Additive Reduction of Alcohol Drinking by 5-HT1A Antagonist WAY 100635 and Serotonin Uptake Blocker Fluoxetine in Alcohol-Preferring P Rats

We found previously that alcohol-preferring (P) rats have fewer serotonin (5-HT) neurons and fibers in key brain regions than alcoholnonpreferring (NP) rats. Because 5-HT uptake blockers increase synaptic 5-HT content and 5-HT_1A receptor antagonists increase 5-HT release by disinhibiting 5-HT autoinnervation, in the present study, our intent was to determine whether increased synaptic 5-HT content and/or 5-HT release in P rats would effectively reduce alcohol consumption. In experiment 1, the 5-HT antagonist WAY 100635 (WAY) was tested on adult female P rats maintained on 24-hr free-choice access to ethanol (10% v/v) and water. Twice daily doses of WAY (0.05, 0.1, 0.5, and 1.0 mg/kg, subcutaneously) were administered to each rat in a counterbalanced order. Baseline ethanol intake, derived from the mean ethanol intakes of the three previous non-drug days, was approximately 8 g/kg/day. Results indicated that 0.05,0.1, and 0.5 mg/kg doses of WAY reduced 24-hr ethanol drinking by 25-30% ( p < 0.01) without affecting 24-hr water intake or body weight In the second experiment, the effects of WAY (0.5 mg/kg), fluoxetine (1.0 mg/kg), or a combination of both were tested in another group of female P rats. WAY and fluoxetine, each alone, reduced ethanol drinking by around 20% and, when combined, decreased ethanol intake by 50%, whereas the body weight and the total fluid intake were not significantly affected. Taken together, these results indicate that both fluoxetine and WAY preferentially reduce ethanol drinking in the P line of rats and, when administered together, reduce ethanol intake in an additive manner. It is proposed that coadministration of these two compounds with distinct mechanisms of action may be a new strategy for reducing alcohol intake.     

Effect of ethanol on hypothalamic opioid peptides, enkephalin, and dynorphin: relationship with circulating triglycerides

BACKGROUND: Recent evidence has demonstrated that ethanol intake can stimulate the expression and production of the feeding-stimulatory peptide, galanin (GAL), in the hypothalamic paraventricular nucleus (PVN), and that PVN injection of this peptide, in turn, can increase the consumption of ethanol. To test the hypothesis that other feeding-related systems are involved in ethanol intake, this study examined the effect of ethanol on the hypothalamic opioid peptides, enkephalin (ENK), and dynorphin (DYN). METHOD: Adult, male Sprague-Dawley rats were trained to voluntarily drink increasing concentrations of ethanol, up to 9% v/v, on a 12-hour access schedule or were given a single injection of ethanol (10% v/v) versus saline vehicle. The effect of ethanol on GAL, ENK, and DYN mRNA was measured using real-time quantitative polymerase chain reaction and radiolabeled in situ hybridization, while radioimmunoassay was used to measure peptide levels. In addition to blood alcohol, circulating levels of triglycerides (TG), leptin, and insulin were also measured. RESULTS: The data demonstrated that: (1) rats voluntarily drinking 9% v/v ethanol (approximately 2.0 g/kg/d) show a significant increase in GAL, ENK, and DYN mRNA in the PVN compared with water-drinking rats; (2) voluntary consumption of ethanol also increases peptide levels of ENK and DYN in the PVN; (3) acute injection of 10% ethanol (1.0 g/kg of 10% v/v) similarly increases the expression of GAL, ENK, and DYN in the PVN; and (4) ethanol consumption and injection, while having little effect on leptin and insulin, consistently increase circulating levels of TG as well as alcohol, both of which are strongly, positively correlated with peptide expression in the PVN. CONCLUSIONS: These findings, together with published studies, suggest a possible role for hypothalamic opioid peptides in the drinking of ethanol. Based on evidence that dietary fat and lipid injections stimulate the PVN peptides and injection of the opiates and GAL increase ethanol intake, it is proposed that both TG and alcohol in the circulation, which are elevated by the ingestion or injection of ethanol, are involved in stimulating these peptides in the PVN, which in turn promote further consumption of ethanol.     

