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Background Environmental lead ( Pb) exposure and alcohol abuse pose significant public health problems for our society. One of the proposed mechanisms of action of the developmental neurotoxicant Pb is related to its ability to affect antioxidant enzymes, including catalase ( CAT). Ethanol's ( Et OH) motivational effects are postulated to be mediated by the CAT-dependent acetaldehyde generated in the brain. The current study sought to investigate the role of this enzyme in the elevated EtOH intake previously reported in perinatally Pb-exposed rats. Methods Thirty-five-day-old male Wistar rats exposed to 220 ppm Pb during gestation and lactation were offered escalating Et OH solutions (2 to 10%) or water, 2 h/d for 28 days. Once baseline 10% Et OH intake was achieved, they were injected with (i) saline ( SAL), (ii) 3-amino 1,2,4 triazole (aminotriazole [AT], a CAT inhibitor, 250 mg/kg intraperitoneally [i.p.], 5 hours before the last 8 Et OH intake sessions), or (iii) 3-nitropropionic acid (3 NPA; a CAT activator, 20 mg/kg subcutaneously [s.c.], 45 minutes before the last 4 Et OH intake sessions). Rats were then sacrificed, blood collected, and brain regions harvested for CAT activity determination. Additional studies evaluated Et OH intake and CAT activity in response to 10 and 30 mg/kg 3 NPA. Both 3 NPA and AT were evaluated for striatal cytotoxicity. Results We observed that AT pretreatment blunted the increased Et OH intake, as well as the elevated CAT activity in blood, cerebellum, and hippocampus evidenced in the developmentally Pb-exposed rats that have consumed Et OH. Conversely, 20 mg/kg 3 NPA further increased voluntary Et OH intake in these animals as compared with controls, concomitantly with a slight elevation in CAT activity both in blood and in the striatum, associated with no changes in striatal cytotoxicity. Conclusions These results suggest a participation of CAT, and possibly acetaldehyde, in Pb-induced high Et OH intake, and open up new avenues to elucidate the mechanism that underlies the Pb and Et OH interaction. [ABSTRACT FROM AUTHOR]

Background In our companion article, we examined the role of MyD88-dependent signaling in ethanol (EtOH) consumption in mice lacking key components of this inflammatory pathway and observed differential effects on drinking. Here, we studied the role of these same signaling components in the acute sedative, intoxicating, and physiological effects of EtOH. Toll-like receptor 4 (TLR4) has been reported to strongly reduce the duration of EtOH-induced sedation, although most studies do not support its direct involvement in EtOH consumption. We examined TLR4 and other MyD88 pathway molecules to determine signaling specificity in acute EtOH-related behaviors. We also studied other GABAergic sedatives to gauge the EtOH specificity and potential role for GABA in EtOH's sedative and intoxicating effects in the mutant mice. Methods Loss of righting reflex ( LORR) and recovery from motor incoordination were studied following acute injection of EtOH or other sedative drugs in male and female control C57 BL/6J mice versus mice lacking CD14, TLR2, TLR4 (C57 BL/10ScN), or MyD88. We also examined EtOH-induced hypothermia and blood EtOH clearance in these mice. Results Male and female mice lacking TLR4 or MyD88 showed reduced duration of EtOH-induced LORR and faster recovery from EtOH-induced motor incoordination in the rotarod test. MyD88 knockout mice had slightly faster recovery from EtOH-induced hypothermia compared to control mice. None of the mutants differed from control mice in the rate of blood EtOH clearance. All of the mutants showed similar decreases in the duration of gaboxadol-induced LORR, but only mice lacking TLR4 were less sensitive to the sedative effects of pentobarbital. Faster recovery from diazepam-induced motor impairment was observed in CD14, TLR4, and MyD88 null mice of both sexes. Conclusions TLR4 and MyD88 were key mediators of the sedative and intoxicating effects of EtOH and GABAergic sedatives, indicating a strong influence of TLR4-MyD88 signaling on GABAergic function. Despite the involvement of TLR4 in EtOH's acute behaviors, it did not regulate EtOH consumption in any drinking model as shown in our companion article. Collectively, our studies demonstrate differential effects of TLR-MyD88 components in the acute versus chronic actions of EtOH. [ABSTRACT FROM AUTHOR]

