Prazosin Prevents Increased Anxiety Behavior That Occurs in Response to Stress During Alcohol Deprivations.

AIMS: Stress-induced anxiety is a risk factor for relapse to alcohol drinking. The aim of this study was to test the hypothesis that the central nervous system (CNS)-active α1-adrenergic receptor antagonist, prazosin, would block the stress-induced increase in anxiety that occurs during alcohol deprivations. METHODS: Selectively bred male alcohol-preferring (P) rats were given three cycles of 5 days of ad libitum voluntary alcohol drinking interrupted by 2 days of alcohol deprivation, with or without 1 h of restraint stress 4 h after the start of each of the first two alcohol deprivation cycles. Prazosin (1.0 or 1.5 mg/kg, IP) or vehicle was administered before each restraint stress. Anxiety-like behavior during alcohol deprivation following the third 5-day cycle of alcohol drinking (7 days after the most recent restraint stress ± prazosin treatment) was measured by performance in an elevated plus-maze and in social approach/avoidance testing. RESULTS: Rats that received constant alcohol access, or alcohol access and deprivations without stress or prazosin treatments in the first two alcohol deprivations did not exhibit augmented anxiety-like behavior during the third deprivation. In contrast, rats that had been stressed during the first two alcohol deprivations exhibited increased anxiety-like behavior (compared with control rats) in both anxiety tests during the third deprivation. Prazosin given before stresses in the first two cycles of alcohol withdrawal prevented increased anxiety-like behavior during the third alcohol deprivation. CONCLUSION: Prazosin treatment before stresses experienced during alcohol deprivations may prevent the increased anxiety during subsequent deprivation/abstinence that is a risk factor for relapse to alcohol drinking. SHORT SUMMARY: Administration of prazosin before stresses during repetitive alcohol deprivations in male alcohol-preferring (P) rats prevents increased anxiety during a subsequent deprivation without further prazosin treatment. Prazosin treatment during repeated alcohol deprivations may prevent the increased anxiety that is a risk factor for relapse to alcohol drinking.

Prazosin + Naltrexone Decreases Alcohol Drinking More Effectively Than Does Either Drug Alone in P Rats with a Protracted History of Extensive Voluntary Alcohol Drinking, Dependence, and Multiple Withdrawals.

BACKGROUND: Prazosin (PRZ; an α1 -adrenergic receptor antagonist) and naltrexone (NTX; a nonspecific opioid receptor antagonist) each decrease alcohol drinking when administered to rats selectively bred for high voluntary alcohol drinking (alcohol-preferring or "P"), and the combination of PRZ + NTX decreases alcohol drinking more effectively than does either drug alone. As drug responsiveness can depend on history of alcohol drinking and dependence, we investigated whether various schedules of PRZ and NTX administration, alone or in combination, are effective in decreasing alcohol drinking in male P rats with a history of protracted voluntary alcohol drinking, dependence, and repeated withdrawals closely resembling human alcoholism. METHODS: Male P rats became alcohol-dependent during 1 year of ad libitum 24 h/d access to food, water, and 20% alcohol with repetitive temporary alcohol withdrawals. Four sequential studies then addressed effects of oral PRZ (2 mg/kg) and NTX (10 mg/kg), alone or together, on alcohol drinking during: (i) daily alcohol access with daily drug treatment, (ii) intermittent alcohol access with daily drug treatment, (iii) intermittent alcohol access with occasional drug treatment, and (iv) postdeprivation reinstatement of alcohol access. RESULTS: The combination of PRZ + NTX consistently suppressed alcohol drinking during daily or intermittent alcohol access conditions and when drug treatment was either daily or occasional. PRZ + NTX was consistently more effective than either drug alone. The reduction in alcohol drinking was not due to sedation, motor effects, or malaise. CONCLUSIONS: Both daily and "as-needed" treatment with PRZ + NTX are highly effective in suppressing daily, intermittent, and postdeprivation alcohol drinking in male P rats with a protracted history of alcohol dependence and repeated withdrawals. This drug combination may be especially effective for treating individuals with long histories of heavy alcohol abuse, dependence, and repeated relapse, as commonly encountered in clinical practice.

Prazosin Reduces Alcohol Intake in an Animal Model of Alcohol Relapse.

