Effects of an Orexin-2 Receptor Antagonist on Sleep and Event-Related Oscillations in Female Rats Exposed to Chronic Intermittent Ethanol During Adolescence.

BACKGROUND: Alcohol use is on the rise among women in the United States which is especially concerning since women who drink have a higher risk of alcohol-related problems. Orexin (hypocretin) receptor antagonists may have some therapeutic value for alcohol-induced insomnia; however, the use of this class of drugs following female adolescent binge drinking is limited. The current study will address whether adolescent intermittent ethanol (AIE) in female rats can result in lasting changes in sleep pathology and whether orexin-targeted treatment can alleviate these deficits. METHODS: Following a 5-week AIE vapor model, young adult rats were evaluated on waking event-related oscillations (EROs) and EEG sleep. Subsequently, AIE rats were treated with orexin receptor 2 (OX2 R) antagonist (MK-1064; 10, 20mg/kg) to test for modifications in sleep pathology and waking ERO. RESULTS: Female AIE rats exhibited lasting changes in sleep compared to controls. This was demonstrated by increased fragmentation of slow wave sleep (SWS) and rapid eye movement sleep, as well as reductions in delta and theta power during SWS. There was no impact of AIE on waking EROs. Acute MK-1064 hastened SWS onset and increased the number of SWS episodes, without increasing sleep fragmentation in AIE and controls. While treatment with MK-1064 did not impact sleep EEG spectra, waking ERO energy was increased in delta, theta, and beta frequency bands. CONCLUSIONS: These results demonstrate that AIE can produce lasting changes in sleep in female rats, highly similar to what we previously found in males. Additionally, while the OX2 R antagonist promoted sleep in both alcohol-exposed and unexposed rats, it did not reverse most of the alcohol-induced disruptions in sleep. Thus, OX2 R antagonism may serve as a potential therapeutic strategy for the treatment of insomnia, but not the specific signs of alcohol-induced insomnia.

Time Course of Blood and Brain Cytokine/Chemokine Levels Following Adolescent Alcohol Exposure and Withdrawal in Rats.

BACKGROUND: Adolescence is a critical period for neural development, and alcohol exposure during adolescence can lead to an elevated risk for health consequences as well as alcohol use disorders. Clinical and experimental data suggest that chronic alcohol exposure may produce immunomodulatory effects that can lead to the activation of pro-inflammatory cytokine pathways as well as microglial markers. The present study evaluated, in brain and blood, the effects of adolescent alcohol exposure and withdrawal on microglia and on the most representative pro- and anti-inflammatory cytokines and major chemokines that can contribute to the establishing of a neuroinflammatory environment. METHODS: Wistar rats (males, n = 96) were exposed to ethanol (EtOH) vapors, or air control, for 5 weeks over adolescence (PD22-PD58). Brains and blood samples were collected at 3 time points: (i) after 35 days of vapor/air exposure (PD58); (ii) after 1 day of withdrawal (PD59), and (iii) 28 days after withdrawal (PD86). The ionized calcium-binding adapter molecule 1 (Iba-1) was used to index microglial activation, and cytokine/chemokine responses were analyzed using magnetic bead panels. RESULTS: After 35 days of adolescent vapor exposure, a significant increase in Iba-1 immunoreactivity was seen in amygdala, frontal cortex, hippocampus, and substantia nigra. However, Iba-1 density returned to control levels at both 1 day and 28 days of withdrawal except in the hippocampus where Iba-1 density was significantly lower than controls. In serum, adolescent EtOH exposure induced a reduction in IL-13 and an increase in fractalkine at day 35. After 1 day of withdrawal, IL-18 was reduced, and IP-10 was elevated, whereas both IP-10 and IL-10 were elevated at 28 days following withdrawal. In the frontal cortex, adolescent EtOH exposure induced an increase in IL-1ß at day 35, and 28 days of withdrawal, and IL-10 was increased after 28 days of withdrawal. CONCLUSION: These data demonstrate that EtOH exposure during adolescence produces significant microglial activation; however, inflammatory markers seen in the blood appear to differ from those observed in the brain.

Effect of Gabapentin on Sleep and Event-Related Oscillations (EROs) in Rats Exposed to Chronic Intermittent Ethanol Vapor and Protracted Withdrawal.

