in hippocampal microvasculature of chronicallystressed rats no effect of fluoxetine treatmentrdquo Hippocampusvol 20 no 1 pp 174ndash185 2010

[7] V H Shende C R Pryce D Ruedi-Bettschen T C Rae andJ Opacka-Juffry ldquoEvidence for hippocampal remodeling as along term effect of early life stress in the rat model of earlydeprivationrdquo Recent Developments in Brain Research vol 1 pp1ndash19 2012

[8] A Jackowski T D Perera C G Abdallah et al ldquoEarly-lifestress corpus callosum development hippocampal volumet-rics and anxious behavior in male nonhuman primatesrdquo Psy-chiatry ResearchmdashNeuroimaging vol 192 no 1 pp 37ndash44 2011

[9] C Pittenger and R S Duman ldquoStress depression and neuro-plasticity a convergence of mechanismsrdquoNeuropsychopharma-cology vol 33 no 1 pp 88ndash109 2008

[10] C L Coe M Kramer B Czeh et al ldquoPrenatal stress diminishesneurogenesis in the dentate gyrus of juvenile Rhesus monkeysrdquoBiological Psychiatry vol 54 no 10 pp 1025ndash1034 2003

[11] J D Funkhouser K J Peevey P B Mockridge and E RHughes ldquoDistribution of dexamethasone between mother andfetus after maternal administrationrdquo Pediatric Research vol 12no 11 pp 1053ndash1056 1978

[12] D Roberts and S Dalziel ldquoAntenatal corticosteroids for accel-erating fetal lung maturation for women at risk of pretermbirthrdquo Cochrane Database of Systematic Reviews vol 3 ArticleID CD004454 2006

[13] T F Yeh Y J Lin H C Lin et al ldquoOutcomes at school ageafter postnatal dexamethasone therapy for lung disease of pre-maturityrdquoTheNew England Journal of Medicine vol 350 no 13pp 1304ndash1313 2004

[14] J P Empana M M Anceschi I Szabo E V Cosmi G Breartand P Truffert ldquoAntenatal corticosteroids policies in 14 Euro-pean countries factors associated with multiple courses TheEURAIL surveyrdquo Acta Paediatrica International Journal of Pae-diatrics vol 93 no 10 pp 1318ndash1322 2004

[15] C Romagnoli E Zecca R Luciano G Torrioli and G Tor-torolo ldquoA three year follow up of preterm infants after moder-ately early treatment with dexamethasonerdquo Archives of Diseasein Childhood Fetal and Neonatal Edition vol 87 no 1 pp F55ndashF58 2002

[16] A Charil D P Laplante C Vaillancourt and S King ldquoPrenatalstress and brain developmentrdquo Brain Research Reviews vol 65no 1 pp 56ndash79 2010

[17] D Tijsseling L D E Wijnberger J B Derks et al ldquoEffects ofantenatal glucocorticoid therapy on hippocampal histology ofpreterm infantsrdquoPLoSONE vol 7 no 3 Article ID e33369 2012

[18] C W Noorlander D Tijsseling E V S Hessel et al ldquoAntenatalglucocorticoid treatment affects hippocampal development inmicerdquo PLoS ONE vol 9 no 1 Article ID e85671 2014

[19] S McArthur E McHale and G E Gillies ldquoThe size and distri-bution of midbrain dopaminergic populations are permanentlyaltered by perinatal glucocorticoid exposure in a sex- region-and time-specific mannerrdquo Neuropsychopharmacology vol 32no 7 pp 1462ndash1476 2007

[20] S McArthur E McHale J W Dalley J C Buckingham andG E Gillies ldquoAlteredmesencephalic dopaminergic populationsin adulthood as a consequence of brief perinatal glucocorticoidexposurerdquo Journal of Neuroendocrinology vol 17 no 8 pp 475ndash482 2005

[21] ETheogaraj CD JohnH C Christian J FMorris S F Smithand J C Buckingham ldquoPerinatal glucocorticoid treatmentproduces molecular functional and morphological changes inthe anterior pituitary gland of the adultmale ratrdquoEndocrinologyvol 146 no 11 pp 4804ndash4813 2005

[22] G Paxinos and C WatsonThe Rat Brain in Stereotaxic Coordi-nates Elsevier Academic Press 5th edition 2005

[23] M Leventopoulos D Ruedi-Bettschen I Knuesel J Feldon CR Pryce and J Opacka-Juffry ldquoLong-term effects of early lifedeprivation on brain glia in Fischer ratsrdquo Brain Research vol1142 no 1 pp 119ndash126 2007

[24] M J West L Slomianka and H J G Gundersen ldquoUnbiasedstereological estimation of the total number of neurons in

8 Neural Plasticity

the subdivisions of the rat hippocampus using the optical frac-tionatorrdquo Anatomical Record vol 231 no 4 pp 482ndash497 1991

[25] D T Theodosis D A Poulain and S H R Oliet ldquoActivity-dependent structural and functional plasticity of astrocyte-neuron interactionsrdquo Physiological Reviews vol 88 no 3 pp983ndash1008 2008

[26] H Uno L Lohmiller C Thieme et al ldquoBrain damage inducedby prenatal exposure to dexamethasone in fetal rhesus maca-ques I Hippocampusrdquo Developmental Brain Research vol 53no 2 pp 157ndash167 1990

[27] R Q Carlos F J Seidler and T A Slotkin ldquoFetal dexametha-sone exposure altersmacromolecular characteristics of rat braindevelopment a critical period for regionally selective altera-tionsrdquo Teratology vol 46 no 1 pp 45ndash59 1992

[28] E R de Kloet ldquoStress in the brainrdquo European Journal of Phar-macology vol 405 pp 187ndash198 2000

[29] S G Matthews ldquoAntenatal glucocorticoids and the developingbrain mechanisms of actionrdquo Seminars in Neonatology vol 6no 4 pp 309ndash317 2001

[30] L J Abernethy M Palaniappan and R W I Cooke ldquoQuanti-tative magnetic resonance imaging of the brain in survivors ofvery low birth weightrdquo Archives of Disease in Childhood vol 87no 4 pp 279ndash283 2002

[31] E B Isaacs A LucasW K Chong et al ldquoHippocampal volumeand everyday memory in children of very low birth weightrdquoPediatric Research vol 47 no 6 pp 713ndash720 2000

[32] B S Peterson B Vohr L H Staib et al ldquoRegional brain volumeabnormalities and long-term cognitive outcome in preterminfantsrdquo Journal of the American Medical Association vol 284no 15 pp 1939ndash1947 2000

[33] B P Murphy T E Inder P S Huppi et al ldquoImpaired cerebralcortical graymatter growth after treatmentwith dexamethasonefor neonatal chronic lung diseaserdquo Pediatrics vol 107 no 2 pp217ndash221 2001

[34] P Yossuck M Kraszpulski and A K Salm ldquoPerinatal corti-costeroid effect on amygdala and hippocampus volume duringbrain development in the rat modelrdquo Early Human Develop-ment vol 82 no 4 pp 267ndash272 2006

[35] D Rice and S Barone Jr ldquoCritical periods of vulnerability forthe developing nervous system evidence from humans andanimal modelsrdquo Environmental Health Perspectives vol 108 no3 pp 511ndash533 2000

[36] C S Woolley E Gould and B S McEwen ldquoExposure toexcess glucocorticoids alters dendritic morphology of adulthippocampal pyramidal neuronsrdquo Brain Research vol 531 no1-2 pp 225ndash231 1990

[37] R M Sapolsky H Uno C S Rebert and C E Finch ldquoHippo-campal damage associatedwith prolonged glucocorticoid expo-sure in primatesrdquoThe Journal of Neuroscience vol 10 no 9 pp2897ndash2902 1990

[38] F Ohl T Michaelis G K Vollmann-Honsdorf C Kirschbaumand E Fuchs ldquoEffect of chronic psychosocial stress and long-term cortisol treatment on hippocampus-mediated memoryand hippocampal volume a pilot-study in tree shrewsrdquo Psy-choneuroendocrinology vol 25 no 4 pp 357ndash363 2000

[39] C Nosarti M H S Al-Asady S Frangou A L Stewart L Rif-kin and R M Murray ldquoAdolescents who were born very pre-term have decreased brain volumesrdquo Brain vol 125 no 7 pp1616ndash1623 2002

[40] L E Haynes M R Griffiths R E Hyde D J Barber and I JMitchell ldquoDexamethasone induces limited apoptosis and exten-sive sublethal damage to specific subregions of the striatum andhippocampus implications for mood disordersrdquo Neurosciencevol 104 no 1 pp 57ndash69 2001

[41] N Sousa M M Paula-Barbosa and O F X Almeida ldquoLigandand subfield specificity of corticoid-induced neuronal loss inthe rat hippocampal formationrdquoNeuroscience vol 89 no 4 pp1079ndash1087 1999

[42] K L Crossin M-H Tai L A Krushel V P Mauro and G MEdelman ldquoGlucocorticoid receptor pathways are involved in theinhibition of astrocyte proliferationrdquo Proceedings of the NationalAcademy of Sciences of the United States of America vol 94 no6 pp 2687ndash2692 1997

[43] M Sabolek A Herborg J Schwarz and A Storch ldquoDexam-ethasone blocks astroglial differentiation from neural precursorcellsrdquo NeuroReport vol 17 no 16 pp 1719ndash1723 2006

[44] N R Nichols J N Masters and C E Finch ldquoChanges in geneexpression in hippocampus in response to glucocorticoids andstressrdquo Brain Research Bulletin vol 24 no 5 pp 659ndash662 1990

[45] N R Nichols H H Osterburg J N Masters S L Millarand C E Finch ldquoMessenger RNA for glial fibrillary acidicprotein is decreased in rat brain following acute and chroniccorticosterone treatmentrdquo Molecular Brain Research vol 7 no1 pp 1ndash7 1990

[46] N J Laping N R Nichols J R Day and C E Finch ldquoCortico-sterone differentially regulates the bilateral response of astro-cyte mRNAs in the hippocampus to entorhinal cortex lesions inmale ratsrdquoMolecular Brain Research vol 10 no 4 pp 291ndash2971991

[47] S Tsuneishi S Takada T Motoike T Ohashi K Sano andH Nakamura ldquoEffects of dexamethasone on the expression ofmyelin basic protein proteolipid protein and glial fibrillaryacidic protein genes in developing rat brainrdquo DevelopmentalBrain Research vol 61 no 1 pp 117ndash123 1991

[48] J P OrsquoCallaghan R E Brinton and B S McEwen ldquoGlucocor-ticoids regulate the synthesis of glial fibrillary acidic protein inintact and adrenalectomized rats but do not affect its expressionfollowing brain injuryrdquo Journal of Neurochemistry vol 57 no 3pp 860ndash869 1991

[49] M V Sofroniew and H V Vinters ldquoAstrocytes biology andpathologyrdquoActa Neuropathologica vol 119 no 1 pp 7ndash35 2010

[50] S Maccari M Darnaudery S Morley-Fletcher A R Zuena CCinque and O van Reeth ldquoPrenatal stress and long-termconsequences implications of glucocorticoid hormonesrdquo Neu-roscience and Biobehavioral Reviews vol 27 no 1-2 pp 119ndash1272003

[51] C R Pryce Y Aubert CMaier P C Pearce and E Fuchs ldquoThedevelopmental impact of prenatal stress prenatal dexametha-sone and postnatal social stress on physiology behaviour andneuroanatomy of primate offspring Studies in rhesus macaqueand common marmosetrdquo Psychopharmacology vol 214 no 1pp 33ndash53 2011

Submit your manuscripts athttpwwwhindawicom

Neurology Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Alzheimerrsquos DiseaseHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

ScientificaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Research and TreatmentSchizophrenia

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Neural Plasticity

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Parkinsonrsquos Disease

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Research and TreatmentAutism

Sleep DisordersHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Neuroscience Journal

Epilepsy Research and TreatmentHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Psychiatry Journal

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Computational and Mathematical Methods in Medicine

Depression Research and TreatmentHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Brain ScienceInternational Journal of

StrokeResearch and TreatmentHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Neurodegenerative Diseases

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Journal of

Cardiovascular Psychiatry and NeurologyHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Neural Plasticity 3

(a) (b)

Control

(c)

Dex-treated

(d)

Figure 1 Dorsal hippocampal volume HX stain (a) and GFAP-positive astroglial cell count (b) estimated by means of stereology Changesin astroglial morphology in response to AGT GFAP-positive astroglial cells in control (c) and Dex-treated (d) male rats (dorsal hippocampaldentate gyrus area) GFAP-positive primary process length is reduced in Dex-treated male rats when compared with control (d) versus (c)Astroglial processes observed at 100x magnification

Germany) An unbiased estimation of total number of cellswas achieved by choosing every 10th section according to thesystematic random sampling procedure [24] yielding 8 sec-tions per animal A pilot study was undertaken to determinethe following parameters A sampling area of 1602500120583m2was found to be optimal for this study Dissector height was8 120583m and a 1 120583m guard zone at the top and bottom part of thesection was excluded from the analysis at every step A fixedcounting frame of 50 times 50 and a variable sampling grid size(119909-119910-axis) of 150 times 150 CA1 150 times 150 CA2 300 times 300 CA3and 300 times 300 DG were used for the dorsal hippocampus(Figure 1(b)) and 150 times 150 was used for both basolateralamygdala and nucleus accumbens These sampling gridsresulted in 100ndash200 counting sites per brain HX and GFAP-positive cells were counted using a 100x Nicon UPlan FLNobjective (NA = 130) which allowed accurate recognitionEach cell was counted according to the unbiased countingrules The efficiency of convenient number of sampled cellsand parameters was checked by estimation of coefficient oferror (CE) and coefficient of variation (CV) [24]

273 AstroglialMorphology Morphological analysiswas per-formed on 80 astrocytes in each brain (10 astrocytes persection) usingNeurolucida software (Version 9MicroBright-Field Inc Germany) for tracing boundaries of the ROIsand astrocytes Astrocytes within the ROI were traced in

a systematic random manner taking care to avoid thosewhich were superimposed upon other astrocytes or bloodvessels The traced astrocytes were then analysed usingNeurolucida explorer software (Version 9 MicroBrightFieldInc Germany) Morphological analysis was performed byobserver who was blind to the experimental groups Primaryprocess length was measured using 100x objective (NiconUPlan FLN NA = 130) and by counting the primary pro-cesses extending directly from the soma in both the lateraland central quadrants of astrocytes in the same sections

28 Statistical Analysis Using SPSS 170 statistical software(SPSS Inc USA) ANOVA with a post hoc Tukey test wasperformed comparing between group differences Indepen-dent 119905-test was performed tomeasure difference between twogroups Statistical significance was set at 119875 lt 005The resultsare expressed as mean plusmn SEM

3 Results

31 Whole Brain Volume and Brain Weight No significanteffects of AGT were observed in the whole brain volume(119875 = 0376) and brain weight (119875 = 0442) (Table 1)

32 VolumeAnalyses AGT rats showed a significant decreasein the absolute volume of dorsal hippocampus by 65

4 Neural Plasticity

Table 1 Effects of AGT exposure on brain weight and volume ofmale adult Sprague Dawley rats

Control Dex-treatedBrain weight (g) 174 plusmn 002 177 plusmn 003Brain volume (mm3) 165018 plusmn 1775 168124 plusmn 2843The brain weight and volume are expressed as mean plusmn SEM (119899 = 6 pergroup) No significant differences between Dex-treated (AGT) and controlwere observed

Table 2 Effects of AGT on the regional brain volume of male adultSprague Dawley rats

VolumeControl Dex-treated

Absolute values (mm3)Dorsal hippocampus 328 plusmn 046 308 plusmn 050lowast

Basolateral amygdala 081 plusmn 002 080 plusmn 002Nucleus accumbens 228 plusmn 006 226 plusmn 007

Relative values (as brain volume)Dorsal hippocampus 199 plusmn 004 183 plusmn 004lowast

Basolateral amygdala 0046 plusmn 0001 0048 plusmn 0001Nucleus accumbens 0138 plusmn 0004 0135 plusmn 0005

The volume of the dorsal hippocampus basolateral amygdala and nucleusin control and dexamethasone-treated groups (119899 = 6 per group) Data areexpressed as mean plusmn SEM lowast119875 lt 005 versus control independent sample 119905-test

versus control (119875 lt 005) When expressed as percentageof the whole brain volume the volume reduction was 8(119875 lt 005) versus control No significant differences wereobserved between the treatment groups in the basolateralamygdala (119875 = 0834) and nucleus accumbens (119875 = 0905)(Table 2)

33 HX Stained Total Cell Numbers AGT did not affect thetotal cell count (HX stain) in the dorsal hippocampal subre-gions CA1 (119875 = 0821) CA2 (119875 = 0975) CA3 (119875 = 0991)and DG (119875 = 0959) basolateral amygdala (119875 = 0180) andthe nucleus accumbens (119875 = 0474) (Table 3)

34 GFAP-Positive Astroglial Cell Numbers There were nosignificant AGT effects on GFAP-positive cell numbers in thedorsal hippocampal subregions CA1 (119875 = 0607) CA2 (119875 =0517) CA3 (119875 = 0500) and DG (119875 = 0424) basolateralamygdala (119875 = 0495) and the nucleus accumbens (119875 =0384) (Table 4)

