Melissa Morales

Effects of ethanol administration on corticosterone levels in adolescent and adult rats

Adolescent humans and rodents have been shown to consume more alcohol than their adult counterparts. Given that corticosterone (CORT) has been shown to be related to the intake of several drugs of abuse, this study assessed the ontogenetic effects of low-moderate doses of ethanol on CORT increases and recovery. Despite no significant differences in baseline (home cage) CORT levels, CORT responses to ethanol were greater in females than in males and in adult females than in adolescent females; males, however, showed less marked age differences in CORT levels after ethanol consumption. Adolescent blood ethanol concentrations (BECs) were lower than those of adults, although these BEC differences appear insufficient to account for the ontogenetic differences in CORT levels. Collectively, these findings suggest that it is unlikely that age differences in CORT elevations provide a major contribution to the ontogenetic differences in alcohol intake seen between adolescents and adults.

Low doses of the NMDA receptor antagonists, MK-801, PEAQX, and ifenprodil, induces social facilitation in adolescent male rats

Adolescents display high levels of interactions with peers relative to other age groups, with these interactions further enhanced by ethanol under some circumstances. Understanding of the neural mechanisms underlying these high levels of social interactions is important given that alcohol use is initiated during adolescence and adolescents tend to report drinking for social reasons. Given that ethanols effects are associated in part with functional antagonism of the NMDA receptor system, the current experiment explored the role of NMDA antagonists for facilitating adolescent social behavior. Adolescent male Sprague-Dawley rats were challenged acutely with either the non-competitive NMDA antagonist, MK-801 (0.01, 0.03mg/kg), the NR2A antagonist, PEAQX (1.25, 3.75mg/kg) or the NR2B antagonist, ifenprodil (0.75, 2.25mg/kg) 30min prior to a 10-min social interaction test. All compounds generally increased overall social activity (i.e., sum of social investigation, contact behavior, and play), with ifenprodil also significantly enhancing play and social contact behaviors. Although the frequencies of peer-directed social behaviors were typically greater following administration with these NMDA antagonists, social preference, indexed via the number of crossovers to the side with the partner relative to crossovers away, was significantly reduced in MK-801 and PEAQX-treated rats. None of these changes were associated with concomitant alterations in overall locomotor activity under these test circumstances. These data support the suggestion that the increases in social interactions observed in adolescents following acute ethanol may be driven in part by NMDA receptor antagonism - particularly of the NR2B subunit - given that ifenprodil stimulated social behavior in a manner similar to that produced by low doses of ethanol.

Stress, kappa manipulations, and aversive effects of ethanol in adolescent and adult male rats

Elevated ethanol use during adolescence, a potentially stressful developmental period, is accompanied by insensitivity to many aversive effects of ethanol relative to adults. Given evidence that supports a role for stress and the kappa opioid receptor (KOR) system in mediating aversive properties of ethanol and other drugs, the present study assessed the role of KOR antagonism by nor-binaltorphimine (nor-BNI) on ethanol-induced conditioned taste aversion (CTA) in stressed (exposed to repeated restraint) and non-stressed male rats (Experiment 1), with half of the rats pretreated with nor-BNI before stressor exposure. In Experiment 2, CTA induced by the kappa agonist U62,066 was also compared in stressed and non-stressed adolescents and adults. A highly palatable solution (chocolate Boost) was used as the conditioned stimulus (CS), thereby avoiding the need for water deprivation to motivate consumption of the CS during conditioning. No effects of stress on ethanol-induced CTA were found, with all doses eliciting aversions in adolescents and adults in both stress conditions. However, among stressed subjects, adults given nor-BNI before the repeated stressor displayed blunted ethanol aversion relative to adults given saline at that time. This effect of nor-BNI was not seen in adolescents, findings that support a differential role for the KOR involvement in ethanol CTA in stressed adolescents and adults. Results from Experiment 2 revealed that all doses of U62,066 elicited aversions in non-stressed animals of both ages that were attenuated in stressed animals, findings that support a modulatory role for stress in aversive effects of KOR activation. Collectively, these results suggest that although KOR sensitivity appears to be reduced in stressed subjects, this receptor system does not appear to contribute to age differences in ethanol-induced CTA under the present test circumstances.

Effects of the kappa opioid receptor antagonist, nor‐binaltorphimine, on ethanol intake: Impact of age and sex

The kappa opioid receptor (KOR) antagonist, nor-binaltorphimine (nor-BNI), was used to investigate the role of the KOR system in mediating ethanol intake. On P25 (adolescent) or P67 (adult) male and female rats were individually housed and given ad libitum access to food and water. The experimental procedure was initiated on P28 or P70: animals were given 30 min/day access to a 10% ethanol/supersaccharin solution every other day (3 baseline exposures). On the day after the final baseline test, rats were injected with nor-BNI (0, 2.5, 5, 10 mg/kg), with testing initiated 24 hr later (30-min access every other day, 3 test exposures). Nor-BNI (10 mg/kg) increased ethanol intake in adult males, whereas the same dose decreased intake in adult females, suggesting pronounced sex differences in KOR-associated mediation of ethanol intake in adulthood. There was no impact of nor-BNI in adolescent animals of either sex, suggesting that the KOR may play less of a role in modulating ethanol intake during adolescence.