BACKGROUND: Disturbances in sleep architecture, especially reductions in slow-wave sleep (SWS), are symptoms commonly observed in individuals with alcohol use disorders. Recent clinical trials have demonstrated that the anticonvulsant and analgesic drug gabapentin may have therapeutic value in normalizing sleep quality in recovering alcoholics. However, the brain mechanisms underlying this improvement in sleep following gabapentin treatment remain unknown. METHODS: In this study, adult Wistar rats were exposed to 8 weeks of chronic intermittent ethanol [EtOH] vapor (blood EtOH concentrations averaged 128.2 ± 17.4 mg/dl) or control conditions and then withdrawn. Sleep electroencephalograms [EEGs] and event-related oscillations (EROs) were evaluated at baseline prior to EtOH exposure and 24 hours following EtOH withdrawal. Four weeks following EtOH withdrawal the effects of saline and 2 doses of gabapentin (30, 120 mg/kg), on EROs and sleep EEGs, were evaluated. RESULTS: As compared to baseline, 24 hours following alcohol withdrawal SWS became fragmented as indexed by a significant increase in the number and a decrease in the duration of SWS episodes. Compared to controls, the EtOH-exposed group had more ERO energy in the beta frequency band in the parietal cortex. Gabapentin induced a dose-dependent decrease in the latency to the first SWS episode, and a reduction in sleep fragmentation. Gabapentin also produced a dose-dependent increase in ERO energy in the control group that was significantly attenuated in the EtOH-exposed group in the theta, and beta frequency bands. CONCLUSIONS: Taken together, these studies suggest that gabapentin can reverse some of the alcohol-induced sleep and EEG deficits but does not eliminate all of the enduring brain effects of EtOH exposure.

Decreases in energy and increases in phase locking of event-related oscillations to auditory stimuli occur during adolescence in human and rodent brain.

Synchrony of phase (phase locking) of event-related oscillations (EROs) within and between different brain areas has been suggested to reflect communication exchange between neural networks and as such may be a sensitive and translational measure of changes in brain remodeling that occur during adolescence. This study sought to investigate developmental changes in EROs using a similar auditory event-related potential (ERP) paradigm in both rats and humans. Energy and phase variability of EROs collected from 38 young adult men (aged 18-25 years), 33 periadolescent boys (aged 10-14 years), 15 male periadolescent rats [at postnatal day (PD) 36] and 19 male adult rats (at PD103) were investigated. Three channels of ERP data (frontal cortex, central cortex and parietal cortex) were collected from the humans using an 'oddball plus noise' paradigm that was presented under passive (no behavioral response required) conditions in the periadolescents and under active conditions (where each subject was instructed to depress a counter each time he detected an infrequent target tone) in adults and adolescents. ERPs were recorded in rats using only the passive paradigm. In order to compare the tasks used in rats to those used in humans, we first studied whether three ERO measures [energy, phase locking index (PLI) within an electrode site and phase difference locking index (PDLI) between different electrode sites] differentiated the 'active' from 'passive' ERP tasks. Secondly, we explored our main question of whether the three ERO measures differentiated adults from periadolescents in a similar manner in both humans and rats. No significant changes were found in measures of ERO energy between the active and passive tasks in the periadolescent human participants. There was a smaller but significant increase in PLI but not PDLI as a function of active task requirements. Developmental differences were found in energy, PLI and PDLI values between the periadolescents and adults in both the rats and the human participants. Neuronal synchrony as indexed by PLI and PDLI was significantly higher to the infrequent (target) tone compared to the frequent (nontarget) tone in all brain sites in all of the regions of interest time-frequency intervals. Significantly higher ERO energy and significantly lower synchrony was seen in the periadolescent humans and rats compared to their adult counterparts. Taken together these findings are consistent with the hypothesis that adolescent remodeling of the brain includes decreases in energy and increases in synchrony over a wide frequency range both within and between neuronal networks and that these effects are conserved over evolution.

Event-related potential responses to the acute and chronic effects of alcohol in adolescent and adult Wistar rats.