The effects of neuropeptide S on ethanol drinking and other related behaviors in alcohol-preferring and -nonpreferring rats

Background.  Neuropeptide S (NPS) is a 20-amino-acid peptide, identified in the brain and periphery, that is reported to regulate arousal, anxiety, and feeding behavior. Studies were conducted to determine whether this peptide would alter ethanol intake, sucrose intake, anxiety, and general motor activity in alcohol-preferring (P) and -nonpreferring (NP) rats.
Methods.  Experiment 1 : P and NP rats were given 8 weeks of continuous access to ethanol (15% w/v) and water. All rats were implanted with a cannula aimed at either the left or right lateral ventricle and 1 week later were infused with NPS (0.075, 0.3, 1.2 nmol) or artificial cerebrospinal fluid (aCSF) and tested for ethanol, food, and water intake. Experiment 2 : The same doses of NPS were administered to a group of P rats and intake of 2.5% (w/v) sucrose was measured. Experiment 3 : Infusions of NPS (1.2 nmol) or aCSF were administered to P rats prior to a 5-minute test on an elevated plus maze. Experiment 4 : Ethanol naive P and NP rats were infused with NPS (0.075, 0.15, 0.3, 0.6, and 1.2 nmol) or aCSF prior to a 20-minute test in activity monitors.
Results.  NPS reduced ethanol intake in P, but not in NP rats. It did not influence sucrose solution intake in P rats. However, an increase in food intake was seen in both rat lines following lower doses of the peptide. NPS did neither alter anxiety-like behavior in the elevated plus maze test nor was there an effect on general motor activity; however, there was an increase in the amount of time spent in the center of the activity monitors following infusions of 0.6 nmol of NPS in P, but not in NP rats, indicating anxioltyic actions of the peptide.
Conclusions.  These data suggest a role for NPS in the modulation of ethanol drinking and possibly anxiety-like behavior in rats selectively bred for high alcohol drinking.     

Role of cholecystokinin in the control of food intake

CCK appears to regulate short-term control of food intake by acting as a satiety signal. Larger doses of CCK may decrease food intake by aversive actions (malaise, nausea, cramps), presumably by effects on gastrointestinal motility. In rats and most likely humans CCK is released from the upper intestine after a mixed meal and appears to activate afferent vagal fibers by causing pyloric contraction with resultant gastric distention or directly binding to the gastric afferent vagus which courses to the nucleus solitarius with further projections to the paraventricular nucleus and ultimately the ventromedial hypothalamus. Peripherally released CCK may also bind to CNS receptors in the area postrema overlying the nucleus solitarius. Central nervous system CCK released from the paraventricular nucleus may also exert a satiety effect. The satiety effect of CCK appears to be a physiologic action of the peptide since antibodies to CCK and CCK receptor antagonists can increase food intake. CCK is probably just one of several satiety signals but can cause a profound decrease in food intake when administered exogenously in pharmacologic doses. Administration of exogenous CCK, as well as endogenous CCK released by oral protease inhibitors, can decrease food intake in humans. Studies designed to examine the effect of chronic administration of CCK on food intake will be necessary to determine if the peptide has a role in the management of obesity and bulimia.     

Central leptin and cholecystokinin in gastroprotection against ethanol-induced damage

