prenatal oxazepam enhances mouse maternal aggression in the offspring, without modifying acute...

Neurotoxicology and Teratology, Vol. 13, pp. 75-81. © Pergamon Press pie, 1991. Printed in the U.S.A. 0892-0362/91 $3.00 + .00

Prenatal Oxazepam Enhances Mouse Maternal Aggression

in the Offspring, Without Modifying Acute Chlordiazepoxide Effects

G I O V A N N I L A V I O L A , L U I G I D E A C E T I S , G I O R G I O B I G N A M I A N D E N R I C O A L L E V A

Section o f Behavioural Pathophysiology, Laboratorio di Fisiopatologia di Organo e di Sistema Istituto Superiore di Sanitft, Viale Regina Elena, 299, 1-00161 Roma, Italy

Rece ived 1 February 1990

LAVIOLA, G., L. DE ACETIS, G. BIGNAMI AND E. ALLEVA. Prenatal oxazepam enhances mouse maternal aggression in the offspring, without modifying acute chlordiazepoxide effects. NEUROTOXICOL TERATOL 13(1) 75-81, 1991.--In the rat, behavioral changes during lactation are in several ways similar to those produced by benzodiazepines (BDZ). Moreover, an increased activity at the GABA/BDZ receptor complex has been found in both conditions. We tested the hypothesis that early manipulation of this neurocbemical system by prenatal BDZ exposure should affect typical responses of lactating dams, such as mammal aggression. Outbred CD-1 mice were treated with either oxazepam (15 mg/kg PO twice/day on days 12-16 of fetal life) or vehicle and fostered at birth to untreated dams. Female offspring were subsequently mated at the young-adult stage and used to assess maternal aggressive responses towards a male intruder. In a first 5-rain test on postpartum day 6, the prenatal oxazepam animals showed a reduced Latency to the First Attack, a markedly enhanced frequency of several offensive scores (such as Fighting Episodes, Attacks, and Offensive Upright, On Top, and Kicking Postures), a decrease of Submissive Postures and a reduced duration of time spent lying still (Out of the Nest). The tests were repeated 48 h later after IP treatment by either chlordiazepoxide (10 mg/kg) or saline. The drug significantly enhanced locomotor activity as well as the frequency of Fighting Episodes and of Attacks, while decreasing the number of Submissive Postures and the time spent On Nest. These effects were not significantly modified by prenatal oxazepam exposure. This suggests that long-term and acute effects of benzodiazepines are produced either by changes in different regulatory systems or by different types of changes in the same system.

Prenatal oxazepam exposure Adult social behavior Mammal aggression Chlordiazepoxide challenge Mouse

ONE way to evaluate the nature and specificity of the proactive developmental effects of any given type of agent is to study behavioral responses modulated by those mechanisms that are known to be affected in the presence of the agent. From this viewpoint, it appears that the study of maternal behavior of altricial rodents, such as mice and rats, can contribute significant information on the nature of long-term effects produced by early benzodiazepine (BDZ) exposure. In fact, lactating animals show behavioral changes that are in many ways similar to those produced by acute BDZ treatment (19), and the latter can further magnify some of these changes (39,54).

Specifically, mother rats eat more, are less fearful or anxious [see e.g., (11)], and develop a strong tendency to attack conspecifics (18). Rats also freeze less in response to a sudden environmental change (i.e., an auditory signal) during lactation than during other stages of the reproductive cycle (20). Such a profile suggests a functional shift in defense strategy from fear-emotionality responses to more effective goal-directed responses aimed at offspring protection.

At a neurochemical level, it has been suggested on the basis of psychopharmacological observations that activity at the GABA/

benzodiazepine receptor complex in specific brain areas might be enhanced during rat motherhood (18, 19, 45, 46). However, a recent study by the same group measured ligand binding to several sites on the complex and found no differences between maternal lactating and virgin rats (11). GABA in cerebrospinal fluid is considerably elevated in lactating rats, and pup removal de- creases GABA levels within six hours (45). Moreover, the activity of glutamic acid decarboxylase (a key enzyme in GABA biosynthesis) is increased in the mediobasal hypothalamus of lac- taring rats (46).