35 Astroglia (GFAP-Positive) Cell Morphology AGT didnot affect the numbers of primary processes in the regionanalysed (Tables 5 and 6) and it had no significant effectson the astroglial morphology in the nonhippocampal regions(Table 5 all group differences at 119875 lt 005)

However AGT affected the length of primary processesin GFAP-positive cells within the hippocampus (Figures 1(a)and 1(b)) it led to a significant reduction in the total length ofastrocytic primary processes in the hippocampal subregionsby 32 in CA1 (119875 lt 005) 50 in CA3 (119875 lt 0001) and by

Table 3 Effects of AGT on the number of total (hematoxylin-stained) and GFAP-positive cellsin dorsal hippocampus of maleadult Sprague Dawley rats

Subregion Control Dex-treatedHematoxylin stained cell numbers estimated per region (times103)

CA1 7068 plusmn 88 7141 plusmn 297CA2 959 plusmn 34 958 plusmn 2CA3 4913 plusmn 372 4907 plusmn 326DG 7914 plusmn 316 7891 plusmn 306

GFAP +ve cell numbers estimated per region (times103)CA1 2194 plusmn 6 2136 plusmn 91CA2 268 plusmn 11 258 plusmn 1CA3 1109 plusmn 61 1044 plusmn 69DG 201 plusmn 47 1953 plusmn 51The numbers of total and GFAP-positive cells are shown for control anddexamethasone-treated groups (119899 = 6 per group) Data are expressed asmean plusmn SEM No significant differences between Dex-treated (AGT) andcontrol were observed

Table 4 Effects of AGT on the number of total and GFAP-positivecells in basolateral amygdala and nucleus accumbens region of cellsin male adult Sprague Dawley rats

Subregion Control Dex-treatedHematoxylin stained cell numbers estimated per region (times103)

Basolateral amygdala 5286 plusmn 34 5879 plusmn 24Nucleus accumbens 14658 plusmn 68 13961 plusmn 64

GFAP +ve cell numbers estimated per region (times103)Basolateral amygdala 2368 plusmn 19 2571 plusmn 21Nucleus accumbens 2073 plusmn 18 1828 plusmn 20Thenumbers of total andGFAP-positive cells in control and dexamethasone-treated groups (119899 = 6 per group) Data are expressed as mean plusmn SEMNo significant differences between Dex-treated (AGT) and control wereobserved

Table 5 Effects of AGT on themorphology of GFAP-positive astro-glia in the basolateral amygdala and nucleus accumbens region ofmale adult Sprague Dawley rats

GFAP-astroglia morphologyControl Dex-treated

Total primary process length (120583m)Basolateral amygdala 57618 plusmn 5826 61235 plusmn 5989Nucleus accumbens 40334 plusmn 4039 38901 plusmn 4892

Primary process mean lengthBasolateral amygdala 268 plusmn 16 275 plusmn 21Nucleus accumbens 227 plusmn 18 208 plusmn 17

Number of primary processesBasolateral amygdala 213 plusmn 114 221 plusmn 91Nucleus accumbens 178 plusmn 85 185 plusmn 106

The total primary process length mean primary process length and num-bers of primary processes of GFAP-positive cells in control and dexametha-sone-treated groups (119899 = 6 per group) Data are expressed as mean plusmn SEMNo significant differences between Dex-treated (AGT) and control wereobserved

Neural Plasticity 5

Table 6 Effects of AGT on GFAP-positive astroglial primaryprocess numbers in male adult Sprague Dawley rats

GFAP-astroglia morphologyControl Dex-treated

Number of primary processCA1 362 plusmn 361 3233 plusmn 44CA2 301 plusmn 262 311 plusmn 264CA3 339 plusmn 169 3057 plusmn 81DG 3217 plusmn 143 3148 plusmn 47

The numbers of primary processes of GFAP-positive cells in control anddexamethasone-treated groups (119899= 6 per group) Data are expressed asmeanplusmn SEM No significant differences between Dex-treated (AGT) and controlwere observed

25 in DG (119875 lt 001) (Figure 2) AGT-dependent changeswere also observed in the mean length of astrocytic primaryprocesses reductions by 25 in CA1 (119875 lt 001) 45 in CA3(119875 lt 0001) and 25 in DG (119875 lt 001) (Figure 3)

4 Discussion

The present study was focused on the effects of AGT oncentral astroglia and regional volume of the hippocampusamygdala and nucleus accumbens as the areas of the limbicsystem implicated in response to stress and corticosteroids [23 9] AGTwas administered to pregnant dams noninvasivelythrough drinkingwater to avoid the stress of animal handlingand injections Of the brain areas tested in adult male rats thehippocampus responded to AGT with a long-term reductionin its volume and changes in astroglial morphology It is plau-sible that the observed impoverishment of astrocytic primarybranches underpins the hippocampal volume reduction aswe found no evidence of cell loss To our knowledge suchAGT-dependent long-term hippocampal remodelling withimplications for astroglia has not been reported before

Astroglia were visualised by means of the commonlyused GFAP immunohistochemistry GFAP is an intermediatefilament protein present in cell bodies and primary branchesalthough fine distal processes show little or no GFAP [25]Although GFAP staining is selective to astrocytes it does notlabel all astrocytes in the brain Accepting these limitationsof GFAP immunohistochemistry the present study focusedon the length and numbers of primary astrocytic processeswhere GFAP is reliably detectable [25] Stereology was ourmethod of choice as it offers an unbiased approach to regionalbrain volumetry and morphometry [24]

Previous volumetric studies have considered specificbrain regions expressing their volumes in absolute termswith no reference to the total brain volume which itself isa variable Thus the present application of pQCT to measuretotal brain volume adds the much needed rigour to volu-metric assessment where regional volumes need to be scaledto the total brain volume This is particularly important asperinatal corticosteroid exposure can lead to a decrease inbrain weightvolume For example a significant brain weightreduction has been reported in rhesus monkeys treatedprenatally with corticosteroids [26] and repeated doses of

lowast

lowastlowast

lowastlowastlowast

1200000

1100000

1000000

900000

800000

700000

600000

500000

400000

300000

200000

100000

000

CA1 CA2 CA3 DG

TreatmentControlDEX

Mea

n pr

imar

y pr

oces

s tot

al le

ngth

(120583m

)

Dorsal hippocampal subregions

Figure 2 Effect of prenatal dexamethasone treatment on the totallength of primary astrocytic processes in hippocampal subregionsof male adult Sprague Dawley rats The total primary process lengthofGFAP-positive cells in control and dexamethasone-treated groups(119899 = 6 per group) Data are expressed as mean plusmn SEM lowast119875 lt 005lowastlowast

119875 lt 001 and lowastlowastlowast119875 lt 0001 versus control independent sample119905-test

lowastlowast

lowastlowast

lowastlowastlowast

500

000

1000

1500

2000

2500

3000

CA1 CA2 CA3 DG

TreatmentControlDEX

Mea

n pr

imar

y pr

oces

s mea

n le

ngth

(120583m

)

Dorsal hippocampal subregions

Figure 3 Effect of prenatal dexamethasone treatment on the meanlength of primary astrocytic processes in hippocampal subregions ofmale adult Sprague Dawley rats The mean primary process lengthofGFAP-positive cells in control and dexamethasone-treated groups(119899 = 6 per group) Data are expressed as mean plusmn SEM lowastlowast119875 lt 001and lowastlowastlowast119875 lt 0001 versus control independent sample 119905-test

6 Neural Plasticity

dexamethasone at E17ndash19 have resulted in a significant brainweight reduction in newborn rats [27] The present studyfound no AGT effects on the whole brain weight and volumethe latter measured by pQCT One of the factors which affectthe brain weight and volume is the amount of corticosteroidadministered In the present study a relatively small dosewas given which as discussed previously [19] is in the rangeused in perinatal medicine and mimics the glucocorticoidactivity that is required for normal lung maturation duringlate gestation

Within the developing brain the limbic system richlyexpressing glucocorticoid and mineralocorticoid receptors[28] is sensitive to the endogenous and exogenous corti-costeroids which can affect these brain areas with long-term consequences for their structure and function [29]Reductions in the regional brain volume of the hippocampusand amygdala have been reported in preterm infants [30ndash32] and a cortical brain volume reduction of 35 has beenobserved in premature infants treated with dexamethasone[33] On the other hand there have been reports of no effectof single betamethasone antenatal dose on regional brainvolumes particularly within the limbic system [34]

In the present study the hippocampus was significantlyaffected by AGT in line with the theory that this regiondevelops primarily during the foetal period [35] althoughhippocampal volume losses have also been observed fol-lowing administration of corticosteroids in adult monkeysand rats [36 37] No changes were seen in the basolateralamygdala and nucleus accumbens volume which suggests aregion-selective treatment effect consistent with the densityof glucocorticoid receptor distribution [28]

A transient reduction in hippocampal volume has beenobserved in adolescent but not adult mice exposed to a clin-ically relevant dose of dexamethasone at E15 [18] The extentof hippocampal volume loss is normally mildmoderate Thepresent reduction of the hippocampal volume by 8 fits inthat range while being statistically significant It is compara-ble with some previous reports on stresssteroid-dependentlosses in the hippocampal volume across several speciesThusCoe et al [10] have found hippocampal volume reductions by12 and 10 in the offspring of rhesus monkeys exposed toearly life stress In tree shrews four weeksrsquo cortisol treatmentor psychosocial stress has resulted in a hippocampal volumeloss by 5ndash10 [38] Chronic social defeat stress has led toa similarly mild yet significant hippocampal volume lossin adult rats [6] Human adolescents who were born verypreterm have been found to have a hippocampal volume lossin the range of 12 to 156 [39]

What causes the hippocampal volume losses in responsesto stressglucocorticoids There is a range of factors such asneuronal andor glial cell death reduction of cell size includ-ing atrophy of neuronal dendrites andor glial branchesreduced water content or impaired neurogenesis [1] andorreduced microvasculature [6] The present study did not findcell losses in the brain areas tested and focused on astrogliabecause of their major role in maintaining brain plasticityat the level of synapses and cells [25] Currently published

studies on the mechanisms underlying glucocorticoid-induced brain volume reductions have focused on neuronsAs a result neuronal cell loss has been well researched anddocumented and various dexamethasone treatments havebeen found to lead to neuronal loss in rats [40 41] and pri-mates [26 37] mostly in the hippocampal CA3 and dentategyrus Less attention has been given to astroglial cells despitetheir role in the maintenance of brain homeostasis andplasticity [25]

The impoverishment in astroglial morphology reportedin the present study is consistent with the known glia-inhibitory effects of exogenous glucocorticoids or stress invitro [42 43] and in vivo [44] Treatment with corticosteroneor synthetic glucocorticoids such as dexamethasone has beenknown to inhibit GFAP expression in the neonatal andadult rat brain [45ndash48] thus suggesting that astrocytes canbe affected by glucocorticoid overload The very fact thatdexamethasone can affect GFAP expression raises a validmethodological consideration as to what extent the presentobservations of changes in the morphology of GFAP-positiveastroglia are affected bymethodological biaswhen comparingbetween the groups Central astroglia are heterogeneous[25 49] and using GFAP as the only immunohistochemicalmarker brings in an intrinsic limitation shared by many cur-rent studies as critically reviewed elsewhere [49] Howeverthe present GFAP responses to AGT appear to be selective tothe hippocampus where astroglial primary branches reducedin length but no GFAP-positive cell losses were found Sucha discrete response of GFAP immunoreactivity implies thatdexamethasone administered at the present lowdose asAGTdid not induce a global effect in GFAP expression Thisnot only highlights the unique susceptibility of hippocampalregions but also suggests that the changes observed here aretrue manifestations of impoverished hippocampal astroglialmorphology in response to AGT

It is increasingly evident that nongeneticenvironmentalfactors acting in early life result in long-term alterations ofthe physiological systems affected Perinatal programmingandor plasticity of the physiological system under extremeconditions such as ldquotoxicrdquo stress or AGT can result in long-term neurobehavioural abnormalities [3 29 50] and there isgrowing evidence that prenatal or early postnatal exposure tostressors affects the long-term brain functioning [51]

To conclude this study reports significant maladaptivechanges in astroglia in the hippocampal subregions CA1 CA3and dentate gyrus as a long-term effect of AGT It is plausibleto assume that the reductions in astrocytic primary branchescontribute to the losses in the hippocampal volume Theyare also likely to affect astroglial function and neuron-gliainteractions as retraction of primary astroglial processes candisadvantage local networks formed with glia and neuronsThe observed dexamethasone-induced impoverishment ofastroglial morphology can be treated as a demonstration ofglial maladaptive plasticity a phenomenon that deservesmore research in the context of effects of corticosteroidoverload and stress-related pathologies Epigenetics of thisphenomenon should be of interest considering the clinicaluse of dexamethasone

Neural Plasticity 7

Abbreviations

AGT Antenatal glucocorticoid treatmentDex DexamethasoneGD Gestational dayGFAP Glial fibrillary acidic proteinHX HematoxylinMDD Major depressive disorderPBS Phosphate-buffered salinePTSD Posttraumatic stress disorderROI Region of interest

Conflict of Interests

The authors declare that there is no conflict of interestsregarding the publication of this paper

Acknowledgments

This work was supported by the Wellcome Trust Grant (no086871Z08Z) to G E Gillies Vishvesh H Shende was aPhD student sponsored by the University of RoehamptonThe authors thank Dr Todd Rae (Roehampton) for trainingVishvesh H Shende in the pQCT technique

References

[1] B Czeh and P J Lucassen ldquoWhat causes the hippocampalvolume decrease in depression Are neurogenesis glial changesand apoptosis implicatedrdquo European Archives of Psychiatry andClinical Neuroscience vol 257 no 5 pp 250ndash260 2007

[2] P J Lucassen J Pruessner N Sousa et al ldquoNeuropathology ofstressrdquo Acta Neuropathologica vol 127 no 1 pp 109ndash135 2014

[3] B S McEwen L Eiland R G Hunter andMMMiller ldquoStressand anxiety structural plasticity and epigenetic regulation as aconsequence of stressrdquo Neuropharmacology vol 62 no 1 pp3ndash12 2012

[4] M A Mehta N I Golembo C Nosarti et al ldquoAmygdala hip-pocampal and corpus callosum size following severe earlyinstitutional deprivation the English and Romanian Adopteesstudy pilotrdquo Journal of Child Psychology and Psychiatry andAllied Disciplines vol 50 no 8 pp 943ndash951 2009

[5] T Frodl and V OrsquoKeane ldquoHow does the brain deal with cumu-lative stress A review with focus on developmental stress HPAaxis function and hippocampal structure in humansrdquo Neurobi-ology of Disease vol 52 pp 24ndash37 2013

[6] B Czeh N Abumaria R Rygula and E Fuchs ldquoQuantita-tive changes in hippocampal microvasculature of chronicallystressed rats no effect of fluoxetine treatmentrdquo Hippocampusvol 20 no 1 pp 174ndash185 2010

[7] V H Shende C R Pryce D Ruedi-Bettschen T C Rae andJ Opacka-Juffry ldquoEvidence for hippocampal remodeling as along term effect of early life stress in the rat model of earlydeprivationrdquo Recent Developments in Brain Research vol 1 pp1ndash19 2012

[8] A Jackowski T D Perera C G Abdallah et al ldquoEarly-lifestress corpus callosum development hippocampal volumet-rics and anxious behavior in male nonhuman primatesrdquo Psy-chiatry ResearchmdashNeuroimaging vol 192 no 1 pp 37ndash44 2011

[9] C Pittenger and R S Duman ldquoStress depression and neuro-plasticity a convergence of mechanismsrdquoNeuropsychopharma-cology vol 33 no 1 pp 88ndash109 2008

[10] C L Coe M Kramer B Czeh et al ldquoPrenatal stress diminishesneurogenesis in the dentate gyrus of juvenile Rhesus monkeysrdquoBiological Psychiatry vol 54 no 10 pp 1025ndash1034 2003

[11] J D Funkhouser K J Peevey P B Mockridge and E RHughes ldquoDistribution of dexamethasone between mother andfetus after maternal administrationrdquo Pediatric Research vol 12no 11 pp 1053ndash1056 1978

[12] D Roberts and S Dalziel ldquoAntenatal corticosteroids for accel-erating fetal lung maturation for women at risk of pretermbirthrdquo Cochrane Database of Systematic Reviews vol 3 ArticleID CD004454 2006

[13] T F Yeh Y J Lin H C Lin et al ldquoOutcomes at school ageafter postnatal dexamethasone therapy for lung disease of pre-maturityrdquoTheNew England Journal of Medicine vol 350 no 13pp 1304ndash1313 2004

[14] J P Empana M M Anceschi I Szabo E V Cosmi G Breartand P Truffert ldquoAntenatal corticosteroids policies in 14 Euro-pean countries factors associated with multiple courses TheEURAIL surveyrdquo Acta Paediatrica International Journal of Pae-diatrics vol 93 no 10 pp 1318ndash1322 2004

[15] C Romagnoli E Zecca R Luciano G Torrioli and G Tor-torolo ldquoA three year follow up of preterm infants after moder-ately early treatment with dexamethasonerdquo Archives of Diseasein Childhood Fetal and Neonatal Edition vol 87 no 1 pp F55ndashF58 2002

[16] A Charil D P Laplante C Vaillancourt and S King ldquoPrenatalstress and brain developmentrdquo Brain Research Reviews vol 65no 1 pp 56ndash79 2010