Background Molecular and behavioral studies support a role for innate immune proinflammatory pathways in mediating the effects of alcohol. Increased levels of Toll-like receptors (TLRs) have been observed in animal models of alcohol consumption and in human alcoholics, and many of these TLRs signal via the MyD88-dependent pathway. We hypothesized that this pathway is involved in alcohol drinking and examined some of its key signaling components. Methods Different ethanol (EtOH)-drinking paradigms were studied in male and female control C57BL/6J mice versus mice lacking CD14, TLR2, TLR4 (C57BL/10ScN), or MyD88. We studied continuous and intermittent access 2-bottle choice (2BC) and 1-bottle and 2BC drinking-in-the-dark (DID) tests as well as preference for saccharin, quinine, and NaCl. Results In the 2BC continuous access test, EtOH intake decreased in male TLR2 knockout (KO) mice, and we previously reported reduced 2BC drinking in male and female CD14 KO mice. In the intermittent access 2BC test, EtOH intake decreased in CD14 KO male and female mice, whereas drinking increased in MyD88 KO male mice. In the 2BC-DID test, EtOH drinking decreased in male and female mice lacking TLR2, whereas drinking increased in MyD88 KO male mice. In the 1-bottle DID test, EtOH intake decreased in female TLR2 KO mice. TLR2 KO and CD14 KO mice did not differ in saccharin preference but showed reduced preference for NaCl. MyD88 KO mice showed a slight reduction in preference for saccharin. Conclusions Deletion of key components of the MyD88-dependent pathway produced differential effects on EtOH intake by decreasing (TLR2 KO and CD14 KO) or increasing (MyD88 KO) drinking, while deletion of TLR4 had no effect. Some of the drinking effects depended on the sex of the mice and/or the EtOH-drinking model. [ABSTRACT FROM AUTHOR]

Background Growing evidence supports a central role for the circadian system in alcohol use disorders, but few studies have examined this relationship during adolescence. In mammals, circadian rhythms are regulated by the suprachiasmatic nucleus, a biological clock whose timing is synchronized (reset) to the environment primarily by light (photic) input. Alcohol (ethanol [EtOH]) disrupts circadian timing in part by attenuating photic phase-resetting responses in adult rodents. However, circadian rhythms change throughout life and it is not yet known whether EtOH has similar effects on circadian regulation during adolescence. Methods General circadian locomotor activity was monitored in male C57BL6/J mice beginning in adolescence (P27) or adulthood (P61) in a 12-hour light, 12-hour dark photocycle for ~2 weeks to establish baseline circadian activity measures. On the day of the experiment, mice received an acute injection of EtOH (1.5 g/kg, i.p.) or equal volume saline 15 minutes prior to a 30-minute light pulse at Zeitgeber Time 14 (2 hours into the dark phase) and then were released into constant darkness (DD) for ~2 weeks to assess phase-resetting responses. Control mice of each age-group received injections but no light pulse prior to DD. Results While adults showed the expected decrease in photic phase-delays induced by acute EtOH, this effect was absent in adolescent mice. Adolescents also showed baseline differences in circadian rhythmicity compared to adults, including advanced photocycle entrainment, larger photic phase-delays, a shorter free-running (endogenous) circadian period, and greater circadian rhythm amplitude. Conclusions Collectively, our results indicate that adolescent mice are less sensitive to the effect of EtOH on circadian photic phase-resetting and that their daily activity rhythms are markedly different than those of adults. [ABSTRACT FROM AUTHOR]

Background Cyclic AMP-dependent protein kinase A ( PKA) signaling is a key target for the action of alcohol and may therefore play a role in the pathophysiology of alcohol withdrawal seizures ( AWSs). Here, we investigated the role of PKA activity with respect to increased seizure susceptibility in rats that were subjected to alcohol withdrawal. Methods Adult male Sprague Dawley rats received 3 daily doses of ethanol (EtOH) (or vehicle) for 4 consecutive days. Rats were then tested for susceptibility to acoustically evoked AWSs 3, 24, and 48 hours after the last alcohol dose. In separate experiments, the inferior colliculus ( IC) was collected at these same time points from rats subjected to alcohol withdrawal and control rats following alcohol withdrawal. PKA activity, catalytic C α ( PKAC α) protein, regulatory RII α ( PKARII α) protein, and RII β ( PKARII β) protein were measured in the IC. Lastly, in situ pharmacological studies were performed to evaluate whether inhibiting PKA activity in the IC suppressed AWSs. Results In the EtOH-treated group, AWSs were observed at the 24-hour time point, but not at the 3-hour or 48-hour time points. In the IC, PKA activity was significantly higher both 3 hours (i.e., before AWS susceptibility) and 24 hours after the last alcohol dose (when AWS susceptibility peaked) than in control rats. Consistent with these findings, protein levels of the PKAC α subunit were significantly increased in the IC both 3 and 24 hours after the last alcohol dose. Lastly, in situ inhibition of PKA activity within the IC suppressed AWSs. Conclusions The increase in PKA activity and PKAC α protein expression in the IC preceded the occurrence of AWSs, and inhibiting PKA activity within the IC suppressed acoustically evoked AWSs. Together, these findings suggest that altered PKA activity plays a key role in the pathogenesis of AWSs. [ABSTRACT FROM AUTHOR]

