Altered corticosterone levels and social play behavior after prolonged maternal separation in adolescent male but not female Wistar rats

Early-life socio-environmental factors are crucial for normal developmental processes; adverse experiences early in life can therefore lead to detrimental effects in several physiological systems. The aim of this study was to examine short-term effects of early adverse experiences in a maternal separation (MS) rodent model. In this study two separation conditions were used: daily 15- (MS15) or 360-min (MS360) separation of the litter from the dam during postnatal day 1–21. In early adolescence, male and female offspring were subjected to a single-isolation procedure with analysis of corticosterone levels prior to and after isolation. In addition, social play behavior was assessed during mid-adolescence. There was a clear difference between male and female offspring in both tests performed. There was no difference in corticosterone levels between the female MS groups, whereas MS360 males showed higher baseline and recovery corticosterone levels than MS15 males. The amount of pinning, a specific social play behavior, was affected by rearing with MS360 males having a higher frequency than MS15 males, while there was no difference between the female MS groups. The observation that males but not females are affected by MS360 has previously been reported for adult animals, and herein we show that this difference is present already in adolescence. Changes in corticosterone levels and social behavior following early-life adversity have been associated with adult behavioral alterations, and our results confirm that these changes emerge already within adolescence.     

Adolescent binge drinking leads to changes in alcohol drinking, anxiety, and amygdalar corticotropin releasing factor cells in adulthood in male rats

Heavy episodic drinking early in adolescence is associated with increased risk of addiction and other stress-related disorders later in life. This suggests that adolescent alcohol abuse is an early marker of innate vulnerability and/or binge exposure impacts the developing brain to increase vulnerability to these disorders in adulthood. Animal models are ideal for clarifying the relationship between adolescent and adult alcohol abuse, but we show that methods of involuntary alcohol exposure are not effective. We describe an operant model that uses multiple bouts of intermittent access to sweetened alcohol to elicit voluntary binge alcohol drinking early in adolescence (~postnatal days 28-42) in genetically heterogeneous male Wistar rats. We next examined the effects of adolescent binge drinking on alcohol drinking and anxiety-like behavior in dependent and non-dependent adult rats, and counted corticotropin-releasing factor (CRF) cell in the lateral portion of the central amygdala (CeA), a region that contributes to regulation of anxiety- and alcohol-related behaviors. Adolescent binge drinking did not alter alcohol drinking under baseline drinking conditions in adulthood. However, alcohol-dependent and non-dependent adult rats with a history of adolescent alcohol binge drinking did exhibit increased alcohol drinking when access to alcohol was intermittent. Adult rats that binged alcohol during adolescence exhibited increased exploration on the open arms of the elevated plus maze (possibly indicating either decreased anxiety or increased impulsivity), an effect that was reversed by a history of alcohol dependence during adulthood. Finally, CRF cell counts were reduced in the lateral CeA of rats with adolescent alcohol binge history, suggesting semi-permanent changes in the limbic stress peptide system with this treatment. These data suggest that voluntary binge drinking during early adolescence produces long-lasting neural and behavioral effects with implications for anxiety and alcohol use disorders.     

Evaluation of accelerated development of stable alcoholic motivation in rats for studying potential alcohol deterrents

The express technique reflecting an acquisition of a clear alcohol addiction during short-term voluntary alcoholization for further antialcoholic drugs testing was performed in male albino rats. By VARIMAX factor analysis of indexes related with preference of alcohol solutions with different tastes the conditions of short-term (2 months) voluntary alcoholization leading to persistent ethanol intake were studied. Isolation stress inducing a specific alcohol drive was excluded from rearing conditions. 0.1% saccharin solution in 15% ethanol was used for alcoholization. Statistical analysis revealed factor of "developed alcohol abuse" which may be detected in conditions of one-trail sweet ethanol intake after 3 days alcohol deprivation (similar to heavy drinking syndrome in humans). Using pharmacological drugs (pyrazidol, piracetam) validity of the method for specific drug design was confirmed.     

