0145-6008/03/2702-0149$03.00/0 ALCOHOLISM: CLINICAL AND EXPERIMENTAL RESEARCH
Vol. 27, No. 2 February 2003
Neuropeptide Y and Alcoholism: Genetic, Molecular, and Pharmacological Evidence Subhash C. Pandey, Lucinda G. Carr, Markus Heilig, Erkki Ilveskoski, and Todd E. Thiele
This article presents the proceedings of a symposium presented at the combined meeting of the Research Society on Alcoholism and the International Society for Biomedical Research on Alcoholism, held in San Francisco, CA, in June 2002. The organizers and chairpersons were Subhash C. Pandey and Todd E. Thiele. The presentations were (1) Altered ethanol-induced sedation and ethanol drinking in mutant mice lacking specific NPY receptor, by Todd E. Thiele; (2) NPY in P and NP rats: polymorphism and mRNA expression, by Lucinda G. Carr; (3) The cAMP-dependent PKA in the central amygdala regulates alcohol intake through NPY gene, by Subhash C. Pandey; (4) Involvement of NPY in alcohol dependence: from animal models to human genetics, by Markus Heilig; and (5) Association of neuropeptide Y polymorphism with the occurrence of type 1 and type 2 alcoholism, by Erkki Ilveskoski. Key Words: Neuropeptide Y, Knock-Out, CREB, Polymorphism, Alcoholism.
N
EUROPEPTIDE Y (NPY) is a 36 –amino acid neuromodulator and is one of the most abundant peptides in the brain (Wettstein et al., 1995). NPY acts through at least five receptor subtypes—namely, the Y1, Y2, Y4, Y5, and Y6 receptors—all of which are functionally coupled to the cyclic adenosine monophosphate (cAMP) second-messenger pathway (Palmiter et al., 1998). NPY has been implicated in several behaviors, including anxiety and depression (Heilig and Widerlov, 1995). During recent years, a growing body of genetic, molecular, and pharmacological evidence has accumulated to suggest that NPY is an important neurobiological substrate for the predisposition to alcohol-seeking behaviors. Rats selectively bred for alcohol preference have altered levels of NPY in several brain regions compared with alcoholnonpreferring (NP) rats (Ehlers et al., 1998; Hwang et al., 1999), and genetic linkage analysis of alcohol-preferring (P) rats identified a chromosomal region that includes the NPY
From the Department of Psychiatry, Psychiatric Institute, University of Illinois, and the Veterans Affairs Chicago Health Care System (West Side Division), Chicago, Illinois (SCP); Indiana University School of Medicine, Indianapolis, Indiana (LGC); Division of Psychiatry, Neurotec Department, Karolinska Institute, Huddinge Hospital, Stockholm, Sweden (MH); Department of Forensic Medicine, Medical School, University of Tampere, Helsinki, Finland (EI); and Department of Psychology, University of North Carolina, Chapel Hill, North Carolina (TET). Received for publication September 19, 2002; accepted October 4, 2002. Supported by NIH Grants AA10005 and AA13341 and a VA merit grant (SCP); NIH Grants AA10707 and AA07611 (LGC); Swedish Medical Research Council Grant K2001-21X-10872 (MH); The Finnish Foundation of Alcohol Research (EI); and NIH Grants AA00258 and AA13573 (TET). Reprint requests: Subhash C. Pandey, PhD, Department of Psychiatry, University of Illinois at Chicago, and VA Chicago Health Care System (West Side Division), 820 S. Damen Ave. (M/C 151), Chicago, IL 60612; Fax: 312-569-8114; E-mail:
[email protected]. Copyright © 2003 by the Research Society on Alcoholism. DOI: 10.1097/01.ALC.0000052706.21367.0E Alcohol Clin Exp Res, Vol 27, No 2, 2003: pp 149–154