Background Novelty-seeking behavior is related to the reward system in the brain and can predict the potential for addiction. Alcohol use is prevalent in HIV-1-infected patients and adversely affects antiretroviral medication. The difference in vulnerability to alcohol addiction between HIV-1-infected and noninfected populations has not been fully investigated. This study was designed to determine whether HIV-1 proteins alter the effects of ethanol (EtOH) on novelty-seeking behavior using the HIV-1 transgenic ( HIV-1Tg) rat as the study model and to examine the molecular mechanisms responsible for this behavior. Methods Both HIV-1Tg and F344 control rats were tested for baseline novelty-seeking behavior, then received either EtOH (1 g/kg) at a concentration of 20% v/v or saline treatment for 13 days, and then were retested for novelty seeking. Quantitative real-time polymerase chain reaction was conducted to examine the differences in expression of 65 genes implicated in novelty seeking and alcohol addiction between strains and treatment groups. Results The HIV-1 proteins significantly enhanced baseline novelty-seeking behaviors in both the hole-board and open-field tests. Chronic EtOH treatment significantly increased baseline novelty-seeking behavior in both strains, but the effects of EtOH appeared to be more robust and prominent in HIV-1Tg rats. Strain-specific patterns of altered gene expression were observed for dopaminergic, cholinergic, and glutamatergic signaling in the nucleus accumbens, suggesting the effects of HIV-1 proteins on the brain's reward system. Chronic EtOH treatment was shown to greatly modulate the effects of HIV-1 proteins in these neurotransmitter systems. Conclusions Taken together, our findings indicate that HIV-1 proteins could modify novelty-seeking behavior at the gene expression level, and EtOH treatment may enhance this behavior in both strains but to a greater extent in HIV-1Tg rats. [ABSTRACT FROM AUTHOR]

Background A number of studies have shown that acetaldehyde synthesized in the brain is necessary to induce ethanol (EtOH) reinforcement in naïve animals ( acquisition phase). However, after chronic intake is achieved ( maintenance phase), EtOH intake becomes independent of acetaldehyde generation or its levels. Glutamate has been reported to be associated with the maintenance of chronic EtOH intake. The levels of brain extracellular glutamate are modulated by 2 glial processes: glutamate reabsorption via an Na+-glutamate transporter ( GLT1) and a cystine-glutamate exchanger. Chronic EtOH intake lowers GLT1 levels and increases extracellular glutamate. The administration of N-acetyl cysteine ( NAC), a precursor of cystine, has been shown to reduce the relapse of several drugs of abuse, while NAC has not been tested on chronic EtOH intake or on EtOH's influence on the motivation for another drug. These were investigated in the present study. Methods (i) Rats bred for their high EtOH intake were allowed access to 10% EtOH and water up to 87 days. NAC was administered (30 and 60 mg/kg daily, intraperitoneally) for 14 consecutive days, either during the acquisition phase or the maintenance phase of EtOH drinking. (ii) In additional experiments, rats were allowed EtOH (10%) and water access for 61 days, after which EtOH was replaced by saccharin (0.3%) to determine both if chronic EtOH consumption influences saccharin intake and whether NAC modifies the post chronic EtOH saccharin intake. Results NAC did not influence the acquisition ('first hit') of chronic EtOH intake, but greatly inhibited (60 to 70%; p < 0.0001) EtOH intake when NAC was administered to animals that were consuming EtOH chronically. NAC did not influence saccharin intake in naïve animals. In animals that had consumed EtOH chronically and were thereafter offered a saccharin solution (0.3%), saccharin intake increased over 100% versus that of EtOH-untreated animals, an effect that was fully suppressed by NAC. Conclusions N-acetyl cysteine, a drug approved for use in humans, markedly reduces chronic EtOH intake and abolishes the increased intake of saccharin stimulated by chronic EtOH drinking. [ABSTRACT FROM AUTHOR]