Behavioral profiles and stress-induced corticosteroid secretion in male Wistar rats subjected to short and prolonged periods of maternal separation

Early life experiences are important for the development of neurobiobehavioral mechanisms and subsequent establishment of mental functions. In experimental animals, early life experiences can be studied using the maternal separation model. Maternal separation has been described to induce neurobiological changes and thus affect brain function, mental state and behavior. We have established a protocol in order to study the effects of repeated short and prolonged periods of maternal separation during the postnatal period on adult neurochemistry, voluntary ethanol intake and behavior. In the present experiment, we focus on the long-term effects of maternal separation on exploration and risk assessment behavior as well corticosteroid secretion. Rat pups were assigned to 15 min (MS15) or 360 min (MS360) of daily maternal separation and normal animal facility rearing (AFR) during postnatal days 1–21. To establish the adult behavioral profile in male rats, three tests were used: the Concentric Square Field (CSF), the Open Field (OF) and the Elevated Plus-maze (EPM). No differences between the three experimental groups were found in the traditional OF and EPM tests. The CSF test indicated that the MS360 rats were more explorative and expressed an altered risk assessment and risk-taking profile. In response to a restraint stress, MS360 rats had a blunted corticosterone release in contrast to MS15 and AFR rats. In contrast to previous results, the outcome of the present investigation does not support the notion that a prolonged period of maternal separation results in an adult phenotype characterized by an increased emotional reactivity.     

Behavioral profiles and stress-induced corticosteroid secretion in male Wistar rats subjected to short and prolonged periods of maternal separation

Early life experiences are important for the development of neurobiobehavioral mechanisms and subsequent establishment of mental functions. In experimental animals, early life experiences can be studied using the maternal separation model. Maternal separation has been described to induce neurobiological changes and thus affect brain function, mental state and behavior. We have established a protocol in order to study the effects of repeated short and prolonged periods of maternal separation during the postnatal period on adult neurochemistry, voluntary ethanol intake and behavior. In the present experiment, we focus on the long-term effects of maternal separation on exploration and risk assessment behavior as well corticosteroid secretion. Rat pups were assigned to 15 min (MS15) or 360 min (MS360) of daily maternal separation and normal animal facility rearing (AFR) during postnatal days 1-21. To establish the adult behavioral profile in male rats, three tests were used: the Concentric Square Field (CSF), the Open Field (OF) and the Elevated Plus-maze (EPM). No differences between the three experimental groups were found in the traditional OF and EPM tests. The CSF test indicated that the MS360 rats were more explorative and expressed an altered risk assessment and risk-taking profile. In response to a restraint stress, MS360 rats had a blunted corticosterone release in contrast to MS15 and AFR rats. In contrast to previous results, the outcome of the present investigation does not support the notion that a prolonged period of maternal separation results in an adult phenotype characterized by an increased emotional reactivity.     

Doctors' drinking habits and consumption of alcohol.

Alcohol consumption and drinking habits among Finnish doctors were studied as part of a survey of stress and burnout. A questionnaire containing 99 questions or groups of questions was sent to all 3496 practising doctors aged under 66 randomly selected from the registry of the Finnish Medical Association. Altogether 2671 doctors (76%) responded; this sample was representative of the Finnish medical profession. The average weekly consumption of alcohol during the past year and various aspects of drinking behaviour were assessed, and the presence or absence of symptoms and diseases often encountered among heavy drinkers and addicts was determined. The data were analysed separately for men and women, for those aged less than or equal to 40 and greater than 40, and for the men with high and low alcohol consumption and with high and low scores on the index of drinking habits. Selected variables related to work, stress, and coping were correlated with alcohol consumption and drinking behaviour. The median consumption of alcohol among male doctors was 4876 g (6.2 litres) and among female doctors 2226 g (2.8 litres) of absolute alcohol per person per year and was higher in those aged over 40. Beer was most commonly drunk by men and wine by women. Increased alcohol consumption was associated with older age, disappointment with career, heavy smoking, use of benzodiazepines, stress and burnout symptoms, suicidal thoughts, general dissatisfaction, and diseases related to alcohol. Drinking habits were heavier among doctors working in community health centres, those taking long sick leaves, younger doctors disappointed with their careers or the atmosphere at work, and older doctors immersed in their work. Alcohol consumption among doctors seems to be higher than that of the general population in Finland, and heavy drinking seems to be associated with stress and burnout.     

Differential Patterns of Alcohol Consumption and Dopamine-2 Receptor Binding in Wistar-Kyoto and Wistar Rats

The Wistar-Kyoto (WKY) rat strain has been described as an animal model of depressive behavior that consumes significantly greater amounts of alcohol compared to the Wistar (WIS) rat strain. Since the mesolimbic dopamine (DA) type-2 (D2) receptors mediate reward-related behaviors, the present study measured the binding of [¹²⁵I]-Iodosulpiride to D2 receptors in the brains of WKY versus WIS rats following 24 days of voluntary alcohol or water consumption. Alcohol consuming WKY rats showed a significant increase in D2 receptor binding in several regions of the mesolimbic and nigrostriatal systems. In contrast, alcohol consuming WIS rats showed a reduction in D2 receptor binding in DA cell body areas. The differential regulation of D2 receptors by voluntary alcohol consumption in the two rat strains suggests that D2 receptor mediated neurotransmission may be playing a role in the increased alcohol drinking behavior reported in WKY rats.     

