Effects of prenatal alcohol exposure on behavior of aged rats

Pregnant rats were intubated with alcohol (3.5 g/kg, twice daily) on gestation days 11-21. Control animals were intubated with an isocaloric sucrose solution and were pair-fed and pair-watered to alcohol-treated dams. At birth, offspring were placed with non-treated surrogate dams. When animals were slightly more than 1 year of age, they were tested for passive avoidance learning, spontaneous alternation and activity. Groups did not differ in passive avoidance learning or spontaneous alteration but animals prenatally exposed to alcohol were more active than controls. Additional studies showed that this increased activity was not affected by testing animals in the presence of environmental stimuli such as objects which could be manipulated, or by odors from mouse shavings from male and female mice.     

Differential effects of long-chain alcohols in long- and short-sleep mice

Observations that the long-sleep (LS) and short-sleep (SS) mouse lines differ in their depressant response to barbiturates, and that the difference between lines becomes greater as lipid solubility increases, prompted this investigation of the effects of alcohols that differ in lipid solubility. Results indicate that LS and SS mice differ significantly in their sleep time responses to propanol, butanol, and 3-methyl butanol, as well as ethanol: their hypothermic responses showed a similar pattern, but only the response to ethanol differed significantly between lines. For both sleep time and hypothermia, the difference between lines decreased with increasing lipid solubility. In all cases, the LS mice were more sensitive than the SS to the depressant effects of the alcohol. Similar ratios of SS:LS waking brain ethanol and butanol levels indicated that CNS sensitivity to long-chain alcohols is similar to that for ethanol. A pharmacokinetic study revealed higher ethanol levels for LS than for SS mice at all time points in blood, fat, and brain body compartments. Blood ethanol elimination curves showed that the SS mice eliminate ethanol at a faster rate than do the LS.     

Cocaine actions, brain levels and receptors in selected lines of mice

The effects of cocaine (15 mg/kg IP) versus IP saline on open-field behaviors were evaluated using a crossover design in long-sleep (LS) and short-sleep (SS) mice. Under treatment order 1, mice received saline injection on day 1 followed 24 h later by cocaine (saline-cocaine, S-C). Under treatment order 2, animals received cocaine on day 1 and saline on day 2 (cocaine-saline, C-S). Immediately following injection, animals were placed into an automated open-field apparatus with behavioral samples taken at 5-min intervals for 30 min. The behaviors measured were distance traveled, stereotypy and time spent in proximity to the margins of the test apparatus (thigmotaxis). Cocaine increased locomotor activity in both lines of mice, with S-C producing more pronounced initial activation than C-S in LS mice. Compared to S-C, C-S also increased thigmotaxis, an effect more pronounced in SS mice. In a separate experiment, brain cocaine levels were measured in brains of adapted and nonadapted LS and SS mice 5 min following injection of 15 mg/kg cocaine. Regardless of order, SS mice had significantly higher brain cocaine levels than did LS mice. Mazindol and cocaine binding studies in the forebrain indicated higher B_max values for both ligands in LS compared to SS mice. The results of this study indicate that genetically based differences in cocaine receptors as well astreatment order contribute to behavioral actions of cocaine.     

Morphine withdrawal in mice selectively bred for differential sensitivity to ethanol

Mice which have been selectively bred for differences in sensitivity to acute doses of alcohol have also been shown to differ in severity of seizures upon withdrawal from chronic alcohol administration. We investigated the responsiveness of these mice to withdrawal from chronic morphine treatment. Mice were made dependent on morphine via pellet implantation, and withdrawal was precipitated with naloxone challenge. Mice which are less sensitive to the hypnotic effects of ethanol (short sleep: SS) displayed more jumping and wet dog shakes during withdrawal than did the more senstive long sleep (LS) mice. In addition, the amount of jumping was dependent on the dose of naloxone in both lines. Differences between lines in naloxone precipitated withdrawal may reflect differences in alterations in extrapyramidal dopaminergic activity, but other substrates for the observed differences cannot be discounted. Finally, the observed difference between SS and LS mice in severity of morphine withdrawal parallels the previously reported difference between these lines in seizure severity during withdrawal from alcohol.     