[17] D Tijsseling L D E Wijnberger J B Derks et al ldquoEffects ofantenatal glucocorticoid therapy on hippocampal histology ofpreterm infantsrdquoPLoSONE vol 7 no 3 Article ID e33369 2012

[18] C W Noorlander D Tijsseling E V S Hessel et al ldquoAntenatalglucocorticoid treatment affects hippocampal development inmicerdquo PLoS ONE vol 9 no 1 Article ID e85671 2014

[19] S McArthur E McHale and G E Gillies ldquoThe size and distri-bution of midbrain dopaminergic populations are permanentlyaltered by perinatal glucocorticoid exposure in a sex- region-and time-specific mannerrdquo Neuropsychopharmacology vol 32no 7 pp 1462ndash1476 2007

[20] S McArthur E McHale J W Dalley J C Buckingham andG E Gillies ldquoAlteredmesencephalic dopaminergic populationsin adulthood as a consequence of brief perinatal glucocorticoidexposurerdquo Journal of Neuroendocrinology vol 17 no 8 pp 475ndash482 2005

[21] ETheogaraj CD JohnH C Christian J FMorris S F Smithand J C Buckingham ldquoPerinatal glucocorticoid treatmentproduces molecular functional and morphological changes inthe anterior pituitary gland of the adultmale ratrdquoEndocrinologyvol 146 no 11 pp 4804ndash4813 2005

[22] G Paxinos and C WatsonThe Rat Brain in Stereotaxic Coordi-nates Elsevier Academic Press 5th edition 2005

[23] M Leventopoulos D Ruedi-Bettschen I Knuesel J Feldon CR Pryce and J Opacka-Juffry ldquoLong-term effects of early lifedeprivation on brain glia in Fischer ratsrdquo Brain Research vol1142 no 1 pp 119ndash126 2007

[24] M J West L Slomianka and H J G Gundersen ldquoUnbiasedstereological estimation of the total number of neurons in

8 Neural Plasticity

the subdivisions of the rat hippocampus using the optical frac-tionatorrdquo Anatomical Record vol 231 no 4 pp 482ndash497 1991

[25] D T Theodosis D A Poulain and S H R Oliet ldquoActivity-dependent structural and functional plasticity of astrocyte-neuron interactionsrdquo Physiological Reviews vol 88 no 3 pp983ndash1008 2008

[26] H Uno L Lohmiller C Thieme et al ldquoBrain damage inducedby prenatal exposure to dexamethasone in fetal rhesus maca-ques I Hippocampusrdquo Developmental Brain Research vol 53no 2 pp 157ndash167 1990

[27] R Q Carlos F J Seidler and T A Slotkin ldquoFetal dexametha-sone exposure altersmacromolecular characteristics of rat braindevelopment a critical period for regionally selective altera-tionsrdquo Teratology vol 46 no 1 pp 45ndash59 1992

[28] E R de Kloet ldquoStress in the brainrdquo European Journal of Phar-macology vol 405 pp 187ndash198 2000

[29] S G Matthews ldquoAntenatal glucocorticoids and the developingbrain mechanisms of actionrdquo Seminars in Neonatology vol 6no 4 pp 309ndash317 2001

[30] L J Abernethy M Palaniappan and R W I Cooke ldquoQuanti-tative magnetic resonance imaging of the brain in survivors ofvery low birth weightrdquo Archives of Disease in Childhood vol 87no 4 pp 279ndash283 2002

[31] E B Isaacs A LucasW K Chong et al ldquoHippocampal volumeand everyday memory in children of very low birth weightrdquoPediatric Research vol 47 no 6 pp 713ndash720 2000

[32] B S Peterson B Vohr L H Staib et al ldquoRegional brain volumeabnormalities and long-term cognitive outcome in preterminfantsrdquo Journal of the American Medical Association vol 284no 15 pp 1939ndash1947 2000

[33] B P Murphy T E Inder P S Huppi et al ldquoImpaired cerebralcortical graymatter growth after treatmentwith dexamethasonefor neonatal chronic lung diseaserdquo Pediatrics vol 107 no 2 pp217ndash221 2001

[34] P Yossuck M Kraszpulski and A K Salm ldquoPerinatal corti-costeroid effect on amygdala and hippocampus volume duringbrain development in the rat modelrdquo Early Human Develop-ment vol 82 no 4 pp 267ndash272 2006

[35] D Rice and S Barone Jr ldquoCritical periods of vulnerability forthe developing nervous system evidence from humans andanimal modelsrdquo Environmental Health Perspectives vol 108 no3 pp 511ndash533 2000

[36] C S Woolley E Gould and B S McEwen ldquoExposure toexcess glucocorticoids alters dendritic morphology of adulthippocampal pyramidal neuronsrdquo Brain Research vol 531 no1-2 pp 225ndash231 1990

[37] R M Sapolsky H Uno C S Rebert and C E Finch ldquoHippo-campal damage associatedwith prolonged glucocorticoid expo-sure in primatesrdquoThe Journal of Neuroscience vol 10 no 9 pp2897ndash2902 1990

[38] F Ohl T Michaelis G K Vollmann-Honsdorf C Kirschbaumand E Fuchs ldquoEffect of chronic psychosocial stress and long-term cortisol treatment on hippocampus-mediated memoryand hippocampal volume a pilot-study in tree shrewsrdquo Psy-choneuroendocrinology vol 25 no 4 pp 357ndash363 2000

[39] C Nosarti M H S Al-Asady S Frangou A L Stewart L Rif-kin and R M Murray ldquoAdolescents who were born very pre-term have decreased brain volumesrdquo Brain vol 125 no 7 pp1616ndash1623 2002

[40] L E Haynes M R Griffiths R E Hyde D J Barber and I JMitchell ldquoDexamethasone induces limited apoptosis and exten-sive sublethal damage to specific subregions of the striatum andhippocampus implications for mood disordersrdquo Neurosciencevol 104 no 1 pp 57ndash69 2001

[41] N Sousa M M Paula-Barbosa and O F X Almeida ldquoLigandand subfield specificity of corticoid-induced neuronal loss inthe rat hippocampal formationrdquoNeuroscience vol 89 no 4 pp1079ndash1087 1999

[42] K L Crossin M-H Tai L A Krushel V P Mauro and G MEdelman ldquoGlucocorticoid receptor pathways are involved in theinhibition of astrocyte proliferationrdquo Proceedings of the NationalAcademy of Sciences of the United States of America vol 94 no6 pp 2687ndash2692 1997

[43] M Sabolek A Herborg J Schwarz and A Storch ldquoDexam-ethasone blocks astroglial differentiation from neural precursorcellsrdquo NeuroReport vol 17 no 16 pp 1719ndash1723 2006

[44] N R Nichols J N Masters and C E Finch ldquoChanges in geneexpression in hippocampus in response to glucocorticoids andstressrdquo Brain Research Bulletin vol 24 no 5 pp 659ndash662 1990

[45] N R Nichols H H Osterburg J N Masters S L Millarand C E Finch ldquoMessenger RNA for glial fibrillary acidicprotein is decreased in rat brain following acute and chroniccorticosterone treatmentrdquo Molecular Brain Research vol 7 no1 pp 1ndash7 1990

[46] N J Laping N R Nichols J R Day and C E Finch ldquoCortico-sterone differentially regulates the bilateral response of astro-cyte mRNAs in the hippocampus to entorhinal cortex lesions inmale ratsrdquoMolecular Brain Research vol 10 no 4 pp 291ndash2971991

[47] S Tsuneishi S Takada T Motoike T Ohashi K Sano andH Nakamura ldquoEffects of dexamethasone on the expression ofmyelin basic protein proteolipid protein and glial fibrillaryacidic protein genes in developing rat brainrdquo DevelopmentalBrain Research vol 61 no 1 pp 117ndash123 1991

[48] J P OrsquoCallaghan R E Brinton and B S McEwen ldquoGlucocor-ticoids regulate the synthesis of glial fibrillary acidic protein inintact and adrenalectomized rats but do not affect its expressionfollowing brain injuryrdquo Journal of Neurochemistry vol 57 no 3pp 860ndash869 1991

[49] M V Sofroniew and H V Vinters ldquoAstrocytes biology andpathologyrdquoActa Neuropathologica vol 119 no 1 pp 7ndash35 2010

[50] S Maccari M Darnaudery S Morley-Fletcher A R Zuena CCinque and O van Reeth ldquoPrenatal stress and long-termconsequences implications of glucocorticoid hormonesrdquo Neu-roscience and Biobehavioral Reviews vol 27 no 1-2 pp 119ndash1272003

[51] C R Pryce Y Aubert CMaier P C Pearce and E Fuchs ldquoThedevelopmental impact of prenatal stress prenatal dexametha-sone and postnatal social stress on physiology behaviour andneuroanatomy of primate offspring Studies in rhesus macaqueand common marmosetrdquo Psychopharmacology vol 214 no 1pp 33ndash53 2011

Submit your manuscripts athttpwwwhindawicom

Neurology Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Alzheimerrsquos DiseaseHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

ScientificaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Research and TreatmentSchizophrenia

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Neural Plasticity

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Parkinsonrsquos Disease

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Research and TreatmentAutism

Sleep DisordersHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Neuroscience Journal

Epilepsy Research and TreatmentHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Psychiatry Journal

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Computational and Mathematical Methods in Medicine

Depression Research and TreatmentHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Brain ScienceInternational Journal of

StrokeResearch and TreatmentHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Neurodegenerative Diseases

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

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Cardiovascular Psychiatry and NeurologyHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

4 Neural Plasticity

Table 1 Effects of AGT exposure on brain weight and volume ofmale adult Sprague Dawley rats

Control Dex-treatedBrain weight (g) 174 plusmn 002 177 plusmn 003Brain volume (mm3) 165018 plusmn 1775 168124 plusmn 2843The brain weight and volume are expressed as mean plusmn SEM (119899 = 6 pergroup) No significant differences between Dex-treated (AGT) and controlwere observed

Table 2 Effects of AGT on the regional brain volume of male adultSprague Dawley rats

VolumeControl Dex-treated

Absolute values (mm3)Dorsal hippocampus 328 plusmn 046 308 plusmn 050lowast

Basolateral amygdala 081 plusmn 002 080 plusmn 002Nucleus accumbens 228 plusmn 006 226 plusmn 007

Relative values (as brain volume)Dorsal hippocampus 199 plusmn 004 183 plusmn 004lowast

Basolateral amygdala 0046 plusmn 0001 0048 plusmn 0001Nucleus accumbens 0138 plusmn 0004 0135 plusmn 0005

The volume of the dorsal hippocampus basolateral amygdala and nucleusin control and dexamethasone-treated groups (119899 = 6 per group) Data areexpressed as mean plusmn SEM lowast119875 lt 005 versus control independent sample 119905-test

versus control (119875 lt 005) When expressed as percentageof the whole brain volume the volume reduction was 8(119875 lt 005) versus control No significant differences wereobserved between the treatment groups in the basolateralamygdala (119875 = 0834) and nucleus accumbens (119875 = 0905)(Table 2)

33 HX Stained Total Cell Numbers AGT did not affect thetotal cell count (HX stain) in the dorsal hippocampal subre-gions CA1 (119875 = 0821) CA2 (119875 = 0975) CA3 (119875 = 0991)and DG (119875 = 0959) basolateral amygdala (119875 = 0180) andthe nucleus accumbens (119875 = 0474) (Table 3)

34 GFAP-Positive Astroglial Cell Numbers There were nosignificant AGT effects on GFAP-positive cell numbers in thedorsal hippocampal subregions CA1 (119875 = 0607) CA2 (119875 =0517) CA3 (119875 = 0500) and DG (119875 = 0424) basolateralamygdala (119875 = 0495) and the nucleus accumbens (119875 =0384) (Table 4)

35 Astroglia (GFAP-Positive) Cell Morphology AGT didnot affect the numbers of primary processes in the regionanalysed (Tables 5 and 6) and it had no significant effectson the astroglial morphology in the nonhippocampal regions(Table 5 all group differences at 119875 lt 005)

However AGT affected the length of primary processesin GFAP-positive cells within the hippocampus (Figures 1(a)and 1(b)) it led to a significant reduction in the total length ofastrocytic primary processes in the hippocampal subregionsby 32 in CA1 (119875 lt 005) 50 in CA3 (119875 lt 0001) and by

Table 3 Effects of AGT on the number of total (hematoxylin-stained) and GFAP-positive cellsin dorsal hippocampus of maleadult Sprague Dawley rats

Subregion Control Dex-treatedHematoxylin stained cell numbers estimated per region (times103)

CA1 7068 plusmn 88 7141 plusmn 297CA2 959 plusmn 34 958 plusmn 2CA3 4913 plusmn 372 4907 plusmn 326DG 7914 plusmn 316 7891 plusmn 306

GFAP +ve cell numbers estimated per region (times103)CA1 2194 plusmn 6 2136 plusmn 91CA2 268 plusmn 11 258 plusmn 1CA3 1109 plusmn 61 1044 plusmn 69DG 201 plusmn 47 1953 plusmn 51The numbers of total and GFAP-positive cells are shown for control anddexamethasone-treated groups (119899 = 6 per group) Data are expressed asmean plusmn SEM No significant differences between Dex-treated (AGT) andcontrol were observed

Table 4 Effects of AGT on the number of total and GFAP-positivecells in basolateral amygdala and nucleus accumbens region of cellsin male adult Sprague Dawley rats

Subregion Control Dex-treatedHematoxylin stained cell numbers estimated per region (times103)

Basolateral amygdala 5286 plusmn 34 5879 plusmn 24Nucleus accumbens 14658 plusmn 68 13961 plusmn 64

GFAP +ve cell numbers estimated per region (times103)Basolateral amygdala 2368 plusmn 19 2571 plusmn 21Nucleus accumbens 2073 plusmn 18 1828 plusmn 20Thenumbers of total andGFAP-positive cells in control and dexamethasone-treated groups (119899 = 6 per group) Data are expressed as mean plusmn SEMNo significant differences between Dex-treated (AGT) and control wereobserved

Table 5 Effects of AGT on themorphology of GFAP-positive astro-glia in the basolateral amygdala and nucleus accumbens region ofmale adult Sprague Dawley rats

GFAP-astroglia morphologyControl Dex-treated

Total primary process length (120583m)Basolateral amygdala 57618 plusmn 5826 61235 plusmn 5989Nucleus accumbens 40334 plusmn 4039 38901 plusmn 4892

Primary process mean lengthBasolateral amygdala 268 plusmn 16 275 plusmn 21Nucleus accumbens 227 plusmn 18 208 plusmn 17

Number of primary processesBasolateral amygdala 213 plusmn 114 221 plusmn 91Nucleus accumbens 178 plusmn 85 185 plusmn 106

The total primary process length mean primary process length and num-bers of primary processes of GFAP-positive cells in control and dexametha-sone-treated groups (119899 = 6 per group) Data are expressed as mean plusmn SEMNo significant differences between Dex-treated (AGT) and control wereobserved

Neural Plasticity 5

Table 6 Effects of AGT on GFAP-positive astroglial primaryprocess numbers in male adult Sprague Dawley rats

GFAP-astroglia morphologyControl Dex-treated

Number of primary processCA1 362 plusmn 361 3233 plusmn 44CA2 301 plusmn 262 311 plusmn 264CA3 339 plusmn 169 3057 plusmn 81DG 3217 plusmn 143 3148 plusmn 47

The numbers of primary processes of GFAP-positive cells in control anddexamethasone-treated groups (119899= 6 per group) Data are expressed asmeanplusmn SEM No significant differences between Dex-treated (AGT) and controlwere observed

25 in DG (119875 lt 001) (Figure 2) AGT-dependent changeswere also observed in the mean length of astrocytic primaryprocesses reductions by 25 in CA1 (119875 lt 001) 45 in CA3(119875 lt 0001) and 25 in DG (119875 lt 001) (Figure 3)

4 Discussion

The present study was focused on the effects of AGT oncentral astroglia and regional volume of the hippocampusamygdala and nucleus accumbens as the areas of the limbicsystem implicated in response to stress and corticosteroids [23 9] AGTwas administered to pregnant dams noninvasivelythrough drinkingwater to avoid the stress of animal handlingand injections Of the brain areas tested in adult male rats thehippocampus responded to AGT with a long-term reductionin its volume and changes in astroglial morphology It is plau-sible that the observed impoverishment of astrocytic primarybranches underpins the hippocampal volume reduction aswe found no evidence of cell loss To our knowledge suchAGT-dependent long-term hippocampal remodelling withimplications for astroglia has not been reported before

Astroglia were visualised by means of the commonlyused GFAP immunohistochemistry GFAP is an intermediatefilament protein present in cell bodies and primary branchesalthough fine distal processes show little or no GFAP [25]Although GFAP staining is selective to astrocytes it does notlabel all astrocytes in the brain Accepting these limitationsof GFAP immunohistochemistry the present study focusedon the length and numbers of primary astrocytic processeswhere GFAP is reliably detectable [25] Stereology was ourmethod of choice as it offers an unbiased approach to regionalbrain volumetry and morphometry [24]