BACKGROUND: Many alcoholics and heavy drinkers undergo repeated cycles of alcohol abstinence followed by relapse to alcohol drinking; a pattern that contributes to escalated alcohol intake over time. In rodents, alcohol drinking that is interspersed with periods of alcohol deprivation (imposed abstinence) increases alcohol intake during reaccess to alcohol. This is termed the "alcohol deprivation effect" or "ADE" and is a model of alcohol relapse in humans. We have previously reported that prazosin reduces alcohol drinking during both brief and prolonged treatment in rats selectively bred for alcohol preference ("P" rats). This study explores whether prazosin prevents alcohol "relapse" in P rats, as reflected by a reduced or abolished ADE. METHODS: Adult male P rats were given 24-hour access to food and water and scheduled access to alcohol (15 and 30% v/v solutions presented concurrently) for 2 h/d. After 5 weeks, rats underwent imposed alcohol deprivation for 2 weeks, followed by alcohol reaccess for 2 weeks, and this pattern was repeated for a total of 3 cycles. Rats were injected with prazosin (0, 0.5, 1.0, or 2.0 mg/kg body weight, intraperitoneally) once a day for the first 5 days of each alcohol reaccess cycle. RESULTS: Alcohol intake increased on the first day of each alcohol reaccess cycle, demonstrating the formation of an ADE. The ADE was short-lived, lasting only 1 day, during each of the 3 cycles. Prazosin, in all doses tested, prevented the expression of an ADE in all 3 alcohol reaccess cycles. CONCLUSIONS: Prazosin decreases alcohol intake in P rats even in a situation that would be expected to increase alcohol drinking, namely following periods of alcohol deprivation. This suggests that prazosin may be effective in reducing alcohol relapse that often occurs during attempts to achieve permanent alcohol abstinence in treatment-seeking alcoholics and heavy drinkers.

Acoustic startle in alcohol-naïve male rats predicts subsequent voluntary alcohol intake and alcohol preference.

AIMS: Acoustic startle response in rats is used to model sensorimotor reactivity. The aim of the study was to determine whether acoustic startle response in alcohol-naïve rats predicts subsequent increased voluntary alcohol drinking or alcohol preference. METHODS: Startle responses to 90, 95 and 100 decibel (dB) white noise stimuli presented in counterbalanced semi-randomized order were tested in alcohol-naïve young adult male Wistar rats before voluntary alcohol intake was established with an intermittent alcohol access (IAA) model. RESULTS: Startle amplitude in response to 95 or 100 dB stimuli was positively correlated with subsequent alcohol intake and alcohol preference following 3 months of IAA. Rats with high (median split) pre-IAA startle amplitude in response to 95 or 100 dB stimuli developed increased alcohol intake as well as increased alcohol preference following 3 months of IAA, relative to rats with low pre-IAA startle amplitude. CONCLUSION: Startle response to moderate acoustic stimuli can be a predictive index of vulnerability to developing increased alcohol drinking.

The α2-adrenergic receptor agonist, clonidine, reduces alcohol drinking in alcohol-preferring (P) rats.

Evidence suggests that noradrenergic signaling may play a role in mediating alcohol-drinking behavior in both rodents and humans. We have investigated this possibility by administering clonidine to alcohol-drinking rats selectively bred for alcohol preference (P line). Clonidine is an α2-adrenergic receptor agonist which, at low doses, inhibits noradrenergic signaling by decreasing norepinephrine release from presynaptic noradrenergic neurons. Adult male P rats were given 24 h access to food and water and scheduled access to a 15% (v/v) alcohol solution for 2 h daily. Rats received intra-peritoneal (IP) injections with clonidine (0, 10, 20, 40, or 80 µg/kg body weight [BW], 10-11 rats/treatment group) once/day at 30 min prior to onset of the daily 2 h alcohol access period for 2 consecutive days. Clonidine, in doses of 40 or 80 µg/kg BW, significantly reduced alcohol intake on both days of treatment (p<0.001). Two weeks later, rats were treated with clonidine for 5 consecutive days and clonidine, in doses of 40 or 80 µg/kg BW, reduced alcohol intake on all 5 treatment days (p < 0.001). Clonidine did not alter water consumption during the daily 2 h free-choice between alcohol and water. In a separate group of male P rats, clonidine (40 µg/kg BW) suppressed intake of a saccharin solution (0.04 g/L). These results are consistent with and complement our previous findings that the α1-adrenergic receptor antagonist, prazosin, decreases voluntary alcohol drinking in alcohol-preferring rats, but suggests that effects of clonidine may not be specific for alcohol. The results suggest that although activation of the noradrenergic system plays an important role in mediating voluntary alcohol drinking, care is needed in selecting which drugs to use to suppress central noradrenergic signaling in order to maximize the selectivity of the drugs for treating alcohol-use disorders.