Concurrent exposure to aluminum and stress during pregnancy in rats: Effects on postnatal development and behavior of the offspring

The present study was conducted to assess the potential combined influence of maternal restraint stress and aluminum (Al) exposure on postnatal development and behavior in the offspring of exposed rats. Female rats were concurrently exposed to 0 (control group), 50 or 100 mg/kg/day of Al administered as Al nitrate nonahydrate in drinking water with citric acid (355 or 710 mg/kg/day) for a period of 15 days prior to mating with untreated males. Aluminum exposure was maintained throughout the gestational, lactational and post-weaning periods. On days 6-20 of gestation, one-half of the pregnant animals in each group were restrained for 2 h/day. Food consumption and maternal body weight were decreased in the groups exposed to restraint only or combined with the highest Al dose. All of the animals were allowed to deliver and wean their offspring. The pups were evaluated for physical development and neuromotor maturation. Moreover, open-field activity, passive avoidance, and spatial learning in a water maze were also determined on postnatal days 30, 35 and 60, respectively. Body weight of pups treated with 100 mg/kg/day of Al was decreased relative to controls from postnatal day 12 through 21, sexual maturation was delayed in Al treated females and in males exposed to 100 mg/kg/day. Forelimb grip strength was reduced in males exposed to 100 mg/Al/kg/day and in females exposed to this Al dose plus prenatal restraint. Learning in a passive avoidance task indicated facilitated performance for Al treated rats at 100 mg/kg/day combined with prenatal restraint as evidenced by longer avoidance latencies, while learning in a water maze task showed a shorter latency to find the platform on acquisition day 2 for Al treated rats. However, no effects of Al on water maze performance were detected during the retention probe trial in which the only effect noted was an increase in the platform quadrant swim time for the prenatal restraint group. In general terms, the results of the present study did not show a notable influence of maternal restraint on the Al-induced postnatal developmental and behavioral effects in the offspring of prenatally Al-exposed rats.     

Effects of prenatal rubratoxin-B exposure on behaviors of mouse offspring.

The effects of prenatal rubratoxin-B (RB) exposure on 8 behavioral parameters in JCL:ICR mice were assessed. Pregnant mice were injected intraperitoneally with 0.1 or 0.2 mg/kg/day of RB dissolved in propylene glycol water solution on days 7-9 (Group A) or 10-12 (Group B) of gestation. Controls received the vehicle similarly on days 7-12 of gestation. Before weaning, the offspring of both sexes were examined to test their the surface righting reflex (5 days of age), cliff avoidance response (6 days), negative geotaxis response (7 days), and swimming development (8, 10, and 12 days). After weaning, male animals were examined using the rotarod test (6 weeks of age), the open-field test (7 weeks), the shuttle-box-avoidance-learning test (9 weeks), and the water E-maze test (10 weeks). The preweanling offspring in the 0.2 mg/kg-B group showed significantly lower success rates and longer response times than controls in the cliff-avoidance response. In swimming development, the offspring in the 0.2 mg/kg B group had significantly lower scores than controls for swimming angle at 10 and 12 days of age. The avoidance learning of the mice in all RB-exposed A and B groups was significantly poorer than that of controls. These results indicate that prenatal exposure to RB produced a delay of early response development and impaired learning ability in the offspring of mice exposed to RB during middle pregnancy.     

Developmental effects of perinatal exposure to bisphenol-A and diethylstilbestrol on reproductive organs in female mice