AGE DIFFERENCES IN THE EXPRESSION OF ACUTE AND CHRONIC TOLERANCE TO ETHANOL IN MALE AND FEMALE RATS

BACKGROUND Ontogenetic differences in response to ethanol (EtOH) challenge have been observed under a variety of circumstances, including varying reports of developmental differences in the expression of tolerance to EtOH. The purpose of the present experiment was to further explore potential differences in acute (AT) and chronic (CT) tolerance expression between adolescent and adult, male and female Sprague-Dawley rats, using the social interaction test. METHODS AT and CT to the social suppressing effects of a moderate dose of EtOH was assessed in adolescent and adult rats following intraperitoneal injections of 2.0 g/kg EtOH or saline daily for 10 days. At test, adults and adolescents were challenged with 1.0 or 1.25 g/kg EtOH, respectively, with AT and CT assessed at 5 and 25 minutes postinjection using ratios of impairment to brain ethanol concentrations (BrECs) at each time period (CT) and within-session declines in impairment relative to BrECs (AT). RESULTS In adolescents, 10 days of EtOH pre-exposure resulted in evidence of CT at 25 minutes postinjection, perhaps associated with an enhanced expression of AT. Among adults, signs of CT were seen at 5 minutes postinjection in adults, and may reflect neuroadaptations unassociated with AT, as with evidence of tolerance emerging only in adult control animals repeatedly exposed to saline injection prior to EtOH challenge on test day. Sex differences in tolerance expression were not observed at either age. CONCLUSIONS Our results show ontogenetic differences between adolescents and adults in the short- and long-term neuroadaptations that they express in response to repeated perturbations with EtOH. Together these findings add age of exposure and time of testing within the intoxication period as critical variables to be considered when exploring the complex relationship between AT and CT.

Conditioned taste aversion to ethanol in a social context: impact of age and sex.

Given that human adolescents place a high value on social interactions-particularly while consuming alcohol-the current study utilized a novel social drinking paradigm to examine rewarding and aversive properties of ethanol in non-water deprived rats that were housed and tested in groups of five same-sex littermates. On postnatal day P34 (adolescents) or P69 (adults), rats were habituated to the testing apparatus for 30 min. On the next day, animals were placed into the test apparatus and given 30 min access to a supersaccharin solution (3% sucrose; 0.125% saccharin), followed immediately by an intraperitoneal injection of ethanol (0, 0.25, 0.5, 1.0, 1.5 g/kg). Subsequent intake of the supersacharrin solution was assessed on three consecutive test days. Adolescent males were less sensitive to ethanols aversive effects than adult males, with adolescent males maintaining an aversion on all three test days only at the 1.5 g/kg dose, whereas adults demonstrated aversions across test days to 1 and 1.5 g/kg. Adolescent females maintained aversions to 1 and 1.5 g/kg across days, whereas adult females continued to show an aversion to the 1.5 g/kg dose only. These opposite patterns of sensitivity that emerged among males and females at each age in the propensity to maintain an ethanol-induced taste aversion under social conditions may contribute to age- and sex-related differences in ethanol intake. Testing in social groups may be useful for future work when studying rodent models of adolescent alcohol use given the importance that human adolescents place on drinking in social settings.

Anxiolytic effects of the GABAA receptor partial agonist, L-838,417: Impact of age, test context familiarity, and stress

The partial α2,3,5 GABA(A) receptor agonist, L-838,417 has been reported to have anxiolytic effects in adult rodents. Although maturational differences exist for the GABA(A) receptor subunits, the anxiolytic effects of L-838,417 have not been tested in younger animals. The goal of the present experiments was to determine whether L-838,417 reverses anxiety-like behavior induced by either an unfamiliar environment (Experiment 1) or repeated restraint stress (Experiment 2) differentially in adolescent and adult, male and female Sprague-Dawley rats using a modified social interaction test. In Experiment 1, rats were injected with 0, 0.5, 1.0, 2.0, or 4.0 mg/kg L-838,417, i.p. and tested 30 min later in an unfamiliar test context for 10 min. In Experiment 2, rats were exposed to restraint stress (90 min daily for 5 days). Immediately after the last restraint session, animals were injected with L-838,417 and placed alone for 30 min in the test apparatus to familiarize them to this context prior to the 10 min social interaction test. In Experiment 1, L-838,417 produced anxiolytic effects in adults at 1.0 mg/kg, as indexed by a transformation of social avoidance into preference and an increase in social investigation. In adolescents, a dose of 2.0 mg/kg eliminated social avoidance, but had no anxiolytic effects on social investigation. Testing under familiar circumstances (Experiment 2) after repeated restraint stress eliminated age differences in sensitivity to L-838,417, with 0.5 mg/kg reversing the anxiogenic effects of prior stress regardless of age, but with doses ≥ 1 mg/kg decreasing social investigation, an effect possibly due in part to locomotor-impairing effects of this compound. Although locomotor activity was suppressed in both experiments, higher doses of L-838,417 were necessary to suppress locomotor activity in Experiment 1. Thus, anxiolytic effects of L-838,417 were found to be context-, age-, and stress-dependent.