BACKGROUND: Leptin, a product of the ob gene controlling food intake, has recently been detected in the stomach and shown to be released by cholecystokinin (CCK) and to induce gastroprotection against various noxious agents, but it is not known whether centrally applied leptin influences gastric secretion and mucosal integrity. AIMS: In this study we compared the effects of leptin and CCK-8 applied intracerebroventricularly (i.c.v.) on gastric secretion and gastric mucosal lesions induced by topical application of 75% ethanol. METHODS: Several major series of Wistar rats were used in this study. The effects of leptin or CCK applied i.c.v. on gastric secretion were examined using conscious rats with gastric fistulas. For the studies on gastroprotection the following series of rats were used to determine the effects of: (A) leptin and CCK applied centrally on this protection and the blockade of CCK(A) with loxiglumide (30 mg/kg i.p.) and CCK(B) receptors with RPR 102681 (30 mg/kg i.p.); (B) cutting of vagal nerves; (C) inactivation of sensory nerves by capsaicin (125 mg/kg s.c.); (D) inhibition of calcitonin gene-related peptide (CGRP) receptors with CGRP(8-37) (100 microg/kg i.p.), and (E) suppression of nitric oxide synthase (NOS) with N(G)-nitro-L-arginine methyl ester (L-NAME) (5 mg/kg i.v. ) on ethanol-induced gastric lesions in rats with or without the i.c.v. pretreatment with leptin or CCK-8. Rats were anesthetized 1 h after ethanol administration to measure the gastric blood flow (GBF) and then to determine the area of gastric lesions by planimetry. Blood was withdrawn for the measurement of plasma leptin and gastrin levels by radioimmunoassay and gastric biopsy samples were collected for the determination of cNOS and iNOS mRNA by RT-PCR. RESULTS: Leptin and CCK-8 (0.01-5 microg/kg i.c.v.) dose dependently attenuated gastric lesions induced by 75% ethanol; the doses reducing these lesions by 50% (ED(50)) were 0.8 and 1.2 microg/kg, respectively. The protective effects of leptin and CCK-8 applied i.c. v. were accompanied by a significant rise in plasma leptin level and an increase in GBF. Blockade of CCK(A) receptors with loxiglumide abolished the protective and hyperemic effects of CCK but not those of leptin, while RPR 10268, a specific antagonist of CCK(B) receptors, counteracted leptin-induced protection and the rise in the GBF but failed to influence those afforded by CCK-8. For comparison, pretreatment with peripheral CCK-8 or leptin (10 microg/kg i.p.) causing a similar rise in the plasma leptin level also significantly reduced gastric lesions induced by 75% ethanol. The protective and hyperemic effects of centrally administered leptin were abolished by vagotomy, producing a fall in plasma leptin levels, and significantly attenuated by sensory denervation with capsaicin, by pretreatment with the CGRP antagonist, CGRP(8-37), or with L-NAME. A strong signal for iNOS mRNA was recorded in the gastric mucosa of leptin- and CCK-8-treated animals, whereas cNOS mRNA was unaffected. CONCLUSIONS: (1) Central leptin exerts a potent gastroprotective action at a dose that has no influence on gastric secretion; (2) this protection depends upon CCK(B) receptors, vagal activity and sensory nerves, and involves hyperemia probably mediated by NO, and (3) leptin mimics the gastroprotective effect of CCK and may be implicated in the protective and hyperemic actions of this peptide on the rat stomach.     

Hypericum perforatum CO2 extract and opioid receptor antagonists act synergistically to reduce ethanol intake in alcohol-preferring rats

BACKGROUND: Hypericum perforatum extracts attenuate ethanol intake in alcohol-preferring rats. The opioid receptor antagonists, naloxone and naltrexone, reduce ethanol intake in rats and humans. The combination of different agents that reduce ethanol intake has been proposed as an approach to the pharmacotherapy of alcoholism. This study evaluated the effect on ethanol intake of the combined administration of a CO2 H. perforatum extract and naloxone or naltrexone in genetically selected Marchigian Sardinian alcohol-preferring rats. METHODS: Ten percent (v/v) ethanol intake was offered 2 hr per day at the beginning of the dark phase of the reverse light-dark cycle. H. perforatum CO2 extract was given intragastrically, 1 hr before access to ethanol. Naloxone or naltrexone was given by intraperitoneal injection 10 min before the extract. RESULTS: H. perforatum CO2 extract reduced ethanol intake at 31 or 125 mg/kg, but not 7 mg/kg. These doses neither modified food or water intake during access to ethanol, nor reduce 0.2% saccharin intake. Naloxone reduced ethanol and food intake at 3 or 5 mg/kg, but not 1 mg/kg. When naloxone 1 mg/kg was combined with the three doses of H. perforatum CO2 extract, the attenuation of ethanol intake was more pronounced than that observed after the administration of the extract alone. Alcohol intake was also significantly reduced by 7 mg/kg of H. perforatum CO2 extract combined with naloxone 1 mg/kg. The combined treatments never modified the rat's locomotor activity nor the simultaneous intake of food, water or 0.2% saccharin. Naltrexone reduced ethanol intake at 1 and 3 mg/kg, but not at 0.5 mg/kg. When naltrexone 0.5 mg/kg was combined with H. perforatum CO2 extract 7 mg/kg, ethanol intake was markedly reduced. CONCLUSIONS: These findings provide evidence that H. perforatum CO2 extract and opiate receptor antagonists act synergistically to induce a pronounced and selective reduction of voluntary ethanol consumption in alcohol-preferring rats.     