Reproductive tract development is influenced by estrogen. The aim of this study was to determine the effects of an environmental estrogenic chemical bisphenol-A (BPA) on prenatal and postnatal development of female mouse reproductive organs. In the prenatal treatment group, BPA or the synthetic estrogen diethylstilbestrol (DES) were given by subcutaneous (s.c.) injections to pregnant mice during gestational days 10-18. Some offspring treated prenatally with 10 and 100 mg/kg bw BPA or 0.67 and 67 microg/kg bw DES were ovariectomized at 30 days and sacrificed at 40 days of age. Vaginal smears were examined in the remaining offspring, then these offspring were mated with normal males. Prenatal exposure to 10 mg/kg BPA reduced the number of mice with corpora lutea compared to sesame oil controls at 30 days, but more than 80% of mice from either prenatally exposed BPA group were fertile at 90 days. Mice exposed prenatally to maternal doses of 67 microg/kg DES were sterile and showed ovary-independent vaginal and uterine epithelial stratification; however, mice exposed prenatally to BPA did not show ovary-independent vaginal and uterine changes. The number of offspring and litter sex ratio from mice exposed prenatally to BPA (10 or 100 mg/kg) or 0.67 microg/kg DES were not different compared to controls.In postnatal treatment group, female mice were given s.c. injections of BPA (15 or 150 microg/pup) or DES (0.3 or 3 microg/pup) for 5 days from the day of birth, then some mice were ovariectomized at 30 days and examined at 40 and 90 days. In the remaining mice, vaginal smears were examined from 61 to 90 days and ovarian histology was evaluated at 90 days. Mice exposed postnatally to 150 microg BPA exhibited ovary-independent vaginal epithelial stratification. Postnatal DES (0.3 and 3 microg) treatment also induced ovary-independent vaginal stratification. Polyovular follicles having more than one oocyte in a follicle were induced by postnatal injections of BPA (150 microg) or DES (0.3 or 3 microg) at 30 days. These findings indicate for the first time that a large dose of BPA can induce ovary-independent vaginal epithelial changes when given postnatally but not prenatally.     

Effects of maternal ethanol consumption on the offspring sensory-motor development, ultrasonic vocalization, audiogenic immobility reaction and brain monoamine synthesis

Female rats were given 16% ethanol solution as the sole liquid during the entire period of gestation. At birth the offspring was removed and reared by foster dams consuming normal tap water. The development of sensory motor behaviour and emotional reactions was delayed by 1-2 days in the prenatally ethanol exposed pups as assessed by tests on body righting, acoustic startle response, air righting, rearing and ultrasonic vocalization. In the open-field test the normally occurring behavioural difference between the sexes was not found in the prenatally ethanol exposed pups. Both sexes of the ethanol exposed pups behaved like the female controls suggesting deficient masculinization of the ethanol exposed male pups during foetal age. Biochemical analysis of the brains showed a decreased synthesis of serotonin and dopamine.     

Prenatal choline supplementation mitigates the adverse effects of prenatal alcohol exposure on development in rats

Prenatal alcohol exposure can lead to a range of physical, neurological, and behavioral alterations referred to as fetal alcohol spectrum disorders (FASD). Variability in outcome observed among children with FASD is likely related to various pre- and postnatal factors, including nutritional variables. Choline is an essential nutrient that influences brain and behavioral development. Recent animal research indicates that prenatal choline supplementation leads to long-lasting cognitive enhancement, as well as changes in brain morphology, electrophysiology and neurochemistry. The present study examined whether choline supplementation during ethanol exposure effectively reduces fetal alcohol effects. Pregnant dams were exposed to 6.0 g/kg/day ethanol via intubation from gestational days (GD) 5–20; pair-fed and lab chow controls were included. During treatment, subjects from each group received choline chloride (250 mg/kg/day) or vehicle. Physical development and behavioral development (righting reflex, geotactic reflex, cliff avoidance, reflex suspension and hindlimb coordination) were examined. Subjects prenatally exposed to alcohol exhibited reduced birth weight and brain weight, delays in eye opening and incisor emergence, and alterations in the development of all behaviors. Choline supplementation significantly attenuated ethanol's effects on birth and brain weight, incisor emergence, and most behavioral measures. In fact, behavioral performance of ethanol-exposed subjects treated with choline did not differ from that of controls. Importantly, choline supplementation did not influence peak blood alcohol level or metabolism, indicating that choline's effects were not due to differential alcohol exposure. These data indicate early dietary supplements may reduce the severity of some fetal alcohol effects, findings with important implications for children of women who drink alcohol during pregnancy.     