gene (Bice et al., 1998; Carr et al., 1998). Resistance to the intoxicating effects of ethanol and ethanol consumption was found to be low in transgenic mice that overexpress NPY, whereas mutant mice lacking normal production of NPY were found to show high ethanol drinking and high resistance to ethanol-induced sedation when maintained on a specific genetic background (Thiele et al., 1998, 2000). Finally, central infusion of exogenous NPY has been shown to modulate ethanol drinking under some circumstances (Badia-Elder et al., 2001; Kelley et al., 2001), but not in others (Slawecki et al., 2000). Recent evidence indicates that NPY systems may also be involved in alcohol dependence (Mayfield et al., 2002; Roy and Pandey, 2002; Woldbye et al., 2002). The purpose of this symposium was to bring together investigators to present the most current research implicating the NPY system in alcoholism. It was our intention that this symposium would both provide a framework for a better understanding of the role of NPY in neurobiological responses to ethanol and stimulate interest and further research in this relatively new field of study. The individuals who graciously agreed to participate in this symposium came from outstanding and respected research institutions both in Europe and the United States and presented findings obtained from a range of analytical methods, including genetic, molecular, pharmacological, and behavioral techniques. ALTERED ETHANOL-INDUCED SEDATION AND ETHANOL DRINKING IN MUTANT MICE LACKING SPECIFIC NPY RECEPTOR
Todd E. Thiele The purpose of the studies described by Dr. Todd E. Thiele was to determine the potential role of NPY receptors in modulating voluntary ethanol consumption and neurobiolog149
150
ical responses to ethanol by using mutant mice lacking specific central NPY receptors. They therefore studied alcohol consumption by Y1⫺/⫺ mice (maintained on a pure C57BL/6 inbred background), which completely lack Y1 receptor expression as a result of targeted gene disruption, and by normal Y1⫹/⫹ mice. Y1⫺/⫺ mice grow and reproduce at normal rates despite slightly diminished daily food intake and a reduced refeeding response to starvation. However, these animals develop late-onset obesity due to low energy expenditure (Pedrazzini et al., 1998). It was found that male Y1⫺/⫺ mice showed increased consumption of solutions containing 3, 6, and 10% (v/v) ethanol when compared with wild-type (Y1⫹/⫹) mice. Female Y1⫺/⫺ mice showed increased consumption of a 10% ethanol solution. However, Y1⫺/⫺ mice showed normal consumption of solutions containing either sucrose or quinine. Relative to Y1⫹/⫹ mice, male Y1⫺/⫺ mice were found to be less sensitive to the sedative effects of ethanol 3.5 and 4.0 g/kg body weight, as measured by more rapid recovery from ethanol-induced sleep, even though plasma ethanol levels did not differ significantly between the genotypes after injection of a 3.5 g/kg dose. Finally, male Y1⫺/⫺ mice showed normal ethanol-induced ataxia on the rotarod test after administration of a 2.5 g/kg dose (Thiele et al., 2002). Evidence suggests that the Y2 receptor is a presynaptic autoreceptor and inhibits NPY release (Naveilhan et al., 1998). Dr. Thiele and his group also studied mutant mice lacking the Y2 receptor that were maintained on a mixed 50% 129/SvJ ⫻ 50% BALB/c background. These mice have been shown to have increased food intake, body weight, and fat production but to have a normal response to centrally infused NPY (Naveilhan et al., 1999). It was hypothesized that if presynaptic Y2 receptors are involved with modulating voluntary ethanol consumption and sensitivity, the Y2⫺/⫺ mice should exhibit ethanol-related phenotypes opposite to those found with the Y1⫺/⫺ mice. Thus, an absence of presynaptic inhibition of NPY release in Y2⫺/⫺ mice would augment NPY signaling, rendering mice with a similar phenotype as NPY-overexpressing mice. It was found that relative to wild-type (Y2⫹/⫹) mice, the Y2⫺/⫺ mice drank significantly less of solutions containing 3 and 6% ethanol and had significantly lower ethanol preference ratios at each concentration tested. However, Y2⫺/⫺ mice showed normal consumption of solutions containing either sucrose or quinine, normal time to recover from ethanolinduced sedation after 3.0 or 3.5 g/kg doses, and normal metabolism of ethanol after injection of a 3.0 g/kg dose. