Publications

2005
Hilary J. Little, David N. Stephens, Tamzon L. Ripley, Gilyana Borlikova, Theodora Duka, Manja Schubert, Doris Albrecht, Howard C. Becker, Marcello F. Lopez, Friedbert Weiss, Colin Drummond, Michelle Peoples, and Christopher Cunningham. “Alcohol withdrawal and conditioning.” Alcoholism, Clinical and Experimental Research, 29, 3, Pp. 453–464. Abstract
This review contains the proceedings from a symposium held at the RSA conference in 2003 on "Alcohol Withdrawal and Conditioning." The presentations covered a range of interactions between conditioning and alcohol withdrawal, in both animal behavior and the clinic. Dr. D.N. Stephens first described his studies exploring the consequences of alcohol dependence and repeated experience of withdrawal on the conditioning process. His data suggested that repeated withdrawal from moderate alcohol intake impairs amygdala-dependent mechanisms for learning about aversive events. Dr. H. Becker then detailed studies examining the consequences of repeated ethanol withdrawal experience on subsequent ethanol drinking behavior in mice, and conditions in which motivational properties of odor cues that are associated with different phases of ethanol withdrawal influence such relapse behavior. The data suggested that cues associated with acute withdrawal or "recovery" from withdrawal may serve as modulating factors in influencing subsequent ethanol drinking behavior, and that the timing of the cues determines their consequences. Dr. F. Weiss described recent findings from animal models of relapse that suggested the efficacy of alcohol-associated contextual stimuli in eliciting alcohol-seeking behavior resembles the endurance of conditioned cue reactivity and cue-induced cocaine craving in humans. The interactive effects of stress and ethanol-related environmental stimuli were found to be dependent on concurrent activation of endogenous opioid and corticotropin-releasing factor systems. Conditioning factors (i.e., exposure to drug-associated stimuli) and stress could therefore interact to augment vulnerability to relapse. Dr. C. Drummond then addressed the clinical aspects of conditioning during alcohol withdrawal and described studies showing exposure of alcoholics to alcohol-related cues elicited greater subjective and physiological responses than exposure to neutral cues. The former responsivity showed a relationship with a measure of motivation to drink alcohol. Finally, Dr. C. Cunningham provided a summary of the concepts involved in the presentations and discussed the conditioning processes that affect behavior during and after alcohol withdrawal.
Pamela Metten and John C. Crabbe. “Alcohol withdrawal severity in inbred mouse (Mus musculus) strains.” Behavioral Neuroscience, 119, 4, Pp. 911–925. Abstract
Male mice (Mus musculus) from 15 standard inbred strains were exposed to a nearly constant concentration of ethanol (EtOH) vapor for 72 hr, averaging 1.59 +/- 0.03 mg EtOH/mL blood at withdrawal. EtOH- and air-exposed groups were tested hourly for handling-induced convulsions for 10 hr and at Hours 24 and 25. Strains differed markedly in the severity of withdrawal (after subtraction of control values), and by design these differences were independent of strain differences in EtOH metabolism. Correlation of strain mean withdrawal severity with other responses to EtOH supported previously reported genetic relationships of high EtOH withdrawal with low drinking, high conditioned taste aversion, low tolerance to EtOH-induced hypothermia, and high stimulated activity after low-dose EtOH. Also supported were the positive genetic correlations among EtOH, barbiturate, and benzodiazepine withdrawal. Sensitivity of naive mice to several chemical convulsant-induced seizures was also correlated with EtOH withdrawal.