Background The lung has a highly regulated system of innate immunity to protect itself from inhaled microbes and toxins. The first line of defense is mucociliary clearance, but if invaders overcome this, inflammatory pathways are activated. Toll-like receptors ( TLRs) are expressed on the airway epithelium. Their signaling initiates the inflammatory cascade and leads to production of inflammatory cytokines such as interleukin (IL)-6 and IL-8. We hypothesized that airway epithelial insults, including heavy alcohol intake or smoking, would alter the expression of TLRs on the airway epithelium. Methods Bronchoscopy with bronchoalveolar lavage and brushings of the airway epithelium was performed in otherwise healthy subjects who had normal chest radiographs and spirometry. A history of alcohol use disorders ( AUDs) was ascertained using the Alcohol Use Disorders Identification Test ( AUDIT), and a history of cigarette smoking was also obtained. Age, gender, and nutritional status in all groups were similar. We used real-time polymerase chain reaction (PCR) to quantitate TLR1 to 9 and enzyme-linked immune assay to measure tumor necrosis factor- α, IL-6, and IL-8. Results Airway brushings were obtained from 26 nonsmoking/non- AUD subjects, 28 smoking/non- AUD subjects, 36 smoking/ AUD subjects, and 17 nonsmoking/ AUD subjects. We found that TLR2 is up-regulated in AUD subjects, compared to nonsmoking/non- AUD subjects, and correlated with their AUDIT scores. We also measured a decrease in TLR4 expression in AUD subjects that correlated with AUDIT score. IL-6 and IL-8 were also increased in bronchial washings from AUD subjects. Conclusions We have previously demonstrated in normal human bronchial epithelial cells that in vitro alcohol exposure up-regulates TLR2 through a NO/ cGMP/ PKG-dependent pathway, resulting in up-regulation of inflammatory cytokine production after Gram-positive bacterial product stimulation. Our current translational study confirms that TLR2 is also up-regulated in humans with AUDs. [ABSTRACT FROM AUTHOR]

Background The molecular mechanisms of the acute hypotensive and indirectly assessed cardiac depressant effect of ethanol ( Et OH)-evoked myocardial depression and hypotension in female rats are not known. We tested the hypothesis that a time-dependent myocardial depression caused by Et OH is initiated by its direct and indirect (cardiac vagal dominance) effects and is exacerbated by gradual development of oxidative stress. Methods In conscious female rats, we directly measured left ventricular developed pressure ( LVDP), the maximal rise of ventricular pressure over time (d P/d tmax), blood pressure ( BP), heart rate ( HR), and sympathovagal activity following intragastric Et OH (1 g/kg) or water over 90 minutes. Catalytic activity of acetaldehyde ( ACA)-generating (alcohol dehydrogenase [ADH] and catalase) and eliminating aldehyde dehydrogenase [ALDH2] enzymes along with mediators of oxidative stress were measured in myocardial tissues collected at 30, 60, or 90 minutes after Et OH or water. Results Et OH reduced myocardial function ( LVDP and d P/d tmax) within 5 to 10 minutes before the steady fall in BP in conscious proestrus rats. Further, Et OH shifted the sympathovagal balance, analyzed by spectral analysis of high frequency and low frequency of interbeat intervals, toward vagal dominance. Prior vagal blockade (atropine) or antioxidant (tempol) treatment attenuated Et OH-evoked myocardial depression and hypotension. Ex vivo studies revealed time-dependent: (i) enhancement of ADH, but not ALDH2 activity (indicative of elevated ACA levels), (ii) increases in phosphorylated Akt and ERK1/2, NADPH-oxidase activity, reactive oxygen species, malondialdehyde, and 4-hydroxy-2-nonenal-modified proteins. These molecular responses along with reduced myocardial catalase activity were most evident at 90 minutes post- Et OH when the reductions in cardiac function and BP reached their nadir. Conclusions Vagal dominance and time-dependent myocardial oxidative stress along with the accumulation of cardiotoxic aldehydes mediate Et OH-evoked myocardial dysfunction and hypotension in conscious proestrus female rats. [ABSTRACT FROM AUTHOR]

Background Affiliative social interactions promote alcohol consumption, and alcohol also promotes affiliative behavior. Furthermore, for most species, moderate doses of ethanol ( Et OH) and social affiliation are each rewarding. However, animal studies of drug and Et OH reward typically test individuals in isolation. To address social dimensions of Et OH reward, this study tested Et OH-induced conditioned social preference in male C57 BL/6 mice with ( ORCHX+ T) and without ( ORCHX) testosterone. Methods ORCHX+T males received Et OH (0, 1, 2, or 3 g/kg) intraperitoneally and were paired 4× for 30 minutes each with 1 of 2 stimulus males: with the CS− stimulus male after saline injection and with the CS+ male following Et OH. After pairing, time spent with CS+ and CS− males was measured in a 10-minute test. Results ORCHX+T test males showed conditioned preference for the CS+ male in response to 3 g/kg Et OH (change in preference: +71.3 ± 30.0 s/10 min, p < 0.05), but not for 0, 1, or 2 g/kg. By contrast, ORCHX males did not demonstrate conditioned preference for 3 g/kg Et OH (+16.0 ± 24.3 s/10 min, p > 0.05). In separate groups of mice, stimulus males ( IS+) received Et OH during pairing to determine whether test mice prefer another intoxicated mouse. Both ORCHX+T and ORCHX test mice showed an increase in preference score for the IS+ mouse ( ORCHX+T: +68.1 ± 24.0 seconds; ORCHX: +58.9 ± 19.6 seconds, p < 0.05). Conclusions These data demonstrate that Et OH promotes social preference in male mice, as it does in females. Testosterone enhances this effect. [ABSTRACT FROM AUTHOR]