Differences in sensitivity to ethanol-induced conditioned taste aversions emerge after pre- or post-pubertal gonadectomy in male and female rats

We have previously demonstrated that gonadectomy either prior to (early) or after (late) puberty elevated ethanol consumption in males to levels similar to intact adult females-effects that were attenuated by testosterone replacement. To assess whether alterations in the aversive effects of ethanol might contribute to gonadectomy-associated increases in ethanol intake in males, the present study examined the impact of gonadectomy on conditioned taste aversions (CTA) to ethanol in male and female Sprague-Dawley rats. Animals were gonadectomized, received sham surgery (SH) or non-manipulated (NM) on postnatal (P) day 23 (early) or 67 (late) and tested for CTA to ethanol in adulthood. Water-deprived rats were given 1 hr access every-other-day to 10% sucrose followed by an injection of ethanol (0, 1g/kg) for 5 test sessions. Test data were analyzed to determine the first day significant aversions emerged in each ethanol group (i.e., sucrose intakes significantly less than their saline-injected counterparts). Early gonadectomized males acquired the CTA more rapidly than did early SH and NM males (day 1 vs 3 and 4 respectively), whereas a gonadectomy-associated enhancement in ethanol CTA was not evident in late males. Among females, gonadectomy had little impact on ethanol-induced CTA, with females in all groups showing an aversion by the first or second day, regardless of surgery age. These data suggest that previously observed elevations in ethanol intake induced by either pre- or post-pubertal gonadectomy in males are not related simply to gonadectomy-induced alterations in the aversive effects of ethanol indexed via CTA.

EVIDENCE FOR CONDITIONED PLACE PREFERENCE TO A MODERATE DOSE OF ETHANOL IN ADULT MALE SPRAGUE-DAWLEY RATS

The present series of experiments examined affective properties of a moderate dose of ethanol using the conditioned place preference (CPP) paradigm in ethanol-naïve, adult male Sprague-Dawley rats. The apparatus and the procedure used were both unbiased. In Experiment 1, rats were given four 30 min conditioning sessions with 1.5 g/kg ethanol (i.p.) or an equivalent volume of saline on the paired side. Animals were found to demonstrate CPP to the ethanol-paired side, an unexpected finding at this relatively high dose in rats. To replicate this finding, and to examine the possibility of non-associative conditioning, an unpaired control group was included in Experiment 2. Once again, rats showed a CPP to the side paired with ethanol relative to either control group. Given that testing in an unfamiliar environment typically results in elevated levels of anxiety and that animals in Experiments 1 and 2 were not exposed to the apparatus prior to conditioning, Experiment 3 was conducted to examine the potential role of context unfamiliarity for induction of ethanol CPP in this test situation by varying whether animals were exposed to the apparatus prior to conditioning. In this study, pre-exposure to the CPP apparatus was found to eliminate the CPP to ethanol observed in rats who were not familiarized with the apparatus. Collectively, these studies demonstrate that ethanol-naïve rats can find ethanol reinforcing as indexed by the CPP test, and provide some evidence for the conditions under which this uncommon finding is observed.

Pre-pubertal gonadectomy and the social consequences of acute ethanol in adolescent male and female rats.

It has previously been shown that pre-pubertal or adult gonadectomy (GX) increases ethanol intake in male rats. This study examined whether this sex-selective increase reflects a GX-induced maintenance in males of more adolescent-typical responsiveness to ethanol characterized by enhanced sensitivity to positive (e.g., socially facilitating) and a decreased sensitivity to adverse (e.g., socially inhibitory) effects of ethanol. Male and female Sprague-Dawley rats were pre-pubertally GX, sham (SH)-operated, or non-manipulated (NM) at postnatal day (P) 25. During the late adolescent transition into adulthood (P48 - baseline day), rats were given a saline injection, placed alone into a familiar test apparatus for 30min and then exposed for 10min to an unfamiliar partner of the same age and sex. On the following day (P49), similar testing occurred after administration of 0.5, 0.75, 1.0 or 1.25g/kg ethanol. At baseline, GX males and females displayed higher levels of social activity (especially adolescent-typical play and contact behavior) than SH and NM animals, with GX females displaying greater social activity than GX males. Neither males nor females demonstrated social facilitation at lower ethanol doses, regardless of hormonal status. Whereas the social inhibitory effects of higher doses of ethanol were similar across groups among females, SH males were less sensitive than both GX and NM males to ethanol-induced social inhibition. These results suggest that enhanced ethanol intake in GX males is not related to alterations in sensitivity to ethanols social inhibitory effects. GX, however, results in retention of adolescent-typical social behaviors, with older GX adolescent rats resembling early adolescents in exhibiting elevated social activity-particularly play and contact behavior.