Combining the α1 -adrenergic receptor antagonist, prazosin, with the ß-adrenergic receptor antagonist, propranolol, reduces alcohol drinking more effectively than either drug alone.

BACKGROUND: Evidence suggests that activation of the noradrenergic system may contribute to alcohol drinking in animals and humans. Our previous studies demonstrated that blocking α1 -adrenergic receptors with the antagonist, prazosin, decreased alcohol drinking in rats under various conditions. As noradrenergic activation is also regulated by ß-adrenergic receptors, we now examine the effects of the ß-adrenergic receptor antagonist, propranolol, alone or in combination with prazosin, on alcohol drinking in rats selectively bred for high voluntary alcohol intake and alcohol preference (P line). METHODS: Two studies were conducted with male P rats. In study 1, rats were allowed to become alcohol-dependent during 14 weeks of ad libitum access to food, water, and 20% alcohol, and the effect of propranolol (5 to 15 mg/kg, intraperitoneally [IP]) and prazosin (1 to 2 mg/kg, IP) on alcohol intake during withdrawal was assessed. In study 2, the effect of propranolol (5 mg/kg, IP) and prazosin (2 mg/kg, IP) on alcohol intake following prolonged imposed abstinence was assessed. RESULTS: Alcohol drinking following propranolol treatment was variable, but the combination of propranolol + prazosin consistently suppressed alcohol drinking during both alcohol withdrawal and following prolonged imposed abstinence, and the combination of these 2 drugs was more effective than was treatment with either drug alone. CONCLUSIONS: Treatment with prazosin + propranolol, or a combination of other centrally active α1 - and ß-adrenergic receptor antagonists, may assist in preventing alcohol relapse in some individuals.

Combining naltrexone and prazosin in a single oral medication decreases alcohol drinking more effectively than does either drug alone.

BACKGROUND: Naltrexone (NTX) is underutilized in clinical treatment settings because its efficacy is modest, and it is not effective for all alcoholics and, when it is effective, a significant number of alcoholics fail to maintain initial treatment gains and subsequently relapse to heavy drinking. This has slowed acceptance of NTX by the treatment community, and there is a clear need for additional treatments for alcoholism and alcohol use disorders. Given that NTX and prazosin can each reduce alcohol drinking in rats selectively bred for alcohol preference and high voluntary alcohol drinking (alcohol-preferring "P" rats), we tested whether a combination of NTX + prazosin is more effective in decreasing alcohol drinking than is either drug alone. METHODS: P rats were given access to a 15% (v/v) alcohol solution for 2 hours daily. Rats were fed NTX and prazosin, alone or in combination, prior to onset of the daily 2-hour alcohol access period for 4 weeks and the effect of drug treatment on alcohol and water intake was assessed. RESULTS: During the first week of treatment, neither a low dose of NTX, nor prazosin, was effective in decreasing alcohol intake when each drug was administered alone, but combining the 2 drugs in a single medication significantly reduced alcohol intake. The combination was as effective as was a higher dose of NTX. Using a low dose of NTX in combination with prazosin may reduce the potential for undesirable side effects early in treatment which, in turn, may improve patient compliance and result in a more successful outcome when NTX is used for treating alcoholism and alcohol use disorders. CONCLUSIONS: Combining low-dose NTX and prazosin in a single medication may be more useful than is either drug alone for treating both inpatient and outpatient alcoholics and heavy drinkers early in the treatment process.

Prazosin reduces alcohol drinking throughout prolonged treatment and blocks the initiation of drinking in rats selectively bred for high alcohol intake.