Avoidance facilitation in adult mice by prenatal administration of the nootropic drug oxiracetam

CD-1 mice prenatally exposed to saline solution or to the nootropic drug oxiracetam (50 mg/kg during the whole pregnancy) were tested, when adults, for locomotor activity and for shuttle-box avoidance acquisition. Prenatal drug exposure produced long-lasting effects, evident in mature offspring. At the age of two months, mice prenatally exposed to oxiracetam showed a slight but significant reduction in locomotor activity. At the age of three months, these animals exhibited higher performances than control mice in avoidance acquisition.     

Withdrawal-like signs induced by a single administration of ethanol in mice that differ in ethanol sensitivity

These experiments studied changes produced by a hypnotic dose of ethanol in the LS and SS lines of mice, which differ in ethanol sensitivity. In the first experiment, animals were injected either with ethanol or saline, and activity and seizure susceptibility measured 7–9 h later when blood levels of ethanol would have reached zero. Ethanol-treated mice of both genetic lines were less active in an open field test and more susceptible to clonic convulsions induced by flurothyl than saline-injected controls. There was no difference in the magnitude of these changes in the two lines. In the control condition SS (short-sleep) mice were more active than LS (long-sleep) mice, and more susceptible than LS mice to myoclonic but not to clonic seizures. The effect of the ethanol injection on body temperature was evaluated in separate groups of animals. LS mice showed a more pronounced hypothermia than LS mice when temperature was measured 2 h after injection. Six hours after injection, SS mice exhibited a small but statistically significant overshoot in temperature, after which they again became hypothermic with respect to controls; hyperthermia was not observed in LS animals.     

Can nootropic drugs be effective against the impact of ethanol teratogenicity on cognitive performance?

Rats exposed pre- (PA) and postnatally (PNA) to ethanol at a dose of 1 g/kg for 24 h developed fetal alcohol effects (FAE). This was measured using a condition-reflex method for active avoidance with punishment reinforcement (shuttle-box) in which pronounced learning and memory deficits in 3-month-old rats were found after ethanol exposure (Vaglenova and Petkov, 1998. Fetal alcohol effects in rats exposed pre- and postnatally to a low dose of ethanol. Alcohol. Clin. Exp. Res. 22(3), 697--703). In the present study the effects of piracetam (Pyramem) at a dose of 600 mg/kg body weight, aniracetam at 50 mg/kg, and meclophenoxate (Centrophenoxine) at 100 mg/kg were studied. The drugs were administered orally during 10 days to separate groups of naive and pre- and postnatally exposed to ethanol rats. All the investigated nootropic drugs showed a significant possibility to alleviate learning and memory disability of rats with FAE. Aniracetam was administered to 1-month-old rats, demonstrating a prolonged (2 months) therapeutic effect, observed in rats aged 3 months. As previously reported (Vaglenova and Petkov, 1998), between male rats with FAE and controls, 66 and 33% were 'poor learners', respectively. In all nootropic treatment groups the percentage of 'poor learners' dropped to 28%. The positive effects of piracetam, aniracetam and meclophenoxate suggest that these drugs could be used for both treatment and prophylactic of FAE-connected disturbances of cognition.     

Effects of pentobarbital in mice selected for differential sensitivity to ethanol

Two lines of mice have been genetically selected for differential sensitivity to ethanol. These lines have been designated long sleep (LS) and short sleep (SS) on the basis of their hypnotic response to the ethanol selection dose. Earlier studies of these mice suggested that this difference was limited to alcohols and did not extend to other classes of hypnotics. The present study examined hypnotic and hypothermic responses produced by pentobarbital in recent generations of these mice. Dose-dependent differences in sleep time and in hypothermia were found, with SS mice affected to a greater degree than LS mice. Pharmacokinetic studies showed that the half-life of pentobarbital disappearance from SS blood was twice that reported for SS mice of the 18th generation. The half-life in the LS line had not changed. The volumes of distribution and waking brain concentrations were identical in LS and SS mice. An altered rate of elimination (not differential CNS sensitivity) appeared to be the major factor responsible for the differences observed between these lines.     

