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ORIGINAL ARTICLE

Hyperalgesia and increased sevoflurane minimumalveolar concentration induced by

A randomised experimental study

Mariana Abreu, Delia Aguado, Javier Benito, Javier Garc

Ignacio A. Gomez de Segura

BACKGROUND Perioperative opioids reduce inhalationalanaesthetic requirements. The initial hypoalgesia may, how-ever, be

OBJECTtration inmight be

DESIGN

SETTINMadrid,

ANIMAL

INTERVtration(MNTs)opioid tnorphine10mgkgandMNketamine

MAIN OUTCOMEMEASURES The effect of different opioidtreatments onMAC andMNTwas evaluated using analysis of

LTS All studied opioids produced an immediatetion in sevoflurane MAC, followed by an increase) imtimasreffeine

CLwitndente

she

Eur J Anaesthesiol 2015; 32:232241

From the LDepartmenFaculty of Veterinary Medicine, University of Montreal (UdM), Saint Hyacinthe, Quebec, Canada (JB) and Department of Anaesthesiology and Critical Care, Puerta de HierroUniversity Hospital, Madrid, Spain (JGF)

CorresponPuerta deTel: +34 9

This aComm

Weinin pefamili

0265-0215 2015 Copyright European Society of Anaesthesiology Ece to Dr Ignacio A. Gomez de Segura, Department of Animal Medicine and Surgery, Veterinary Faculty, Complutense University of Madrid (UCM), Avda.rro s/n, 28040 Madrid, Spain943 858; fax: +34 913 943 808; e-mail: iagsegura@vet.ucm.esdenHie13

DOI:10.1097/EJA.0000000000000188aboratory of Respiration Physiology, Carlos Chagas Filho Institute of BiophysicsIntroductionOpioids are widely used analgesics to treat moderate tosevere pain. There is evidence, however, suggesting thatopioids activate not only antinociceptive but also prono-ciceptive mechanisms leading to an increase in painsensitivity, also known as opioid-induced hyperalgesia(OIH). Opioid-induced hyperalgesia has been defined asa state of nociceptive sensitisation caused by exposure to

, CCS, Federal University of Rio de Janeiro, Rio de Janeiro, Brasil (MA),t of Animal Medicine and Surgery, Veterinary Faculty, Complutense University of Madrid (UCM), Madrid, Spain (IAGS, DA), Department of Clinical Sciences.

rticle is accompanied by the following Invitedentary:

broum AA. Role of anaesthetics and opioidsrioperative hyperalgesia: One step towardsarisation. Eur J Anaesthesiol 2015; 32:230231.IVES To determine whether prior opioid adminis-fluences inhalational anaesthetic requirements,whichassociated with opioid-induced hyperalgesia.

A prospective, randomised, experimental study.

G Experimental Surgery, La Paz University Hospital,Spain.

S Seventy-nine adult male Wistar rats.

ENTIONS Sevoflurane minimum alveolar concen-(MAC) and mechanical nociceptive thresholdswere assessed at baseline and 7 days later followingreatment with remifentanil 120mg kg1 h1, bupre-150mg kg1, methadone 8mgkg1 or morphine1. The duration of the effect of remifentanil onMAC

T was evaluated in addition to the preventive effect of10mgkg1 on remifentanil-induced hyperalgesia.

RESUreduc(16%the imthatdecrean incThe eketam

CONatednot upreve

Publiuropean Society of Anaesthesiology. Unauthorn baseline MAC 7 days later (P

Copyright

opioids, whereby a patient receiving opioids for thetreatmestimulat

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Opioid-induced hyperalgesia and minimum alveolar concentration 233nt of pain becomes more sensitive to nociceptiveion.1

opioid receptor system has been suggested asa pivotal role in the development of OIH.2

ne3 and remifentanil4,5 are the drugs most com-elated to OIH, but other opioids such as metha-buprenorphine have also been implicated.69

more, OIH might be long lasting; a single remi-dose has been associated with hyperalgesia inr up to 10 days.10 The central glutaminergicspinal dynorphins, descending facilitation,mechanisms with decreased reuptake andd nociceptive responses have all been hypo-to have a role in the development of OIH.11

yl-D-aspartate (NMDA) receptor activation hasown to be involved in OIH12 and the NMDAist ketamine has been used for its antihyperalge-erties.13

inhalational anaesthesia, opioids are usually coad-red and produce a reduction in anaestheticents. The anaesthetic-sparing action of remi-