Background The cAMP-dependent protein kinase ( PKA) signaling transduction pathway has been shown to play an important role in the modulation of several ethanol (EtOH)-induced behavioral actions. In vivo, short-term exposure to EtOH up-regulates the cAMP-signaling cascade. Interestingly, different Ca2+-dependent cAMP- PKA cascade mediators play a critical role in the neurobehavioral response to EtOH, being of special relevance to the Ca2+-dependent adenylyl cyclases 1 and 8. We hypothesize an intracellular PKA activation elicited by EtOH administration, which may be regulated by a Ca2+-dependent mechanism as an early cellular response. Thus, the present work aims to explore the role of Ca2+ (internal and external) on the EtOH-activated PKA cascade. Methods Swiss male mice received an intraperitoneal injection of EtOH (0 or 4 g/kg), and brains were dissected following a temporal pattern (7, 15, 30, 45, 90, or 120 minutes). Either the enzymatic PKA activity or its fingerprint was analyzed on different brain areas (cortex, hypothalamus, hippocampus, and striatum). To explore the role of Ca2+ on the EtOH-activated PKA cascade, mice were pretreated with diltiazem (0 or 20 mg/kg), dantrolene (0 or 5 mg/kg), or 3,7- Dimethyl-1-(2-propynyl)xanthine (0 or 1 mg/kg) 30 minutes before EtOH (4 g/kg) administration. After 45 minutes of EtOH administration, brains were removed and dissected to measure the PKA activity or its fingerprint. Results Results from these experiments showed an EtOH-dependent activation of PKA in different brain areas. Manipulations involving a disruption of intracellular Ca2+ release from the endoplasmic reticulum resulted in a decreased EtOH-induced activation of PKA. On the contrary, extracellular-to-cytoplasm Ca2+ manipulations did not prevent the PKA activation by EtOH. Conclusions Altogether, these results show the critical role of stored Ca2+ as an intracellular mediator of different neurobiological actions of EtOH and provide further evidence of a possible new target for EtOH within the central nervous system. [ABSTRACT FROM AUTHOR]

Background Ethanol ( Et OH) abuse is a major health and economic concern, particularly for females who appear to be more sensitive to the rewarding effects of Et OH. This study compared sex differences to the rewarding and aversive effects of Et OH using place-conditioning procedures in rats. Methods Separate groups of adult (male, female, ovariectomized [ OVX] female) and adolescent (male and female) rats received Et OH (0, 0.5, 1.0, 2.0, or 2.5 g/kg, intraperitoneal) and were confined to their initially nonpreferred side of our conditioning apparatus for 30 minutes. On alternate days, they received saline and were confined to the other side. Following 5 drug pairings, the rats were retested for preference behavior. Separate cohorts of the same groups of rats were injected with a similar dose range of Et OH, and blood Et OH levels ( BELs) were compared 30 minutes later. Results Et OH produced rewarding or aversive effects in a dose-dependent manner. An intermediate dose of Et OH (1.0 g/kg) produced rewarding effects in adult female, but not in male or OVX female rats, suggesting that ovarian hormones facilitate the rewarding effects of Et OH. Similarly, this intermediate dose of Et OH produced rewarding effects in adolescent female, but not in male rats. The highest dose of Et OH (2.5 g/kg) produced aversive effects that were similar across all adult groups. However, the aversive effects of Et OH were lower in adolescents than adults, suggesting that adolescents are less sensitive to the aversive effects of Et OH. The aversive effects of Et OH did not vary across the estrous cycle in intact adult females. There were also no group differences in BELs, suggesting that our results are not related to Et OH metabolism. Conclusion Our results in rats suggest that human females may be more vulnerable to Et OH abuse due to enhanced rewarding effects of this drug that are mediated by the presence of ovarian hormones. [ABSTRACT FROM AUTHOR]