Effects of paternal alcohol consumption on pregnancy outcome in rats

Male rats were divided into 3 groups and were given ad lib access to a liquid alcohol diet containing 35% ethanol derived calories (EDC) or were pair-fed with an isocaloric control diet, containing 17.5% or 0% EDC, for a minimum of 52 days. A fourth group was fed rat chow ad lib. Males were then mated with non-treated females. Males consuming alcohol did not differ in sexual behavior but were less fertile and sired offspring that were significantly less active than controls and the effects on activity were dose-related. Females sired by males consuming alcohol also performed worse in a two-way shock avoidance learning task although differences in this task were not dose-related. There were no significant effects, however, on body weight of offspring at birth or at 21 days of age or on other behavioral measures such as spontaneous alternation or passive avoidance learning.     

Apomorphine effects on behavioral response to ethanol in mice selectively bred for differential sensitivity to ethanol

Two lines of mice selectively bred for differences in response to a hypnotic dose of ethanol were administered apomorphine alone or in combination with ethanol. When administered by itself, apomorphine produced similar dose-dependent depression of locomotor activity and increases in stereotypy in the two lines. Doses of apomorphine (0.5 microM/kg and 2 microM/kg) thought to bind only presynaptic dopamine receptors blocked the slight locomotor activation to 1.5 g/kg ethanol in the ethanol-sensitive Long-Sleep (LS) mice; in the ethanol-insensitive Short-Sleep (SS) mice which show marked activation to all subhypnotic doses of ethanol, these doses of apomorphine only attenuated the activation. A higher apomorphine dose (8 microM/kg) antagonized the locomotor depressant effects of 2.0 and 2.5 g/kg of ethanol in LS mice but did not alter the shape of the SS ethanol dose response curve for locomotor activity. Apomorphine (2 and 8 microM/kg) potentiated ethanol-induced loss of the righting reflex in LS mice in a dose dependent fashion, but did not alter this soporific effect of ethanol in SS mice. These findings extend the data base suggesting a role for dopamine both in the mechanism(s) differentiating the LS and SS mice and the stimulant and intoxicating properties of ethanol.     

Neonatal cerebellectomy alters ethanol-induced sleep time of short sleep but not long sleep mice

The effects of neonatal cerebellectomy on ethanol-induced sleep times in long sleep (LS) and short sleep (SS) mice were investigated. Cerebellectomy did not alter the ethanol sensitivity of LS animals for loss of righting reflex. In contrast, SS mice became more sensitive to alcohol after cerebellectomy. Even so, large differences were still observed between the alcohol-induced sleep times of cerebellectomized LS and SS mice. The data indicate that, while the cerebellum must have a prominant influence on alcohol sleep time in SS animals, this brain structure is not solely responsible for the observed differences in righting reflex sensitivity to ethanol in these two mouse lines. We postulate the existence of noncerebellar central neurons with differential sensitivities to the depressant effects of ethanol in LS and SS mice.     

Differential modulation of benzodiazepine receptor binding by ethanol in LS and SS mice

The LS and SS lines of mice were initially selected based on sedative responses to ethanol, but have been found to differ in response to a variety of hypnotics and anesthetics. These differences do not appear to be due to pharmacokinetic factors and several lines of evidence suggest involvement of the GABAergic system. To examine an important component of this system, the benzodiazepine receptor, we analyzed benzodiazepine receptor binding in vivo in LS and SS mice, and modulation of receptor binding by three interventions known to increase binding in other strains: pentobarbital, defeat stress, and ethanol. Receptor binding was determined by specific uptake of [3H]-Ro15-1788. Receptor binding was increased in cortex and hippocampus of LS mice compared to SS mice, with the increase in cortex most likely due to increased receptor number rather than a change in apparent affinity. Pentobarbital (30 mg/kg IP) induced similar increases in binding in both lines in several brain regions. Defeat stress caused increased binding in several brain regions of both SS and LS mice, with greater binding in cortex of LS mice. In contrast, ethanol at 3 doses (0.5, 1, and 2 g/kg) led to greater increases in binding in SS mice compared to LS mice in most brain regions. None of the interventions altered nonspecific binding. Ethanol concentrations were slightly greater in plasma and brain of LS mice. These results indicate differences in benzodiazepine receptor binding in LS and SS mice, with differential modulation of binding by ethanol but not by pentobarbital or stress. These differences may contribute to differential pharmacodynamic responses in the two lines of mice.     