Irreversible Exocrine Pancreatic Insufficiency in Alcoholic Rats Without Chronic Pancreatitis After Alcohol Withdrawal

Background: Long‐term alcohol consumption alone did not cause chronic pancreatitis (CP) but impaired exocrine pancreatic function. This study is to explore the reversibility of exocrine pancreatic insufficiency in the abstinent rats and its mechanism. Methods: Forty‐eight healthy male Wistar rats were divided randomly into 4 groups: 6‐month control, 6‐month ethanol, 9‐month control, and 9‐month ethanol + withdrawal. Morphological changes of pancreatic acinar cells were observed. Pancreatic amylase and lipase were measured using an automatic biochemical analyzer. Free fatty acid (FFA) in rat intestinal chyme was measured. Cholecystokinin (CCK) levels were determined by radioimmunoassay. The expression of CCK‐A receptors was quantitatively analyzed by Western blot. Results: Alcohol‐induced ultramicrostructure changes of pancreatic acinar cells, including lipid droplets, myelinoid inclusion bodies, dilated rough endoplasmic reticulums, and diminished zymogen granules, were not attenuated after alcohol abstinence. The outputs of amylase and lipase, FFA content in intestinal chyme, and the intestinal and the pancreatic CCK levels in rats were reduced after chronic alcohol intake and were still lower than the control after cessation of alcohol use. Chronic ethanol intake or abstinence did not induce any change in the expression of CCK‐A receptors. Conclusions: Exocrine pancreatic insufficiency was irreversible in alcoholic rats without CP after alcohol withdrawal. It may be attributed to reduced pancreatic CCK, long‐standing fatty infiltration, ultramicrostructure injuries in pancreatic acinar cells, and aging.     

Time-dependent effects of maximal doses of cerulein on the secretory response of the rat pancreas

The present study was performed to determine the time-dependent effects of chronic administration of maximal doses of cerulein on the secretory function of the rat pancreas. Four groups of rats, one treated with 0.9% NaCl (control) and the others with cerulein (2, 5, and 10 μg/kg twice a day ip, respectively) were used. After a treatment period of 15, 30, or 60 d, eight rats were taken from each group, the pancreatic bile duct was surgically cannulated, and the pancreatic juice was collected after CCK or secretin stimulation. Then the rats were killed and the pancreas was removed and weighed. Administration of 2 μg/kg of cerulein for 15 d induced a significant increase (p<0.05) in the volume of pancreatic juice in response to hormonal stimulation. The enzyme and bicarbonate outputs increased only in response to CCK, not to secretin. The above-mentioned increases are significant only when related to body weight, not to pancreatic weight. There was good correlation between pancreatic weight and the increase in the secretory parameters. The secretion pattern was not significantly modified by higher doses of cerulein nor by longer duration of treatment. We conclude that 1) increase in pancreatic secretory function induced by 2 μg/kg of cerulein is not enhanced by higher doses of peptide and 2) the functional capacity of the acinar cells is not affected by the length of treatment. These data are of interest as experimental backing to therapeutic use of CCK-like peptides in pancreatic exocrine insufficiency.     

Quantitative comparison of the effects of cholecystokinin, secretin, and pentagastrin on gastrointestinal myoelectric activity in the conscious fasted dog.

The effects on gastrointestinal myoelectric activity of infused pentagastrin, cholecystokinin (CCK), and secretin at physiological doses were studied in live dogs with implanted serosal electrodes during 56 six-hour studies. Pentagastrin dose-dependently increased gastric and duodenal slow-wave frequencies; secretin and CCK did not. Pentagastrin and CCK diminished the incidence of fasting migrating myoelectric complexes (MMCs), but MMCs were abolished only in the proximal small intestine. Pentagastrin infusion was not reflected in an increased number of spikes, whereas CCK induced a dose-dependent increase in jejunal spike activity. Secretin dose-dependently decreased duodenal and jejunal spike incidence without a marked effect on MMC incidence. Analysis of patterns of spike activity showed significant dose-dependent changes with all three peptides. The different effects of pentagastrin and CCK on spike activity in these studies may have been a consequence of pentagastrin-stimulated gastric acid secretion. None of the three peptides produced a pattern of myoelectric activity which closely resembled that seen on feeding; since, unlike food, all three peptides had little or no effect on the distal small intestine, it seems unlikely that combinations of these peptides are responsible for the change induced by food. The failure of these peptides to abolish fasting patterns in the distal intestine suggests a possible mechanism for some types of post-vagotomy dysfunction.     