Previous volumetric studies have considered specificbrain regions expressing their volumes in absolute termswith no reference to the total brain volume which itself isa variable Thus the present application of pQCT to measuretotal brain volume adds the much needed rigour to volu-metric assessment where regional volumes need to be scaledto the total brain volume This is particularly important asperinatal corticosteroid exposure can lead to a decrease inbrain weightvolume For example a significant brain weightreduction has been reported in rhesus monkeys treatedprenatally with corticosteroids [26] and repeated doses of

lowast

lowastlowast

lowastlowastlowast

1200000

1100000

1000000

900000

800000

700000

600000

500000

400000

300000

200000

100000

000

CA1 CA2 CA3 DG

TreatmentControlDEX

Mea

n pr

imar

y pr

oces

s tot

al le

ngth

(120583m

)

Dorsal hippocampal subregions

Figure 2 Effect of prenatal dexamethasone treatment on the totallength of primary astrocytic processes in hippocampal subregionsof male adult Sprague Dawley rats The total primary process lengthofGFAP-positive cells in control and dexamethasone-treated groups(119899 = 6 per group) Data are expressed as mean plusmn SEM lowast119875 lt 005lowastlowast

119875 lt 001 and lowastlowastlowast119875 lt 0001 versus control independent sample119905-test

lowastlowast

lowastlowast

lowastlowastlowast

500

000

1000

1500

2000

2500

3000

CA1 CA2 CA3 DG

TreatmentControlDEX

Mea

n pr

imar

y pr

oces

s mea

n le

ngth

(120583m

)

Dorsal hippocampal subregions

Figure 3 Effect of prenatal dexamethasone treatment on the meanlength of primary astrocytic processes in hippocampal subregions ofmale adult Sprague Dawley rats The mean primary process lengthofGFAP-positive cells in control and dexamethasone-treated groups(119899 = 6 per group) Data are expressed as mean plusmn SEM lowastlowast119875 lt 001and lowastlowastlowast119875 lt 0001 versus control independent sample 119905-test

6 Neural Plasticity

dexamethasone at E17ndash19 have resulted in a significant brainweight reduction in newborn rats [27] The present studyfound no AGT effects on the whole brain weight and volumethe latter measured by pQCT One of the factors which affectthe brain weight and volume is the amount of corticosteroidadministered In the present study a relatively small dosewas given which as discussed previously [19] is in the rangeused in perinatal medicine and mimics the glucocorticoidactivity that is required for normal lung maturation duringlate gestation

Within the developing brain the limbic system richlyexpressing glucocorticoid and mineralocorticoid receptors[28] is sensitive to the endogenous and exogenous corti-costeroids which can affect these brain areas with long-term consequences for their structure and function [29]Reductions in the regional brain volume of the hippocampusand amygdala have been reported in preterm infants [30ndash32] and a cortical brain volume reduction of 35 has beenobserved in premature infants treated with dexamethasone[33] On the other hand there have been reports of no effectof single betamethasone antenatal dose on regional brainvolumes particularly within the limbic system [34]

In the present study the hippocampus was significantlyaffected by AGT in line with the theory that this regiondevelops primarily during the foetal period [35] althoughhippocampal volume losses have also been observed fol-lowing administration of corticosteroids in adult monkeysand rats [36 37] No changes were seen in the basolateralamygdala and nucleus accumbens volume which suggests aregion-selective treatment effect consistent with the densityof glucocorticoid receptor distribution [28]

A transient reduction in hippocampal volume has beenobserved in adolescent but not adult mice exposed to a clin-ically relevant dose of dexamethasone at E15 [18] The extentof hippocampal volume loss is normally mildmoderate Thepresent reduction of the hippocampal volume by 8 fits inthat range while being statistically significant It is compara-ble with some previous reports on stresssteroid-dependentlosses in the hippocampal volume across several speciesThusCoe et al [10] have found hippocampal volume reductions by12 and 10 in the offspring of rhesus monkeys exposed toearly life stress In tree shrews four weeksrsquo cortisol treatmentor psychosocial stress has resulted in a hippocampal volumeloss by 5ndash10 [38] Chronic social defeat stress has led toa similarly mild yet significant hippocampal volume lossin adult rats [6] Human adolescents who were born verypreterm have been found to have a hippocampal volume lossin the range of 12 to 156 [39]

What causes the hippocampal volume losses in responsesto stressglucocorticoids There is a range of factors such asneuronal andor glial cell death reduction of cell size includ-ing atrophy of neuronal dendrites andor glial branchesreduced water content or impaired neurogenesis [1] andorreduced microvasculature [6] The present study did not findcell losses in the brain areas tested and focused on astrogliabecause of their major role in maintaining brain plasticityat the level of synapses and cells [25] Currently published

studies on the mechanisms underlying glucocorticoid-induced brain volume reductions have focused on neuronsAs a result neuronal cell loss has been well researched anddocumented and various dexamethasone treatments havebeen found to lead to neuronal loss in rats [40 41] and pri-mates [26 37] mostly in the hippocampal CA3 and dentategyrus Less attention has been given to astroglial cells despitetheir role in the maintenance of brain homeostasis andplasticity [25]

The impoverishment in astroglial morphology reportedin the present study is consistent with the known glia-inhibitory effects of exogenous glucocorticoids or stress invitro [42 43] and in vivo [44] Treatment with corticosteroneor synthetic glucocorticoids such as dexamethasone has beenknown to inhibit GFAP expression in the neonatal andadult rat brain [45ndash48] thus suggesting that astrocytes canbe affected by glucocorticoid overload The very fact thatdexamethasone can affect GFAP expression raises a validmethodological consideration as to what extent the presentobservations of changes in the morphology of GFAP-positiveastroglia are affected bymethodological biaswhen comparingbetween the groups Central astroglia are heterogeneous[25 49] and using GFAP as the only immunohistochemicalmarker brings in an intrinsic limitation shared by many cur-rent studies as critically reviewed elsewhere [49] Howeverthe present GFAP responses to AGT appear to be selective tothe hippocampus where astroglial primary branches reducedin length but no GFAP-positive cell losses were found Sucha discrete response of GFAP immunoreactivity implies thatdexamethasone administered at the present lowdose asAGTdid not induce a global effect in GFAP expression Thisnot only highlights the unique susceptibility of hippocampalregions but also suggests that the changes observed here aretrue manifestations of impoverished hippocampal astroglialmorphology in response to AGT

It is increasingly evident that nongeneticenvironmentalfactors acting in early life result in long-term alterations ofthe physiological systems affected Perinatal programmingandor plasticity of the physiological system under extremeconditions such as ldquotoxicrdquo stress or AGT can result in long-term neurobehavioural abnormalities [3 29 50] and there isgrowing evidence that prenatal or early postnatal exposure tostressors affects the long-term brain functioning [51]

To conclude this study reports significant maladaptivechanges in astroglia in the hippocampal subregions CA1 CA3and dentate gyrus as a long-term effect of AGT It is plausibleto assume that the reductions in astrocytic primary branchescontribute to the losses in the hippocampal volume Theyare also likely to affect astroglial function and neuron-gliainteractions as retraction of primary astroglial processes candisadvantage local networks formed with glia and neuronsThe observed dexamethasone-induced impoverishment ofastroglial morphology can be treated as a demonstration ofglial maladaptive plasticity a phenomenon that deservesmore research in the context of effects of corticosteroidoverload and stress-related pathologies Epigenetics of thisphenomenon should be of interest considering the clinicaluse of dexamethasone

Neural Plasticity 7

Abbreviations

AGT Antenatal glucocorticoid treatmentDex DexamethasoneGD Gestational dayGFAP Glial fibrillary acidic proteinHX HematoxylinMDD Major depressive disorderPBS Phosphate-buffered salinePTSD Posttraumatic stress disorderROI Region of interest

Conflict of Interests

The authors declare that there is no conflict of interestsregarding the publication of this paper

Acknowledgments

This work was supported by the Wellcome Trust Grant (no086871Z08Z) to G E Gillies Vishvesh H Shende was aPhD student sponsored by the University of RoehamptonThe authors thank Dr Todd Rae (Roehampton) for trainingVishvesh H Shende in the pQCT technique

References

[1] B Czeh and P J Lucassen ldquoWhat causes the hippocampalvolume decrease in depression Are neurogenesis glial changesand apoptosis implicatedrdquo European Archives of Psychiatry andClinical Neuroscience vol 257 no 5 pp 250ndash260 2007

[2] P J Lucassen J Pruessner N Sousa et al ldquoNeuropathology ofstressrdquo Acta Neuropathologica vol 127 no 1 pp 109ndash135 2014

[3] B S McEwen L Eiland R G Hunter andMMMiller ldquoStressand anxiety structural plasticity and epigenetic regulation as aconsequence of stressrdquo Neuropharmacology vol 62 no 1 pp3ndash12 2012

[4] M A Mehta N I Golembo C Nosarti et al ldquoAmygdala hip-pocampal and corpus callosum size following severe earlyinstitutional deprivation the English and Romanian Adopteesstudy pilotrdquo Journal of Child Psychology and Psychiatry andAllied Disciplines vol 50 no 8 pp 943ndash951 2009

[5] T Frodl and V OrsquoKeane ldquoHow does the brain deal with cumu-lative stress A review with focus on developmental stress HPAaxis function and hippocampal structure in humansrdquo Neurobi-ology of Disease vol 52 pp 24ndash37 2013

[6] B Czeh N Abumaria R Rygula and E Fuchs ldquoQuantita-tive changes in hippocampal microvasculature of chronicallystressed rats no effect of fluoxetine treatmentrdquo Hippocampusvol 20 no 1 pp 174ndash185 2010

[7] V H Shende C R Pryce D Ruedi-Bettschen T C Rae andJ Opacka-Juffry ldquoEvidence for hippocampal remodeling as along term effect of early life stress in the rat model of earlydeprivationrdquo Recent Developments in Brain Research vol 1 pp1ndash19 2012

[8] A Jackowski T D Perera C G Abdallah et al ldquoEarly-lifestress corpus callosum development hippocampal volumet-rics and anxious behavior in male nonhuman primatesrdquo Psy-chiatry ResearchmdashNeuroimaging vol 192 no 1 pp 37ndash44 2011

[9] C Pittenger and R S Duman ldquoStress depression and neuro-plasticity a convergence of mechanismsrdquoNeuropsychopharma-cology vol 33 no 1 pp 88ndash109 2008

[10] C L Coe M Kramer B Czeh et al ldquoPrenatal stress diminishesneurogenesis in the dentate gyrus of juvenile Rhesus monkeysrdquoBiological Psychiatry vol 54 no 10 pp 1025ndash1034 2003

[11] J D Funkhouser K J Peevey P B Mockridge and E RHughes ldquoDistribution of dexamethasone between mother andfetus after maternal administrationrdquo Pediatric Research vol 12no 11 pp 1053ndash1056 1978

[12] D Roberts and S Dalziel ldquoAntenatal corticosteroids for accel-erating fetal lung maturation for women at risk of pretermbirthrdquo Cochrane Database of Systematic Reviews vol 3 ArticleID CD004454 2006

[13] T F Yeh Y J Lin H C Lin et al ldquoOutcomes at school ageafter postnatal dexamethasone therapy for lung disease of pre-maturityrdquoTheNew England Journal of Medicine vol 350 no 13pp 1304ndash1313 2004

[14] J P Empana M M Anceschi I Szabo E V Cosmi G Breartand P Truffert ldquoAntenatal corticosteroids policies in 14 Euro-pean countries factors associated with multiple courses TheEURAIL surveyrdquo Acta Paediatrica International Journal of Pae-diatrics vol 93 no 10 pp 1318ndash1322 2004

[15] C Romagnoli E Zecca R Luciano G Torrioli and G Tor-torolo ldquoA three year follow up of preterm infants after moder-ately early treatment with dexamethasonerdquo Archives of Diseasein Childhood Fetal and Neonatal Edition vol 87 no 1 pp F55ndashF58 2002

[16] A Charil D P Laplante C Vaillancourt and S King ldquoPrenatalstress and brain developmentrdquo Brain Research Reviews vol 65no 1 pp 56ndash79 2010

[17] D Tijsseling L D E Wijnberger J B Derks et al ldquoEffects ofantenatal glucocorticoid therapy on hippocampal histology ofpreterm infantsrdquoPLoSONE vol 7 no 3 Article ID e33369 2012

[18] C W Noorlander D Tijsseling E V S Hessel et al ldquoAntenatalglucocorticoid treatment affects hippocampal development inmicerdquo PLoS ONE vol 9 no 1 Article ID e85671 2014

[19] S McArthur E McHale and G E Gillies ldquoThe size and distri-bution of midbrain dopaminergic populations are permanentlyaltered by perinatal glucocorticoid exposure in a sex- region-and time-specific mannerrdquo Neuropsychopharmacology vol 32no 7 pp 1462ndash1476 2007

[20] S McArthur E McHale J W Dalley J C Buckingham andG E Gillies ldquoAlteredmesencephalic dopaminergic populationsin adulthood as a consequence of brief perinatal glucocorticoidexposurerdquo Journal of Neuroendocrinology vol 17 no 8 pp 475ndash482 2005

[21] ETheogaraj CD JohnH C Christian J FMorris S F Smithand J C Buckingham ldquoPerinatal glucocorticoid treatmentproduces molecular functional and morphological changes inthe anterior pituitary gland of the adultmale ratrdquoEndocrinologyvol 146 no 11 pp 4804ndash4813 2005

[22] G Paxinos and C WatsonThe Rat Brain in Stereotaxic Coordi-nates Elsevier Academic Press 5th edition 2005

[23] M Leventopoulos D Ruedi-Bettschen I Knuesel J Feldon CR Pryce and J Opacka-Juffry ldquoLong-term effects of early lifedeprivation on brain glia in Fischer ratsrdquo Brain Research vol1142 no 1 pp 119ndash126 2007

[24] M J West L Slomianka and H J G Gundersen ldquoUnbiasedstereological estimation of the total number of neurons in

8 Neural Plasticity

the subdivisions of the rat hippocampus using the optical frac-tionatorrdquo Anatomical Record vol 231 no 4 pp 482ndash497 1991

[25] D T Theodosis D A Poulain and S H R Oliet ldquoActivity-dependent structural and functional plasticity of astrocyte-neuron interactionsrdquo Physiological Reviews vol 88 no 3 pp983ndash1008 2008

[26] H Uno L Lohmiller C Thieme et al ldquoBrain damage inducedby prenatal exposure to dexamethasone in fetal rhesus maca-ques I Hippocampusrdquo Developmental Brain Research vol 53no 2 pp 157ndash167 1990

[27] R Q Carlos F J Seidler and T A Slotkin ldquoFetal dexametha-sone exposure altersmacromolecular characteristics of rat braindevelopment a critical period for regionally selective altera-tionsrdquo Teratology vol 46 no 1 pp 45ndash59 1992

[28] E R de Kloet ldquoStress in the brainrdquo European Journal of Phar-macology vol 405 pp 187ndash198 2000

[29] S G Matthews ldquoAntenatal glucocorticoids and the developingbrain mechanisms of actionrdquo Seminars in Neonatology vol 6no 4 pp 309ndash317 2001

[30] L J Abernethy M Palaniappan and R W I Cooke ldquoQuanti-tative magnetic resonance imaging of the brain in survivors ofvery low birth weightrdquo Archives of Disease in Childhood vol 87no 4 pp 279ndash283 2002

[31] E B Isaacs A LucasW K Chong et al ldquoHippocampal volumeand everyday memory in children of very low birth weightrdquoPediatric Research vol 47 no 6 pp 713ndash720 2000

[32] B S Peterson B Vohr L H Staib et al ldquoRegional brain volumeabnormalities and long-term cognitive outcome in preterminfantsrdquo Journal of the American Medical Association vol 284no 15 pp 1939ndash1947 2000

[33] B P Murphy T E Inder P S Huppi et al ldquoImpaired cerebralcortical graymatter growth after treatmentwith dexamethasonefor neonatal chronic lung diseaserdquo Pediatrics vol 107 no 2 pp217ndash221 2001

[34] P Yossuck M Kraszpulski and A K Salm ldquoPerinatal corti-costeroid effect on amygdala and hippocampus volume duringbrain development in the rat modelrdquo Early Human Develop-ment vol 82 no 4 pp 267ndash272 2006

[35] D Rice and S Barone Jr ldquoCritical periods of vulnerability forthe developing nervous system evidence from humans andanimal modelsrdquo Environmental Health Perspectives vol 108 no3 pp 511ndash533 2000

[36] C S Woolley E Gould and B S McEwen ldquoExposure toexcess glucocorticoids alters dendritic morphology of adulthippocampal pyramidal neuronsrdquo Brain Research vol 531 no1-2 pp 225ndash231 1990

[37] R M Sapolsky H Uno C S Rebert and C E Finch ldquoHippo-campal damage associatedwith prolonged glucocorticoid expo-sure in primatesrdquoThe Journal of Neuroscience vol 10 no 9 pp2897ndash2902 1990

[38] F Ohl T Michaelis G K Vollmann-Honsdorf C Kirschbaumand E Fuchs ldquoEffect of chronic psychosocial stress and long-term cortisol treatment on hippocampus-mediated memoryand hippocampal volume a pilot-study in tree shrewsrdquo Psy-choneuroendocrinology vol 25 no 4 pp 357ndash363 2000

[39] C Nosarti M H S Al-Asady S Frangou A L Stewart L Rif-kin and R M Murray ldquoAdolescents who were born very pre-term have decreased brain volumesrdquo Brain vol 125 no 7 pp1616ndash1623 2002