Background Hydrogen peroxide (H2O2) is the cosubstrate used by the enzyme catalase to form Compound I (the catalase-H2O2 system), which is the major pathway for the conversion of ethanol (EtOH) into acetaldehyde in the brain. This centrally formed acetaldehyde has been shown to be involved in some of the psychopharmacological effects induced by EtOH in rodents, including voluntary alcohol intake. It has been observed that different levels of this enzyme in the central nervous system (CNS) result in variations in the amount of EtOH consumed. This has been interpreted to mean that the brain catalase-H2O2 system, by determining EtOH metabolism, mediates alcohol self-administration. To date, however, the role of H2O2 in voluntary EtOH drinking has not been investigated. Methods In the present study, we explored the consequence of a reduction in cerebral H2O2 levels in volitional EtOH ingestion. With this end in mind, we injected mice of the C57BL/6J strain intraperitoneally with the H2O2 scavengers alpha-lipoic acid (LA; 0 to 50 mg/kg) or ebselen (Ebs; 0 to 25 mg/kg) 15 or 60 minutes, respectively, prior to offering them an EtOH (10%) solution following a drinking-in-the-dark procedure. The same procedure was followed to assess the selectivity of these compounds in altering EtOH intake by presenting mice with a (0.1%) solution of saccharin. In addition, we indirectly tested the ability of LA and Ebs to reduce brain H2O2 availability. Results The results showed that both LA and Ebs dose-dependently reduced voluntary EtOH intake, without altering saccharin consumption. Moreover, we demonstrated that these treatments decreased the central H2O2 levels available to catalase. Conclusions Therefore, we propose that the amount of H2O2 present in the CNS, by determining brain acetaldehyde formation by the catalase-H2O2 system, could be a factor that determines an animal's propensity to consume EtOH. [ABSTRACT FROM AUTHOR]

Background We tested the hypothesis that alterations of the phosphorylation/dephosphorylation profile of mitogen-activated protein kinases ( MAPKs) in the rostral ventrolateral medulla ( RVLM) underlie the pressor response elicited by ethanol ( Et OH) microinjection into the RVLM of spontaneously hypertensive rats ( SHRs). The studies were extended to determine whether acetaldehyde ( ACA), the primary oxidative product of Et OH, replicates the molecular effects of Et OH within the RVLM and the consequent pressor response. Methods Effects of Et OH or ACA on blood pressure ( BP) were evaluated in the absence or presence of selective JNK ( SP600125), ERK ( PD98059), p38 ( SB203580), or ser/thr phosphatases (okadaic acid [ OKA]) inhibitor. Results Intra- RVLM Et OH (10 μg/rat) or ACA (2 μg/rat) caused a similar ERK2-dependent pressor response because Et OH or ACA-evoked increases in BP and in RVLM p- ERK2 level were abolished after pharmacologic inhibition of ERK phosphorylation. SP600125 abrogated the pressor action of Et OH, but not ACA, thus implicating JNK in Et OH action on BP. Despite Et OH enhancement of p38 phosphorylation, pharmacological studies argued against a causal role for this kinase in Et OH-evoked pressor response. RVLM phosphatase catalytic activity was not influenced by Et OH or ACA. Interestingly, pharmacologic phosphatase inhibition ( OKA), which increased RVLM p- ERK2 and BP, abrogated the pressor effect of subsequently administered Et OH or ACA. Conclusions Enhancement of RVLM ERK2 phosphorylation constitutes a major molecular mechanism for the pressor response elicited by intra- RVLM Et OH or its metabolite, ACA, in conscious SHRs. Further, RVLM kinases dephosphorylation does not contribute to intra- RVLM Et OH- or ACA-evoked pressor response. [ABSTRACT FROM AUTHOR]

Background The main system of central ethanol (EtOH) oxidation is mediated by the enzyme catalase. By reacting with H2 O2, brain catalase forms compound I (the catalase- H2 O2 system), which is able to oxidize EtOH to acetaldehyde ( ACD) in the brain. We have previously shown that ACD regulates EtOH motivational properties and possesses reinforcing effects by itself. In this study, we investigate the effects of alpha-lipoic acid ( ALA), a scavenging agent for H2 O2, on oral EtOH self-administration. Methods To this end, we trained Wistar rats to orally self-administer EtOH (10%) by nose poking. The effect of intraperitoneal pretreatment with ALA was evaluated during (i) maintenance of EtOH self-administration, (ii) EtOH self-administration under a progressive ratio ( PR) schedule of reinforcement, and (iii) oral EtOH priming to induce reinstatement of EtOH seeking behavior. Moreover, we tested the effect of ALA on saccharin (0.05%) reinforcement, as assessed by oral self-administration. Results The results indicate that ALA dose-dependently reduced the maintenance, the break point of EtOH self-administration under a PR and the reinstatement of EtOH seeking behavior without suppressing saccharin self-administration. Conclusions These results support that ALA may have a potential use in alcoholism treatment. [ABSTRACT FROM AUTHOR]