Neuropeptide Y reduces oral ethanol intake in alcohol-preferring (P) rats following a period of imposed ethanol abstinence

BACKGROUND: Intracerebroventricular infusion of NPY has been shown to reduce ethanol intake in alcohol-preferring (P) rats in a limited access procedure. The purpose of the present investigation was to extend this finding to a two-bottle free-choice continuous access procedure in groups of rats that either did or did not undergo a period of imposed ethanol abstinence and ethanol reinstatement. METHODS: In experiment 1, female P rats were given 6 weeks of continuous access to ethanol (8% w/v) and water. Ethanol was removed for a period of 2 weeks during which the rats were surgically implanted with a cannula into the lateral ventricle. Following the ethanol abstinence period and immediately before ethanol reinstatement, rats received a single infusion of either artificial cerebrospinal fluid or NPY (10 microg). Ethanol and water intake was measured at both 4 hr and 24 hr after infusion, and 24-hr intake measures were taken daily for 13 postinfusion days. Experiment 2 was run in parallel with experiment 1, with the exception that rats did not undergo a period of imposed ethanol abstinence. Also, food intake was measured 4 and 24 hr after infusion. RESULTS: Following 2 weeks of imposed ethanol abstinence (experiment 1), NPY suppressed ethanol intake through postinfusion day 2. After uninterrupted continuous access to ethanol (experiment 2), NPY suppressed ethanol intake to a lesser extent and this effect lasted only 24 hr. NPY increased food intake at the 4-hr but not the 24-hr measure. CONCLUSIONS: Previous findings that central administration of NPY suppresses ethanol intake in P rats are extended by this study to a continuous access procedure, and the effect is amplified following a period of imposed ethanol abstinence. This effect of NPY compares favorably to results obtained with other treatments tested in similar animal models and provides support for a role of NPY in an allostasis model of addiction.     

Suppression of Alcohol Intake after Administration of the Chinese Herbal Medicine, NPI-028, and Its Derivatives

The Chinese herbal medicine, NPI-028, has been used for centuries in China to counteract alcohol intoxication. The present study used a number of different experimental conditions to determine whether NPI-028 and its derivatives might selectively influence alcohol intake in rodents that naturally exhibit high alcohol intakes. It was determined that intraperitoneal (IP) injections of NPI-028 (0.5, 0.75, and 1.0 g/kg) suppressed alcohol intake by up to 30% in both alcohol-preferring P and Fawn-Hooded (FH) rats during a continuous access schedule. These injections did not significantly affect food or water intakes, nor did the highest dose of NPI-028 (1 g/kg) alter blood ethanol levels after an IP injection of 2.5 g/kg of ethanol. In P rats, it was found that NPI-028 was orally active with the dose of 1.5 g/kg having a greater effect on ethanol intake than the 1.0 g/kg dose; once again, food and water intakes were not significantly altered. In FH rats maintained on a limited access schedule (1 hr/day), alcohol intake was completely abolished by 1.5 g/kg of NPI-028. Chronic IP administration of NPI-028 (0.75 g/kg) for four consecutive days in FH rats maintained on a continuous access schedule did not lead to any diminution of its alcohol-suppressant effects. Thus, NPI-028 has significant effects on alcohol intake without much effect on water and food intake, and tolerance does not readily develop to these effects. The IP administration of a partially purified extract (NPI-031) of NPI-028, obtained by countercurrent chromatography, also dose-dependently suppressed ethanol intake in FH rats, but the highest dose (200 mg/kg) also significantly decreased food intake. Finally, the IP administration of puerarin (NPI-31G), an isoflavone isolated from NPI-031 by countercurrent chromatography, significantly reduced ethanol intake in FH rats without affecting food or water intake. Therefore, NPI-028 and one of its pure components, NPI-031G, selectively reduced ethanol intake in alcohol-preferring rats.     

Attenuation of Alcohol Preference in Alcohol-Preferring Rats by a Novel TRH Analogue, TA-0910

Experiments were performed to characterize the acute effect different doses of a novel thyrotropin-releasing hormone (TRH) analogue (TA-0910) on ethanol intake in rats. Selectively bred alcohol-preferring (P) rats received a single intraperitoneal injection of normal saline or 0.083, 0.25 and 0.75 mg/kg of TA-0910 at 9:30 AM, and their consumption of ethanol, water, and food was measured for hr. TA-0910 dose-dependently attenuated ethanol intake and commensurately increased water consumption. Only the highest dose TA-0910 increased the total caloric intake. TA-0910 did not affect the pharmacokinetics of ethanol. These findings indicate involvement of TRH systems in ethanol preference and suggest that centrally acting TRH analogues may be therapeutic in the treatment of alcoholism.     