As concerns hormonal factors that may be involved in the changes in GABA/BDZ mechanisms during lactation, the ovarian sex steroids estradiol and progesterone are known to affect GABA/ BDZ receptors in several brain areas (32). Moreover, progesterone metabolites have been shown to increase the binding of the BDZ flunitrazepam to benzodiazepine sites, and to be potent barbiturate-like ligands of the GABA receptor complex (33), while progesterone produces anxiolytic effects in the female, but not in the male rat (48). On the other hand, it is known that acute stress rapidly and reversibly affects both BDZ (23,35) and GABA (6) binding in the brain. Since fear/anxiety patterns are a function of

75

76 LAVIOLA, DE ACETIS, BIGNAMI AND ALLEVA

the whole adaptive response displayed by a rodent in a social interaction, particularly in the case of aggression (47), the various physiological changes apparently fit well with the behavioral ad- aptations that occur at the time of parental care.

In the present study, the behavioral repertoire of female mice prenatally exposed to BDZ was assessed during a specific reproductive stage, i.e., the early postpartum period. The attention was focussed on a particular form of mouse aggressive behavior, namely, maternal aggression, which is known to be enhanced by acute BDZ treatments [(36, 39, 54); for more details see the Dis- cussion section]. The BDZ agent (oxazepam) and dosing schedule for prenatal treatment were the same as those used in several previous studies in the present long-term project after the assess- ment of dose-response relations [(2-4, 29); see the Method section]. It was also deemed advisable to assess whether or not prenatal BDZ exposure can produce long-term changes in reactivity to the same type of agent. For this purpose chlordiazepoxide, a BDZ whose effects on various types of aggressive responses have been thoroughly studied [for adult male aggression see (47); for female aggression see (39,54)], was used as an acute drug challenge prior to maternal aggression tests.

METHOD

Animals, Breeding, and Prenatal Treatment

Mice of an outbred Swiss-derived strain (CD-1) weighing 25- 27 g were purchased from a commercial breeder (Charles River Italia, 1-22050 Calco). Upon arrival at the laboratory, the animals were housed in an air-conditioned room (temperature 21---I°C, relative humidity 60___ 10%) with lights on from 9.30 p.m. to 9.30 a.m. Males and nulliparous females were housed separately in groups of 8-10 in 42 x 27 x 15 cm Plexiglas boxes with saw- dust as bedding and a metal top. Pellet food (enriched standard diet purchased from Piccioni, 1-25100 Brescia) and tap water were continuously available. After 2-3 weeks, breeding pairs were formed and housed in 33 x 13 x 14 cm boxes. The females were inspected twice daily at 9 a.m. and 8 p.m. for delivery (postnatal day 1). The stud was removed ten days after the finding of the plug. All litters were reduced at birth to four males and two females and fostered to untreated dams of the same strain that had given birth to healthy litters within 24 h.

Oxazepam (Agrar, 1-00195 Roma) was suspended in a 0.5% solution of sodium carboxymethylcellulose (Fluka AG, Switzer- land) in water. Females were treated per os twice daily (between 9 and 10 a.m. and between 7 and 8 p.m.) on pregnancy days 12-16. Dams received either oxazepam (15 mg/kg in each treatment) in a volume of 0.01 ml per g body weight or vehicle (N = 16 per group). This treatment schedule, used in several previous studies (2-4, 29), was originally adopted on the basis of pharma- codynamic considerations, of preliminary behavioral assessments, and of developmental data from an initial multidose experiment that covered the 5-50 mg/kg range [see particularly (4)].

Dams remained with pups until weaning, which took place on postnatal day 21. When weaned, groups of four female individu- als were randomly formed and maintained in 4 2 x 27 x 15 cm Plexiglas boxes. When adult (i.e., 10 weeks of age), the female offspring were mated with a sexually experienced adult male (breeding procedures were as reported above). After male removal, pregnant females were housed individually. At delivery the length of pregnancy, the number of viable pups, and sex-ratio were recorded, and all litters were culled to six male pups. No additional material was provided for nest building; nests were lo- cated in most instances under the metallic food holder.

Maternal Aggression Tests Aggressive behavior testing was carried out on postnatal day

6 (day of delivery was considered as day 1), when maternal aggression is expected to be at a high level [(44,51); however, the pattern of maternal aggression does not undergo significant changes between postpartum day 3 and 9 (43,53)]. Adult male mice of the same strain, individually housed for 1-2 weeks in 33 x 13 x 14 cm boxes, were used as intruders. All males had previous mating experience, although not in the 30 days before testing. The male, previously marked by a black spot on its back, was introduced for 5 min into the home cage of the female in the presence of her litter.