BACKGROUND: This study explored the hypothesis that adolescent ethanol (EtOH) exposure may cause long-lasting changes in EtOH sensitivity by exploring the age-related effects of acute alcohol on intoxication and on event-related potential (ERP) responses to acoustic stimuli in EtOH-naïve adolescent and adult male Wistar rats and in adult rats that were exposed to chronic EtOH/control conditions during adolescence. METHODS: EtOH-naïve adolescent (postnatal day 32 [PD32]) and adult male rats (PD99) were included in the first study. In a second study, rats were exposed to 5 weeks of EtOH vapor (blood EtOH concentrations at 175 mg%) or air from PD24 to 59 and allowed to mature until PD90. In both studies, rats were implanted with cortical recording electrodes, and the effects of acute EtOH (0.0, 1.5, and 3.0 g/kg) on behavioral and ERP responses were assessed. RESULTS: Adolescents were found to have higher amplitude and longer latency P3a and P3b components at baseline as compared to adult rats, and EtOH was found to produce a robust dose-dependent increase in the latency of the P3a and P3b components of the auditory ERP recorded in cortical sites in both adolescents and adults. However, EtOH produced significantly larger delays in P3a and P3b latencies in adults as compared to adolescents. Acute EtOH administration was also found to produce a robust dose-dependent increase in the latency of the P3a and P3b components in adult animals exposed to EtOH vapor as adolescents and air exposed controls; however, larger acute EtOH-induced increases in P3a and P3b latencies were seen in controls as compared to adolescent vapor exposed rats. CONCLUSIONS: Adolescent rats have a less intense P3 latency response to acute EtOH administration when compared to adult rats. Exposure to chronic EtOH during adolescence can cause "retention" of the adolescent phenotype of reduced P3 latency sensitivity to EtOH.

Hypermorphic mutation of the voltage-gated sodium channel encoding gene Scn10a causes a dramatic stimulus-dependent neurobehavioral phenotype.

The voltage-gated sodium channel Na(v)1.8 is known to function in the transmission of pain signals induced by cold, heat, and mechanical stimuli. Sequence variants of human Na(v)1.8 have been linked to altered cardiac conduction. We identified an allele of Scn10a encoding the α-subunit of Na(v)1.8 among mice homozygous for N-ethyl-N-nitrosourea-induced mutations. The allele creates a dominant neurobehavioral phenotype termed Possum, characterized by transient whole-body tonic immobility induced by pinching the skin at the back of the neck ("scruffing"). The Possum mutation enhanced Na(v)1.8 sodium currents and neuronal excitability and heightened sensitivity of mutants to cold stimuli. Striking electroencephalographic changes were observed concomitant with the scruffing-induced behavioral change. In addition, electrocardiography demonstrated that Possum mice exhibited marked sinus bradycardia and R-R variability upon scruffing, abrogated by infusion of atropine. However, atropine failed to prevent or mitigate the tonic immobility response. Hyperactive sodium conduction via Na(v)1.8 thus leads to a complex neurobehavioral phenotype, which resembles catatonia in schizophrenic humans and tonic immobility in other mammals upon application of a discrete stimulus; no other form of mechanosensory stimulus could induce the immobility phenotype. Our data confirm the involvement of Na(v)1.8 in transducing pain initiated by cold and additionally implicate Na(v)1.8 in previously unknown functions in the central nervous system and heart.

Social zeitgeber and sleep loss as risk factors for suicide in American Indian adolescents.

American Indians / Alaska Natives (AI/AN) bear a high burden of suicide, the reasons for which are not completely understood, and rates can vary by tribal group and location. This article aims to identify circumstances reported by a community group of American Indian adolescent participants to be associated with their depression and/or suicide. American Indian adolescents (n = 360) were recruited from contiguous reservations and were assessed with a semi-structured diagnostic interview. Twenty percent of the adolescents reported suicidal thoughts (ideation, plans), an additional 8% reported a history of suicide attempts, and three deaths due to suicide were reported. Suicidal behaviors and major depressive disorder (MDD) co-occurred and were more common among female adolescents. The distressing events that adolescents most often reported were: death in the family, family disruption, peer relationship problems, and school problems. All of these events were significantly associated with suicidal behaviors, however those with suicidal acts were more likely to report death in the family. Those with MDD but no suicidal behaviors were more likely to report disruptions in the family. Disruptions in falling asleep were also associated with suicidal behaviors and having experienced a death in the family. Disruptions in important relationships, particularly through death or divorce, may be interpreted as a loss or disruption in "social zeitgebers" that may in turn disturb biological rhythms, such as sleep, thus potentially increase the risk for MDD and/or suicide. Prevention programs aimed at ameliorating the impact of disruptions in important relationships may potentially reduce suicidal behaviors in AI/AN adolescents.

Peri-adolescent ethanol vapor exposure produces reductions in hippocampal volume that are correlated with deficits in prepulse inhibition of the startle.