Intermittent access to 20% ethanol induces high ethanol consumption in Long-Evans and Wistar rats

Background: There has been some difficulty getting standard laboratory rats to voluntarily consume large amounts of ethanol without the use of initiation procedures. It has previously been shown that standard laboratory rats will voluntarily consume high levels of ethanol if given intermittent‐access to 20% ethanol in a 2‐bottle‐choice setting [ Wise, Psychopharmacologia 29 (1973), 203 ]. In this study, we have further characterized this drinking model. Methods: Ethanol‐naïve Long–Evans rats were given intermittent‐access to 20% ethanol (three 24‐hour sessions per week). No sucrose fading was needed and water was always available ad libitum. Ethanol consumption, preference, and long‐term drinking behaviors were investigated. Furthermore, to pharmacologically validate the intermittent‐access 20% ethanol drinking paradigm, the efficacy of acamprosate and naltrexone in decreasing ethanol consumption were compared with those of groups given continuous‐access to 10 or 20% ethanol, respectively. Additionally, ethanol consumption was investigated in Wistar and out‐bred alcohol preferring (P) rats following intermittent‐access to 20% ethanol. Results: The intermittent‐access 20% ethanol 2‐bottle‐choice drinking paradigm led standard laboratory rats to escalate their ethanol intake over the first 5 to 6 drinking sessions, reaching stable baseline consumption of high amounts of ethanol (Long–Evans: 5.1 ± 0.6; Wistar: 5.8 ± 0.8 g/kg/24 h, respectively). Furthermore, the cycles of excessive drinking and abstinence led to an increase in ethanol preference and increased efficacy of both acamprosate and naltrexone in Long–Evans rats. P‐rats initiate drinking at a higher level than both Long–Evans and Wistar rats using the intermittent‐access 20% ethanol paradigm and showed a trend toward a further escalation in ethanol intake over time (mean ethanol intake: 6.3 ± 0.8 g/kg/24 h). Conclusion: Standard laboratory rats will voluntarily consume ethanol using the intermittent‐access 20% ethanol drinking paradigm without the use of any initiation procedures. This model promises to be a valuable tool in the alcohol research field.     

Bombesin stimulates prolactin and growth hormone release by pituitary cells in culture

Bombesin (BBS) has been previously shown to stimulate the secretion of PRL and GH in steroid-primed rats. To determine whether these effects were mediated by the central nervous system or were due to direct action on the pituitary gland, we studied the interaction of BBS with GH4C1 cells, a clonal strain of rat pituitary cells which synthesizes and secretes PRL and GH. The addition of 100 nM BBS to GH4C1 cells for 60 min increased PRL release to 140 +/- 3% of the control value (mean +/- SE) and GH release to 133 +/- 5% of the control value. Stimulation of hormone secretion was observed within 15 min of treatment with 100 nM BBS and continued for at least 2 h. Half-maximal stimulation of PRL release occurred with 0.5 nM BBS, and a maximal effect was observed with 10 nM peptide. The BBS analogs ranatensin, litorin, and [Tyr4]BBS, each at a concentration of 100 nM, caused the same stimulation of PRL release as maximal concentrations of BBS itself. BBS stimulated hormone release selectively in two of five different clonal pituitary cell strains examined. Pretreatment of GH4C1 cells with 1 nM estradiol and/or 100 nM insulin resulted in more powerful stimulation of PRL release by both TRH and BBS. When epidermal growth factor and vasoactive intestinal peptide were added simultaneously with BBS, PRL release was greater than in the presence of either peptide alone. In contrast, the stimulatory effects of TRH and BBS were not additive. Somatostatin inhibited both basal and stimulated PRL release. Thus, low concentrations of BBS can directly stimulate PRL and GH release by a clonal pituitary cell strain in culture. These results suggest that BBS may stimulate PRL and GH secretion in vivo by direct action on the pituitary gland.     