During the test session, the animals were continuously video- taped using a Sony VO-5630 apparatus equipped with CH-1400CE videocameras for red lights. Recordings were scored by an ob- server blind to the assignment of animals to different groups, ac- cording to a score list of elements of female behavior, most of which are defined in previous reports (14, 42, 53): 1) Latency to the First Attack, that is, the time (in seconds) elapsed from the introduction of the intruder to the first biting attack by the female; 2) number and duration of Fighting Episodes (bouts of aggressive encounter involving actual body contact, often followed by a bite, separated from a successive episode by at least 4 s, irrespec- tive of the subject which initiates the aggressive intercourse); 3) number of Attacks by the female; 4) number of Tail Rattling Ep- isodes; 5) number of Upright Offensive Postures that may or may not be accompanied by boxing; 6) number of On Top Postures occurring when the female holds down the opponent; 7) frequency of Kicking (female kicks with a hindleg at the intruder, irrespec- tive of the posture of the latter); 8) number of Evade Episodes; 9) number of crouching Submissive Postures; 10) frequency and duration of episodes of On Nest (female in the nest nursing pups); 11) frequency and duration of episodes of Out Nest (female lying still out of the nest).

The locomotor activity of the female mice during the 5-min testing period was also scored by measuring the number of Cross- ings. For this purpose, the floor of the cage was subdivided by four black vertical lines; passing a line with the head and the forepaws was the response criterion.

Chlordiazepoxide Challenge

On lactation day 8, eight females out of the 16 in each prenatal treatment group were randomly assigned to one of two treatment conditions (injections IP, in a volume of 0.01 mug body weight): saline solution (NaC1 0.9%) or chlordiazepoxide HC1 (CDP) (Agrar, 1-00195 Roma). Drug dosage (10 mg/kg) was cho- sen on the basis of literature data (38,53) supplemented by indications obtained in preliminary tests using mice of the same age, sex and strain, and a 5-20 mg/kg dosage range. Upon injection, the animals were immediately returned to their respective home boxes. Thirty min later, the dams underwent the same maternal aggression test of postnatal day 6.

All tests were performed in dim red light between 9.30 and 12 a.m., i.e., during the animals' most active period in the initial hours of the dark phase of the 24-h lighting regime. The design of the experiments followed as closely as possible the method- ological recommendations of the Collaborative Behavioral Tera- tology Study group (27), and the ethical recommendations of Huntingford (22) and Bateson (5).

Design and Statistical Analysis

Separate ANOVAs considering prenatal exposure (vehicle vs. oxazepam) and treatment before test (saline vs. chlordiazepoxide) were used. To avoid the bias due to litter effects (8,10), one or at most two animals from the same litter were used.

PRENATAL OXAZEPAM AND MATERNAL AGGRESSION 77

c= 10'

[ ] Prenatal vehicle

• Prenatal oxazepam

FIG. 1. Frequencies of selected behavioral responses recorded during 5- min maternal aggression tests (nursing female mice exposed to male intruders in the presence of their litters on postpartum day 6). The dams had been exposed prenatally to either oxazepam administered to their mothers (15 mg/kg twice/day on pregnancy days 12-16) or to vehicle. Data are means (S.E.M.) of 16 animals.

RESULTS

Body Weight and Reproductive Performance

After weaning, growth rates of drug-treated and control offspring were very similar. Thus, by 10 weeks, female controls weighed 41.44 ± 0.5 g and oxazepam females weighed 40.25 ± 1.1 g. Length of pregnancy was 18.0 ± 0.16 and 18.3 ± 0.17 days for vehicle and for oxazepam females, respectively. The offspring sex-ratio (No. of males/total No. of pups) was 0 .50±4 .24 and 0.46 ± 2.24, respectively. The mean number of pups born viable

was 12.0±0.47 and 13.1 ±0.59, respectively. As a whole, no significant effects of prenatal exposure were evident.