BACKGROUND: This study examined whether prazosin reduces alcohol drinking over the course of prolonged treatment and whether it blocks the initiation of alcohol drinking in rats with a genetic predisposition toward high alcohol drinking, that is alcohol-preferring (P) rats. METHODS: In study one, alcohol-experienced P rats that had been drinking alcohol for 2 h/d for several months were treated daily with prazosin (0, 0.5, 1.0, or 2.0 mg/kg body weight [BW]) for 7 weeks. In study two, alcohol-naïve P rats were treated daily with prazosin (0, 1.0, or 2.0 mg/kg BW) for 2 weeks prior to, or concomitantly with, the initiation of alcohol access and throughout 3 weeks of alcohol availability. Prazosin treatment and alcohol access were then discontinued for 2 weeks followed by reinstatement of alcohol access without prazosin treatment for 4 weeks, followed by resumption of daily prazosin treatment (2.0 mg/kg BW) for 3 weeks. RESULTS: Prazosin reduced alcohol drinking throughout 7 weeks of treatment in P rats accustomed to drinking alcohol. Following termination of prazosin treatment, alcohol drinking slowly returned to pretreatment baseline. Reduced alcohol intake was accompanied by increased water intake. In alcohol-naïve P rats, prazosin administration prior to the first opportunity to drink alcohol and throughout 3 weeks of alcohol access retarded acquisition of alcohol drinking and reduced the amount of alcohol consumed. When prazosin was administered concomitantly with the first opportunity to drink alcohol, it abolished acquisition of alcohol drinking. Discontinuation of prazosin treatment allowed expression of a genetic predisposition toward high alcohol drinking to gradually emerge. Prazosin retained the ability to reduce alcohol intake with repeated treatments. CONCLUSIONS: Prazosin decreased alcohol drinking during prolonged treatment and may be useful for treating alcoholism and alcohol-use disorders. Prazosin may also be useful for deterring the initiation of drinking in individuals with a family history of alcoholism.

The α1-adrenergic receptor antagonist, doxazosin, reduces alcohol drinking in alcohol-preferring (P) Rats.

BACKGROUND: Evidence supports a role for the noradrenergic system in alcohol drinking in animals and humans. Our previous studies demonstrated the efficacy of prazosin, an α1-adrenergic antagonist, in decreasing alcohol drinking in rat models of alcohol dependence. Prazosin has also been shown to decrease alcohol drinking in treatment-seeking alcohol-dependent men. Clinically, the use of prazosin is limited by the requirement for multiple daily administrations, whereas doxazosin, a structurally similar α1-adrenergic antagonist, requires only once-daily dosing. In this study, we tested the hypothesis that doxazosin, like prazosin, would decrease alcohol drinking in rats selectively bred for alcohol preference (P line). METHODS: Adult male P rats were given 2 h/d scheduled access to a 2-bottle choice (15% v/v alcohol vs. water) session 5 d/wk (M-F), with food and water available ad libitum 24 h/d. Rats were injected with doxazosin (0 to 10 mg/kg, IP) 40 minutes prior to initiation of the alcohol access session in 3 trials (of 3, 5, and 5 consecutive days) each separated by 5 to 8 weeks. The third trial included 1 day without alcohol access (for locomotor testing), and 1 day of a single hour of alcohol access (for plasma alcohol determination). RESULTS: Doxazosin significantly reduced alcohol intake in all 3 trials. The 5 mg/kg dose consistently reduced alcohol intake, increased water drinking, did not affect locomotor activity, and resulted in lower plasma alcohol concentrations, suggesting that the doxazosin-induced reduction in alcohol drinking was not dependent on a motor impairment or an alteration in alcohol clearance. CONCLUSIONS: Doxazosin decreases voluntary alcohol consumption by male alcohol-preferring (P) rats, supporting a role for the noradrenergic system in alcohol drinking in P rats and suggesting that doxazosin could potentially be an effective once-daily pharmacotherapeutic agent for the treatment of alcohol use disorders.

Effects of prazosin, an α1-adrenergic receptor antagonist, on the seeking and intake of alcohol and sucrose in alcohol-preferring (P) rats.