Fulton T. Crews, Tracey Buckley, Peter R. Dodd, Gabriele Ende, Nina Foley, Clive Harper, Jun He, David Innes, El-Wui Loh, Adolph Pfefferbaum, Jian Zou, and Edith V. Sullivan. “Alcoholic neurobiology: changes in dependence and recovery.” Alcoholism, Clinical and Experimental Research, 29, 8, Pp. 1504–1513. Abstract
This article presents the proceedings of a symposium held at the meeting of the International Society for Biomedical Research on Alcoholism (ISBRA) in Mannheim, Germany, in October, 2004. Chronic alcoholism follows a fluctuating course, which provides a naturalistic experiment in vulnerability, resilience, and recovery of human neural systems in response to presence, absence, and history of the neurotoxic effects of alcoholism. Alcohol dependence is a progressive chronic disease that is associated with changes in neuroanatomy, neurophysiology, neural gene expression, psychology, and behavior. Specifically, alcohol dependence is characterized by a neuropsychological profile of mild to moderate impairment in executive functions, visuospatial abilities, and postural stability, together with relative sparing of declarative memory, language skills, and primary motor and perceptual abilities. Recovery from alcoholism is associated with a partial reversal of CNS deficits that occur in alcoholism. The reversal of deficits during recovery from alcoholism indicates that brain structure is capable of repair and restructuring in response to insult in adulthood. Indirect support of this repair model derives from studies of selective neuropsychological processes, structural and functional neuroimaging studies, and preclinical studies on degeneration and regeneration during the development of alcohol dependence and recovery form dependence. Genetics and brain regional specificity contribute to unique changes in neuropsychology and neuroanatomy in alcoholism and recovery. This symposium includes state-of-the-art presentations on changes that occur during active alcoholism as well as those that may occur during recovery-abstinence from alcohol dependence. Included are human neuroimaging and neuropsychological assessments, changes in human brain gene expression, allelic combinations of genes associated with alcohol dependence and preclinical studies investigating mechanisms of alcohol induced neurotoxicity, and neuroprogenetor cell expansion during recovery from alcohol dependence.
Amanda L. Sharpe, Natalia O. Tsivkovskaia, and Andrey E. Ryabinin. “Ataxia and c-Fos expression in mice drinking ethanol in a limited access session.” Alcoholism, Clinical and Experimental Research, 29, 8, Pp. 1419–1426. Abstract
BACKGROUND: Although previous murine studies have demonstrated ethanol self-administration resulting in blood ethanol concentrations (BECs) believed to be pharmacologically relevant, to our knowledge, no study reported to date has demonstrated intoxication via ataxia after self-administration. Thus, the goal of this study was to demonstrate ataxia and to examine changes in c-Fos expression in mice after self-administration of intoxicating doses of ethanol. METHODS: Male C57BL/6J mice were trained to drink a 10% ethanol solution during daily 30-min limited access sessions. Mice were exposed to increasing concentrations of ethanol until a 10% ethanol solution was reached. BEC and ataxia, measured as foot slips off of a balance beam, were examined after the limited access self-administration session. In a separate experiment, various brain structures from mice drinking water or ethanol were examined for changes in c-Fos expression two hr after the limited access session. RESULTS: Mice drank between 1.5 and 2 g/kg of 10% ethanol during the daily 30-min session. BECs for these mice 15 min after the limited access session ranged between 0.52 and 2.13 mg/ml. A significant increase in foot slips off a balance beam was seen immediately after ethanol consumption during the limited access session. Among mice drinking ethanol, an increase in c-Fos expression was seen in the Edinger-Westphal nucleus, and a decrease in c-Fos expression was seen in the cingulate cortex, ventral tegmental area, lateral and medial septum, CA1 region of the hippocampus, and basolateral amygdala. CONCLUSIONS: After this procedure in mice, BECs are achieved that are in a range considered pharmacologically relevant and intoxicating. Significant ataxia was observed after ethanol self-administration. Brain regions showing changes in c-Fos expression after voluntary intoxication were similar to those previously reported, suggesting that these brain regions are involved in regulating behavioral effects of alcohol intoxication.
Adam Z. Weitemier and Andrey E. Ryabinin. “Brain Region–Specific Regulation of Urocortin 1 Innervation and Corticotropin-Releasing Factor Receptor Type 2 Binding by Ethanol Exposure.” Alcoholism: Clinical and Experimental Research, 29, 9, Pp. 1610–1620. Publisher's Version Abstract
Background: Ethanol administration and consumption selectively activates the urocortin 1 (Ucn1)-expressing neurons of the Edinger-Westphal nucleus. We investigated whether repeated ethanol exposure affects Ucn1 and Ucn1-responsive corticotropin-releasing factor type-2 receptors (CRF2). Methods: Male C57BL/6J and DBA/2J mice were exposed to 2 g/kg ethanol via intraperitoneal injection once per day for 14, seven, or zero days. Ucn1 immunoreactivity was measured in the lateral septum, dorsal raphe, and Edinger-Westphal nucleus. In a separate experiment, C57BL/6J mice were exposed to ethanol for seven, one, or zero days, and CRF2 receptor binding was measured in the lateral septum and dorsal raphe by receptor autoradiography. Results: Ethanol exposure induced parallel changes in Ucn1 immunoreactive terminal fibers in the lateral septum and dorsal raphe of both strains. Seven ethanol exposures but not one ethanol exposure significantly increased CRF2 receptor binding in the dorsal raphe and slightly increased CRF2 receptor binding in the lateral septum. Conclusions: These results provide evidence that the Ucn1/CRF2 receptor system can be modified by ethanol exposure. They additionally suggest that this system may be involved in behavioral changes during alcoholism.