The role of early life experience and species differences in alcohol intake in microtine rodents

Social relationships have important effects on alcohol drinking. There are conflicting reports, however, about whether early-life family structure plays an important role in moderating alcohol use in humans. We have previously modeled social facilitation of alcohol drinking in peers in socially monogamous prairie voles. We have also modeled the effects of family structure on the development of adult social and emotional behaviors. Here we assessed whether alcohol intake would differ in prairie voles reared by both parents compared to those reared by a single mother. We also assessed whether meadow voles, a closely related species that do not form lasting reproductive partnerships, would differ in alcohol drinking or in the effect of social influence on drinking. Prairie voles were reared either bi-parentally (BP) or by a single mother (SM). BP- and SM-reared adult prairie voles and BP-reared adult meadow voles were given limited access to a choice between alcohol (10%) and water over four days and assessed for drinking behavior in social and non-social drinking environments. While alcohol preference was not different between species, meadow voles drank significantly lower doses than prairie voles. Meadow voles also had significantly higher blood ethanol concentrations than prairie voles after receiving the same dose, suggesting differences in ethanol metabolism. Both species, regardless of rearing condition, consumed more alcohol in the social drinking condition than the non-social condition. Early life family structure did not significantly affect any measure. Greater drinking in the social condition indicates that alcohol intake is influenced similarly in both species by the presence of a peer. While the ability of prairie voles to model humans may be limited, the lack of differences in alcohol drinking in BP- and SM-reared prairie voles lends biological support to human studies demonstrating no effect of single-parenting on alcohol abuse.     

DRD1 5'UTR variation, sex and early infant stress influence ethanol consumption in rhesus macaques

The mesolimbic dopamine system plays an important role in mediating a variety of behaviors and is involved in mediating the reinforcing effects of ethanol. Genes encoding dopamine receptor subtypes are thus good candidate loci for understanding the genetic etiologies of susceptibility to alcohol dependence and its antecedent behavioral phenotypes. We tested whether variation in DRD1 influences alcohol consumption in rhesus macaques and whether its influence is mediated by sex and early rearing experience. We genotyped a single nucleotide polymorphism (−111 G/T) in the 5'UTR of DRD1 in 96 subjects raised with their mothers until 6 months of age ( n = 43) or in peer-only groups ( n = 53). As young adults they underwent a 7-week voluntary ethanol consumption experiment. anova revealed a significant main effect of sex ( F _1,95 = 6.3, P = 0.014) and an interaction between genotype, sex and rearing on ethanol consumption ( F _7,95 = 4.63, P = 0.0002). Maternally deprived males heterozygous for the T allele consumed significantly more ethanol ( P > t ≤ 0.0001) than the other subgroups. Maternal deprivation can produce individuals that are anxious and impulsive, both of which are known risk factors for alcohol dependence. Our work demonstrates a potential role for the dopamine D1 receptor gene in modulating alcohol consumption, especially in the context of early environmental stress.     

Fischer Rats Consume 20% Ethanol in a Long-Term Intermittent-Access Two-Bottle-Choice Paradigm

The 20% ethanol intermittent-access (IAE) two-bottle-choice drinking procedure has been shown to produce high voluntary ethanol consumption in a number of rat strains. For this study, we applied this procedure to male Fischer (F344) rats, a strain previously reported to exhibit low levels of ethanol consumption. We also subjected these animals to a two-week ethanol-deprivation-period to see if they would exhibit an alcohol deprivation effect (ADE) signified by a transient increase in alcohol consumption following deprivation. Our data show a separation between high and low consuming animals within this strain, with high-consumers exhibiting an escalation in consumption. In contrast, Fischer rats did not show a significant separation between high and low consumers or any significant escalation in consumption, using the 20% ethanol continuous-access two-bottle-choice drinking protocol. Following the two-week deprivation period, animals in the high (but not the low) IAE group exhibited the transient increase in ethanol consumption and preference typically associated with an ADE. Together, the data suggest that the intermittent access protocol is a useful protocol for increasing ethanol consumption.     