[40] L E Haynes M R Griffiths R E Hyde D J Barber and I JMitchell ldquoDexamethasone induces limited apoptosis and exten-sive sublethal damage to specific subregions of the striatum andhippocampus implications for mood disordersrdquo Neurosciencevol 104 no 1 pp 57ndash69 2001

[41] N Sousa M M Paula-Barbosa and O F X Almeida ldquoLigandand subfield specificity of corticoid-induced neuronal loss inthe rat hippocampal formationrdquoNeuroscience vol 89 no 4 pp1079ndash1087 1999

[42] K L Crossin M-H Tai L A Krushel V P Mauro and G MEdelman ldquoGlucocorticoid receptor pathways are involved in theinhibition of astrocyte proliferationrdquo Proceedings of the NationalAcademy of Sciences of the United States of America vol 94 no6 pp 2687ndash2692 1997

[43] M Sabolek A Herborg J Schwarz and A Storch ldquoDexam-ethasone blocks astroglial differentiation from neural precursorcellsrdquo NeuroReport vol 17 no 16 pp 1719ndash1723 2006

[44] N R Nichols J N Masters and C E Finch ldquoChanges in geneexpression in hippocampus in response to glucocorticoids andstressrdquo Brain Research Bulletin vol 24 no 5 pp 659ndash662 1990

[45] N R Nichols H H Osterburg J N Masters S L Millarand C E Finch ldquoMessenger RNA for glial fibrillary acidicprotein is decreased in rat brain following acute and chroniccorticosterone treatmentrdquo Molecular Brain Research vol 7 no1 pp 1ndash7 1990

[46] N J Laping N R Nichols J R Day and C E Finch ldquoCortico-sterone differentially regulates the bilateral response of astro-cyte mRNAs in the hippocampus to entorhinal cortex lesions inmale ratsrdquoMolecular Brain Research vol 10 no 4 pp 291ndash2971991

[47] S Tsuneishi S Takada T Motoike T Ohashi K Sano andH Nakamura ldquoEffects of dexamethasone on the expression ofmyelin basic protein proteolipid protein and glial fibrillaryacidic protein genes in developing rat brainrdquo DevelopmentalBrain Research vol 61 no 1 pp 117ndash123 1991

[48] J P OrsquoCallaghan R E Brinton and B S McEwen ldquoGlucocor-ticoids regulate the synthesis of glial fibrillary acidic protein inintact and adrenalectomized rats but do not affect its expressionfollowing brain injuryrdquo Journal of Neurochemistry vol 57 no 3pp 860ndash869 1991

[49] M V Sofroniew and H V Vinters ldquoAstrocytes biology andpathologyrdquoActa Neuropathologica vol 119 no 1 pp 7ndash35 2010

[50] S Maccari M Darnaudery S Morley-Fletcher A R Zuena CCinque and O van Reeth ldquoPrenatal stress and long-termconsequences implications of glucocorticoid hormonesrdquo Neu-roscience and Biobehavioral Reviews vol 27 no 1-2 pp 119ndash1272003

[51] C R Pryce Y Aubert CMaier P C Pearce and E Fuchs ldquoThedevelopmental impact of prenatal stress prenatal dexametha-sone and postnatal social stress on physiology behaviour andneuroanatomy of primate offspring Studies in rhesus macaqueand common marmosetrdquo Psychopharmacology vol 214 no 1pp 33ndash53 2011

Submit your manuscripts athttpwwwhindawicom

Neurology Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Alzheimerrsquos DiseaseHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

ScientificaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Research and TreatmentSchizophrenia

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Neural Plasticity

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Parkinsonrsquos Disease

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Research and TreatmentAutism

Sleep DisordersHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Neuroscience Journal

Epilepsy Research and TreatmentHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Psychiatry Journal

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Computational and Mathematical Methods in Medicine

Depression Research and TreatmentHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Brain ScienceInternational Journal of

StrokeResearch and TreatmentHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Neurodegenerative Diseases

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Journal of

Cardiovascular Psychiatry and NeurologyHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Neural Plasticity 5

Table 6 Effects of AGT on GFAP-positive astroglial primaryprocess numbers in male adult Sprague Dawley rats

GFAP-astroglia morphologyControl Dex-treated

Number of primary processCA1 362 plusmn 361 3233 plusmn 44CA2 301 plusmn 262 311 plusmn 264CA3 339 plusmn 169 3057 plusmn 81DG 3217 plusmn 143 3148 plusmn 47

The numbers of primary processes of GFAP-positive cells in control anddexamethasone-treated groups (119899= 6 per group) Data are expressed asmeanplusmn SEM No significant differences between Dex-treated (AGT) and controlwere observed

25 in DG (119875 lt 001) (Figure 2) AGT-dependent changeswere also observed in the mean length of astrocytic primaryprocesses reductions by 25 in CA1 (119875 lt 001) 45 in CA3(119875 lt 0001) and 25 in DG (119875 lt 001) (Figure 3)

4 Discussion

The present study was focused on the effects of AGT oncentral astroglia and regional volume of the hippocampusamygdala and nucleus accumbens as the areas of the limbicsystem implicated in response to stress and corticosteroids [23 9] AGTwas administered to pregnant dams noninvasivelythrough drinkingwater to avoid the stress of animal handlingand injections Of the brain areas tested in adult male rats thehippocampus responded to AGT with a long-term reductionin its volume and changes in astroglial morphology It is plau-sible that the observed impoverishment of astrocytic primarybranches underpins the hippocampal volume reduction aswe found no evidence of cell loss To our knowledge suchAGT-dependent long-term hippocampal remodelling withimplications for astroglia has not been reported before

Astroglia were visualised by means of the commonlyused GFAP immunohistochemistry GFAP is an intermediatefilament protein present in cell bodies and primary branchesalthough fine distal processes show little or no GFAP [25]Although GFAP staining is selective to astrocytes it does notlabel all astrocytes in the brain Accepting these limitationsof GFAP immunohistochemistry the present study focusedon the length and numbers of primary astrocytic processeswhere GFAP is reliably detectable [25] Stereology was ourmethod of choice as it offers an unbiased approach to regionalbrain volumetry and morphometry [24]

Previous volumetric studies have considered specificbrain regions expressing their volumes in absolute termswith no reference to the total brain volume which itself isa variable Thus the present application of pQCT to measuretotal brain volume adds the much needed rigour to volu-metric assessment where regional volumes need to be scaledto the total brain volume This is particularly important asperinatal corticosteroid exposure can lead to a decrease inbrain weightvolume For example a significant brain weightreduction has been reported in rhesus monkeys treatedprenatally with corticosteroids [26] and repeated doses of

lowast

lowastlowast

lowastlowastlowast

1200000

1100000

1000000

900000

800000

700000

600000

500000

400000

300000

200000

100000

000

CA1 CA2 CA3 DG

TreatmentControlDEX

Mea

n pr

imar

y pr

oces

s tot

al le

ngth

(120583m

)

Dorsal hippocampal subregions

Figure 2 Effect of prenatal dexamethasone treatment on the totallength of primary astrocytic processes in hippocampal subregionsof male adult Sprague Dawley rats The total primary process lengthofGFAP-positive cells in control and dexamethasone-treated groups(119899 = 6 per group) Data are expressed as mean plusmn SEM lowast119875 lt 005lowastlowast

119875 lt 001 and lowastlowastlowast119875 lt 0001 versus control independent sample119905-test

lowastlowast

lowastlowast

lowastlowastlowast

500

000

1000

1500

2000

2500

3000

CA1 CA2 CA3 DG

TreatmentControlDEX

Mea

n pr

imar

y pr

oces

s mea

n le

ngth

(120583m

)

Dorsal hippocampal subregions

Figure 3 Effect of prenatal dexamethasone treatment on the meanlength of primary astrocytic processes in hippocampal subregions ofmale adult Sprague Dawley rats The mean primary process lengthofGFAP-positive cells in control and dexamethasone-treated groups(119899 = 6 per group) Data are expressed as mean plusmn SEM lowastlowast119875 lt 001and lowastlowastlowast119875 lt 0001 versus control independent sample 119905-test

6 Neural Plasticity

dexamethasone at E17ndash19 have resulted in a significant brainweight reduction in newborn rats [27] The present studyfound no AGT effects on the whole brain weight and volumethe latter measured by pQCT One of the factors which affectthe brain weight and volume is the amount of corticosteroidadministered In the present study a relatively small dosewas given which as discussed previously [19] is in the rangeused in perinatal medicine and mimics the glucocorticoidactivity that is required for normal lung maturation duringlate gestation

Within the developing brain the limbic system richlyexpressing glucocorticoid and mineralocorticoid receptors[28] is sensitive to the endogenous and exogenous corti-costeroids which can affect these brain areas with long-term consequences for their structure and function [29]Reductions in the regional brain volume of the hippocampusand amygdala have been reported in preterm infants [30ndash32] and a cortical brain volume reduction of 35 has beenobserved in premature infants treated with dexamethasone[33] On the other hand there have been reports of no effectof single betamethasone antenatal dose on regional brainvolumes particularly within the limbic system [34]

In the present study the hippocampus was significantlyaffected by AGT in line with the theory that this regiondevelops primarily during the foetal period [35] althoughhippocampal volume losses have also been observed fol-lowing administration of corticosteroids in adult monkeysand rats [36 37] No changes were seen in the basolateralamygdala and nucleus accumbens volume which suggests aregion-selective treatment effect consistent with the densityof glucocorticoid receptor distribution [28]

A transient reduction in hippocampal volume has beenobserved in adolescent but not adult mice exposed to a clin-ically relevant dose of dexamethasone at E15 [18] The extentof hippocampal volume loss is normally mildmoderate Thepresent reduction of the hippocampal volume by 8 fits inthat range while being statistically significant It is compara-ble with some previous reports on stresssteroid-dependentlosses in the hippocampal volume across several speciesThusCoe et al [10] have found hippocampal volume reductions by12 and 10 in the offspring of rhesus monkeys exposed toearly life stress In tree shrews four weeksrsquo cortisol treatmentor psychosocial stress has resulted in a hippocampal volumeloss by 5ndash10 [38] Chronic social defeat stress has led toa similarly mild yet significant hippocampal volume lossin adult rats [6] Human adolescents who were born verypreterm have been found to have a hippocampal volume lossin the range of 12 to 156 [39]

What causes the hippocampal volume losses in responsesto stressglucocorticoids There is a range of factors such asneuronal andor glial cell death reduction of cell size includ-ing atrophy of neuronal dendrites andor glial branchesreduced water content or impaired neurogenesis [1] andorreduced microvasculature [6] The present study did not findcell losses in the brain areas tested and focused on astrogliabecause of their major role in maintaining brain plasticityat the level of synapses and cells [25] Currently published

studies on the mechanisms underlying glucocorticoid-induced brain volume reductions have focused on neuronsAs a result neuronal cell loss has been well researched anddocumented and various dexamethasone treatments havebeen found to lead to neuronal loss in rats [40 41] and pri-mates [26 37] mostly in the hippocampal CA3 and dentategyrus Less attention has been given to astroglial cells despitetheir role in the maintenance of brain homeostasis andplasticity [25]

The impoverishment in astroglial morphology reportedin the present study is consistent with the known glia-inhibitory effects of exogenous glucocorticoids or stress invitro [42 43] and in vivo [44] Treatment with corticosteroneor synthetic glucocorticoids such as dexamethasone has beenknown to inhibit GFAP expression in the neonatal andadult rat brain [45ndash48] thus suggesting that astrocytes canbe affected by glucocorticoid overload The very fact thatdexamethasone can affect GFAP expression raises a validmethodological consideration as to what extent the presentobservations of changes in the morphology of GFAP-positiveastroglia are affected bymethodological biaswhen comparingbetween the groups Central astroglia are heterogeneous[25 49] and using GFAP as the only immunohistochemicalmarker brings in an intrinsic limitation shared by many cur-rent studies as critically reviewed elsewhere [49] Howeverthe present GFAP responses to AGT appear to be selective tothe hippocampus where astroglial primary branches reducedin length but no GFAP-positive cell losses were found Sucha discrete response of GFAP immunoreactivity implies thatdexamethasone administered at the present lowdose asAGTdid not induce a global effect in GFAP expression Thisnot only highlights the unique susceptibility of hippocampalregions but also suggests that the changes observed here aretrue manifestations of impoverished hippocampal astroglialmorphology in response to AGT

It is increasingly evident that nongeneticenvironmentalfactors acting in early life result in long-term alterations ofthe physiological systems affected Perinatal programmingandor plasticity of the physiological system under extremeconditions such as ldquotoxicrdquo stress or AGT can result in long-term neurobehavioural abnormalities [3 29 50] and there isgrowing evidence that prenatal or early postnatal exposure tostressors affects the long-term brain functioning [51]

To conclude this study reports significant maladaptivechanges in astroglia in the hippocampal subregions CA1 CA3and dentate gyrus as a long-term effect of AGT It is plausibleto assume that the reductions in astrocytic primary branchescontribute to the losses in the hippocampal volume Theyare also likely to affect astroglial function and neuron-gliainteractions as retraction of primary astroglial processes candisadvantage local networks formed with glia and neuronsThe observed dexamethasone-induced impoverishment ofastroglial morphology can be treated as a demonstration ofglial maladaptive plasticity a phenomenon that deservesmore research in the context of effects of corticosteroidoverload and stress-related pathologies Epigenetics of thisphenomenon should be of interest considering the clinicaluse of dexamethasone

Neural Plasticity 7

Abbreviations

AGT Antenatal glucocorticoid treatmentDex DexamethasoneGD Gestational dayGFAP Glial fibrillary acidic proteinHX HematoxylinMDD Major depressive disorderPBS Phosphate-buffered salinePTSD Posttraumatic stress disorderROI Region of interest

Conflict of Interests

The authors declare that there is no conflict of interestsregarding the publication of this paper

Acknowledgments

This work was supported by the Wellcome Trust Grant (no086871Z08Z) to G E Gillies Vishvesh H Shende was aPhD student sponsored by the University of RoehamptonThe authors thank Dr Todd Rae (Roehampton) for trainingVishvesh H Shende in the pQCT technique

References

[1] B Czeh and P J Lucassen ldquoWhat causes the hippocampalvolume decrease in depression Are neurogenesis glial changesand apoptosis implicatedrdquo European Archives of Psychiatry andClinical Neuroscience vol 257 no 5 pp 250ndash260 2007

[2] P J Lucassen J Pruessner N Sousa et al ldquoNeuropathology ofstressrdquo Acta Neuropathologica vol 127 no 1 pp 109ndash135 2014

[3] B S McEwen L Eiland R G Hunter andMMMiller ldquoStressand anxiety structural plasticity and epigenetic regulation as aconsequence of stressrdquo Neuropharmacology vol 62 no 1 pp3ndash12 2012

[4] M A Mehta N I Golembo C Nosarti et al ldquoAmygdala hip-pocampal and corpus callosum size following severe earlyinstitutional deprivation the English and Romanian Adopteesstudy pilotrdquo Journal of Child Psychology and Psychiatry andAllied Disciplines vol 50 no 8 pp 943ndash951 2009

[5] T Frodl and V OrsquoKeane ldquoHow does the brain deal with cumu-lative stress A review with focus on developmental stress HPAaxis function and hippocampal structure in humansrdquo Neurobi-ology of Disease vol 52 pp 24ndash37 2013

[6] B Czeh N Abumaria R Rygula and E Fuchs ldquoQuantita-tive changes in hippocampal microvasculature of chronicallystressed rats no effect of fluoxetine treatmentrdquo Hippocampusvol 20 no 1 pp 174ndash185 2010

[7] V H Shende C R Pryce D Ruedi-Bettschen T C Rae andJ Opacka-Juffry ldquoEvidence for hippocampal remodeling as along term effect of early life stress in the rat model of earlydeprivationrdquo Recent Developments in Brain Research vol 1 pp1ndash19 2012

[8] A Jackowski T D Perera C G Abdallah et al ldquoEarly-lifestress corpus callosum development hippocampal volumet-rics and anxious behavior in male nonhuman primatesrdquo Psy-chiatry ResearchmdashNeuroimaging vol 192 no 1 pp 37ndash44 2011

[9] C Pittenger and R S Duman ldquoStress depression and neuro-plasticity a convergence of mechanismsrdquoNeuropsychopharma-cology vol 33 no 1 pp 88ndash109 2008

[10] C L Coe M Kramer B Czeh et al ldquoPrenatal stress diminishesneurogenesis in the dentate gyrus of juvenile Rhesus monkeysrdquoBiological Psychiatry vol 54 no 10 pp 1025ndash1034 2003

[11] J D Funkhouser K J Peevey P B Mockridge and E RHughes ldquoDistribution of dexamethasone between mother andfetus after maternal administrationrdquo Pediatric Research vol 12no 11 pp 1053ndash1056 1978

[12] D Roberts and S Dalziel ldquoAntenatal corticosteroids for accel-erating fetal lung maturation for women at risk of pretermbirthrdquo Cochrane Database of Systematic Reviews vol 3 ArticleID CD004454 2006

[13] T F Yeh Y J Lin H C Lin et al ldquoOutcomes at school ageafter postnatal dexamethasone therapy for lung disease of pre-maturityrdquoTheNew England Journal of Medicine vol 350 no 13pp 1304ndash1313 2004