Maternal Aggression Towards a Male Intruder (Lactation Day 6, No Acute Treatment)

Results are shown in Fig. 1 (response frequencies) and Table 1 (response durations). Relative to the vehicle controls, the animals prenatally exposed to oxazepam showed a significant increase in frequency of Fighting Episodes, F(1,30)=19.31, p<0.001, Attacks, F(1,30)= 13.24, p<0.001, Offensive Upright Postures, F( I ,30)= 8.83, p<0.005, On Top Postures, F(1,30)= 6.34, p<0.01, and Kicking Postures, F(1,30)=4.40, p<0.05. Correspondingly, Submissive Postures were adopted less fre- quently by oxazepam than by control mice, F(1,30)= 10.61, p<0.005. The change in the frequency of Evade behaviors, which was small and apparently in the direction of an increase, missed statistical significance, F(1,30) = 3.10, p =0.08. Frequencies of both On Nest and Out Nest Episodes did not differ significantly between the two groups. The data on Tail Rattling were excluded from the analysis, since the number of these responses was too small for a reliable evaluation.

Relative to the vehicle controls, prenatal oxazepam mice also showed a shorter Latency to First Attack, F(1,30)=5.85, p<0.05, a higher duration of Fighting Episodes, F(1,30) = 17.06, p<0.001, but a shorter duration of Out Nest Episodes, F(1,30) = 4.00, p<0.05. The latter difference can be easily explained by the fact that oxazepam animals were less prone to lie still, and also more often engaged in aggressive intercourses. No significant difference in On Nest duration was found. The changes so far described were unlikely to be due to changes in overall activity, since no difference in the number of Crossings was found (data on the left in Fig. 3).

Chlordiazepoxide Effects on Maternal Aggression Towards a Male Intruder at Lactation Day 8

The data from the retest with or without the chlordiazepoxide (10 mg/kg) challenge are reported in Table 1 and in Figs. 2 and 3. The differences due to prenatal treatment were less marked than in the previous tests, but essentially in the same direction. Prenatal oxazepam mice showed a higher frequency of Offensive Upright Postures, F(1,28) = 8.86, p<0.005, and a shorter Latency to the First Attack, F(1,28)=4.93, p<0.05. Three other scores

TABLE 1

DURATION (S) OF SELECTED BEHAVIORAL RESPONSES IN MOUSE MATERNAL AGGRESSION TESTS*

No-Drug State (Lactation day 6) CDP Challenge (Lactation day 8) Prenatal Vehicle Prenatal Oxazepam Prenatal Vehicle Prenatal Oxazepam

Saline CDP Saline CDP (N-- 16) (N = 16) (N = 8) (N = 8) (N = 8) (N = 8)

Latency to 137.7 (-+37.1) 36.8 (--- 19.0) 263.1 (-+36.8) 153.1 (-+55.7) 87.0 (_-.43.6) 115.5 (-+53.9) First Attack

Durationof 3.4 (-+1.3) 19.6 (-+3.7) 0.25(---0.2) 2.5 (-+1.9) 1.3 (-+0.2) 8.7 (-+3.7) Fighting Episodes

Duration of 21.3 (-+5.9) 19.4 (-+3.9) 159.9 (_+25.4) 21.5 (-+31.4) 139.5 (-+38.2) 55.9 (-+24.1) On Nest

Durationof 35.5 (-+7.48) 15.9 (--+6.33) 21.9 (-+5.6) 65.1(-+20.3) 37.3(-+29.6) 33.4(-+13.7) Out Nest

*For other indications see legends of Fig. 1 (tests on lactation day 6) and Fig. 2 (tests on lactation day 8).

78 LAVIOLA, DE ACETIS, BIGNAMI AND ALLEVA

100

day 6 day 8

SALINE COP

to

to to

c,. t . j

50

0

[ ~ Prenatal vehicle • Prenatal oxazepam

FIG. 2. Mean number of Crossings recorded during 5-min tests of agonistic confrontation towards male intruders by nursing female mice prenatally exposed to either vehicle or oxazepam (postnatal day 6 on the left; postnatal day 8--CDP challenge--on the right). Data are means (S.E.M.) of 16 or 8 animals, respectively, and refer to the same animals of Fig. 1.

(Fighting, Attacks, and Kicking) were higher in oxazepam than in vehicle mice, but the differences just missed statistical significance, F 's(1 ,28)=3.61, 3.61, 3.67, respectively; p ' s = 0 . 0 6 .