, as determined by a reduction of the minimumconcentration (MAC), has been recently shownly decrease in rats due to an acute toleranceThis suggests that increasing doses of the opioidired to maintain a similar anaesthetic level. As study suggested a potential relationshipthe hyperalgesic action of remifentanil given

fusion, and its effect on the MAC.15

othesised that other opioids would also increaseonal anaesthetic requirements secondary to OIHnner similar to remifentanil. The aim of thisas, therefore, to determine whether remifentanil,rphine, methadone and morphine administrationan increase in the MAC associated with OIH,ation of these effects and whether ketaminetration could prevent them.

als and methodsdy was approved by the institutional animal carecommittee [Ethics Animal Experimentation

ttee (CEA) of the University Complutense,Spain, Chairperson Prof D. Jose Francisco

Fernandez. CEA approval reference date: 298] and the care of animal and licensing guidelineshich the study was performed were in accordancee ARRIVE (Animals in Research: Reporting Inperiments) statement.

-two adult male Wistar rats (Charles Riverories, Barcelona, Spain), of mean (SD) weightg were used. The animals were housed in groupsr cage (Macrolon Type IV; Souralit 3000, Souralit

cyclemeanFoodAnimSpainmentat 9Surg

AnaedeterRatschamLabo(3 lmVapowas(Terven,recumintousedAfterby degauzgraphdead

HearsaturdioxiouslyElecandrecorto demonimeanTherard P

To dusedthe ficoncMACchanvapoequilsevoflthe n

Thewereprevilonggen, European Society of Anaesthesiology. Unauthoights on between 8 : 00 a.m. and 8 : 00 p.m.) atD) 22 (2)8C and 40 to 70% relative humidity.A03 maintenance diet for rodents; ScientificFood & Engineering, SAFE, Panlab, Barcelona,and water were available ad libitum. All experi-ere performed during the light period startinga.m. (Microsurgery Laboratory, ExperimentalDepartment).

esia and minimum alveolar concentrationnationre placed in a methacrylate anaesthetic inductionr into which 8% sevoflurane (Sevorane; Abbottories, Madrid, Spain) was delivered in oxygen1) via a sevoflurane vapouriser (Sevorane Drager000; Lubeck, Germany). Endotracheal intubationformed using a 14-gauge polyethylene cathetero Surflo i.v. catheter; Terumo Europe NV, Leu-lgium) with the animal positioned in sternalence. A flexible blunt-tip wire guide was insertedtrachea with the aid of an otoscope and this wasguide the endotracheal tube into the trachea.e correct position of the catheter was ascertainedrmining gas movement through the catheter withreads and exhaled carbon dioxide with a capno-small T-piece breathing system with minimumace was connected.

nd respiratory rates, arterial haemoglobin oxygenn (via pulse oximetry) and end-tidal carbonand sevoflurane concentrations were continu-easured (Datex-Ohmeda SA5 monitor; General, Helsinki, Finland). Heart and respiratory ratesd-tidal carbon dioxide concentrations wered 60 s prior to tail clamping at each step usedrmine the MAC value. Rectal temperature wased and maintained between 378C and 388C byof a water-circulating warming blanket (Heaty PumpModel TP-220; Gaymar Industries, Orch-, New York, USA) and a standard heating light.

rminate the MAC, intratracheal gas sampling wasmeasure anaesthetic gas concentrations. BeforeMAC determination, the end-tidal anaestheticration was adjusted to 2.4%, a value close to thereviously reported for the rat.16 After every stepin the anaesthetic concentration delivered by theser to each rat, at least 10min were allowed foration (steady-state end-tidal concentration ofane) before obtaining gas samples and applyingious stimulus.

AC and any reduction produced by the opioidsstablished according to the method describedsly.1618 A noxious stimulus was applied with amostat (230mm Doyen forceps; Martin, Tutlin-rmany) clamped to the first ratchet lock on the

Eur J Anaesthesiol 2015; 32:232241

rized reproduction of this article is prohibited.

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rats tail for 60 s, or until a positive response was observed,immedithe tracthe tailimal tonegativepositivegross pand/or bresponsmovemchewingthe sevdecrema posititration wnegativetration mpermittlowestDetermat 650mto the bformula(760mmMAC(700mm(760mm

BehavioTo minmentalthe tesobtainetion we

Von Frey

The hinstimulatmethacrations),von Fr(ElectroLife ScBefore sfor 15remainements ofilamen60 g. Thwere apwithdresideredstimulubetweenan interdrawal

positive responses determined the threshold for eachurereeconhoan

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234 Abreu et al.