[14] J P Empana M M Anceschi I Szabo E V Cosmi G Breartand P Truffert ldquoAntenatal corticosteroids policies in 14 Euro-pean countries factors associated with multiple courses TheEURAIL surveyrdquo Acta Paediatrica International Journal of Pae-diatrics vol 93 no 10 pp 1318ndash1322 2004

[15] C Romagnoli E Zecca R Luciano G Torrioli and G Tor-torolo ldquoA three year follow up of preterm infants after moder-ately early treatment with dexamethasonerdquo Archives of Diseasein Childhood Fetal and Neonatal Edition vol 87 no 1 pp F55ndashF58 2002

[16] A Charil D P Laplante C Vaillancourt and S King ldquoPrenatalstress and brain developmentrdquo Brain Research Reviews vol 65no 1 pp 56ndash79 2010

[17] D Tijsseling L D E Wijnberger J B Derks et al ldquoEffects ofantenatal glucocorticoid therapy on hippocampal histology ofpreterm infantsrdquoPLoSONE vol 7 no 3 Article ID e33369 2012

[18] C W Noorlander D Tijsseling E V S Hessel et al ldquoAntenatalglucocorticoid treatment affects hippocampal development inmicerdquo PLoS ONE vol 9 no 1 Article ID e85671 2014

[19] S McArthur E McHale and G E Gillies ldquoThe size and distri-bution of midbrain dopaminergic populations are permanentlyaltered by perinatal glucocorticoid exposure in a sex- region-and time-specific mannerrdquo Neuropsychopharmacology vol 32no 7 pp 1462ndash1476 2007

[20] S McArthur E McHale J W Dalley J C Buckingham andG E Gillies ldquoAlteredmesencephalic dopaminergic populationsin adulthood as a consequence of brief perinatal glucocorticoidexposurerdquo Journal of Neuroendocrinology vol 17 no 8 pp 475ndash482 2005

[21] ETheogaraj CD JohnH C Christian J FMorris S F Smithand J C Buckingham ldquoPerinatal glucocorticoid treatmentproduces molecular functional and morphological changes inthe anterior pituitary gland of the adultmale ratrdquoEndocrinologyvol 146 no 11 pp 4804ndash4813 2005

[22] G Paxinos and C WatsonThe Rat Brain in Stereotaxic Coordi-nates Elsevier Academic Press 5th edition 2005

[23] M Leventopoulos D Ruedi-Bettschen I Knuesel J Feldon CR Pryce and J Opacka-Juffry ldquoLong-term effects of early lifedeprivation on brain glia in Fischer ratsrdquo Brain Research vol1142 no 1 pp 119ndash126 2007

[24] M J West L Slomianka and H J G Gundersen ldquoUnbiasedstereological estimation of the total number of neurons in

8 Neural Plasticity

the subdivisions of the rat hippocampus using the optical frac-tionatorrdquo Anatomical Record vol 231 no 4 pp 482ndash497 1991

[25] D T Theodosis D A Poulain and S H R Oliet ldquoActivity-dependent structural and functional plasticity of astrocyte-neuron interactionsrdquo Physiological Reviews vol 88 no 3 pp983ndash1008 2008

[26] H Uno L Lohmiller C Thieme et al ldquoBrain damage inducedby prenatal exposure to dexamethasone in fetal rhesus maca-ques I Hippocampusrdquo Developmental Brain Research vol 53no 2 pp 157ndash167 1990

[27] R Q Carlos F J Seidler and T A Slotkin ldquoFetal dexametha-sone exposure altersmacromolecular characteristics of rat braindevelopment a critical period for regionally selective altera-tionsrdquo Teratology vol 46 no 1 pp 45ndash59 1992

[28] E R de Kloet ldquoStress in the brainrdquo European Journal of Phar-macology vol 405 pp 187ndash198 2000

[29] S G Matthews ldquoAntenatal glucocorticoids and the developingbrain mechanisms of actionrdquo Seminars in Neonatology vol 6no 4 pp 309ndash317 2001

[30] L J Abernethy M Palaniappan and R W I Cooke ldquoQuanti-tative magnetic resonance imaging of the brain in survivors ofvery low birth weightrdquo Archives of Disease in Childhood vol 87no 4 pp 279ndash283 2002

[31] E B Isaacs A LucasW K Chong et al ldquoHippocampal volumeand everyday memory in children of very low birth weightrdquoPediatric Research vol 47 no 6 pp 713ndash720 2000

[32] B S Peterson B Vohr L H Staib et al ldquoRegional brain volumeabnormalities and long-term cognitive outcome in preterminfantsrdquo Journal of the American Medical Association vol 284no 15 pp 1939ndash1947 2000

[33] B P Murphy T E Inder P S Huppi et al ldquoImpaired cerebralcortical graymatter growth after treatmentwith dexamethasonefor neonatal chronic lung diseaserdquo Pediatrics vol 107 no 2 pp217ndash221 2001

[34] P Yossuck M Kraszpulski and A K Salm ldquoPerinatal corti-costeroid effect on amygdala and hippocampus volume duringbrain development in the rat modelrdquo Early Human Develop-ment vol 82 no 4 pp 267ndash272 2006

[35] D Rice and S Barone Jr ldquoCritical periods of vulnerability forthe developing nervous system evidence from humans andanimal modelsrdquo Environmental Health Perspectives vol 108 no3 pp 511ndash533 2000

[36] C S Woolley E Gould and B S McEwen ldquoExposure toexcess glucocorticoids alters dendritic morphology of adulthippocampal pyramidal neuronsrdquo Brain Research vol 531 no1-2 pp 225ndash231 1990

[37] R M Sapolsky H Uno C S Rebert and C E Finch ldquoHippo-campal damage associatedwith prolonged glucocorticoid expo-sure in primatesrdquoThe Journal of Neuroscience vol 10 no 9 pp2897ndash2902 1990

[38] F Ohl T Michaelis G K Vollmann-Honsdorf C Kirschbaumand E Fuchs ldquoEffect of chronic psychosocial stress and long-term cortisol treatment on hippocampus-mediated memoryand hippocampal volume a pilot-study in tree shrewsrdquo Psy-choneuroendocrinology vol 25 no 4 pp 357ndash363 2000

[39] C Nosarti M H S Al-Asady S Frangou A L Stewart L Rif-kin and R M Murray ldquoAdolescents who were born very pre-term have decreased brain volumesrdquo Brain vol 125 no 7 pp1616ndash1623 2002

[40] L E Haynes M R Griffiths R E Hyde D J Barber and I JMitchell ldquoDexamethasone induces limited apoptosis and exten-sive sublethal damage to specific subregions of the striatum andhippocampus implications for mood disordersrdquo Neurosciencevol 104 no 1 pp 57ndash69 2001

[41] N Sousa M M Paula-Barbosa and O F X Almeida ldquoLigandand subfield specificity of corticoid-induced neuronal loss inthe rat hippocampal formationrdquoNeuroscience vol 89 no 4 pp1079ndash1087 1999

[42] K L Crossin M-H Tai L A Krushel V P Mauro and G MEdelman ldquoGlucocorticoid receptor pathways are involved in theinhibition of astrocyte proliferationrdquo Proceedings of the NationalAcademy of Sciences of the United States of America vol 94 no6 pp 2687ndash2692 1997

[43] M Sabolek A Herborg J Schwarz and A Storch ldquoDexam-ethasone blocks astroglial differentiation from neural precursorcellsrdquo NeuroReport vol 17 no 16 pp 1719ndash1723 2006

[44] N R Nichols J N Masters and C E Finch ldquoChanges in geneexpression in hippocampus in response to glucocorticoids andstressrdquo Brain Research Bulletin vol 24 no 5 pp 659ndash662 1990

[45] N R Nichols H H Osterburg J N Masters S L Millarand C E Finch ldquoMessenger RNA for glial fibrillary acidicprotein is decreased in rat brain following acute and chroniccorticosterone treatmentrdquo Molecular Brain Research vol 7 no1 pp 1ndash7 1990

[46] N J Laping N R Nichols J R Day and C E Finch ldquoCortico-sterone differentially regulates the bilateral response of astro-cyte mRNAs in the hippocampus to entorhinal cortex lesions inmale ratsrdquoMolecular Brain Research vol 10 no 4 pp 291ndash2971991

[47] S Tsuneishi S Takada T Motoike T Ohashi K Sano andH Nakamura ldquoEffects of dexamethasone on the expression ofmyelin basic protein proteolipid protein and glial fibrillaryacidic protein genes in developing rat brainrdquo DevelopmentalBrain Research vol 61 no 1 pp 117ndash123 1991

[48] J P OrsquoCallaghan R E Brinton and B S McEwen ldquoGlucocor-ticoids regulate the synthesis of glial fibrillary acidic protein inintact and adrenalectomized rats but do not affect its expressionfollowing brain injuryrdquo Journal of Neurochemistry vol 57 no 3pp 860ndash869 1991

[49] M V Sofroniew and H V Vinters ldquoAstrocytes biology andpathologyrdquoActa Neuropathologica vol 119 no 1 pp 7ndash35 2010

[50] S Maccari M Darnaudery S Morley-Fletcher A R Zuena CCinque and O van Reeth ldquoPrenatal stress and long-termconsequences implications of glucocorticoid hormonesrdquo Neu-roscience and Biobehavioral Reviews vol 27 no 1-2 pp 119ndash1272003

[51] C R Pryce Y Aubert CMaier P C Pearce and E Fuchs ldquoThedevelopmental impact of prenatal stress prenatal dexametha-sone and postnatal social stress on physiology behaviour andneuroanatomy of primate offspring Studies in rhesus macaqueand common marmosetrdquo Psychopharmacology vol 214 no 1pp 33ndash53 2011

Submit your manuscripts athttpwwwhindawicom

Neurology Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Alzheimerrsquos DiseaseHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

ScientificaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Research and TreatmentSchizophrenia

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Neural Plasticity

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Parkinsonrsquos Disease

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Research and TreatmentAutism

Sleep DisordersHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Neuroscience Journal

Epilepsy Research and TreatmentHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Psychiatry Journal

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Computational and Mathematical Methods in Medicine

Depression Research and TreatmentHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Brain ScienceInternational Journal of

StrokeResearch and TreatmentHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Neurodegenerative Diseases

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Journal of

Cardiovascular Psychiatry and NeurologyHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

6 Neural Plasticity

dexamethasone at E17ndash19 have resulted in a significant brainweight reduction in newborn rats [27] The present studyfound no AGT effects on the whole brain weight and volumethe latter measured by pQCT One of the factors which affectthe brain weight and volume is the amount of corticosteroidadministered In the present study a relatively small dosewas given which as discussed previously [19] is in the rangeused in perinatal medicine and mimics the glucocorticoidactivity that is required for normal lung maturation duringlate gestation

Within the developing brain the limbic system richlyexpressing glucocorticoid and mineralocorticoid receptors[28] is sensitive to the endogenous and exogenous corti-costeroids which can affect these brain areas with long-term consequences for their structure and function [29]Reductions in the regional brain volume of the hippocampusand amygdala have been reported in preterm infants [30ndash32] and a cortical brain volume reduction of 35 has beenobserved in premature infants treated with dexamethasone[33] On the other hand there have been reports of no effectof single betamethasone antenatal dose on regional brainvolumes particularly within the limbic system [34]

In the present study the hippocampus was significantlyaffected by AGT in line with the theory that this regiondevelops primarily during the foetal period [35] althoughhippocampal volume losses have also been observed fol-lowing administration of corticosteroids in adult monkeysand rats [36 37] No changes were seen in the basolateralamygdala and nucleus accumbens volume which suggests aregion-selective treatment effect consistent with the densityof glucocorticoid receptor distribution [28]

A transient reduction in hippocampal volume has beenobserved in adolescent but not adult mice exposed to a clin-ically relevant dose of dexamethasone at E15 [18] The extentof hippocampal volume loss is normally mildmoderate Thepresent reduction of the hippocampal volume by 8 fits inthat range while being statistically significant It is compara-ble with some previous reports on stresssteroid-dependentlosses in the hippocampal volume across several speciesThusCoe et al [10] have found hippocampal volume reductions by12 and 10 in the offspring of rhesus monkeys exposed toearly life stress In tree shrews four weeksrsquo cortisol treatmentor psychosocial stress has resulted in a hippocampal volumeloss by 5ndash10 [38] Chronic social defeat stress has led toa similarly mild yet significant hippocampal volume lossin adult rats [6] Human adolescents who were born verypreterm have been found to have a hippocampal volume lossin the range of 12 to 156 [39]

What causes the hippocampal volume losses in responsesto stressglucocorticoids There is a range of factors such asneuronal andor glial cell death reduction of cell size includ-ing atrophy of neuronal dendrites andor glial branchesreduced water content or impaired neurogenesis [1] andorreduced microvasculature [6] The present study did not findcell losses in the brain areas tested and focused on astrogliabecause of their major role in maintaining brain plasticityat the level of synapses and cells [25] Currently published

studies on the mechanisms underlying glucocorticoid-induced brain volume reductions have focused on neuronsAs a result neuronal cell loss has been well researched anddocumented and various dexamethasone treatments havebeen found to lead to neuronal loss in rats [40 41] and pri-mates [26 37] mostly in the hippocampal CA3 and dentategyrus Less attention has been given to astroglial cells despitetheir role in the maintenance of brain homeostasis andplasticity [25]

The impoverishment in astroglial morphology reportedin the present study is consistent with the known glia-inhibitory effects of exogenous glucocorticoids or stress invitro [42 43] and in vivo [44] Treatment with corticosteroneor synthetic glucocorticoids such as dexamethasone has beenknown to inhibit GFAP expression in the neonatal andadult rat brain [45ndash48] thus suggesting that astrocytes canbe affected by glucocorticoid overload The very fact thatdexamethasone can affect GFAP expression raises a validmethodological consideration as to what extent the presentobservations of changes in the morphology of GFAP-positiveastroglia are affected bymethodological biaswhen comparingbetween the groups Central astroglia are heterogeneous[25 49] and using GFAP as the only immunohistochemicalmarker brings in an intrinsic limitation shared by many cur-rent studies as critically reviewed elsewhere [49] Howeverthe present GFAP responses to AGT appear to be selective tothe hippocampus where astroglial primary branches reducedin length but no GFAP-positive cell losses were found Sucha discrete response of GFAP immunoreactivity implies thatdexamethasone administered at the present lowdose asAGTdid not induce a global effect in GFAP expression Thisnot only highlights the unique susceptibility of hippocampalregions but also suggests that the changes observed here aretrue manifestations of impoverished hippocampal astroglialmorphology in response to AGT

It is increasingly evident that nongeneticenvironmentalfactors acting in early life result in long-term alterations ofthe physiological systems affected Perinatal programmingandor plasticity of the physiological system under extremeconditions such as ldquotoxicrdquo stress or AGT can result in long-term neurobehavioural abnormalities [3 29 50] and there isgrowing evidence that prenatal or early postnatal exposure tostressors affects the long-term brain functioning [51]

To conclude this study reports significant maladaptivechanges in astroglia in the hippocampal subregions CA1 CA3and dentate gyrus as a long-term effect of AGT It is plausibleto assume that the reductions in astrocytic primary branchescontribute to the losses in the hippocampal volume Theyare also likely to affect astroglial function and neuron-gliainteractions as retraction of primary astroglial processes candisadvantage local networks formed with glia and neuronsThe observed dexamethasone-induced impoverishment ofastroglial morphology can be treated as a demonstration ofglial maladaptive plasticity a phenomenon that deservesmore research in the context of effects of corticosteroidoverload and stress-related pathologies Epigenetics of thisphenomenon should be of interest considering the clinicaluse of dexamethasone

Neural Plasticity 7

Abbreviations

AGT Antenatal glucocorticoid treatmentDex DexamethasoneGD Gestational dayGFAP Glial fibrillary acidic proteinHX HematoxylinMDD Major depressive disorderPBS Phosphate-buffered salinePTSD Posttraumatic stress disorderROI Region of interest

Conflict of Interests

The authors declare that there is no conflict of interestsregarding the publication of this paper

Acknowledgments

This work was supported by the Wellcome Trust Grant (no086871Z08Z) to G E Gillies Vishvesh H Shende was aPhD student sponsored by the University of RoehamptonThe authors thank Dr Todd Rae (Roehampton) for trainingVishvesh H Shende in the pQCT technique

References

[1] B Czeh and P J Lucassen ldquoWhat causes the hippocampalvolume decrease in depression Are neurogenesis glial changesand apoptosis implicatedrdquo European Archives of Psychiatry andClinical Neuroscience vol 257 no 5 pp 250ndash260 2007

[2] P J Lucassen J Pruessner N Sousa et al ldquoNeuropathology ofstressrdquo Acta Neuropathologica vol 127 no 1 pp 109ndash135 2014

[3] B S McEwen L Eiland R G Hunter andMMMiller ldquoStressand anxiety structural plasticity and epigenetic regulation as aconsequence of stressrdquo Neuropharmacology vol 62 no 1 pp3ndash12 2012

[4] M A Mehta N I Golembo C Nosarti et al ldquoAmygdala hip-pocampal and corpus callosum size following severe earlyinstitutional deprivation the English and Romanian Adopteesstudy pilotrdquo Journal of Child Psychology and Psychiatry andAllied Disciplines vol 50 no 8 pp 943ndash951 2009