BACKGROUND: Epidemiological studies suggest that excessive alcohol consumption is prevalent among adolescents and may have lasting neurobehavioral consequences. The use of animal models allows for the separation of the effects of adolescent ethanol (EtOH) exposure from genetic background and other environmental insults. In this study, the effects of moderate EtOH vapor exposure, during adolescence, on structural diffusion tensor imaging (DTI) and behavioral measures were evaluated in adulthood. METHODS: A total of 53 Wistar rats were received at postnatal day (PD) 21 and were randomly assigned to EtOH vapor (14 hours on/10 hours off/day) or air exposure for 35 days from PD 23 to 58 (average blood ethanol concentration: 169 mg%). Animals were received in 2 groups that were subsequently sacrificed at 2 time points following withdrawal from EtOH vapor: (i) at 72 days of age, 2 weeks following withdrawal or (ii) at day 128, 10 weeks following withdrawal. In the second group, behavior in the light/dark box and prepulse inhibition (PPI) of the startle was also evaluated. Fifteen animals in each group were scanned, postmortem, for structural DTI. RESULTS: There were no significant differences in body weight between EtOH and control animals. Volumetric data demonstrated that total brain, hippocampal, corpus callosum but not ventricular volume were significantly larger in the 128-day-sacrificed animals as compared to the 72 day animals. The hippocampus was smaller and the ventricles larger at 128 days as compared to 72 days, in the EtOH-exposed animals, leading to a significant group × time effect. EtOH-exposed animals sacrificed at 128 days also had diminished PPI, and more rears in the light box were significantly correlated with hippocampal size. CONCLUSIONS: These studies demonstrate that DTI volumetric measures of hippocampus are significantly impacted by age and peri-adolescent EtOH exposure and withdrawal in Wistar rats.

Adolescent alcohol exposure increases orexin-A/hypocretin-1 in the anterior hypothalamus.

Adolescence is a time of marked changes in sleep, neuromaturation, and alcohol use. While there is substantial evidence that alcohol disrupts sleep and that disrupted sleep may play a role in the development of alcohol use disorders (AUD), there is very little known about the brain mechanisms underlying this phenomenon. The orexin (also known as hypocretin) system is fundamental for a number of homeostatic mechanisms, including the initiation and maintenance of wakefulness that may be impacted by adolescent alcohol exposure. The current study investigated the impact of adolescent ethanol exposure on adult orexin-A/hypocretin-1 immunoreactive (orexin-A + IR) cells in hypothalamic nuclei in two models of adolescent intermittent ethanol (AIE) exposure. Both models assess adult hypothalamic orexin following either an AIE vapor exposure paradigm, or an AIE intragastric gavage paradigm during adolescence. Both AIE exposure models found that binge levels of ethanol intoxication during adolescence significantly increased adult orexin-A + IR expression in the anterior hypothalamic nucleus (AHN). Further, both AIE models found no change in orexin-A + IR in the posterior hypothalamic area (PH), perifornical nucleus (PeF), dorsomedial hypothalamic nucleus dorsal part (DMD) or lateral hypothalamic area (LH). However, AIE vapor exposure reduced orexin-A + IR in the paraventricular nucleus (PVN), but AIE gavage exposure did not. These findings suggest that the AHN orexinergic system is increased in adults following binge-like patterns of intoxication during adolescence. Altered adult AHN orexin could contribute to long-lasting changes in sleep.

Delta Event-Related Oscillations Are Related to a History of Extreme Binge Drinking in Adolescence and Lifetime Suicide Risk.

Alcohol exposure typically begins in adolescence, and heavy binge drinking is associated with health risk behaviors. Event-related oscillations (EROs) may represent sensitive biomarkers or endophenotypes for early alcohol exposure as well as other risk behaviors such as suicidal thoughts and actions. In this study, young adults (age 18-30 years) of American Indian (AI) (n = 479) and Mexican American (MA) (n = 705) ancestry were clinically assessed, and EROs were generated to happy, sad and neutral faces. Extreme adolescent binge drinking (10+ drinks) was common (20%) in this population of AI/MA and associated with a significantly increased risk of a lifetime history of suicidal acts (SA, suicide attempts, deaths) but not suicidal thoughts (ST, ideation, plans). ST were reported among MA participants, whereas SA were more common among AI young adults. Extreme adolescent binge drinking was also associated with errors in detection of sad and neutral faces, increases in delta ERO energy, and decreases in phase locking (PL), particularly in parietal areas. A lifetime history of ST was associated with increases in delta ERO energy and PL, whereas SA were associated with decreases in both. These studies suggest that ERO measures may represent important potential biomarkers of adolescent extreme binge drinking and risk for suicidal behaviors.