Eur J Aately after the gas sample had been obtained fromhea. The initial stimulus point was 6 cm distal tobase. The tail was always stimulated 1 cm prox-a previous test site, when a previous response was, or 1 cm distal to that point if the response was.19 A positive response was considered to be aurposeful movement of the head, extremitiesody or a clear movement of the spine. A negativee was considered to be either the lack ofent or discreet reactions (grimacing, swallowing,or tail flick). When a negative response occurred,oflurane concentration was reduced by 0.2%ents until a positive response was obtained. Whenve response occurred, the sevoflurane concen-as increased until the positive response became. The MAC was considered to be the concen-id-way between the highest concentration that

ed movement in response to the stimulus and theconcentration that prevented such movement.ination of theMACwas performed in a laboratoryabove sea level and MAC values were corrected

arometric pressure at sea level using the following: MAC (%) at sea level barometric pressureHg); (altitude-adjusted MAC)measured(%) xmeasured ambient barometric pressureHg in Madrid)/barometric pressure at sea levelHg).20

ural tests of nociceptionimise the influence of stress during the experi-procedure, animals were acclimatised for 3 days toting procedure before baseline values wered. Two behavioural tests of mechanical nocicep-re employed: von Frey and RandallSelitto.

test

d paw withdrawal response to von Frey filamention was evaluated.21 Animals were placed in aylate cage with a wire grid bottom (1 cm2 perfor-through which the electronically calibratedey anaesthesiometer filaments were appliednic von Frey Anesthesiometer Model 2393; IITCience Inc., Woodland Hills, California, USA).tarting the test, the animals were left in the cageto 30min for acclimatisation, that is, the ratsd calmed and still, without exploratory move-r defaecation and not resting.21 Eight von Freyts were used and bending force ranged from 0.1 toe filaments (from the thinnest to the thickest)plied to the right foot paw for 7 s or until the ratw its hind limb.2123 Locomotion was not con-as a withdrawal response, and in that case, thes was repeated.24 The stimulus was appliedthree and five times with each filament with

val of 5 s. The lowest force producing a with-response was considered the threshold. Three

meason thtwothresmech

Rand

Mechdallmodelineastarti5minappliincrewhicmeasdiffemeanvaluebetw300 gvaluenocic

OpioreduSelit

Expe

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(after

Oncewastereddeterwas amaticFinapotasanae

Drugclinicof OIon thdeterrando(nEste8mg10mtanilMadfentaappro

naesthesiol 2015; 32:232241

European Society of Anaesthesiology. Unauthorment. The threshold in each animal was basedseparate measurements (5min interval betweensecutive measurements). The mean of theseld values at each time point was considered theical nociceptive thresholds (MNT).25

Selitto test

ical thresholds were quantified using the Ran-litto paw pressure device (Digital Randall-Selitto600; IITC Life Science Inc.),26 which produces aincreasing mechanical force in grams. Beforethe test, the animal was handled for at leastill it was calmed and still. The stimulus wasto the plantar surface of the left hind paw byng the pressure until the rat withdrew its limb,as considered the threshold.23 The threshold wasd two to five times until two consecutive valuesg by no more than 15% were obtained and thef these two values was considered the thresholdthat time point. Aminimum of 5min was allowedconsecutive measures.24 A cut-off pressure of

as used to avoid tissue damage.27 The thresholdeach time point was considered the mechanicaltive threshold.

induced hyperalgesia was confirmed by an in withdrawal thresholds in the Randallr von Frey tests.

ental design and drug groups

f opioids on minimum alveolar concentration

days)

e animals were anaesthetised, a baseline MACermined. Each rat was then randomly adminis-e of the treatments, and 30min later, a newMACnation was started. Seven days later, the MACin determined with the same treatment. A sche-ew of the experimental design is shown in Fig. 1a.rats were euthanised with an overdose of

m chloride given intravenously while still deeplyetised.

ere selected because they are commonly usedy and have been associated with the development3,4,7,8 The dose of each selected opioid was basedlinical dose range for the rat and a previous studyning its MAC-sparing action.28 Animals werely allocated to receive one out of five treatmentser group): buprenorphine 150mg kg1 (Buprex;Laboratories, Barcelona, Spain); methadone

1 (Metasedin; Esteve Laboratories); morphineg1 (Morfina; Braun, Barcelona, Spain); remifen-120mg kg1 h1 (Ultiva; Glaxo-Smith-Kline,Spain); and 0.9% saline (control group). Remi-was administered as an intravenous infusion formately 2 h and the remaining opioids were dilutedized reproduction of this article is prohibited.