[5] T Frodl and V OrsquoKeane ldquoHow does the brain deal with cumu-lative stress A review with focus on developmental stress HPAaxis function and hippocampal structure in humansrdquo Neurobi-ology of Disease vol 52 pp 24ndash37 2013

[6] B Czeh N Abumaria R Rygula and E Fuchs ldquoQuantita-tive changes in hippocampal microvasculature of chronicallystressed rats no effect of fluoxetine treatmentrdquo Hippocampusvol 20 no 1 pp 174ndash185 2010

[7] V H Shende C R Pryce D Ruedi-Bettschen T C Rae andJ Opacka-Juffry ldquoEvidence for hippocampal remodeling as along term effect of early life stress in the rat model of earlydeprivationrdquo Recent Developments in Brain Research vol 1 pp1ndash19 2012

[8] A Jackowski T D Perera C G Abdallah et al ldquoEarly-lifestress corpus callosum development hippocampal volumet-rics and anxious behavior in male nonhuman primatesrdquo Psy-chiatry ResearchmdashNeuroimaging vol 192 no 1 pp 37ndash44 2011

[9] C Pittenger and R S Duman ldquoStress depression and neuro-plasticity a convergence of mechanismsrdquoNeuropsychopharma-cology vol 33 no 1 pp 88ndash109 2008

[10] C L Coe M Kramer B Czeh et al ldquoPrenatal stress diminishesneurogenesis in the dentate gyrus of juvenile Rhesus monkeysrdquoBiological Psychiatry vol 54 no 10 pp 1025ndash1034 2003

[11] J D Funkhouser K J Peevey P B Mockridge and E RHughes ldquoDistribution of dexamethasone between mother andfetus after maternal administrationrdquo Pediatric Research vol 12no 11 pp 1053ndash1056 1978

[12] D Roberts and S Dalziel ldquoAntenatal corticosteroids for accel-erating fetal lung maturation for women at risk of pretermbirthrdquo Cochrane Database of Systematic Reviews vol 3 ArticleID CD004454 2006

[13] T F Yeh Y J Lin H C Lin et al ldquoOutcomes at school ageafter postnatal dexamethasone therapy for lung disease of pre-maturityrdquoTheNew England Journal of Medicine vol 350 no 13pp 1304ndash1313 2004

[14] J P Empana M M Anceschi I Szabo E V Cosmi G Breartand P Truffert ldquoAntenatal corticosteroids policies in 14 Euro-pean countries factors associated with multiple courses TheEURAIL surveyrdquo Acta Paediatrica International Journal of Pae-diatrics vol 93 no 10 pp 1318ndash1322 2004

[15] C Romagnoli E Zecca R Luciano G Torrioli and G Tor-torolo ldquoA three year follow up of preterm infants after moder-ately early treatment with dexamethasonerdquo Archives of Diseasein Childhood Fetal and Neonatal Edition vol 87 no 1 pp F55ndashF58 2002

[16] A Charil D P Laplante C Vaillancourt and S King ldquoPrenatalstress and brain developmentrdquo Brain Research Reviews vol 65no 1 pp 56ndash79 2010

[17] D Tijsseling L D E Wijnberger J B Derks et al ldquoEffects ofantenatal glucocorticoid therapy on hippocampal histology ofpreterm infantsrdquoPLoSONE vol 7 no 3 Article ID e33369 2012

[18] C W Noorlander D Tijsseling E V S Hessel et al ldquoAntenatalglucocorticoid treatment affects hippocampal development inmicerdquo PLoS ONE vol 9 no 1 Article ID e85671 2014

[19] S McArthur E McHale and G E Gillies ldquoThe size and distri-bution of midbrain dopaminergic populations are permanentlyaltered by perinatal glucocorticoid exposure in a sex- region-and time-specific mannerrdquo Neuropsychopharmacology vol 32no 7 pp 1462ndash1476 2007

[20] S McArthur E McHale J W Dalley J C Buckingham andG E Gillies ldquoAlteredmesencephalic dopaminergic populationsin adulthood as a consequence of brief perinatal glucocorticoidexposurerdquo Journal of Neuroendocrinology vol 17 no 8 pp 475ndash482 2005

[21] ETheogaraj CD JohnH C Christian J FMorris S F Smithand J C Buckingham ldquoPerinatal glucocorticoid treatmentproduces molecular functional and morphological changes inthe anterior pituitary gland of the adultmale ratrdquoEndocrinologyvol 146 no 11 pp 4804ndash4813 2005

[22] G Paxinos and C WatsonThe Rat Brain in Stereotaxic Coordi-nates Elsevier Academic Press 5th edition 2005

[23] M Leventopoulos D Ruedi-Bettschen I Knuesel J Feldon CR Pryce and J Opacka-Juffry ldquoLong-term effects of early lifedeprivation on brain glia in Fischer ratsrdquo Brain Research vol1142 no 1 pp 119ndash126 2007

[24] M J West L Slomianka and H J G Gundersen ldquoUnbiasedstereological estimation of the total number of neurons in

8 Neural Plasticity

the subdivisions of the rat hippocampus using the optical frac-tionatorrdquo Anatomical Record vol 231 no 4 pp 482ndash497 1991

[25] D T Theodosis D A Poulain and S H R Oliet ldquoActivity-dependent structural and functional plasticity of astrocyte-neuron interactionsrdquo Physiological Reviews vol 88 no 3 pp983ndash1008 2008

[26] H Uno L Lohmiller C Thieme et al ldquoBrain damage inducedby prenatal exposure to dexamethasone in fetal rhesus maca-ques I Hippocampusrdquo Developmental Brain Research vol 53no 2 pp 157ndash167 1990

[27] R Q Carlos F J Seidler and T A Slotkin ldquoFetal dexametha-sone exposure altersmacromolecular characteristics of rat braindevelopment a critical period for regionally selective altera-tionsrdquo Teratology vol 46 no 1 pp 45ndash59 1992

[28] E R de Kloet ldquoStress in the brainrdquo European Journal of Phar-macology vol 405 pp 187ndash198 2000

[29] S G Matthews ldquoAntenatal glucocorticoids and the developingbrain mechanisms of actionrdquo Seminars in Neonatology vol 6no 4 pp 309ndash317 2001

[30] L J Abernethy M Palaniappan and R W I Cooke ldquoQuanti-tative magnetic resonance imaging of the brain in survivors ofvery low birth weightrdquo Archives of Disease in Childhood vol 87no 4 pp 279ndash283 2002

[31] E B Isaacs A LucasW K Chong et al ldquoHippocampal volumeand everyday memory in children of very low birth weightrdquoPediatric Research vol 47 no 6 pp 713ndash720 2000

[32] B S Peterson B Vohr L H Staib et al ldquoRegional brain volumeabnormalities and long-term cognitive outcome in preterminfantsrdquo Journal of the American Medical Association vol 284no 15 pp 1939ndash1947 2000

[33] B P Murphy T E Inder P S Huppi et al ldquoImpaired cerebralcortical graymatter growth after treatmentwith dexamethasonefor neonatal chronic lung diseaserdquo Pediatrics vol 107 no 2 pp217ndash221 2001

[34] P Yossuck M Kraszpulski and A K Salm ldquoPerinatal corti-costeroid effect on amygdala and hippocampus volume duringbrain development in the rat modelrdquo Early Human Develop-ment vol 82 no 4 pp 267ndash272 2006

[35] D Rice and S Barone Jr ldquoCritical periods of vulnerability forthe developing nervous system evidence from humans andanimal modelsrdquo Environmental Health Perspectives vol 108 no3 pp 511ndash533 2000

[36] C S Woolley E Gould and B S McEwen ldquoExposure toexcess glucocorticoids alters dendritic morphology of adulthippocampal pyramidal neuronsrdquo Brain Research vol 531 no1-2 pp 225ndash231 1990

[37] R M Sapolsky H Uno C S Rebert and C E Finch ldquoHippo-campal damage associatedwith prolonged glucocorticoid expo-sure in primatesrdquoThe Journal of Neuroscience vol 10 no 9 pp2897ndash2902 1990

[38] F Ohl T Michaelis G K Vollmann-Honsdorf C Kirschbaumand E Fuchs ldquoEffect of chronic psychosocial stress and long-term cortisol treatment on hippocampus-mediated memoryand hippocampal volume a pilot-study in tree shrewsrdquo Psy-choneuroendocrinology vol 25 no 4 pp 357ndash363 2000

[39] C Nosarti M H S Al-Asady S Frangou A L Stewart L Rif-kin and R M Murray ldquoAdolescents who were born very pre-term have decreased brain volumesrdquo Brain vol 125 no 7 pp1616ndash1623 2002

[40] L E Haynes M R Griffiths R E Hyde D J Barber and I JMitchell ldquoDexamethasone induces limited apoptosis and exten-sive sublethal damage to specific subregions of the striatum andhippocampus implications for mood disordersrdquo Neurosciencevol 104 no 1 pp 57ndash69 2001

[41] N Sousa M M Paula-Barbosa and O F X Almeida ldquoLigandand subfield specificity of corticoid-induced neuronal loss inthe rat hippocampal formationrdquoNeuroscience vol 89 no 4 pp1079ndash1087 1999

[42] K L Crossin M-H Tai L A Krushel V P Mauro and G MEdelman ldquoGlucocorticoid receptor pathways are involved in theinhibition of astrocyte proliferationrdquo Proceedings of the NationalAcademy of Sciences of the United States of America vol 94 no6 pp 2687ndash2692 1997

[43] M Sabolek A Herborg J Schwarz and A Storch ldquoDexam-ethasone blocks astroglial differentiation from neural precursorcellsrdquo NeuroReport vol 17 no 16 pp 1719ndash1723 2006

[44] N R Nichols J N Masters and C E Finch ldquoChanges in geneexpression in hippocampus in response to glucocorticoids andstressrdquo Brain Research Bulletin vol 24 no 5 pp 659ndash662 1990

[45] N R Nichols H H Osterburg J N Masters S L Millarand C E Finch ldquoMessenger RNA for glial fibrillary acidicprotein is decreased in rat brain following acute and chroniccorticosterone treatmentrdquo Molecular Brain Research vol 7 no1 pp 1ndash7 1990

[46] N J Laping N R Nichols J R Day and C E Finch ldquoCortico-sterone differentially regulates the bilateral response of astro-cyte mRNAs in the hippocampus to entorhinal cortex lesions inmale ratsrdquoMolecular Brain Research vol 10 no 4 pp 291ndash2971991

[47] S Tsuneishi S Takada T Motoike T Ohashi K Sano andH Nakamura ldquoEffects of dexamethasone on the expression ofmyelin basic protein proteolipid protein and glial fibrillaryacidic protein genes in developing rat brainrdquo DevelopmentalBrain Research vol 61 no 1 pp 117ndash123 1991

[48] J P OrsquoCallaghan R E Brinton and B S McEwen ldquoGlucocor-ticoids regulate the synthesis of glial fibrillary acidic protein inintact and adrenalectomized rats but do not affect its expressionfollowing brain injuryrdquo Journal of Neurochemistry vol 57 no 3pp 860ndash869 1991

[49] M V Sofroniew and H V Vinters ldquoAstrocytes biology andpathologyrdquoActa Neuropathologica vol 119 no 1 pp 7ndash35 2010

[50] S Maccari M Darnaudery S Morley-Fletcher A R Zuena CCinque and O van Reeth ldquoPrenatal stress and long-termconsequences implications of glucocorticoid hormonesrdquo Neu-roscience and Biobehavioral Reviews vol 27 no 1-2 pp 119ndash1272003

[51] C R Pryce Y Aubert CMaier P C Pearce and E Fuchs ldquoThedevelopmental impact of prenatal stress prenatal dexametha-sone and postnatal social stress on physiology behaviour andneuroanatomy of primate offspring Studies in rhesus macaqueand common marmosetrdquo Psychopharmacology vol 214 no 1pp 33ndash53 2011

Submit your manuscripts athttpwwwhindawicom

Neurology Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Alzheimerrsquos DiseaseHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

ScientificaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Research and TreatmentSchizophrenia

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Neural Plasticity

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Parkinsonrsquos Disease

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Research and TreatmentAutism

Sleep DisordersHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Neuroscience Journal

Epilepsy Research and TreatmentHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Psychiatry Journal

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Computational and Mathematical Methods in Medicine

Depression Research and TreatmentHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Brain ScienceInternational Journal of

StrokeResearch and TreatmentHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Neurodegenerative Diseases

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Journal of

Cardiovascular Psychiatry and NeurologyHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Neural Plasticity 7

Abbreviations

AGT Antenatal glucocorticoid treatmentDex DexamethasoneGD Gestational dayGFAP Glial fibrillary acidic proteinHX HematoxylinMDD Major depressive disorderPBS Phosphate-buffered salinePTSD Posttraumatic stress disorderROI Region of interest

Conflict of Interests

The authors declare that there is no conflict of interestsregarding the publication of this paper

Acknowledgments

This work was supported by the Wellcome Trust Grant (no086871Z08Z) to G E Gillies Vishvesh H Shende was aPhD student sponsored by the University of RoehamptonThe authors thank Dr Todd Rae (Roehampton) for trainingVishvesh H Shende in the pQCT technique

References

[1] B Czeh and P J Lucassen ldquoWhat causes the hippocampalvolume decrease in depression Are neurogenesis glial changesand apoptosis implicatedrdquo European Archives of Psychiatry andClinical Neuroscience vol 257 no 5 pp 250ndash260 2007

[2] P J Lucassen J Pruessner N Sousa et al ldquoNeuropathology ofstressrdquo Acta Neuropathologica vol 127 no 1 pp 109ndash135 2014

[3] B S McEwen L Eiland R G Hunter andMMMiller ldquoStressand anxiety structural plasticity and epigenetic regulation as aconsequence of stressrdquo Neuropharmacology vol 62 no 1 pp3ndash12 2012

[4] M A Mehta N I Golembo C Nosarti et al ldquoAmygdala hip-pocampal and corpus callosum size following severe earlyinstitutional deprivation the English and Romanian Adopteesstudy pilotrdquo Journal of Child Psychology and Psychiatry andAllied Disciplines vol 50 no 8 pp 943ndash951 2009

[5] T Frodl and V OrsquoKeane ldquoHow does the brain deal with cumu-lative stress A review with focus on developmental stress HPAaxis function and hippocampal structure in humansrdquo Neurobi-ology of Disease vol 52 pp 24ndash37 2013

[6] B Czeh N Abumaria R Rygula and E Fuchs ldquoQuantita-tive changes in hippocampal microvasculature of chronicallystressed rats no effect of fluoxetine treatmentrdquo Hippocampusvol 20 no 1 pp 174ndash185 2010

[7] V H Shende C R Pryce D Ruedi-Bettschen T C Rae andJ Opacka-Juffry ldquoEvidence for hippocampal remodeling as along term effect of early life stress in the rat model of earlydeprivationrdquo Recent Developments in Brain Research vol 1 pp1ndash19 2012

[8] A Jackowski T D Perera C G Abdallah et al ldquoEarly-lifestress corpus callosum development hippocampal volumet-rics and anxious behavior in male nonhuman primatesrdquo Psy-chiatry ResearchmdashNeuroimaging vol 192 no 1 pp 37ndash44 2011

[9] C Pittenger and R S Duman ldquoStress depression and neuro-plasticity a convergence of mechanismsrdquoNeuropsychopharma-cology vol 33 no 1 pp 88ndash109 2008

[10] C L Coe M Kramer B Czeh et al ldquoPrenatal stress diminishesneurogenesis in the dentate gyrus of juvenile Rhesus monkeysrdquoBiological Psychiatry vol 54 no 10 pp 1025ndash1034 2003

[11] J D Funkhouser K J Peevey P B Mockridge and E RHughes ldquoDistribution of dexamethasone between mother andfetus after maternal administrationrdquo Pediatric Research vol 12no 11 pp 1053ndash1056 1978

[12] D Roberts and S Dalziel ldquoAntenatal corticosteroids for accel-erating fetal lung maturation for women at risk of pretermbirthrdquo Cochrane Database of Systematic Reviews vol 3 ArticleID CD004454 2006

[13] T F Yeh Y J Lin H C Lin et al ldquoOutcomes at school ageafter postnatal dexamethasone therapy for lung disease of pre-maturityrdquoTheNew England Journal of Medicine vol 350 no 13pp 1304ndash1313 2004

[14] J P Empana M M Anceschi I Szabo E V Cosmi G Breartand P Truffert ldquoAntenatal corticosteroids policies in 14 Euro-pean countries factors associated with multiple courses TheEURAIL surveyrdquo Acta Paediatrica International Journal of Pae-diatrics vol 93 no 10 pp 1318ndash1322 2004

[15] C Romagnoli E Zecca R Luciano G Torrioli and G Tor-torolo ldquoA three year follow up of preterm infants after moder-ately early treatment with dexamethasonerdquo Archives of Diseasein Childhood Fetal and Neonatal Edition vol 87 no 1 pp F55ndashF58 2002

[16] A Charil D P Laplante C Vaillancourt and S King ldquoPrenatalstress and brain developmentrdquo Brain Research Reviews vol 65no 1 pp 56ndash79 2010

[17] D Tijsseling L D E Wijnberger J B Derks et al ldquoEffects ofantenatal glucocorticoid therapy on hippocampal histology ofpreterm infantsrdquoPLoSONE vol 7 no 3 Article ID e33369 2012