Effect of suvorexant on event-related oscillations and EEG sleep in rats exposed to chronic intermittent ethanol vapor and protracted withdrawal.

STUDY OBJECTIVES: Insomnia is a prominent complaint in patients with alcohol use disorders (AUD). However, despite the importance of sleep in the maintenance of sobriety, treatment options for sleep disturbance associated with a history of AUD are currently limited. Recent clinical trials have demonstrated that suvorexant, a dual Hct/OX receptor antagonist, normalizes sleep in patients with primary insomnia; yet, its potential for the treatment of sleep pathology associated with AUD has not been investigated in either preclinical or clinical studies. METHODS: This study employed a model whereby ethanol vapor exposure or control conditions were administered for 8 weeks to adult rats. Waking event-related oscillations (EROs) and EEG sleep were evaluated at baseline before exposure and again following 24 hr of withdrawal from the exposure. Subsequently, the ability of vehicle (VEH) and two doses (10, 30 mg/kg IP) of suvorexant to modify EROs, sleep, and the sleep EEG was investigated. RESULTS: After 24 hr following EtOH withdrawal, the ethanol-treated group had increases in waking ERO θ and ß activity, more fragmented sleep (shorter duration and increased frequency of slow wave (SW) and rapid eye movement [REM] sleep episodes), and increased θ and ß power in REM and SW sleep. Suvorexant induced a dose-dependent decrease in the latency to REM and SW sleep onsets but also produced REM and SW sleep fragmentation and increased ß energy in waking EROs when compared with VEH. CONCLUSIONS: Taken together, these studies suggest that suvorexant has overall sleep-promoting effects, but it may exacerbate some aspects of sleep and EEG pathology.

CR-19-0950: Event-related responses to alcohol-related stimuli in Mexican-American young adults: Relation to age, gender, comorbidity and "dark side" symptoms.

BACKGROUND: Electrophysiological variables may represent sensitive biomarkers of vulnerability to or endophenotypes for alcohol use disorders (AUD). METHODS: Young adults (age 18-30 yrs, n = 580) of Mexican American heritage were assessed with the Semi-Structured Assessment for the Genetics of Alcoholism and event-related oscillations (EROs) generated in response to a task that used pictures of objects, food, and alcohol-related and non-alcohol-related drinks as stimuli. RESULTS: Decreases in energy in the alpha and beta frequencies and higher phase synchrony within cortical brain areas were seen in response to the alcohol-related as compared to the non-alcohol-related stimuli. Differences in ERO energy and synchrony responses to alcohol-related stimuli were also found as a function of age, sex, AUD status and comorbidity. Age-related decreases in energy and increases in synchrony were found. Females had significantly higher energy and lower synchrony values than males. Participants with AUD had higher synchrony values specifically in the beta frequencies, whereas those with a lifetime diagnosis of conduct disorder and/or antisocial personality disorder had lower alpha power and synchrony, and those with any affective disorder had lower ERO energy in the beta frequencies. Those with substance-associated affective "dark-side" symptoms had slower reaction times to the task, lower energy in the beta frequencies, lower local synchrony in the theta frequencies, and higher long-range synchrony in the delta and beta frequencies. CONCLUSIONS: These findings suggest that EROs recorded to alcohol-related stimuli may be biomarkers of comorbid risk factors, symptoms and disorders associated with AUD that also can differentiate those with "dark-side symptoms".

Time course of microglia activation and brain and blood cytokine/chemokine levels following chronic ethanol exposure and protracted withdrawal in rats.

Alcohol produces complex effects on the immune system. Moderate alcohol use (1-2 drinks per day) has been shown to produce anti-inflammatory responses in human blood monocytes, whereas, the post mortem brains of severe alcoholics show increased immune gene expression and activated microglial markers. The present study was conducted to evaluate the time course of alcohol effects during exposure and after withdrawal, and to determine the relationship between microglial and cytokine responses in brain and blood. Forty-eight adult, male Wistar rats were exposed to chronic ethanol vapors, or air control, for 5 weeks. Following ethanol/air exposure blood and brains were collected at three time points: 1) while intoxicated, following 35 days of air/vapor exposure; 2) following 24 h of withdrawal from exposure, and 3) 28 days after withdrawal. One hemisphere of the brain was flash-frozen for cytokine analysis, and the other was fixed for immunohistochemical analysis. The ionized calcium-binding adapter molecule 1 (Iba-1) was used to evaluate microglia activation at the three time points, and rat cytokine/chemokine Magnetic Bead Panels (Millipore) were used to analyze frontal cortex tissue lysate and serum. Ethanol induced a significant increase in Iba-1 that peaked at day 35, remained significant after 1 day of withdrawal, and was elevated at day 28 in frontal cortex, amygdala, and substantia nigra. Ethanol exposure was associated with a transient reduction of the serum level of the major pro- and anti-inflammatory cytokines and chemokines and a transient increase of effectors of sterile inflammation. Little or no changes in these molecules were seen in the frontal cortex except for HMG1 and fractalkine that were reduced and elevated, respectively, at day 28 following withdrawal. These data show that ethanol exposure produces robust microglial activation; however, measures of inflammation in the blood differ from those in the brain over a protracted time course.