Zachary A. Rodd, Richard L. Bell, Victoria K. McQueen, Michelle R. Davids, Cathleen C. Hsu, James M. Murphy, Ting-Kai Li, Lawrence Lumeng, and William J. McBride. “Chronic ethanol drinking by alcohol-preferring rats increases the sensitivity of the posterior ventral tegmental area to the reinforcing effects of ethanol.” Alcoholism, Clinical and Experimental Research, 29, 3, Pp. 358–366. Abstract
BACKGROUND: The ventral tegmental area (VTA) is involved in regulating ethanol drinking, and the posterior VTA seems to be a neuroanatomical substrate that mediates the reinforcing effects of ethanol in ethanol-naive Wistar and ethanol-naive alcohol-preferring (P) rats. The objective of this study was to test the hypothesis that chronic ethanol drinking increases the sensitivity of the posterior VTA to the reinforcing effects of ethanol. METHODS: Two groups of female P rats (one given water as its sole source of fluid and the other given 24-hr free-choice access to 15% ethanol and water for at least 8 weeks) were stereotaxically implanted with guide cannulae aimed at the posterior VTA. One week after surgery, rats were placed in standard two-lever (active and inactive) operant chambers and connected to the microinfusion system. Depression of the active lever produced the infusion of 100 nl of artificial cerebrospinal fluid (CSF) or ethanol. The ethanol-naive and chronic ethanol-drinking groups were assigned to subgroups to receive artificial CSF or 25, 50, 75, or 125 mg/dl of ethanol (n = 6-9/dose/group) to self-infuse (FR1 schedule) during the 4-hr sessions given every other day. RESULTS: Compared with the infusions of artificial CSF, the control group reliably (p \textless 0.05) self-infused 75 and 125 mg/dl of ethanol but not the lower concentrations. The ethanol-drinking group had significantly (p \textless 0.05) higher self-infusions of 50, 75, and 125 mg/dl of ethanol than artificial CSF during the four acquisition sessions; the number of infusions of all three doses was higher in the ethanol-drinking group than in the ethanol-naive group. Both groups decreased responding on the active lever when artificial CSF was substituted for ethanol, and both groups demonstrated robust reinstatement of responding on the active lever when ethanol was restored. CONCLUSIONS: Chronic ethanol drinking by P rats increased the sensitivity of the posterior VTA to the reinforcing effects of ethanol.
Marisa Roberto, Michal Bajo, Elena Crawford, Samuel G. Madamba, and George R. Siggins. “Chronic Ethanol Exposure and Protracted Abstinence Alter NMDA Receptors in Central Amygdala.” Neuropsychopharmacology, 31, 5, Pp. 988–996. Publisher's Version Abstract
We recently reported that chronic ethanol treatment (CET) and early withdrawal (2–8 h) altered glutamatergic transmission at both pre- and postsynaptic sites in central nucleus of the amygdala (CeA). Acute ethanol (44 mM) inhibited the NMDA receptor (NMDAR)-mediated EPSCs (NMDA-EPSCs) more in CeA neurons from CET rats than from naïve rats and also decreased paired-pulse facilitation (PPF) of NMDA-EPSCs only in CET rats. To determine whether these CET effects persisted after prolonged withdrawal, we recorded intracellularly in rat CeA slices and measured mRNA and protein expression of CeA NMDAR subunits from CET rats and those withdrawn from ethanol for 1 or 2 weeks. At 1 week withdrawal, acute ethanol decreased evoked NMDA-EPSC amplitudes and NMDA currents induced by exogenous NMDA (20%) equally to that in naïve rats, indicating that CET effects on postsynaptic mechanisms reversed 1 week after CET cessation. However, acute ethanol still decreased PPF of NMDA-EPSCs, indicating that the acute ethanol-induced increase in glutamate release in CeA seen in CET rats was still present at this time. CET also significantly increased mRNA levels of NR1 and NR2B NMDAR subunits compared to control rats. At 1 week withdrawal, mRNA levels for NR1 and NR2B subunits were significantly decreased. These changes reversed at 2 weeks withdrawal. In Western blots, a significant increase in protein for all three subunits occurred in CeA from CET rats, but not after 1 and 2 weeks of withdrawal. These data indicate that CET induces reversible neuroadaptations in synaptic function, gene expression, and protein composition of NMDAR at CeA synapses.
Douglas B. Matthews, Sanjiv V. Bhave, John K. Belknap, Cynthia Brittingham, Elissa J. Chesler, Robert J. Hitzemann, Paula L. Hoffmann, Lu Lu, Shannon McWeeney, Michael F. Miles, Boris Tabakoff, and Robert W. Williams. “Complex genetics of interactions of alcohol and CNS function and behavior.” Alcoholism, Clinical and Experimental Research, 29, 9, Pp. 1706–1719. Abstract
This work summarizes the proceedings of a symposium at the 2004 RSA Meeting in Vancouver, Canada. The organizers were R. W. Williams and D. B. Matthews; the Chair was M. F. Miles. The presentations were (1) WebQTL: A resource for analysis of gene expression variation and the genetic dissection of alcohol related phenotypes, by E. J. Chesler, (2) The marriage of microarray and qtl analyses: what's to gain, by J. K. Belknap, (3) Use of WebQTL to identify QTLs associated with footshock stress and ethanol related behaviors, by D. B. Matthews, (4) A high throughput strategy for the detection of quantitative trait genes, by R. J. Hitzemann, and (5) The use of gene arrays in conjunction with transgenic and selected animals to understand anxiety in alcoholism, by. B. Tabakoff.
Monica Lisa Berlanga, Taylor Kathryn Simpson, and Adriana Angelica Alcantara. “Dopamine D5 receptor localization on cholinergic neurons of the rat forebrain and diencephalon: a potential neuroanatomical substrate involved in mediating dopaminergic influences on acetylcholine release.” The Journal of Comparative Neurology, 492, 1, Pp. 34–49. Abstract
The study of dopaminergic influences on acetylcholine release is especially useful for the understanding of a wide range of brain functions and neurological disorders, including schizophrenia, Parkinson's disease, Alzheimer's disease, and drug addiction. These disorders are characterized by a neurochemical imbalance of a variety of neurotransmitter systems, including the dopamine and acetylcholine systems. Dopamine modulates acetylcholine levels in the brain by binding to dopamine receptors located directly on cholinergic cells. The dopamine D5 receptor, a D1-class receptor subtype, potentiates acetylcholine release and has been investigated as a possible substrate underlying a variety of brain functions and clinical disorders. This receptor subtype, therefore, may prove to be a putative target for pharmacotherapeutic strategies and cognitive-behavioral treatments aimed at treating a variety of neurological disorders. The present study investigated whether cholinergic cells in the dopamine targeted areas of the cerebral cortex, striatum, basal forebrain, and diencephalon express the dopamine D5 receptor. These receptors were localized on cholinergic neurons with dual labeling immunoperoxidase or immunofluorescence procedures using antibodies directed against choline acetyltransferase (ChAT) and the dopamine D5 receptor. Results from this study support previous findings indicating that striatal cholinergic interneurons express the dopamine D5 receptor. In addition, cholinergic neurons in other critical brain areas also show dopamine D5 receptor expression. Dopamine D5 receptors were localized on the somata, dendrites, and axons of cholinergic cells in each of the brain areas examined. These findings support the functional importance of the dopamine D5 receptor in the modulation of acetylcholine release throughout the brain.