[18] C W Noorlander D Tijsseling E V S Hessel et al ldquoAntenatalglucocorticoid treatment affects hippocampal development inmicerdquo PLoS ONE vol 9 no 1 Article ID e85671 2014

[19] S McArthur E McHale and G E Gillies ldquoThe size and distri-bution of midbrain dopaminergic populations are permanentlyaltered by perinatal glucocorticoid exposure in a sex- region-and time-specific mannerrdquo Neuropsychopharmacology vol 32no 7 pp 1462ndash1476 2007

[20] S McArthur E McHale J W Dalley J C Buckingham andG E Gillies ldquoAlteredmesencephalic dopaminergic populationsin adulthood as a consequence of brief perinatal glucocorticoidexposurerdquo Journal of Neuroendocrinology vol 17 no 8 pp 475ndash482 2005

[21] ETheogaraj CD JohnH C Christian J FMorris S F Smithand J C Buckingham ldquoPerinatal glucocorticoid treatmentproduces molecular functional and morphological changes inthe anterior pituitary gland of the adultmale ratrdquoEndocrinologyvol 146 no 11 pp 4804ndash4813 2005

[22] G Paxinos and C WatsonThe Rat Brain in Stereotaxic Coordi-nates Elsevier Academic Press 5th edition 2005

[23] M Leventopoulos D Ruedi-Bettschen I Knuesel J Feldon CR Pryce and J Opacka-Juffry ldquoLong-term effects of early lifedeprivation on brain glia in Fischer ratsrdquo Brain Research vol1142 no 1 pp 119ndash126 2007

[24] M J West L Slomianka and H J G Gundersen ldquoUnbiasedstereological estimation of the total number of neurons in

8 Neural Plasticity

the subdivisions of the rat hippocampus using the optical frac-tionatorrdquo Anatomical Record vol 231 no 4 pp 482ndash497 1991

[25] D T Theodosis D A Poulain and S H R Oliet ldquoActivity-dependent structural and functional plasticity of astrocyte-neuron interactionsrdquo Physiological Reviews vol 88 no 3 pp983ndash1008 2008

[26] H Uno L Lohmiller C Thieme et al ldquoBrain damage inducedby prenatal exposure to dexamethasone in fetal rhesus maca-ques I Hippocampusrdquo Developmental Brain Research vol 53no 2 pp 157ndash167 1990

[27] R Q Carlos F J Seidler and T A Slotkin ldquoFetal dexametha-sone exposure altersmacromolecular characteristics of rat braindevelopment a critical period for regionally selective altera-tionsrdquo Teratology vol 46 no 1 pp 45ndash59 1992

[28] E R de Kloet ldquoStress in the brainrdquo European Journal of Phar-macology vol 405 pp 187ndash198 2000

[29] S G Matthews ldquoAntenatal glucocorticoids and the developingbrain mechanisms of actionrdquo Seminars in Neonatology vol 6no 4 pp 309ndash317 2001

[30] L J Abernethy M Palaniappan and R W I Cooke ldquoQuanti-tative magnetic resonance imaging of the brain in survivors ofvery low birth weightrdquo Archives of Disease in Childhood vol 87no 4 pp 279ndash283 2002

[31] E B Isaacs A LucasW K Chong et al ldquoHippocampal volumeand everyday memory in children of very low birth weightrdquoPediatric Research vol 47 no 6 pp 713ndash720 2000

[32] B S Peterson B Vohr L H Staib et al ldquoRegional brain volumeabnormalities and long-term cognitive outcome in preterminfantsrdquo Journal of the American Medical Association vol 284no 15 pp 1939ndash1947 2000

[33] B P Murphy T E Inder P S Huppi et al ldquoImpaired cerebralcortical graymatter growth after treatmentwith dexamethasonefor neonatal chronic lung diseaserdquo Pediatrics vol 107 no 2 pp217ndash221 2001

[34] P Yossuck M Kraszpulski and A K Salm ldquoPerinatal corti-costeroid effect on amygdala and hippocampus volume duringbrain development in the rat modelrdquo Early Human Develop-ment vol 82 no 4 pp 267ndash272 2006

[35] D Rice and S Barone Jr ldquoCritical periods of vulnerability forthe developing nervous system evidence from humans andanimal modelsrdquo Environmental Health Perspectives vol 108 no3 pp 511ndash533 2000

[36] C S Woolley E Gould and B S McEwen ldquoExposure toexcess glucocorticoids alters dendritic morphology of adulthippocampal pyramidal neuronsrdquo Brain Research vol 531 no1-2 pp 225ndash231 1990

[37] R M Sapolsky H Uno C S Rebert and C E Finch ldquoHippo-campal damage associatedwith prolonged glucocorticoid expo-sure in primatesrdquoThe Journal of Neuroscience vol 10 no 9 pp2897ndash2902 1990

[38] F Ohl T Michaelis G K Vollmann-Honsdorf C Kirschbaumand E Fuchs ldquoEffect of chronic psychosocial stress and long-term cortisol treatment on hippocampus-mediated memoryand hippocampal volume a pilot-study in tree shrewsrdquo Psy-choneuroendocrinology vol 25 no 4 pp 357ndash363 2000

[39] C Nosarti M H S Al-Asady S Frangou A L Stewart L Rif-kin and R M Murray ldquoAdolescents who were born very pre-term have decreased brain volumesrdquo Brain vol 125 no 7 pp1616ndash1623 2002

[40] L E Haynes M R Griffiths R E Hyde D J Barber and I JMitchell ldquoDexamethasone induces limited apoptosis and exten-sive sublethal damage to specific subregions of the striatum andhippocampus implications for mood disordersrdquo Neurosciencevol 104 no 1 pp 57ndash69 2001

[41] N Sousa M M Paula-Barbosa and O F X Almeida ldquoLigandand subfield specificity of corticoid-induced neuronal loss inthe rat hippocampal formationrdquoNeuroscience vol 89 no 4 pp1079ndash1087 1999

[42] K L Crossin M-H Tai L A Krushel V P Mauro and G MEdelman ldquoGlucocorticoid receptor pathways are involved in theinhibition of astrocyte proliferationrdquo Proceedings of the NationalAcademy of Sciences of the United States of America vol 94 no6 pp 2687ndash2692 1997

[43] M Sabolek A Herborg J Schwarz and A Storch ldquoDexam-ethasone blocks astroglial differentiation from neural precursorcellsrdquo NeuroReport vol 17 no 16 pp 1719ndash1723 2006

[44] N R Nichols J N Masters and C E Finch ldquoChanges in geneexpression in hippocampus in response to glucocorticoids andstressrdquo Brain Research Bulletin vol 24 no 5 pp 659ndash662 1990

[45] N R Nichols H H Osterburg J N Masters S L Millarand C E Finch ldquoMessenger RNA for glial fibrillary acidicprotein is decreased in rat brain following acute and chroniccorticosterone treatmentrdquo Molecular Brain Research vol 7 no1 pp 1ndash7 1990

[46] N J Laping N R Nichols J R Day and C E Finch ldquoCortico-sterone differentially regulates the bilateral response of astro-cyte mRNAs in the hippocampus to entorhinal cortex lesions inmale ratsrdquoMolecular Brain Research vol 10 no 4 pp 291ndash2971991

[47] S Tsuneishi S Takada T Motoike T Ohashi K Sano andH Nakamura ldquoEffects of dexamethasone on the expression ofmyelin basic protein proteolipid protein and glial fibrillaryacidic protein genes in developing rat brainrdquo DevelopmentalBrain Research vol 61 no 1 pp 117ndash123 1991

[48] J P OrsquoCallaghan R E Brinton and B S McEwen ldquoGlucocor-ticoids regulate the synthesis of glial fibrillary acidic protein inintact and adrenalectomized rats but do not affect its expressionfollowing brain injuryrdquo Journal of Neurochemistry vol 57 no 3pp 860ndash869 1991

[49] M V Sofroniew and H V Vinters ldquoAstrocytes biology andpathologyrdquoActa Neuropathologica vol 119 no 1 pp 7ndash35 2010

[50] S Maccari M Darnaudery S Morley-Fletcher A R Zuena CCinque and O van Reeth ldquoPrenatal stress and long-termconsequences implications of glucocorticoid hormonesrdquo Neu-roscience and Biobehavioral Reviews vol 27 no 1-2 pp 119ndash1272003

[51] C R Pryce Y Aubert CMaier P C Pearce and E Fuchs ldquoThedevelopmental impact of prenatal stress prenatal dexametha-sone and postnatal social stress on physiology behaviour andneuroanatomy of primate offspring Studies in rhesus macaqueand common marmosetrdquo Psychopharmacology vol 214 no 1pp 33ndash53 2011

Submit your manuscripts athttpwwwhindawicom

Neurology Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Alzheimerrsquos DiseaseHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

ScientificaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Research and TreatmentSchizophrenia

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Neural Plasticity

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Parkinsonrsquos Disease

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Research and TreatmentAutism

Sleep DisordersHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Neuroscience Journal

Epilepsy Research and TreatmentHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Psychiatry Journal

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Computational and Mathematical Methods in Medicine

Depression Research and TreatmentHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Brain ScienceInternational Journal of

StrokeResearch and TreatmentHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Neurodegenerative Diseases

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Journal of

Cardiovascular Psychiatry and NeurologyHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

8 Neural Plasticity

the subdivisions of the rat hippocampus using the optical frac-tionatorrdquo Anatomical Record vol 231 no 4 pp 482ndash497 1991

[25] D T Theodosis D A Poulain and S H R Oliet ldquoActivity-dependent structural and functional plasticity of astrocyte-neuron interactionsrdquo Physiological Reviews vol 88 no 3 pp983ndash1008 2008

[26] H Uno L Lohmiller C Thieme et al ldquoBrain damage inducedby prenatal exposure to dexamethasone in fetal rhesus maca-ques I Hippocampusrdquo Developmental Brain Research vol 53no 2 pp 157ndash167 1990

[27] R Q Carlos F J Seidler and T A Slotkin ldquoFetal dexametha-sone exposure altersmacromolecular characteristics of rat braindevelopment a critical period for regionally selective altera-tionsrdquo Teratology vol 46 no 1 pp 45ndash59 1992

[28] E R de Kloet ldquoStress in the brainrdquo European Journal of Phar-macology vol 405 pp 187ndash198 2000

[29] S G Matthews ldquoAntenatal glucocorticoids and the developingbrain mechanisms of actionrdquo Seminars in Neonatology vol 6no 4 pp 309ndash317 2001

[30] L J Abernethy M Palaniappan and R W I Cooke ldquoQuanti-tative magnetic resonance imaging of the brain in survivors ofvery low birth weightrdquo Archives of Disease in Childhood vol 87no 4 pp 279ndash283 2002

[31] E B Isaacs A LucasW K Chong et al ldquoHippocampal volumeand everyday memory in children of very low birth weightrdquoPediatric Research vol 47 no 6 pp 713ndash720 2000

[32] B S Peterson B Vohr L H Staib et al ldquoRegional brain volumeabnormalities and long-term cognitive outcome in preterminfantsrdquo Journal of the American Medical Association vol 284no 15 pp 1939ndash1947 2000

[33] B P Murphy T E Inder P S Huppi et al ldquoImpaired cerebralcortical graymatter growth after treatmentwith dexamethasonefor neonatal chronic lung diseaserdquo Pediatrics vol 107 no 2 pp217ndash221 2001

[34] P Yossuck M Kraszpulski and A K Salm ldquoPerinatal corti-costeroid effect on amygdala and hippocampus volume duringbrain development in the rat modelrdquo Early Human Develop-ment vol 82 no 4 pp 267ndash272 2006

[35] D Rice and S Barone Jr ldquoCritical periods of vulnerability forthe developing nervous system evidence from humans andanimal modelsrdquo Environmental Health Perspectives vol 108 no3 pp 511ndash533 2000

[36] C S Woolley E Gould and B S McEwen ldquoExposure toexcess glucocorticoids alters dendritic morphology of adulthippocampal pyramidal neuronsrdquo Brain Research vol 531 no1-2 pp 225ndash231 1990

[37] R M Sapolsky H Uno C S Rebert and C E Finch ldquoHippo-campal damage associatedwith prolonged glucocorticoid expo-sure in primatesrdquoThe Journal of Neuroscience vol 10 no 9 pp2897ndash2902 1990

[38] F Ohl T Michaelis G K Vollmann-Honsdorf C Kirschbaumand E Fuchs ldquoEffect of chronic psychosocial stress and long-term cortisol treatment on hippocampus-mediated memoryand hippocampal volume a pilot-study in tree shrewsrdquo Psy-choneuroendocrinology vol 25 no 4 pp 357ndash363 2000

[39] C Nosarti M H S Al-Asady S Frangou A L Stewart L Rif-kin and R M Murray ldquoAdolescents who were born very pre-term have decreased brain volumesrdquo Brain vol 125 no 7 pp1616ndash1623 2002

[40] L E Haynes M R Griffiths R E Hyde D J Barber and I JMitchell ldquoDexamethasone induces limited apoptosis and exten-sive sublethal damage to specific subregions of the striatum andhippocampus implications for mood disordersrdquo Neurosciencevol 104 no 1 pp 57ndash69 2001

[41] N Sousa M M Paula-Barbosa and O F X Almeida ldquoLigandand subfield specificity of corticoid-induced neuronal loss inthe rat hippocampal formationrdquoNeuroscience vol 89 no 4 pp1079ndash1087 1999

[42] K L Crossin M-H Tai L A Krushel V P Mauro and G MEdelman ldquoGlucocorticoid receptor pathways are involved in theinhibition of astrocyte proliferationrdquo Proceedings of the NationalAcademy of Sciences of the United States of America vol 94 no6 pp 2687ndash2692 1997

[43] M Sabolek A Herborg J Schwarz and A Storch ldquoDexam-ethasone blocks astroglial differentiation from neural precursorcellsrdquo NeuroReport vol 17 no 16 pp 1719ndash1723 2006

[44] N R Nichols J N Masters and C E Finch ldquoChanges in geneexpression in hippocampus in response to glucocorticoids andstressrdquo Brain Research Bulletin vol 24 no 5 pp 659ndash662 1990

[45] N R Nichols H H Osterburg J N Masters S L Millarand C E Finch ldquoMessenger RNA for glial fibrillary acidicprotein is decreased in rat brain following acute and chroniccorticosterone treatmentrdquo Molecular Brain Research vol 7 no1 pp 1ndash7 1990

[46] N J Laping N R Nichols J R Day and C E Finch ldquoCortico-sterone differentially regulates the bilateral response of astro-cyte mRNAs in the hippocampus to entorhinal cortex lesions inmale ratsrdquoMolecular Brain Research vol 10 no 4 pp 291ndash2971991

[47] S Tsuneishi S Takada T Motoike T Ohashi K Sano andH Nakamura ldquoEffects of dexamethasone on the expression ofmyelin basic protein proteolipid protein and glial fibrillaryacidic protein genes in developing rat brainrdquo DevelopmentalBrain Research vol 61 no 1 pp 117ndash123 1991

[48] J P OrsquoCallaghan R E Brinton and B S McEwen ldquoGlucocor-ticoids regulate the synthesis of glial fibrillary acidic protein inintact and adrenalectomized rats but do not affect its expressionfollowing brain injuryrdquo Journal of Neurochemistry vol 57 no 3pp 860ndash869 1991

[49] M V Sofroniew and H V Vinters ldquoAstrocytes biology andpathologyrdquoActa Neuropathologica vol 119 no 1 pp 7ndash35 2010

[50] S Maccari M Darnaudery S Morley-Fletcher A R Zuena CCinque and O van Reeth ldquoPrenatal stress and long-termconsequences implications of glucocorticoid hormonesrdquo Neu-roscience and Biobehavioral Reviews vol 27 no 1-2 pp 119ndash1272003

[51] C R Pryce Y Aubert CMaier P C Pearce and E Fuchs ldquoThedevelopmental impact of prenatal stress prenatal dexametha-sone and postnatal social stress on physiology behaviour andneuroanatomy of primate offspring Studies in rhesus macaqueand common marmosetrdquo Psychopharmacology vol 214 no 1pp 33ndash53 2011

Submit your manuscripts athttpwwwhindawicom

Neurology Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Alzheimerrsquos DiseaseHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

ScientificaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Research and TreatmentSchizophrenia

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Neural Plasticity

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Parkinsonrsquos Disease

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Research and TreatmentAutism

Sleep DisordersHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Neuroscience Journal

Epilepsy Research and TreatmentHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Psychiatry Journal

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Computational and Mathematical Methods in Medicine

Depression Research and TreatmentHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Brain ScienceInternational Journal of

StrokeResearch and TreatmentHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Neurodegenerative Diseases

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Journal of

Cardiovascular Psychiatry and NeurologyHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Submit your manuscripts athttpwwwhindawicom

Neurology Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Alzheimerrsquos DiseaseHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

ScientificaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Research and TreatmentSchizophrenia

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Neural Plasticity

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Parkinsonrsquos Disease

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Research and TreatmentAutism

Sleep DisordersHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Neuroscience Journal

Epilepsy Research and TreatmentHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Psychiatry Journal

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Computational and Mathematical Methods in Medicine

Depression Research and TreatmentHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Brain ScienceInternational Journal of

StrokeResearch and TreatmentHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Neurodegenerative Diseases

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Journal of

Cardiovascular Psychiatry and NeurologyHindawi Publishing Corporationhttpwwwhindawicom Volume 2014