Electrophysiological effects of dizocilpine (MK-801) in adult rats exposed to ethanol during adolescence.

BACKGROUND: Despite evidence showing persistent changes in N-methyl-D-aspartate (NMDA)-receptor function following ethanol (EtOH) exposure, the contribution of NMDA systems to the long-term neurophysiological consequences of adolescent EtOH exposure is unclear. The aims of this study were the following: (1) to determine whether adolescent EtOH exposure produces neurophysiological changes after a prolonged withdrawal period in adult rats and (2) to assess protracted alterations in neurophysiological responses to the NMDA antagonist MK-801 in adult rats exposed to EtOH during adolescence. METHODS: Adolescent male Wistar rats were exposed to EtOH vapor for 12 h/d for 5 weeks. The effects of MK-801 (0.0 to 0.1 mg/kg, intraperitoneally) on the electroencephalogram (EEG) and auditory event-related potentials (ERPs) were assessed after 8 weeks of abstinence from EtOH. RESULTS: Experiments in aim 1 revealed that adolescent EtOH exposure reduced EEG variability in the frontal cortex in the 4 to 6 Hz band but had no effect on cortical and hippocampal EEG power and ERPs. Experiments in aim 2 showed that MK-801 significantly reduced EEG power in the parietal cortex (4 to 6 Hz, 6 to 8 Hz, 8 to 16 Hz, 16 to 32 Hz) and hippocampus (16 to 32 Hz) and EEG variability in the parietal cortex (6 to 8 Hz, 16 to 32 Hz) following adolescent EtOH exposure. MK-801 produced a significant decrease in hippocampal EEG variability (4 to 6 Hz, 8 to 16 Hz, 16 to 32 Hz) in control, but not in EtOH-exposed rats. MK-801 reduced frontal P1 ERP amplitude and latency in response to the rare tone in EtOH-exposed rats compared to controls. In contrast, MK-801 significantly reduced P3 ERP amplitude and latency in control, but not in EtOH-exposed rats. CONCLUSIONS: The effects of MK-801 on hippocampal EEG variability and P3 ERP amplitude and latency are significantly attenuated after a prolonged withdrawal period following adolescent EtOH exposure. However, the inhibitory effects of MK-801 on cortical and hippocampal EEG power were enhanced in rats exposed to EtOH during adolescence. Taken together, these data suggest protracted changes in NMDA systems following adolescent EtOH exposure.

Intermittent voluntary ethanol consumption combined with ethanol vapor exposure during adolescence increases drinking and alters other behaviors in adulthood in female and male rats.

Epidemiological studies suggest that binge drinking is prevalent among adolescents, and may result in neurobehavioral consequences. Animal models provide the experimental control to investigate the consequences of "binge" alcohol exposure during this neurodevelopmental epoch. The current study used an animal model that combined an intermittent pattern of alcohol vapor exposure with voluntary drinking of 20% unsweetened alcohol in adolescent male and female Wistar rats (postnatal day [PD] 22-62), in order to test for potential differences in behavioral changes, ethanol drinking, and hypocretin/orexin (Hcrt/OX) signaling associated with exposure status. Two weeks after discontinuation of the alcohol vapor exposure and drinking during adolescence, rats were tested in adulthood for anxiety-like behaviors using a modified open-field conflict task, pre-pulse facilitation of startle response, light/dark box, and marble burying test. Adolescent alcohol exposure led to overall decreased startle response and increased behavioral arousal in the light/dark chamber during adulthood. Additionally, male rats demonstrated more disinhibited behavior during the conflict task compared to females, and female rats exhibited more rearing behavior during the light/dark test. Rats were also given a 2-bottle choice test that resulted in adolescent alcohol-exposed rats drinking significantly more alcohol in adulthood. Further, female rats also consumed more alcohol in adulthood compared to males. Estrous cycle phase did not account for any of the sex differences observed in the behavioral measures. Histological results indicated that adolescent alcohol did not alter Hcrt/OX-1 or Hcrt/OX-2 receptor mRNA expression levels in adult rats compared to control adults. However, female rats expressed a higher level of Hcrt/OX-1 and Hcrt/OX-2 receptor mRNA in the frontal cortex compared to males. These data suggest that our current model of intermittent ethanol exposure in adolescence can modestly affect both behavior and future consumption of alcohol and that Hcrt/OX receptor signaling differs between males and females.