BACKGROUND: Previous studies show that prazosin, an α(1) -adrenergic receptor antagonist, decreases alcohol drinking in animal models of alcohol use and dependence [Rasmussen et al. (2009) Alcohol Clin Exp Res 3:264-272; Walker et al. (2008) Alcohol 42:91-97] and in alcohol-dependent men [Simpson et al. (2009) Alcohol Clin Exp Res 33:255-263]. This study extended these findings by using a paradigm that allows for separate assessment of prazosin on motivation to seek versus consume alcohol or sucrose in selectively bred rats. METHODS: Alcohol-preferring (P) rats were trained to complete an operant response that resulted in access to either 2% sucrose or 10% alcohol. A 4-week Seeking Test Phase examined responding in single, weekly extinction sessions when no reinforcer could be obtained. A 4-week Drinking Test Phase consisted of rats lever-pressing to "pay" a specified amount up front to gain access to unlimited alcohol (or sucrose) for a 20-minute period. On Seeking and Drinking test days, prazosin (0.0, 0.5, 1.0, and 1.5 mg/kg) was administered intraperitoneally 30 minutes prior to behavioral sessions. RESULTS: Rats were self-administering an average of 0.9 (±0.09) g/kg alcohol on vehicle test day and had pharmacologically relevant blood ethanol concentrations. Prazosin significantly decreased alcohol seeking at all doses tested. The highest dose of prazosin also increased the latency to first response for alcohol and decreased alcohol intake. While sucrose-seeking and intake were similarly affected by prazosin, the high dose of prazosin did not increase response latency. CONCLUSIONS: These findings are consistent with and extend previous research and suggest that prazosin decreases motivation to initiate and engage in alcohol consumption. The specificity of prazosin in attenuating the initiation of alcohol- but not sucrose-seeking suggests that this effect is not because of prazosin-induced motor-impairment or malaise. Together with previous findings, these data suggest that prazosin may be an effective pharmacotherapy, with specific application in people that drink excessively or have a genetic predisposition to alcohol abuse.

The alpha1-adrenergic receptor antagonist, prazosin, reduces alcohol drinking in alcohol-preferring (P) rats.

BACKGROUND: Preliminary evidence suggest that noradrenergic signaling may play a role in mediating alcohol drinking behavior in both humans and rats. Accordingly, we tested the hypothesis that blockade of alpha(1)-adrenergic receptors will suppress alcohol drinking in rats selectively bred for alcohol preference (P line). METHODS: Adult male P rats were given 24-hour access to food and water and scheduled access to a 15% (v/v) alcohol solution for 2 hours daily. Rats were injected IP with the alpha(1)-adrenergic receptor antagonist, prazosin (0, 0.5, 1.0, 1.5, or 2.0 mg/kg body weight), once a day at 15 minutes prior to onset of the daily 2-hour 2-bottle choice, alcohol versus water, access period for 2 consecutive days and then 3 weeks later for 5 consecutive days. RESULTS: Prazosin significantly reduced (p < 0.01) alcohol intake during the initial 2 daily administrations, and this reduction of alcohol intake was maintained for 5 consecutive days by daily prazosin treatment in the subsequent more prolonged trial (p < 0.05). The prazosin-induced reduction of alcohol intake was not dependent upon drug-induced motor impairment since increases in water drinking (p < 0.05) were exhibited during the 2-hour access periods during both 2- and 5-day prazosin treatment. CONCLUSIONS: The results indicate that the noradrenergic system plays a role in mediating alcohol drinking in rats of the P line and suggest that prazosin--a safe, well-characterized, and well-tolerated drug--may be an effective pharmacotherapeutic agent for the treatment of alcohol use disorders.

alpha1-noradrenergic receptor antagonism blocks dependence-induced increases in responding for ethanol.

The purpose of this study was to test the hypothesis that blockade of alpha1-adrenergic receptors may suppress the excessive ethanol consumption associated with acute withdrawal in ethanol-dependent rats. Following the acquisition and stabilization of operant ethanol self-administration in male Wistar rats, dependence was induced in half the animals by subjecting them to a 4-week intermittent vapor exposure period in which animals were exposed to ethanol vapor for 14h/day. Subsequent to dependence induction, the effect of alpha1-noradrenergic receptor antagonist prazosin (0.0, 0.25, 0.5, 1, 1.5, and 2.0mg/kg IP) was tested on operant responding for ethanol in vapor-exposed and control rats during acute withdrawal. In ethanol-dependent animals, prazosin significantly suppressed responding at the 1.5 and 2.0mg/kg doses, whereas only the 2.0mg/kg dose was effective in nondependent animals, identifying an increase in the sensitivity to prazosin in dependent animals. Conversely, at the lowest dose tested (0.25mg/kg), prazosin increased responding in nondependent animals, which is consistent with the effect of anxiolytics on ethanol self-administration in nondependent animals. None of the doses tested reliably affected concurrent water self-administration. These results suggest the involvement of the noradrenergic system in the excessive alcohol drinking seen during acute withdrawal in ethanol-dependent rats.

Acoustic startle amplitude predicts vulnerability to develop post-traumatic stress hyper-responsivity and associated plasma corticosterone changes in rats.

Following exposure to trauma, a vulnerable sub-population of individuals develops post-traumatic stress disorder (PTSD) with characteristic persistent autonomic hyper-responsivity, associated increased startle response, and commonly altered hypothalamo-pituitary-adrenal regulation. A goal of this investigation was to identify a predictive marker for this vulnerability. Previous investigators have developed a model for PTSD in which male mice were exposed to a single brief episode of inescapable footshock followed by 1-min contextual reminders of this trauma at weekly intervals for 6 weeks. Exposure to these reminders induced a progressive and persistent increase in the amplitude of acoustic startle consistent with the persistently increased acoustic startle of individuals exhibiting PTSD. We adapted this model to adult male Wistar rats, with added characterization of initial (pre-trauma) startle response. After one episode of inescapable footshock (10 s, 2 mA) or control treatment followed by six weekly 1-min contextual reminders, acoustic startle was re-tested. Data were analyzed after dividing rats within each treatment into LOW vs MID vs HIGH (33% in each group) pre-treatment startle responders. Rats which exhibited pre-treatment LOW- and MID-range acoustic startle responses did not develop increased acoustic startle responses following subsequent traumatic stress+reminders ([TS+R]) treatment. However, rats which exhibited HIGH pre-treatment startle responses exhibited further significant (p<0.01) [TS+R]-induced persistent enhancement of this already elevated startle response. Furthermore, rats exhibiting HIGH pre-treatment startle responses were also the only subgroup which exhibited increased basal plasma corticosterone levels following [TS+R] treatment. These results suggest that initial pre-stress acoustic startle response can identify subgroups of rats which are predisposed to, or resistant to, developing a PTSD-like syndrome following subsequent trauma.

Alcoholism, craving, and hormones: the role of leptin, ghrelin, prolactin, and the pro-opiomelanocortin system in modulating ethanol intake.

Evidence is growing that appetite regulating peptides such as leptin and ghrelin, but also other hormones including prolactin are altered in alcoholism. The brain pro-opiomelanocortin (POMC) system which has important mediating roles in alcohol intake also has important functions in prolactin regulation and energy homeostasis. Furthermore, it has been demonstrated to be functionally integrated with leptin regulation. The satiety factor leptin seems to be counteracted by the gut-derived peptide ghrelin which increases hunger and food intake. Consequently, the POMC system may have a role in integrating regulation of alcohol effects and these seemingly disparate regulatory systems. The goal of this mini-review is to discuss the results of some recent investigations of the potential interactions of these systems with acute and chronic alcohol responses.

Olanzapine-induced weight gain and increased visceral adiposity is blocked by melatonin replacement therapy in rats.

The atypical antipsychotic drug olanzapine increases body weight and visceral adiposity in schizophrenia. In rats, aging-associated increased body weight and visceral adiposity are reversed by administration of the pineal hormone melatonin. We asked if melatonin similarly would reverse olanzapine-induced increased weight and visceral adiposity in rats. Four groups (n=11/group) of female rats (240-250 g) were treated for 8 weeks with olanzapine, melatonin, olanzapine+melatonin, or vehicle alone in drinking water. Body weight and food and water consumption were determined weekly, locomotor activity at weeks 3 and 6, and nocturnal plasma melatonin concentration at week 7. At week 8, the rats were killed and visceral (perirenal, retroperitoneal, omental, and mesenteric) fat pads dissected and weighed. Olanzapine treatment reduced nocturnal plasma melatonin by 55% (p<0.001), which was restored to control levels by olanzapine+melatonin. Body weight increased 18% in rats treated with olanzapine alone, but only 10% with olanzapine+melatonin, 5% with melatonin alone, and 7% with vehicle control. Body weight and visceral fat pad weight increases in rats treated with olanzapine alone were greater than in each of the other three groups (all p<0.01), which were not significantly different. These results suggest that olanzapine-induced increases in body weight and visceral adiposity may be at least in part secondary to olanzapine-induced reduction of plasma melatonin levels, and that melatonin may be useful for the management of olanzapine-induced weight gain in humans.