Acute chlordiazepoxide treatment significantly increased the number of Fighting Episodes, F(1,28) = 5.20, p<0.05, of At- tacks, F(1,28) = 5.20, p<0.05 , and of Crossings, F(1,28) = 3.96, p<0.05 (see Fig. 3), while decreasing the frequency of Submis- sive Postures, F(1,28)= 4.33, p<0.05. No significant chlordiazepoxide effects on Offensive Uptight Postures, On Top, Kicking, Evade, On Nest, and Out Nest were observed. Chlordiazepoxide significantly increased the duration of Fighting Episodes, F(1,28) =

5.06, p<0.05, and decreased On Nest duration, F(1,28)=4.04, p<0.05, while Latency to the First Attack and duration of Out Nest were unaffected.

Some of the effects of acute chlordiazepoxide, particularly those on Fighting Episodes, Attacks, Uptight Offensive Postures and Crossings, were apparently more marked in prenatal oxazepam than in vehicle mice. These differences, however, were not supported by ANOVA results since no significant interactions were found between the prenatal and the acute treatment variables.

Overall, comparisons between the data from the saline mice in these tests and those of the tests performed two days earlier indi- cate that aggressive responses were reduced from the first to the second exposure to male intruders. Such a reduction was more marked in oxazepam than in vehicle mice, which was obviously related to the much higher aggression scores in the former than in the latter animals in the first test.

Finally, the results so far described were apparently not bi- assed by possible confounds originating with the male intruder. Specifically, in no case was infanticide by the intruder observed; dams were constantly successful in preventing the intruder from entering the nest area. Retaliatory attacks and mating attempts occurred very rarely, without differences between the various groups.

DISCUSSION

The data reported here clearly show specific changes in the behavioral repertoire of lactating female mice after prenatal BDZ treatment. In summary, nearly all indexes of maternal aggression towards a male intruder were elevated, while the frequency of Submissive Postures was decreased. On the other hand, there were no changes in the time spent with pups in the nest, which ex- cludes substantial modifications of maternal care.

The specificity of the proactive oxazepam effects is further confirmed by the absence of changes in locomotor activity. This apparently denies that the enhanced aggression can be ascribed to "hyperarousal," an explanation that has been used to account for prenatal diazepam effects in rats (34). It seems rather that the treatment can produce specific changes in the mechanisms that

SALINE COP SALINE COP SALINE EDP SALINE CDP SALINE CDP

J J J i L. ~ Fighting Attacks Upright Offensive On lop Kick=ng

,,... episodes postures

~ 5 [ ] Prenatal vehicle

• Prenatal oxazepam

0 ~ ITI I ' ~ Evade Submissive On Nest Out Nest

postures

FIG. 3. Data obtained by retesting on postnatal day 8 the same nursing mice of Fig. 1. One-half of the animals in each of the original groups (prenatal oxazepam or vehicle) was injected with chlordiazepoxide (10 mg/kg IP, CDP) 30 min before testing, while the other half was treated with the same volume of saline.


modulate various components of the aggressive/defensive repertoire including species-specific aggression-inhibiting displays, In particular, goal-directed, offspring protective, responses (Attacks and Offensive Upright, On Top, and Kicking Postures) were fa- vored at the expense of the fear-emotionality components of the behavioral repertoire.

These preliminary inferences need to be further discussed in relation to the mechanisms which are involved in modulation of various types of aggressive responses and in production of BDZ effects [for a study of the analogies and differences between rat and mouse maternal aggression, see (17)]. Rodent maternal aggression has been shown to be a testicular-hormone-independent form of aggressive behavior, which is characteristic of both go- nadally intact and ovariectomized adult females (21). Moreover, the unique hormonal status of lactating females [i.e., high prolactin levels (7)] is quite different from that of the males, which are usually employed when studying the determinants and mechanisms of aggressive responses. From a behavioral viewpoint, stimuli that are necessary to initiate maternal aggression (e.g., suckling) do not play a role in male aggression [for more infor- marion on the role of environmental and organismic variables in maternal aggression, see (1, 13, 15, 37, 41, 49)].

Although there is evidence against a role of CNS mechanisms relying on BDZ receptors in the modulation of maternal aggression (38), ample data point to a specificity of acute BDZ effects on maternal aggressive responses. Up to a certain dose, these effects are clearly proaggressive [(36, 39, 40, 54) and present study], while a shift from facilitation to reduction of aggression can occur at different dose levels, depending apparently on the baseline. Specificity is supported by the lack of substantial effects on the pattern of pup care and self-directed activities (body care). Quite interestingly, an increase in the level of "introductory" social exploration parallels the increase in aggressive behavior (52). Overall, acute BDZ effects are best accounted for by postu- lating a subtle change in the balance between different responses that can be triggered by the approach/avoidance conflict created by the testing situation, apparently without significant modifications in the structure of maternal agonistic behavior (41, 43, 52).

It appears at this point that there are interesting analogies between the long-term effects of prenatal BDZ exposure on maternal aggression and those of acute treatments given shortly before testing. It should also be noted that perinatal exposure to diazepam produces an increase of aggressive responses at the adult stage of male rats (12,16), while the effects on other components of the emotional repertoire appear to be variable; for example, after neonatal BDZ there were no changes in several tests of anxiety, but the effects of an anxiogenic drug such as yohimbine in a social interaction test were reduced (12). Overall, this suggests that in spite of the differences between male and maternal aggression (see above) the two repertoires have at least some of the mechanisms in common, particularly those which are sensitive to early influences (see later).

As concerns the mechanisms by which long-term BDZ effects may be produced, the data extend previous models based on the finding of specific changes in sensory functions, such as modifications of stimulus interactions in the triggering of startle responses (26). In general, these changes have been shown to be mainly in the domain of arousal-attention and stress-related phe-

nomena [see also (24, 31, 34)]. Therefore, it would be useful to assess whether or not prenatal BDZ exposure produces specific changes in sensitivity to stimuli that normally trigger (or favor), or vice versa inhibit, the display of various components of the maternal repertoire, including aggressive responses towards male intruders.

As pointed out in the Introduction, lactating altricial rodents show changes in CSF GABA levels that must be taken into account when attempting to explain the behavioral changes (45,46). On the other hand, the activity of GABA-synthesizing enzyme (GAD) was either unmodified or increased in two experiments using different daily doses of prenatal diazepam (2.5 and 1 mg/ kg daily, respectively), and different fostering procedures (cross- fostering versus no cross-fostering of newborn rat pups to untreated dams) (25,50). While the functional links between the components of the large GABA/BDZ receptor complex are still poorly understood, it may have at least heuristic value to focus the attention on possible alterations in the development of such a system when attempting to understand the effects of early BDZ exposure.

From this viewpoint, one must also consider the fact that the long-term BDZ effects on maternal aggression were not accompanied by a change in the proaggressive effect of an acute BDZ challenge. A caveat here must be that the latter effect was assessed in a retest that showed a lowered baseline of aggressive responding. Moreover, only one dose of the acute BDZ treatment was used since a dose-response evaluation would have required a much larger number of litters in both original groups (prenatal BDZ and control).

With these limitations, the profile so far discussed can be viewed in the light of the fact that distinct BDZ receptors exist in the brain, for which benzodiazepines have equal affinity. An ad- equate functional interpretation of the respective roles of these receptor systems is still lacking; however, it is known that their developmental trends are different. Specifically, Type I receptors show only neonatal proliferation, while Type II receptors (that is, those which are part of the GABA/BDZ receptor complex) show mostly prenatal proliferation (28,30), making it likely that only the latter are affected by prenatal BDZ treatment. The data, however, also show regional variation in receptor profile development (9,28). Therefore, any hypothesis concerning differential effects of prenatal BDZ treatments can only be highly tentative.

It is reasonable to speculate at this point that the effects of early influences (including prenatal BDZ exposure) and those of events occurring at the mature stage (including acute BDZ treatments) may be mediated either by changes in different functional systems or by different types of changes within the same system. These eventualities must be taken into account in further attempts to clarify the role of BDZ mechanisms in behavior modulation and the nature of possible differences between long-term and acute BDZ effects.

ACKNOWLEDGEMENTS

This research was supported as part of the Sub-project on Neurobe- havioral Pathophysiology (Project on Non-infectious Pathology) of the Istituto Superiore di Santii~, and by grant No. 89.04677.CTO4 from the Consiglio Nazionale delle Ricerche. We acknowledge Dr. Flavia Chia- rotti for expert statistical advice.

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prenatal oxazepam enhances mouse maternal aggression in the offspring, without modifying acute...

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