Acute low-level alcohol consumption reduces phase locking of event-related oscillations in rodents.

Event-related oscillations (EROs) are rhythmic changes that are evoked by a sensory and/or cognitive stimulus that can influence the dynamics of the EEG. EROs are defined by the decomposition of the EEG signal into magnitude (energy) and phase information and can be elicited in both humans and animals. EROs have been linked to several relevant genes associated with ethanol dependence phenotypes in humans and are altered in selectively bred alcohol-preferring rats. However, pharmacological studies are only beginning to emerge investigating the impact low intoxicating doses of ethanol can have on event-related neural oscillations. The main goal of this study was to investigate the effects of low levels of voluntary consumption of ethanol, in rats, on phase locking of EROs in order to give further insight into the acute intoxicating effects of ethanol on the brain. To this end, we allow rats to self-administer unsweetened 20% ethanol over 15 intermittent sessions. This method results in a stable low-dose consumption of ethanol. Using an auditory event-related potential "oddball" paradigm, we investigated the effects of alcohol on the phase variability of EROs from electrodes implanted into the frontal cortex, dorsal hippocampus, and amygdala. We found that intermittent ethanol self-administration was sufficient to produce a significant reduction in overall intraregional synchrony across all targeted regions. These data suggest that phase locking of EROs within brain regions known to be impacted by alcohol may represent a sensitive biomarker of low levels of alcohol intoxication.

Alcohol drinking during adolescence increases consumptive responses to alcohol in adulthood in Wistar rats.

Binge drinking and the onset of alcohol-use disorders usually peak during the transition between late adolescence and early adulthood, and early adolescent onset of alcohol consumption has been demonstrated to increase the risk for alcohol dependence in adulthood. In the present study, we describe an animal model of early adolescent alcohol consumption where animals drink unsweetened and unflavored ethanol in high concentrations (20%). Using this model, we investigated the influence of drinking on alcohol-related appetitive behavior and alcohol consumption levels in early adulthood. Further, we also sought to investigate whether differences in alcohol-related drinking behaviors were specific to exposure in adolescence versus exposure in adulthood. Male Wistar rats were given a 2-bottle choice between 20% ethanol and water in one group and between two water bottles in another group during their adolescence (Postnatal Day [PD] 26-59) to model voluntary drinking in adolescent humans. As young adults (PD85), rats were trained in a paradigm that provided free access to 20% alcohol for 25 min after completing up to a fixed-ratio (FR) 16 lever press response. A set of young adult male Wistar rats was exposed to the same paradigm using the same time course, beginning at PD92. The results indicate that adolescent exposure to alcohol increased consumption of alcohol in adulthood. Furthermore, when investigating differences between adolescent high and low drinkers in adulthood, high consumers continued to drink more alcohol, had fewer FR failures, and faster completion of FR schedules in adulthood, whereas the low consumers were no different from controls. Rats exposed to ethanol in young adulthood also increased future intake, but there were no differences in any other components of drinking behavior. Both adolescent- and adult-exposed rats did not exhibit an increase in lever pressing during the appetitive challenge session. These data indicate that adolescent and early adult alcohol exposure can increase consumptive aspects of drinking but that adolescent exposure may preferentially influence the motivation to drink.

Sleep Quality in an Adult American Indian Community Sample.