Differences in basal and ethanol-induced levels of opioid peptides in Wistar rats from five different suppliers

One major cause for discrepancies in results from animal experimental studies is the use of different animal strains and suppliers. We have previously reported that Wistar rats from five different suppliers display profound differences in ethanol intake and behavior. One of the neurobiological processes that could be underlying these differences is the endogenous opioid system, which has been implicated in the rewarding and reinforcing effects of alcohol. We therefore hypothesized that the differences between the supplier groups would also be evident in the endogenous opioid system. Radioimmunoassay was used to determine the levels of the opioid peptides Met-enkephalin-Arg⁶Phe⁷ and dynorphin B in several brain areas of ethanol-drinking and ethanol naïve Wistar rats from five different suppliers. In the ethanol naïve animals, differences between the supplier groups were found in the pituitary gland, hypothalamus, frontal cortex, dorsal striatum and hippocampus. In the ethanol-drinking rats, differences were found in the same structures, with the addition of medial prefrontal cortex and substantia nigra. Correlations between ethanol intake and peptide levels were also found in several of the areas examined. The structures in which differences were found have all been implicated in the transition from drug use to addiction and these differences may lead to different propensities and vulnerability to this transition. Because the endogenous opioids have been suggested to be involved in a number of neurobiological disorders the results do not only have implications for research on alcohol or drug addiction, but many other fields as well.     

Renal artery stenosis: an example of how the periphery can modulate voluntary alcohol drinking

Voluntary alcohol drinking, using a free-access procedure, was studied in rats three weeks after unilateral renal artery stenosis was produced by applying a 0.20 mm solid silver clip to the left renal artery (Two-Kidney, One-Clip, Hypertension). The group of animals with the arterial clip drank significantly less alcohol and more water than a sham-operated group. The two groups, however, drank similar amounts of a palatable glucose solution, suggesting that they did not differ in taste sensitivity. Blood alcohol levels measured for six hours following a 2.5 g/kg intraperitoneal injection of alcohol indicated that the difference in alcohol consumption between the two groups could not be accounted for in terms of differences in drug disposition or metabolism. These findings provide an example of a previously unknown type of control which originates in the periphery and contributes to the regulation of voluntary alcohol drinking.     

Immunohistochemical and biochemical evidence for the presence of the pentapeptide met-enkephalin and the heptapeptide met-enkephalin-Arg6-Phe7 but not the octapeptide met-enkephalin-Arg6-Gly7-Leu8 in amphibian chromaffin cells

Using specific antibodies to met-enkephalin, met-enkephalin-Arg⁶-Phe⁷ and met-enkephalin-Arg⁶-Gly⁷-Leu⁸, we have studied the distribution of these opioid peptides in the frog adrenal gland by means of the indirect immunofluorescence technique. Bright staining of all chromaffin cells was observed by application of met-enkephalin and met-enkephalin-Arg⁶-Phe⁷ antisera. No nerve endings could be detected. A few chromaffin cells were weakly stained by met-enkephalin-Arg⁶-Gly⁷-Leu⁸ antiserum. Using a specific radioimmunoassay for met-enkephalin, the dilution curve of frog adrenal extracts was parallel to that of synthetic met-enkephalin. The concentration of met-enkephalin-like material measured in crude acetic extracts of frog adrenals (2.31 ± 0.16 pmol/mg w. wt) was high when compared to those reported for most mammalian species. No leu-enkephalin and virtually no met-enkephalin-Arg⁶-Gly⁷-Leu⁸ were detected by the corresponding radioimmunoassays. Reverse phase HPLC analysis revealed that oxidized met-enkephalin was the main form of met-enkephalin detected in acetic extracts of frog adrenals. HPLC separation showed the presence of a peptide co-eluting with synthetic met-enkephalin-Arg⁶-Phe⁷. Higher molecular weight forms were also separated by HPLC. These results show the presence of both met-enkephalin and the heptapeptide met-enkephalin-Arg⁶-Phe⁷ and the lack of met-enkephalin-Arg⁶-Gly⁷-Leu⁸ in frog chromaffin cells.     