Timothy Donohue, Paula L. Hoffman, and Boris Tabakoff. “Effect of ethanol on DARPP-32 phosphorylation in transgenic mice that express human type VII adenylyl cyclase in brain.” Alcoholism, Clinical and Experimental Research, 29, 3, Pp. 310–316. Abstract
BACKGROUND: Dopamine and cyclic adenosine monophosphate-regulated phosphoprotein of molecular weight 32 kDa (DARPP-32) is a bidirectional signaling protein found in dopaminergically innervated brain areas. The characteristics and direction of DARPP-32 effects are regulated by phosphorylation of this protein. Phosphorylation of DARPP-32 on threonine-34 (T34) is regulated through the activation of dopamine (D1) receptors and stimulation of adenylyl cyclase (AC) and protein kinase A activity and by calcineurin. Phosphorylation of DARPP-32 on threonine-75 (T75) is regulated by cyclin-dependent kinase 5 and protein phosphatase 2A. DARPP-32 has been implicated in the motivational effects of ethanol. METHODS: The authors characterized transgenic mice that overexpress an ethanol-sensitive isoform of AC (AC7) in brain by measuring basal and ethanol-modulated DARPP-32 phosphorylation. Phosphorylated and total DARPP-32 were measured by immunoblotting in brain areas associated with the motivational and anxiolytic effects of ethanol (nucleus accumbens, striatum, and amygdala). RESULTS: AC7 transgenic mice had higher basal levels of T34 DARPP-32 than wild-type mice in striatum and amygdala, whereas basal levels of T75 DARPP-32 did not differ between wild-type and transgenic mice. Ethanol administration increased T34 DARPP-32 in nucleus accumbens and amygdala (but not in the striatum) of wild-type and transgenic mice (with a greater effect in amygdala of transgenic mice than wild-type mice). Ethanol administration increased T75 DARPP-32 in amygdala of only the wild-type mice and in nucleus accumbens and striatum of both the transgenic and wild-type mice. CONCLUSIONS: The effect of ethanol on the balance of DARPP-32 phosphorylation, especially in amygdala of wild-type versus transgenic mice, may contribute to differential motivational effects of ethanol in these animals.
Helen J. K. Sable, Zachary A. Rodd, Richard L. Bell, Jonathan A. Schultz, Larry Lumeng, and William J. McBride. “Effects of ethanol drinking on central nervous system functional activity of alcohol-preferring rats.” Alcohol, 35, 2, Pp. 129–135. Publisher's Version Abstract
The [14C]-2-deoxyglucose (2-DG) technique was used to assess the rates of local cerebral glucose utilization (LCGU) in key limbic, cerebral cortical, hippocampal, basal ganglionic, and subcortical regions of alcohol-preferring (P) rats following chronic 24-h free-choice ethanol drinking. Adult male P rats were submitted to (1) 8 continuous weeks of two-bottle access to 15% ethanol and water (E-C group); (2) 8 weeks of identical two-bottle access followed by 2 weeks of ethanol deprivation (E-D group); (3) cycles of 2 weeks of two-bottle ethanol access and 2 weeks of deprivation, repeated for four cycles (E-RD group); or (4) water only treatment [ethanol-naive group (E-N group)]. A single pulse of [14C]-2-DG (125 μCi/kg) was administered via a venous catheter, and timed arterial blood samples were collected over 45 min and later assayed for plasma glucose and [14C]-2-DG concentrations. Quantitative autoradiography was used to determine [14C] densities, and LCGU values were calculated. With the exception of a few small differences in the hippocampus, no significant differences were found in any of the central nervous system (CNS) regions examined among the four experimental groups of P rats. Animals in the E-D group had lower LCGU rates in the anterior hippocampal CA1 subregion than animals in the E-N, E-C, and E-RD groups. In the anterior hippocampal CA3 subregion and the anterior hippocampal dentate gyrus, the E-D group had significantly lower LCGU rates than the E-RD group. Overall, the results of this study indicate that 24-h ethanol-drinking experience has little effect on CNS functional neuronal activity in P rats.