STUDY OBJECTIVES: Epidemiological studies have found that insufficient sleep (< 7 h/night) is more common among American Indians/Alaska Natives (AI/AN). In this study we sought to identify specific demographic, clinical, and cultural factors that may be associated with reduced sleep quality in an American Indian community sample. METHODS: Information on demography along with personal medical, psychiatric, and drinking history was obtained using the Semi-Structured Assessment for the Genetics of Alcoholism (SSAGA). Sleep quality was assessed by the Pittsburgh Sleep Quality Index (PSQI). RESULTS: The adult participants (n = 386, 54% women) had a mean ± standard deviation age of 31.35 ± 14.4 y. Higher degrees of AI ancestry, but not cultural identification, being older than 30 y, and having a high school diploma all were factors predictive of having a short sleep duration (< 6 h). The global score on the PSQI was significantly higher in those participants with a lifetime diagnosis of substance use disorders, anxiety disorders, and affective disorders. Alcohol use disorders and affective disorders were significant predictors of sleep latency whereas anxiety and affective disorders were correlated with waking more often in the night/early morning. Nicotine dependence was associated with having trouble breathing, and alcohol use disorders and anxiety disorders with bad dreams. CONCLUSIONS: Alcohol use disorders are associated with poorer quality of sleep in this population and substance use disorders were associated with different aspects of sleep than anxiety and depressive disorders. These findings add to the understanding of the interactions between sleep and substance use, anxiety, and affective disorders in an understudied and underserved population.

Low voltage alpha EEG phenotype is associated with reduced amplitudes of alpha event-related oscillations, increased cortical phase synchrony, and a low level of response to alcohol.

Low voltage EEG (LVEEG) is a heritable phenotype that differs depending on ancestral heritage, yet its impact on brain networks and cognition remain relatively unexplored. In this study we assessed energy and task related phase locking of event-related oscillation (EROs), behavioral responses, measures of IQ and personality, and expected responses to alcohol in a large sample of individuals with LVEEG compared to those with higher voltage variants. Participants (n=762) were recruited from a Native American community and completed a diagnostic interview, the Quick Test, the Subjective High Assessment Scale Expectation Version (SHAS-E) and the Maudsley Personality Inventory. Clinical and spectral analyzed EEGs were collected for determination of the presence of a LVEEG variant. EROs were generated using a facial expression recognition task. Participants with LVEEG (n=451) were significantly more likely to be older, married and have higher degrees of Native American heritage but did not differ in gender, income or education. Individuals with LVEEG were also found to have decreased energy in their alpha EROs, increased phase locking between stimulus trials, and increased phase-locking between cortical brain areas. No significant differences in the cognitive tests, personality variables or alcohol dependence or anxiety diagnoses were found, however, individuals with LVEEG did report a larger number of drinks ever consumed in a 24-h period and a less intense expected response to alcohol. These data suggest that alpha power in the resting EEG is highly associated with energy and cortical connectivity measures generated by event-related stimuli, as well as potentially increased risk for alcohol use.

Characterization of the sleep-wake patterns in mice lacking fatty acid amide hydrolase.

STUDY OBJECTIVES: Oleamide and anandamide are fatty acid amides implicated in the regulatory mechanisms of sleep processes. However, due to their prompt catabolism by fatty acid amide hydrolase (FAAH), their pharmacologic and behavioral effects, in vivo, disappear rapidly. To determine if, in the absence of FAAH, the hypnogenic fatty acid amides induce an increase of sleep, we characterized the sleep-wake patters in FAAH-knockout mice [FAAH (-/-)] before and after sleep deprivation. DESIGN: FAAH (-/-), FAAH (+/-), and FAAH (+/+) mice were implanted chronically for sleep, body temperature (Tb), and locomotor activity (LMA) recordings. Sleep-wake states were recorded during a 24-hour baseline session followed by 8 hours of sleep deprivation. Recovery recordings were done during the 16 hours following sleep deprivation. Total amount of wake, slow-wave sleep, and rapid eye movement sleep were calculated and compared between genotypes. The electroencephalographic spectral analysis was performed by fast Fourier transform analysis. Telemetry recordings of Tb and LMA were carried out continuously during 4 days under baseline conditions. SETTING: N/A. PATIENTS OR PARTICIPANTS: FAAH (-/-) mice and their heterozygote (+/-) and control (+/+) littermates were used. INTERVENTIONS: Sleep deprivation. MEASUREMENTS AND RESULTS: FAAH (-/-) mice possess higher values of slow-wave sleep and more intense episodes of slow-wave sleep than do control littermates under baseline conditions that are not related to differences in Tb and LMA. A rebound of slow-wave sleep and rapid eye movement sleep as well an increase in the levels of slow-wave activity were observed after sleep deprivation in all genotypes. CONCLUSION: These findings support the role of fatty acid amides as possible modulators of sleep and indicate that the homeostatic mechanisms of sleep in FAAH (-/-) mice are not disrupted.