Circadian and genetic influences on tissue sensitivity and sleep time to ethanol in LS and SS mice

Circadian variations in response to ethanol were studied in long-sleep (LS) and short-sleep (SS) mice. Each LS animal received a 2.5 g/kg intraperitoneal ethanol injection, while the SS animals were injected with 4.1 or 5.0 g/kg. Different groups of mice were assessed for sleep time, waking blood alcohol concentration (BAC), and waking brain ethanol concentration (BREC) at 03.00, 09.00, 15.00, or 21.00 hr. Sleep times, waking BACs, and waking BRECs showed circadian variations in the LS mice. SS animals given the 4.1 g/kg dose showed circadian variations for waking BAC and waking BREC, but not for sleep time. The observed variations in the physiological parameters for these animals may have been confounded by a short sleep time so that they reflected circadian variations in drug absorption and/or distribution rather than in CNS sensitivity. SS mice given the 5.0 g/kg dose slept longer than those given the 4.1 g/kg dose and did not show circadian variations for sleep time, waking BAC, or waking BREC. These results suggest both circadian and genetic influences on tissue sensitivity to ethanol.     

Effect of taurine on ethanol-induced sleep time in mice genetically bred for differences in ethanol sensitivity

Long Sleep (LS) and Short Sleep (SS) mice were used in this study to investigate the interaction between ethanol and taurine. Sleep time (hypnosis) was selected as an index of ethanol-induced central nervous system depression. In order to achieve a similar degree of central nervous system depression with ethanol, SS and LS mice received 5.3 and 3.0 g/kg, IP, of ethanol, respectively. When taurine (7.5, 15 and 25 mumol/kg) was administered intracerebroventricularly (ICV) to LS and SS mice immediately after regaining the righting reflex following ethanol injection, a return to sleep time was produced. This effect of taurine was immediate in onset and occurred in a dose-dependent fashion. LS mice exhibited a greater effect from taurine administration than SS mice. In another experiment LS and SS mice were given ICV TAG, a taurine antagonist (6-aminomethyl-3-methyl-4H-1,2,4-benzothiadiazine-1,1-dioxide HCl), which significantly reduced the effect of taurine to produce a return to sleep time in the presence of ethanol. TAG did not affect ethanol-induced sleep time. In control experiments, in the absence of ethanol, neither taurine (25 mumol/kg, ICV) nor TAG (1 mumol/kg, ICV) caused a significant loss of the righting reflex (sleep time). When pentobarbital (50 mg/kg, IP) was injected instead of ethanol in the sleep time experiments, taurine (7.5, 15 and 25 mumol/kg, ICV) produced a return to sleep time in LS and SS mice that resembled the effect of taurine with ethanol in SS mice. These results indicate that taurine (ICV) can enhance the central depressant action of ethanol and pentobarbital and that the greatest effect of taurine occurred with LS mice in the presence of ethanol. It is possible that taurine may have some role in the central nervous system depressant properties of ethanol.     

Prenatal alcohol exposure and offspring hyperactivity: effects of physostigmine and neostigmine

Rats were fed a liquid diet containing ethanol from days 6-19 of gestation. Controls were pair-fed the same diet with sucrose substituted for ethanol, or received ad lib chow and water. The activity of the offspring was observed at 10, 16, 22 or 28 days-of-age. Overall, offspring exposed to alcohol prenatally were hyperactive compared to controls at 16 and 22 days, but not at 10 or 28 days. Administration of physostigmine had no effect on the activity of any group at 10 days. At 16 days, physostigmine enhanced the activity of the alcohol group, had no effect on the sucrose group, but depressed the activity of the chow group. At 22 days it led to a dose-related decrease in activity in controls but had no effect on the activity of the alcohol treated pups. At 28 days, physostigmine decreased the activity of all three groups. Administration of the quaternary derivative, neostigmine, indicated that the effects of scopolamine at 22 days were probably central in origin. These data indicate that a putative cholinergic/inhibitory system becomes functional in control pups before 22 days, but in pups exposed to alcohol prenatally development is delayed by a number of days.