Vladimir I. Chefer, Traci Czyzyk, Elizabeth A. Bolan, Jose Moron, John E. Pintar, and Toni S. Shippenberg. “Endogenous kappa-opioid receptor systems regulate mesoaccumbal dopamine dynamics and vulnerability to cocaine.” The Journal of Neuroscience: The Official Journal of the Society for Neuroscience, 25, 20, Pp. 5029–5037. Abstract
Genetic and pharmacological approaches were used to examine kappa-opioid receptor (KOR-1) regulation of dopamine (DA) dynamics in the nucleus accumbens and vulnerability to cocaine. Microdialysis revealed that basal DA release and DA extraction fraction (Ed), an indirect measure of DA uptake, are enhanced in KOR-1 knock-out mice. Analysis of DA uptake revealed a decreased Km but unchanged Vmax in knock-outs. Knock-out mice exhibited an augmented locomotor response to cocaine, which did not differ from that of wild-types administered a behavioral sensitizing cocaine treatment. The ability of cocaine to increase DA was enhanced in knock-outs, whereas c-fos induction was decreased. Although repeated cocaine administration to wild types produced behavioral sensitization, knock-outs exhibited no additional enhancement of behavior. Administration of the long-acting KOR antagonist nor-binaltorphimine to wild-type mice increased DA dynamics. However, the effects varied with the duration of KOR-1 blockade. Basal DA release was increased whereas Ed was unaltered after 1 h blockade. After 24 h, release and Ed were increased. The behavioral and neurochemical effects of cocaine were enhanced at both time points. These data demonstrate the existence of an endogenous KOR-1 system that tonically inhibits mesoaccumbal DA neurotransmission. Its loss induces neuroadaptations characteristic of "cocaine-sensitized" animals, indicating a critical role of KOR-1 in attenuating responsiveness to cocaine. The increased DA uptake after pharmacological inactivation or gene deletion highlights the plasticity of mesoaccumbal DA neurons and suggests that loss of KOR-1 and the resultant disinhibition of DA neurons trigger short- and long-term DA transporter adaptations that maintain normal DA levels, despite enhanced release.
V. F. Turek and A. E. Ryabinin. “Ethanol versus lipopolysaccharide-induced hypothermia: involvement of urocortin.” Neuroscience, 133, 4, Pp. 1021–1028. Abstract
The urocortin1 (Ucn1) neurons of the mid-brain-localized Edinger-Westphal nucleus (EW) are robustly responsive to ethanol (EtOH) administration, and send projections to the dorsal raphe nucleus (DRN), which contains corticotropin-releasing factor type 2 receptors (CRF2) that are responsive to Ucn1. In addition, the DRN has been shown to be involved in regulation of body temperature, a function greatly affected by EtOH administration. The goal of the present study was to identify the role that the urocortinergic projections from the EW to the DRN have in mediating EtOH-induced and lipopolysaccharide (LPS)-induced hypothermia. Male C57BL6/J mice were used. Groups of mice underwent cannulation of the DRN, and then received i.p. injections of EtOH (2g/kg) or LPS (600 microg/kg or 400 microg/kg), followed by intra-DRN injections of artificial cerebrospinal fluid (aCSF) or anti-sauvagine (aSVG) (55 pmol), a CRF2 antagonist. Separate groups of mice received single intra-DRN injections of Ucn1 (20 pmol), CRF (20 pmol) or aCSF. For all experiments, core temperatures were monitored rectally every 30 min for several hours post-injection. Both EtOH and LPS induced hypothermia, and aSVG significantly attenuated this effect after EtOH; however, there was no significant attenuation of hypothermia after either dose of LPS. Ucn1 injection also caused hypothermia, while CRF injection did not. These data demonstrate that EtOH-induced hypothermia, but not LPS-induced hypothermia, may involve Ucn1 from EW acting at CRF2 receptors in the DRN.
Justin S. Rhodes, Karyn Best, John K. Belknap, Deborah A. Finn, and John C. Crabbe. “Evaluation of a simple model of ethanol drinking to intoxication in C57BL/6J mice.” Physiology & Behavior, 84, 1, Pp. 53–63. Abstract
Because of intrinsic differences between humans and mice, no single mouse model can represent all features of a complex human trait such as alcoholism. It is therefore necessary to develop partial models. One important feature is drinking to the point where blood ethanol concentration (BEC) reaches levels that have measurable affects on physiology and/or behavior (\textgreater1.0 mg ethanol/ml blood). Most models currently in use examine relative oral self-administration from a bottle containing alcohol versus one containing water (two-bottle preference drinking), or oral operant self-administration. In these procedures, it is not clear when or if the animals drink to pharmacologically significant levels because the drinking is episodic and often occurs over a 24-h period. The aim of this study was to identify the optimal parameters and evaluate the reliability of a very simple procedure, taking advantage of a mouse genotype (C57BL/6J) that is known to drink large quantities of ethanol. We exchanged for the water bottle a solution containing ethanol in tap water for a limited period, early in the dark cycle, in the home cage. Mice regularly drank sufficient ethanol to achieve BEC\textgreater1.0 mg ethanol/ml blood. The concentration of ethanol offered (10%, 20% or 30%) did not affect consumption in g ethanol/kg body weight. The highest average BEC ( approximately 1.6 mg/ml) occurred when the water-to-ethanol switch occurred 3 h into the dark cycle, and when the ethanol was offered for 4 rather than 2 h. Ethanol consumption was consistent within individual mice, and reliably predicted BEC after the period of ethanol access. C57BL/6J mice from three sources provided equivalent data, while DBA/2J mice drank much less than C57BL/6J in this test. We discuss advantages of the model for high-throughput screening assays where the goal is to find other genotypes of mice that drink excessively, or to screen drugs for their efficacy in blocking excessive drinking.