Background Animals that have chronically consumed alcohol and are subsequently deprived of it markedly increase their intake above basal levels when access to alcohol is reinstated. Such an effect, termed the alcohol deprivation effect ( ADE), has been proposed to reflect (i) an obsessive-compulsive behavior, (ii) craving, or (iii) an increased reinforcing value of ethanol ( Et OH). It has been reported that acetaldehyde, a highly reinforcing metabolite of Et OH, is generated in the brain by the action of catalase. Recent studies show that the administration of an anticatalase (sh RNA)-encoding lentiviral vector into the brain ventral tegmental area ( VTA) of naïve rats virtually abolishes (85 to 95%) their Et OH intake. It is hypothesized that the antireinforcing effect of the anticatalase vector will also inhibit the ADE. Methods Two-month-old Wistar-derived UCh B alcohol drinker rats were offered free access to water and 10 and 20% Et OH for 67 days. Thereafter, the animals were deprived of Et OH for 15 days and were subsequently offered access to the Et OH solutions. At the start of the deprivation period, animals were microinjected a single dose of an anticatalase (or control) vector into the VTA. Et OH intake was measured on the first hour of Et OH re-exposure as well as on a 24-hour basis for 7 days. Results A marked ADE was observed when Et OH intake was measured on the first hour or 24 hours following Et OH re-exposure, compared to the corresponding controls. The administration of the anticatalase vector reduced ADE by 60 to 80% ( p < 0.001) on the first hour and by 63 to 80% ( p < 0.001) on the initial 24 hours of Et OH re-exposure (first and second ADE, respectively) without changing the total fluid intake, indicating a specific effect on Et OH drinking. Conclusions Ethanol intake associated with ADE-a binge-like drinking behavior-is markedly inhibited by the administration of an anticatalase vector into the VTA, which blocks the conversion of Et OH into acetaldehyde, strongly suggesting that the marked increased Et OH intake that follows an alcohol deprivation period is mediated by acetaldehyde and its reinforcing metabolite. [ABSTRACT FROM AUTHOR]

Background Published findings indicate that acetaldehyde ( ACD; the first metabolite of ethanol [ Et OH]) and salsolinol ( SAL; formed through the nonenzymatic condensation of ACD and dopamine [ DA]) can be formed following Et OH consumption. Both ACD and SAL exhibit reinforcing properties within the posterior ventral tegmental area (p VTA) and both exhibit an inverted ' U-shaped' dose-response curve. The current study was undertaken to examine the dose-response effects of microinjections of ACD or SAL into the p VTA on DA efflux in the nucleus accumbens shell ( Acb Sh). Methods For the first experiment, separate groups of male Wistar rats received pulse microinjections of artificial cerebrospinal fluid (a CSF) or 12-, 23-, or 90-μM ACD into the p VTA, while extracellular DA levels were concurrently measured in the Acb Sh. The second experiment was similarly conducted, except rats were given microinjections of a CSF or 0.03-, 0.3-, 1.0-, or 3.0-μ M SAL, while extracellular levels of DA were measured in the Acb Sh. Results Both ACD and SAL produced a dose-dependent inverted ' U-shaped' response on DA release in the Acb Sh, with 23-μ M ACD (200% baseline) and 0.3-μ M SAL (300% baseline) producing maximal peak responses with higher concentrations of ACD (90 μ M) and SAL (3.0 μ M) producing significantly lower DA efflux. Conclusions The findings from the current study indicate that local application of intermediate concentrations of ACD and SAL stimulated DA neurons in the p VTA, whereas higher concentrations may be having secondary effects within the p VTA that inhibit DA neuronal activity. The present results parallel the studies on the reinforcing effects of ACD and SAL in the p VTA and support the idea that the reinforcing effects of ACD and SAL within the p VTA are mediated by activating DA neurons. [ABSTRACT FROM AUTHOR]

Acetaldehyde (ACD), the first metabolite of ethanol (EtOH) appears to be involved in many of the psychoactive effects of its parent compound, including EtOH-induced activation of the mesolimbic dopamine (DA) system, thereby suggesting that ACD may participate in EtOH motivational properties. -cysteine (Lcys), a thiol compound sequestering ACD, is able to prevent the behavioral effect of EtOH and ACD. Here we show that the stimulatory effect of both EtOH and ACD on the mesolimbic DA system is prevented by Lcys pretreatment. Male Wistar rats were implanted with a microdialysis probe in the nucleus accumbens shell (NAccs), and pretreated intraperitoneally with Lcys (30 mg/kg) before intragastric administration of EtOH (1 g/kg) or ACD (20 mg/kg) or before intraperitoneal administration of morphine (2.5 mg/kg). Pretreatment with Lcys prevented both EtOH and ACD-induced DA release in the NAccs without influencing morphine-induced DA release, suggesting that Lcys specifically affects EtOH-induced DA release possibly through ACD sequestering. Our results underscore the role of ACD on EtOH-induced stimulation of DA mesoaccumbens system and support the notion that thiol compounds such as Lcys, by modulating EtOH-derived ACD bioavailability, would blunt EtOH rewarding properties. [ABSTRACT FROM AUTHOR]