Effects of Voluntary Alcohol Intake on Risk Preference and Behavioral Flexibility during Rat Adolescence

Alcohol use is common in adolescence, with a large portion of intake occurring during episodes of binging. This pattern of alcohol consumption coincides with a critical period for neurocognitive development and may impact decision-making and reward processing. Prior studies have demonstrated alterations in adult decision-making following adolescent usage, but it remains to be seen if these alterations exist in adolescence, or are latent until adulthood. Here, using a translational model of voluntary binge alcohol consumption in adolescents, we assess the impact of alcohol intake on risk preference and behavioral flexibility during adolescence. During adolescence (postnatal day 30–50), rats were given 1-hour access to either a 10% alcohol gelatin mixture (EtOH) or a calorie equivalent gelatin (Control) at the onset of the dark cycle. EtOH consuming rats were classified as either High or Low consumers based on intake levels. Adolescent rats underwent behavioral testing once a day, with one group performing a risk preference task, and a second group performing a reversal-learning task during the 20-day period of gelatin access. EtOH-High rats showed increases in risk preference compared to Control rats, but not EtOH-Low animals. However, adolescent rats did a poor job of matching their behavior to optimize outcomes, suggesting that adolescents may adopt a response bias. In addition, adolescent ethanol exposure did not affect the animals'' ability to flexibly adapt behavior to changing reward contingencies during reversal learning. These data support the view that adolescent alcohol consumption can have short-term detrimental effects on risk-taking when examined during adolescence, which does not seem to be attributable to an inability to flexibly encode reward contingencies on behavioral responses.     

Adult consequences of late adolescent alcohol consumption: a systematic review of cohort studies

Background

Although important to public policy, there have been no rigorous evidence syntheses of the long-term consequences of late adolescent drinking.

Methods and Findings

This systematic review summarises evidence from general population cohort studies of drinking between 15–19 years old and any subsequent outcomes aged 20 or greater, with at least 3 years of follow-up study. Fifty-four studies were included, of which 35 were assessed to be vulnerable to bias and/or confounding. The principal findings are: (1) There is consistent evidence that higher alcohol consumption in late adolescence continues into adulthood and is also associated with alcohol problems including dependence; (2) Although a number of studies suggest links to adult physical and mental health and social consequences, existing evidence is of insufficient quality to warrant causal inferences at this stage.

Conclusions

There is an urgent need for high quality long-term prospective cohort studies in order to better understand the public health burden that is consequent on late adolescent drinking, both in relation to adult drinking and more broadly. Reducing drinking during late adolescence is likely to be important for preventing long-term adverse consequences as well as protecting against more immediate harms. Please see later in the article for the Editors'' Summary     

Occurrence of met-enkephalin,met-enkephalin-Arg6-Phe7 and met-enkephalin-Arg6-Gly7-Leu8 in gastrin cells of hog antral mucosa

Summary Region-specific antisera to three enkephalins: met-enkephalin, met-enkephalin-Arg⁶-Phe⁷ and met-enkephalin-Arg⁶-Gly⁷-Leu⁸, together with four region specific antisera to progastrin: C-terminal G17 specific, N-terminal G34 specific, cryptic peptides A- and B-specific, were used in immunohistochemical studies of hog antral mucosa. A sub-population (6–10%) of the gastrin-containing endocrine cells (G-cells) was found to react with antisera to met-enkephalin, met-enkephalin-Arg⁶-Phe⁷ and met-enkephalin-Arg⁶-Gly⁷-Leu⁸. About 30% of all the enkephalin-containing cells were identified as G-cells. The results indicate that a fraction of G-cells produces both enkephalin-like peptides and gastrin.     

Cognitive and neurobiological mechanisms of alcohol-related aggression

Alcohol-related violence is a serious and common social problem. Moreover, violent behaviour is much more common in alcohol-dependent individuals. Animal experiments and human studies have provided insights into the acute effect of alcohol on aggressive behaviour and into common factors underlying acute and chronic alcohol intake and aggression. These studies have shown that environmental factors, such as early-life stress, interact with genetic variations in serotonin-related genes that affect serotonergic and GABAergic neurotransmission. This leads to increased amygdala activity and impaired prefrontal function that, together, predispose to both increased alcohol intake and impulsive aggression. In addition, acute and chronic alcohol intake can further impair executive control and thereby facilitate aggressive behaviour.     

Involvement of kappa type opioids on ethanol drinking

The effects of the administration of the kappa agonist dynorphin1-17 and/or the kappa antagonist MR-2266-BS on ethanol preference was investigated using a paradigm by which rats develop alcohol preference. Administration of dynorphin shortly before or after the conditioning session (forced ethanol exposure) failed to affect later ethanol preference. However, dynorphin treatment prior to the first choice session reduced ethanol preference during the three consecutive testing days. This effect was reversed by the simultaneous administration of the kappa antagonist MR-2266-BS. The results of the present study provide further support for evidence of the involvement of dynorphinergic systems on drinking behavior and suggest that kappa-type opioid mechanisms may be involved in the consumption and development of preference to ethanol in rats.