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in 0.9%by the i

Duration

on minim

Rats weRandalland insmined.durationrat wasremifen(n 8120mg kwas 4.8recoverperformto evaludetermiwere reeuthaniintraven

Effects o

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Opioid-induced hyperalgesia and minimum alveolar concentration 235

Fig. 1

D

An

Experimen C)(immediat on m4-week p as(day 0) an tssaline to a total volume of 1ml and administered expe

(b)

(c)

(a)

Anaesthesia

Day 0

MAC

Opioid

MAC30min

Anaesthesia MAC

Remifentanil

MNT

AdaptationMNT

Day 0 3 days

Anaesthesia MACMNT

Day 0 3 days

Remifentanil +/ketamine

AdaptationMNT

tal design. (a) Determination of the minimum alveolar concentration (MAe) MAC reduction at day 0 and after 7 days. (b) Effects of remifentanileriod. After an adaptation period of the rats during 3 days, animals wered 1, 3, 7, 14, 21 and 28 days after remifentanil administration. (c) Effec European Society of Anaesthesiology. Unautho

ntraperitoneal (i.p.) route.

of remifentanil-induced hyperalgesia and effects

um alveolar concentration

re acclimatised over 3 days to the von Frey andSelitto tests. On day 0, rats were anaesthetisedtrumented and the baseline MAC was deter-Remifentanil was selected to determine theof the effects on hyperalgesia and MAC. Eachrandomly administered the treatment (eithertanil or 0.9% saline) intravenously as an infusionper group). The remifentanil dose wasg1 h1, and in both groups, the rate of infusionml kg1 h1 for 2 h. The rats were then allowed tofrom anaesthesia. The same procedures wereed at days 1, 3, 7, 14, 21 and 28 after treatmentate MNT andMAC. Schematic view of theMACnation is shown in Fig. 1b. As baseline valuesgained on day 28 after treatment, the rats weresed with an overdose of potassium chloride givenously while still deeply anaesthetised.

f ketamine on remifentanil-induced hyperalgesia

imum alveolar concentration (after 7 days)

r study was performed wherein remifentanil wastered alone or combined with ketamine, and itsdetermined at 7 days. A schematic view of the

selectedOIH anreceive2h, rem500; Pa(n 7 pintravenmaintaiveins ofcontrol(4.8mlk

StatisticAn n valwas confrom simof the nSolutionSmirnovof the dRandallpared wBonferramongconductvariatiotests. Aental design is shown in Fig. 1c. Remifentanil was

30min

ay 7

Opioid

aesthesia

Anaesthesia MACMNT

Days 1, 7, 14 , 21 and 28

Anaesthesia MACMNT

Day 7

MAC MAC

at baseline and following opioid administration to determineechanical nociceptive thresholds (MNT) and MAC over a

sessed for MNT followed by MAC determinations at baselineof remifentanil and ketamine on MNT and MAC after 7 days.rized reproduction of this article is prohibited.

to determine the blocking action of ketamine ond MAC. Animals were randomly assigned toremifentanil [120mgkg1 h1 intravenously forifentanil (same dose) as well as ketamine (Ketolarrke-Davis-Pfizer, Madrid, Spain] or 0.9% salineer group). Ketamine 10mgkg1 was administeredously 30min before remifentanil infusion andned at 10mgkg1 h1 for 2.5h. The two lateralthe rats tail were used for the infusions.Rats in thegroup received the same volume of salineg1 h1) intravenously for 2 h.

al analysisue of between seven and eight animals per groupsidered adequate on the basis of previous datailar studies in our laboratory and from calculationvalue (nQuery Advisor version 2.0; Statisticals Ltd., Cork, Ireland).16 The Kolmogorovtest was performed to verify normal distributionata. Differences in MAC and the von Frey andSelitto tests produced by treatments were com-ith one-way analysis of variance (ANOVA). Theoni posthoc test was used to compare differencesgroups. An ANOVA for repeated measures wased (drug x time) by comparing the percentage ofn in the MAC, von Frey and RandallSelittoone-way ANOVA was employed to compare both

Eur J Anaesthesiol 2015; 32:232241

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groupsestablisindicatemean (SPSS (vdata are

ResultBuprenbut notadminisrespecti

Mean b(n 35)all groulater in0.9% sa(Fig. 2P 0.00opioids(P> 0.0observe

The imopioidsfentanilanaesthgroups)MAC ddone, m(8), 33 (

iferel.0021muctiothho4).