Restraint stress, lipopolysaccharide (LPS), and ethanol (EtOH) administration have all been found to induce c-Fos in the brain, and to cause hypothermia. The present study was designed to assess whether the c-Fos expression that occurs in the Edinger–Westphal nucleus (EW) after EtOH administration is independent of the hypothermia or any stress effects that occur. To test this, we used restraint stress and LPS in addition to EtOH, and also examined two control areas, the dorsal raphe nucleus (DRN) and the periaqueductal gray (PAG), in addition to EW. Male C57BL6/J mice were used. Groups of mice received intraperitoneal (IP) injections of EtOH (2 g/kg), LPS (600 μg/kg or 50 μg/kg), or saline. A separate group of mice received no injection, but were placed in plastic restrainers for the entirety of the experiment. For all groups, core temperatures were monitored rectally every 30 min for 3 h postinjection, after which, the animals were sacrificed. Then, the number of Fos-positive cells in the brain regions of the EW, DRN, and PAG was quantified. Both EtOH and restraint stress induced a transient hypothermia, where core temperature (Tc) declined immediately and then rose again. Both doses of LPS induced a slower developing, longer lasting hypothermia, while saline had no effect on Tc. Only EtOH induced a significant amount of c-Fos in EW, while both doses of LPS and restraint stress induced c-Fos in DRN, and only restraint stress caused induction in PAG. These data demonstrate that activation of EW after EtOH is unrelated to hypothermia or stress.
Susan E. Bergeson, Ari E. Berman, Peter R. Dodd, Howard J. Edenberg, Robert J. Hitzemann, Joanne M. Lewohl, Kerrie H. Lodowski, and Wolfgang H. Sommer. “Expression Profiling in Alcoholism Research.” Alcoholism, clinical and experimental research, 29, 6, Pp. 1066–1073. Publisher's Version Abstract
This article represents the proceedings of a symposium at the 2004 International Society for Biomedical Research on Alcoholism in Mannheim, Germany, organized and co-chaired by Susan E. Bergeson and Wolfgang Sommer. The presentations and presenter were (1) Gene Expression in Brains of Alcohol-Preferring and Non-Preferring Rats, by Howard J. Edenberg (2) Candidate Treatment Targets for Alcoholism: Leads from Functional Genomics Approaches, by Wolfgang Sommer (3) Microarray Analysis of Acute and Chronic Alcohol Response in Brain, by Susan E. Bergeson (4) On the Integration of QTL and Gene Expression Analysis, by Robert J. Hitzemann (5) Microarray and Proteomic Analysis of the Human Alcoholic Brain, by Peter R. Dodd.
Paula Hoffman and Boris Tabakoff. “Gene expression in animals with different acute responses to ethanol.” Addiction Biology, 10, 1, Pp. 63–69. Abstract
The genetic and environmental contributions to differences in response to ethanol have been examined widely using inbred strains, selected lines and genetically engineered (transgenic and 'knock-out') animals. In addition, recombinant inbred strains have been used to identify QTLs (chromosomal regions) associated with particular responses to ethanol. If the polymorphism that underlies such a QTL is localized within the regulatory region of a gene, it could alter the level or stability of the gene product (transcript). This possibility can be addressed by measuring mRNA levels in brains (or other tissue) of inbred or selected lines of animals using DNA microarray technology. In this paper, we review microarray studies conducted in animals that differ in their responses to ethanol. The results of these studies point out the critical nature of the experimental design, statistical analyses and 'filtering' procedures for producing interpretable data and identifying candidate genes. In particular, the determination of differentially expressed genes between selected lines of animals, and the localization of the differentially expressed genes within QTLs for the selected phenotype, dramatically increases the probability of identifying genes that contribute to that phenotype through differential expression. Microarray analysis can also be used to assess changes in gene expression that accompany transgene introduction and/or gene 'knock-out', which may modulate the influence of the targeted gene on behaviour.