Effects of concurrent access to multiple ethanol concentrations and repeated deprivations on alcohol intake of alcohol-preferring rats

BACKGROUND: The alcohol deprivation effect (ADE) is a temporary increase in the voluntary intake of ethanol solutions following a period of alcohol deprivation. Multiple deprivations can prolong the expression of an ADE. This study examined the effects of initial deprivation length, concurrent exposure to multiple ethanol concentrations, and number of deprivation exposures on the magnitude and duration of the ADE in alcohol-preferring (P) rats. METHODS: Adult female P rats received 24-hr free-choice access to 10, 20, and 30% ethanol and water for 6 weeks. Rats were then randomly assigned to three groups; one group served as a nondeprived control, whereas the other two groups were initially deprived of ethanol for 2 or 8 weeks. The ethanol solutions were restored to both deprived groups for 2 weeks before the groups were deprived of ethanol for another 2 weeks. This cycle was repeated three times for a total of four deprivations. RESULTS: After the initial ethanol deprivation period, both deprived groups displayed a similar 2-fold increased ethanol intake (g/Kg/day) during the initial 24-hr period when ethanol was restored. Both deprived groups showed greater than 2-fold increases in intake of the 20 and 30% ethanol solutions after re-exposure. Ethanol consumption returned to baseline levels within 2 weeks, before the subsequent deprivation period. Multiple deprivations increased the magnitude of the ADE over that observed in the first deprivation during the initial 24-hr period of re-exposure and prolonged the duration of the ADE. In addition, repeated deprivations increased ethanol intake in the first 2-hr period of re-exposure and produced blood ethanol levels in excess of 150 mg/100 ml. CONCLUSIONS: Alterations in the reinforcing and/or aversive effects of alcohol occurred after a single prolonged deprivation and were enhanced with repeated deprivations.     

Effects of MDL 72222, a serotonin3 antagonist, on operant responding for ethanol by alcohol-preferring P rats

BACKGROUND: Previous studies indicated that, under continuous access conditions, the 5-HT3 antagonist MDL 72222 (MDL) effectively reduced ethanol drinking of alcohol-preferring P rats. However, MDL was without effect when similar doses were tested under scheduled access conditions, unless the ethanol access period was randomly presented. This study examined the effects of MDL on operant responding for ethanol and water by adult male alcohol-preferring P rats. METHODS: During the dark cycle, subjects in the first experiment were trained to respond concurrently for 15% ethanol and water on a fixed-ratio 5 (FR-5) and FR-1 schedule of reinforcement, respectively. Approximately 30 min before the 4-hr operant session, rats were injected subcutaneously (sc) with saline or MDL (1, 3, or 5 mg/kg). A second experiment tested the effects of 1 mg/kg MDL on operant responding for 15% ethanol in 1-hr sessions when operant access was given at a fixed time each day (fixed scheduled access, FSA group) or at variable time periods throughout the dark cycle (variable scheduled access, VSA group). RESULTS: In the first experiment, only the 5 mg/kg dose of MDL decreased responding for ethanol (approximately 20%) during the first 30 min of the 4-hr session. This dose also reduced total 4-hr responding for ethanol and water. In the second experiment, the 1 mg/kg dose of MDL had no effect on operant responding by the FSA group, but significantly reduced ethanol responding by the VSA group. CONCLUSIONS: These results suggest that 5-HT3 receptors may be involved in mediating the reinforcing effects of ethanol, and that temporal-environmental cues associated with the presentation of ethanol may be one factor involved in reducing the effectiveness of 5-HT3 antagonists to attenuate ethanol intake.     

CCK-releasing activity of rat intestinal secretion: effect of atropine and comparison with monitor peptide

A bioassay for studying the cholecystokinin (CCK)-releasing activity of intraluminal protease-sensitive bioactive peptides was developed. In conscious rats, bile and pancreatic juice were chronically diverted from the proximal intestine to the ileum to cause chronic stimulation of CCK release and pancreatic protein secretion. CCK-releasing activity of test substances was assayed during transient inhibition of CCK release by intraduodenal sodium taurocholate (78 mumols/h). Intestinal secretion as a source of the putative trypsin-sensitive intestinal CCK-releasing peptide was obtained by rapid intestinal perfusion of isolated Thiry-Vella fistulae of jejunum in conscious rats, collected with or without atropine pretreatment. Partially purified rat pancreatic secretory trypsin inhibitor (PSTI, or "monitor peptide") was compared with ovomucoid trypsin inhibitor (OMTI) and with concentrated jejunal secretions for CCK-releasing activity and trypsin inhibitor activity. Concentrated, heat-treated jejunal secretions were the strongest stimulants of CCK release and pancreatic protein secretion in this model. OMTI had no CCK-releasing activity in this model, whereas a larger amount (approximately 5x, based on trypsin inhibitor activity) of PSTI weakly but significantly stimulated CCK release. CCK-releasing activity manifested by pancreatic protein secretion was equivalent in intestinal washes from atropine-treated and control Thiry-Vella fistula donor rats. Concentrated jejunal secretions had no trypsin inhibitory activity, indicating that the putative intestinal CCK-releasing peptide and "monitor peptide" are different substances.     