Background: The current therapies for alcohol abuse disorders are not effective in all patients, and continued development of pharmacotherapies is needed. One approach that has generated recent interest is the antagonism of ghrelin receptors. Ghrelin is a gut-derived peptide important in energy homeostasis and regulation of hunger. Recent studies have implicated ghrelin in alcoholism, showing altered plasma ghrelin levels in alcoholic patients as well as reduced intakes of alcohol in ghrelin receptor knockout mice and in mice treated with ghrelin receptor antagonists. The aim of this study was to determine the neuroanatomical locus/loci of the effect of ghrelin receptor antagonism on alcohol consumption using the ghrelin receptor antagonist, D-Lys3-GHRP-6. Methods: In Experiment 1 , male C57BL/6J mice were injected with saline 3 hours into the dark cycle and allowed access to 15% (v/v) ethanol or water for 2 hours in a 2-bottle choice experiment. On test day, the mice were injected with either saline or 400 nmol of the ghrelin receptor antagonist, D-Lys3-GHRP-6, and allowed to drink 15% ethanol or water for 4 hours. The preference for alcohol and alcohol intake were determined. In Experiment 2 , the same procedure was followed as in Experiment 1 but mice were only allowed access to a single bottle of 20% ethanol (v/v), and alcohol intake was determined. Blood ethanol levels were analyzed, and immunohistochemistry for c-Fos was carried out to investigate changes in neural activity. To further elucidate the mechanism by which D-Lys3-GHRP-6 affects alcohol intake, in Experiment 3 , the effect of D-Lys3-GHRP-6 on the neural activation induced by intraperitoneal ethanol was investigated. For the c-Fos studies, brain regions containing ghrelin receptors were analyzed, i.e. the perioculomotor urocortin population of neurons (pIIIu), the ventral tegmental area (VTA), and the arcuate nucleus (Arc). In Experiment 4 , to test if blood ethanol concentrations were affected by D-Lys3-GHRP-6, blood samples were taken at 2 time-points after D-Lys3-GHRP-6 pretreatment and systemic ethanol administration. Results: In Experiment 1 , D-Lys3-GHRP-6 reduced preference to alcohol and in a follow-up experiment ( Experiment 2 ) also dramatically reduced alcohol intake when compared to saline-treated mice. The resulting blood ethanol concentrations were lower in mice treated with the ghrelin receptor antagonist. Immunohistochemistry for c-Fos showed fewer immunopositive cells in the pIIIu of the antagonist-treated mice but no difference was seen in the VTA or Arc. In Experiment 3 , D-Lys3-GHRP-6 reduced the induction of c-Fos by intraperitoneal ethanol in the pIIIu but had no effect in the VTA. In the Arc, there was a significant increase in the number of c-Fos immunopositive cells after D-Lys3-GHRP-6 administration, but the antagonist had no effect on ethanol-induced expression of c-Fos. D-Lys3-GHRP-6-pretreatment also did not affect the blood ethanol concentrations observed after a systemic injection of ethanol when compared to saline-pretreated mice ( Experiment 4 ). Conclusions: These findings indicate that the action of ghrelin on the regulation of alcohol consumption may occur via the pIIIu. [ABSTRACT FROM AUTHOR]

Background: Ethanol exposure during early life has been shown to permanently alter the circadian expression of clock regulatory genes and the β-endorphin precursor proopiomelanocortin (POMC) gene in the hypothalamus. Ethanol also alters the stress- and immune-regulatory functions of β-endorphin neurons in laboratory rodents. Our aim was to determine whether the circadian clock regulatory Per2 gene modulates the action of ethanol on β-endorphin neurons in mice. Methods: Per2 mutant ( mPer2 Brdml) and wild type (C57BL/6J) mice were used to determine the effect of Per2 mutation on ethanol-regulated β-endorphin neuronal activity during neonatal period using an in vitro mediobasal hypothalamic (MBH) cell culture model and an in vivo milk formula feeding animal model. The β-endorphin neuronal activity following acute and chronic ethanol treatments was evaluated by measuring the peptide released from cultured cells or peptide levels in the MBH tissues, using enzyme-linked immunosorbent assay (ELISA). Results: Per2 mutant mice showed a higher basal level of β-endorphin release from cultured MBH cells and a moderate increase in the peptide content in the MBH in comparison with control mice. However, unlike wild type mice, Per2 mutant mice showed no stimulatory or inhibitory β-endorphin-secretory responses to acute and chronic ethanol challenges in vitro. Furthermore, Per2 mutant mice, but not wild type mice, failed to show the stimulatory and inhibitory responses of MBH β-endorphin levels to acute and chronic ethanol challenges in vivo. Conclusions: These results suggest for the first time that the Per2 gene may be critically involved in regulating β-endorphin neuronal function. Furthermore, the data revealed an involvement of the Per2 gene in regulating β-endorphin neuronal responses to ethanol. [ABSTRACT FROM AUTHOR]