236 Abreu et al.

Fig. 2

D

Minimum a sremifentan pe(P 0.40 forps). An increase in the MAC was observed 7 daysall groups administered opioids (P 0.03) but notline [MAC 2.3 (0.2)% vs. baseline, P 0.530]). The increase in the MAC [16 (10)%,1] was not different between groups receiving7 days earlier, with a mean value of 2.8 (0.3)%,5) (n 28). This time-related increase was notd in the control group [2 (7)%, P> 0.05] (Fig. 3).

mediate MAC reduction produced by the studied(buprenorphine, methadone, morphine and remi-) was not modified by a previous sevofluraneesia using opioids 7 days earlier (P> 0.05 for all. The MAC reduction observed in the secondetermination following buprenorphine, metha-orphine and remifentanil administration was 326), 34 (7) and 19 (11)%, respectively.

3 with agroupsbut not

The MAnil (absorespectinot at d(7)%] w

Remife(3)% (PFrey; Pon day 7effect oMAC anobservevalues wbaselineand 6 (3(vs. basFig. 3).tests wegroup c

naesthesiol 2015; 32:232241ntanil produced a significant increase in absoluteative MNT (von Frey test) (P 0.010 and1, respectively; Fig. 4), which was observed until(P 0.017) but not at day 28 (P 0.449). Am decrease was observed on day 7 with an of 48 (5)%. A similar reduction was observede RandallSelitto test (absolute and relativelds P 0.007 and P< 0.001, respectively;The maximum decrease was observed on day

Morphine Remifentanil

**

ay 7

ingle dose of buprenorphine, methadone, morphine andr group. Difference vs. control group at the same time pointized reproduction of this article is prohibited.

reduction of 50 (20)% and differences betweenwere observed from day 1 to day 21 (P 0.015),at day 28 (P 0.149).C was increased in rats administered remifenta-lute and relative values P 0.042 and P< 0.001,vely; Fig. 4) from day 1 to day 21 (P 0.013), butay 28 (P 0.110). The maximum increase [19as observed on day 7.

ntanil produced an increase in the MAC of 20 0.007) and a decrease in MNT of 42 (6)% (von 0.001) and 45 (7)% (RandallSelitto; P 0.001)after drug administration. However, a preventivef ketamine on remifentanil-induced effects ondMNT (von Frey and RandallSelitto tests) wasd and no significant differences from baselineere determined. The percentage of change fromvalues was 2 (6)% in the MAC value and 3 (7)%)% with the von Frey and RandallSelitto testseline P 1.000 for both MAC and MNT tests;Similarly, no differences in the MAC and MNTre observed in the remifentanil plus ketamineompared with control (P 1.000; Fig. 3).

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DiscusAs expeately foincreaseconsequthreshoobserve

Opioid-induced hyperalgesia and minimum alveolar concentration 237

Fig. 3

Mechanic(MAC) (cmean (SD0

20

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(%

) Von Frey

(a) European Society of Anaesthesiology. Unautho

sioncted, remifentanil decreased the MAC immedi-llowing its administration but induced a delayedobservable 24 h later. This increase might be aence of OIH, as the decrease in mechanicallds and the increase in MAC were simultaneouslyd when remifentanil was administered; ketamine

blockedincreaselasted utration.phine,increasealthoug

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al nociceptive thresholds in rats [determined with the von Frey (a) and Ran) values from rats determined at baseline and after the administration of eithe). The n value is always 8 rats per group. aDifferences vs. control group atrized reproduction of this article is prohibited.

both phenomena. The magnitude of the MACwas relatively moderate (16%) and consistentlyp to 21 days following remifentanil adminis-Other commonly used opioids, namely buprenor-methadone and morphine produced a similarin the MAC following administration and

h the duration of the effect was only determined

21 28

21 28

Remifentanil

Control

21 28

*

*

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dallSelitto (b) tests] and minimum alveolar concentrationr remifentanil or 0.9% saline (control). Data are expressed assame time point (P

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with reeffect o

Remifethetised

238 Abreu et al.

Fig. 4

Changesconcentraare expre

Eur J A0

20

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%) Von Frey Test

(a) European Society of Anaesthesiology. Unauthor

mifentanil administration, a similar long-lastingf the studied opioids cannot be ruled out.

ntanil use has been associated with OIH in anaes-patients. Total doses of remifentanil of 1.55 and

60mg kg(as dete90min.tered ovpostope

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in mechanical nociceptive thresholds in rats [determined with von Frey (a)tion (MAC) (c) produced by remifentanil, remifentanil and ketamine or 0.9%ssed as mean (SD). aDifferences vs. control group at same time point (P