Cholecystokinin and Bombesin Inhibit Ethanol and Food Intake in Rats Selectively Bred for Ethanol Sensitivity

Cholecystokinin octapeptide (CCK-8) and bombesin tetradecapeptide (BBS-14) are brain-gut neuropeptides shown to inhibit intake and choice of alcohol solutions and foods in a variety of species. Recently, Draski and colleagues selectively bred strains descended from N/Nih outbred Norway rats that differ in sleep time after injection of ethanol. The intake of 5% w/v ethanol, food, and water was measured in these rats with high, low, and control alcohol sensitivity (HAS, US, and CAS), after intraperitoneal injection of randomized sequences of doses of CCK-8 or BBS-14 (0–8 μg/kg). During baseline adaptation to water deprivation-induced consumption of alcohol, LAS rats consumed reliably more ethanol than HAS or CAS rats. Injection of CCK-8 or BBS-14 significantly and equivalently suppressed intake of ethanol and food at 30 min after presentation in each group of rats. Water intake and food intake at 30–60 min following alcohol access was not affected by prior injection of either neuropeptide. Large differences in alcohol neurosensitivity (HAS > CAS > LAS) were observed in these rats' resting behavior for 1 hr after intraperitoneal injection of 1 g/kg of ethanol. These selectively bred alcohol neurosensitivity differences cannot be explained by corresponding differences in sensitivity to the inhibitory behavioral effects of CCK-8 or BBS-14. However, differences in alcohol intake and resting behavior do correspond to artificially selected sensitivities to ethanol's hypnotic effect.     

Time course of elevated ethanol intake in adolescent relative to adult rats under continuous, voluntary-access conditions

BACKGROUND: Adolescence is a period of elevated alcohol consumption in humans as well as in animal models. Previous studies in our laboratory have shown that adolescent Sprague-Dawley rats consume approximately 2 times more ethanol on a gram per kilogram basis than adult animals in a 2-bottle choice free-access situation. The purpose of the present study was to examine the time course and pattern of elevated ethanol intake during adolescence and the adolescent-to-adult transition, contrast this intake with ontogenetic patterns of food and water intake, and determine whether adolescent access to ethanol elevates voluntary consumption of ethanol in adulthood. METHODS: Adolescent [postnatal day (P)27-28] and adult (P69-70) male Sprague-Dawley rats were singly housed with continuous access to both water and 1 of 3 experimental solutions in ball-bearing-containing sipper tubes: unsweetened ethanol (10% v/v), sweetened ethanol (10% v/v+0.1% w/v saccharin), and saccharin alone (0.1% w/v). RESULTS: Ethanol consumption plateaued at approximately 7.5 g/kg/d during the first 2 weeks of measurement (i.e., P28-39) in early adolescence, before declining sharply at approximately P40 to levels that were only modestly elevated compared with adult-typical consumption patterns that were reached by approximately P70. In contrast, intake of food and total calories showed a more gradual decline into adulthood with no distinguishable plateaus in early adolescence. When adolescent-initiated and adult-initiated animals were tested at the same chronological age in adulthood, animals drank similar amounts regardless of the age at which they were first given voluntary access to ethanol. CONCLUSIONS: Taken together, these data suggest that the elevated ethanol intake characteristic of early-to-mid adolescence is not simply a function of adolescent-typical hyperphagia or hyperdipsia, but instead may reflect age-related differences in neural substrates contributing to the rewarding or aversive effects of ethanol, as well as possible modulatory influences of ontogenetic differences in sensitivity to novelty or in ethanol pharmacokinetics. Voluntary home cage consumption of ethanol during adolescence, however, was not found to subsequently elevate ethanol drinking in adulthood.