Chronic daily ethanol and withdrawal: 6. Effects on rat sympathoadrenal activity during "abstinence".

We have reported that repetitive daily ethanol consumption increased anxiety-like behavior in rats 4 weeks after ethanol consumption had ceased, consistent with the persistently increased anxiety exhibited by abstinent alcoholics. Increased anxiety is associated with sympathoadrenal activation, so we have now also investigated ethanol-induced persistent changes in basal and stress-induced plasma epinephrine (E) and norepinephrine (NE) levels. Male Sprague-Dawley rats received liquid diet containing ethanol versus pair-fed isocaloric control liquid diet for 9 weeks. After 5 weeks' subsequent "abstinence" (i.e., no ethanol in the diet), the control rats exhibited low basal plasma E and NE, which were both increased by 150-300% within 5 min after transfer to a novel cage in a novel room, returning toward basal levels within 15 min. "Abstinent" ethanol-treated rats exhibited elevated basal E levels (195% of controls, P<.05), which were not significantly altered by transfer to novel environment; basal NE levels tended (P<.07) to be elevated and likewise were not altered by novel environment. These results suggest that daily ethanol consumption can induce persistent increases in sympathoadrenal activation during subsequent "abstinence," which are relatively refractory to further stimulation.

Chronic daily ethanol and withdrawal: 4. Long-term changes in plasma testosterone regulation, but no effect on GnRH gene expression or plasma LH concentrations.

Although ethanol has been repeatedly demonstrated to inhibit the hypothalamo-pituitary-testes axis by multiple mechanisms, plasma testosterone levels can be normal in alcoholics who do not exhibit severely compromised liver function and even increased in some abstinent alcoholics, suggesting that adaptive changes to chronic alcohol abuse may alter these regulatory mechanisms. To address this variability, we have investigated the effects of chronic ethanol and withdrawal on rat testosterone regulation using a well-characterized liquid diet model that we have previously demonstrated to (1) provide daily oral ethanol consumption that produces behaviorally relevant plasma ethanol levels during the active (awake) stage of the photoperiod; (2) establish physical dependence on ethanol; and (3) produce not only hypothalamo-pituitary-adrenal axis, but also behavioral (anxiety-like behavior, response to novelty, sucrose preference) changes consistent with those of actively drinking and subsequently abstinent alcoholics. The results demonstrate that chronic daily episodes of ethanol consumption and withdrawal by male Sprague-Dawley rats decreased (p < 0.01) plasma testosterone levels late in the afternoon (by 70% relative to ad libitum-fed controls and 63% relative to pair-fed controls), but not in the morning. During gradual cessation of daily ethanol consumption, morning plasma testosterone levels increased, and this 90-115% (p < 0.05) increase was maintained for 3 d after complete cessation of ethanol consumption. Three weeks after cessation of ethanol consumption, plasma testosterone levels were again increased by approx 100% (p < 0.01). Plasma luteinizing hormone (LH) concentrations and anterior hypothalamus/preoptic area gonadotropin-releasing hormone (GnRH) mRNA levels were not altered at any of these time points. Thus, chronic daily ethanol consumption and daily withdrawal induced changes in circulating testosterone regulation that (a) were time of day dependent and (b) included adaptive changes persisting long after consumption of ethanol ceased. Accordingly, resolution of changes in testosterone regulation and their potential roles in alcohol abuse and relapse will require evaluating changes throughout the circadian cycle during, shortly after, and long after active alcohol abuse.

Melatonin effect on rat body weight regulation in response to high-fat diet at middle age.