H. J. Edenberg, W. N. Strother, J. N. McClintick, H. Tian, M. Stephens, R. E. Jerome, L. Lumeng, T.-K. Li, and W. J. McBride. “Gene expression in the hippocampus of inbred alcohol-preferring and -nonpreferring rats.” Genes, Brain, and Behavior, 4, 1, Pp. 20–30. Abstract
The hippocampus is sensitive to the effects of ethanol and appears to have a role in the development of alcohol tolerance. The objective of this study was to test the hypothesis that there are innate differences in gene expression in the hippocampus of inbred alcohol-preferring (iP) and -nonpreferring (iNP) rats that may contribute to differences in sensitivity to ethanol and/or in the development of tolerance. Affymetrix microarrays were used to measure gene expression in the hippocampus of alcohol-naive male iP and iNP rats in two experiments (n=4 and 6 per strain in the two experiments). Combining data from the two experiments, there were 137 probesets representing 129 genes that significantly differed (P \textless or = 0.01); 62 probesets differed at P \textless or = 0.001. Among the 36% of the genes that were expressed more in the iP than iNP rat at this level of significance, many were involved in cell growth and adhesion, cellular stress reduction and anti-oxidation, protein trafficking, regulation of gene expression, synaptic function and metabolism. Among the 64% of the genes that had lower expression in the hippocampus of iP than iNP rats were genes involved in metabolic pathways, cellular signaling systems, protein trafficking, cell death and neurotransmission. Overall, the data indicate that there are significant innate differences in gene expression in the hippocampus between iP and iNP rats, some of which might contribute to the differences observed in the development of alcohol tolerance between the selectively bred P and NP lines.
Justin S. Rhodes and John C. Crabbe. “Gene expression induced by drugs of abuse.” Current Opinion in Pharmacology, 5, 1, Pp. 26–33. Abstract
The transition from infrequent drug use to addiction (i.e. the loss of control over consumption of a drug) probably involves changes in gene expression that restructure neural circuits in the brain. The number of genes that have been demonstrated to change expression in response to drugs has increased rapidly in recent years owing to microarray technology, which allows measurement of thousands of genes at one time. It is now important to identify which of these changes are causally related to the compulsive behavior associated with drug addiction, and which are non-specific changes related to general features of arousal or other physiological responses (e.g. stress, altered body temperature or energy metabolism).
Karen K. Szumlinski, Kevin D. Lominac, Erik B. Oleson, Jennifer K. Walker, Ashley Mason, Marlin H. Dehoff, Matthias Klugmann, Matthias Klugman, Stephanie Cagle, Kristine Welt, Matthew During, Paul F. Worley, Lawrence D. Middaugh, and Peter W. Kalivas. “Homer2 is necessary for EtOH-induced neuroplasticity.” The Journal of Neuroscience: The Official Journal of the Society for Neuroscience, 25, 30, Pp. 7054–7061. Abstract
Homer proteins are integral to the assembly of proteins regulating glutamate signaling and synaptic plasticity. Constitutive Homer2 gene deletion [knock-out (KO)] and rescue with adeno-associated viral (AAV) transfection of Homer2b was used to demonstrate the importance of Homer proteins in neuroplasticity produced by repeated ethanol (EtOH) administration. Homer2 KO mice avoided drinking high concentrations of EtOH and did not develop place preference or locomotor sensitization after repeated EtOH administration. The deficient behavioral plasticity to EtOH after Homer2 deletion was paralleled by a lack of augmentation in the rise in extracellular dopamine and glutamate elicited by repeated EtOH injections. The genotypic differences in EtOH-induced change in behavior and neurochemistry were essentially reversed by AAV-mediated transfection of Homer2b into accumbens cells including, differences in EtOH preference, locomotor sensitization, and EtOH-induced elevations in extracellular glutamate and dopamine. These data demonstrate a necessary and active role for accumbens Homer2 expression in regulating EtOH-induced behavioral and cellular neuroplasticity.

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