We previously demonstrated that daily melatonin administration to middle-aged rats to restore youthful plasma melatonin levels also decreased body weight, visceral fat, plasma leptin, and plasma insulin to more youthful levels, without detectable changes in consumption of chow diet. We now evaluate: (a) whether melatonin alters consumption of a more precisely quantifiable liquid diet similar in high-fat content to the typical American diet; (b) differences between melatonin-induced endocrine responses in the fasted vs fed state; and (c) time course of these responses. Ten-month-old male Sprague- Dawley rats received liquid diet containing either 0.2 micro g/mL melatonin (MELATONIN) or vehicle (CONTROL) (n = 14/treatment); the diet was available throughout each night, but was removed for the final 10 h of each daytime. MELATONIN rats gained 4% body weight during the first 2 wk and then stabilized, whereas CONTROL rats continued to gain for an additional week, achieving 8% gain (p < 0.05 vs MELATONIN). During the first 3 wk, afternoon tail-blood leptin, but not insulin, levels decreased in melatonin-treated rats (p < 0.05 vs CONTROL). After 8 wk, half of the rats were killed at the midpoint of the dark period (NIGHT; fed) and half at the end of the light period (DAYTIME; fasted). NIGHT but not DAYTIME plasma leptin levels were decreased in MELATONIN rats, whereas DAYTIME but not NIGHT plasma insulin levels were decreased (p < 0.05 vs CONTROL). Melatonin treatment did not alter cumulative food consumption. Thus, melatonin decreased weight gain in response to high-fat diet, decreased plasma leptin levels within 3 wk-before decreasing plasma insulin-and exerted these metabolic effects independent of total food consumption.

Melatonin effects on metabolism independent of gonad function.

We previously demonstrated that daily melatonin administration to middle-aged rats, restoring nocturnal plasma melatonin to young adult levels, decreased body weight and suppressed visceral fat and plasma leptin. In some species, metabolic and some neuronal responses to melatonin are mediated or dependent at least in part on gonadal steroid levels. Thus, melatonin-induced changes in gonadal steroid secretion may have mediated the aging-dependent melatonin-induced metabolic responses in our previous studies. To address this issue, melatonin (0.4 micro g/mL) or vehicle (0.01% ethanol) was administered for 10 wk in the drinking water of both castrate and sham-operated Sprague-Dawley male rats, starting 1 mo after surgery at 9 mo of age. Melatonin treatment decreased (p < 0.05) body weight in sham-operated rats by 7 +/- 2% relative to control (n = 7/treatment), comparable to our previous results; melatonin likewise decreased (p < 0.05) body weight in castrate rats by 6 +/- 2% relative to control (n = 7/treatment). Melatonin treatment also decreased both intraabdominal fat and plasma leptin levels in both intact and castrate rats, with no significant differences of percentage suppression in the intact versus castrate rats. These results demonstrate that suppression of body weight, visceral adiposity, and plasma leptin levels by daily melatonin administration to middle-aged rats was independent of gonadal function.

Suppression of hypothalamic pro-opiomelanocortin (POMC) gene expression by daily melatonin supplementation in aging rats.

Both plasma melatonin levels and hypothalamic arcuate nucleus pro-opiomelanocortin (POMC) (biosynthetic precursor to the endogenous opioid ss-endorphin and other opiomelanocortins) mRNA content decrease with aging. To test whether the decline in melatonin is responsible for the decline in POMC mRNA, we investigated the effects of daily melatonin treatment on hypothalamic POMC mRNA content in middle-aged and older Sprague-Dawley rats. Daily nocturnal melatonin treatment (50 microg kg bw(-1) night(-1), in the night-time drinking water) for 7 months, starting at 13 months of age, did not significantly alter female arcuate nucleus POMC mRNA content determined at the end of the light period (i.e., before nightly melatonin administration), but suppressed (24%, P < 0.05) POMC mRNA content at the end of the dark period (i.e., following melatonin administration). Likewise, nocturnal administration of 50 or 500 microg melatonin kg bw(-1) night(-1) to male rats for 7 months suppressed (31 or 28%, respectively; P < 0.05) POMC mRNA content at the middle of the dark period at 20 months of age. Finally, 10 wk administration of 30 microg melatonin kg bw(-1) day(-1) suppressed (31%, P < 0.01) POMC mRNA content in middle-aged male rats killed at the end of the dark period. Melatonin treatments did not significantly alter estradiol or testosterone levels. Thus, moderate-dosage nocturnal melatonin supplementation suppressed nocturnal hypothalamic POMC gene expression in both middle-aged males and females, suggesting that melatonin supplementation during aging decreases, rather than increases, forebrain opiomelanocortinergic activity. These POMC responses were apparently not dependent on gonadal steroid responses and did not become refractory to melatonin treatment maintained until old age.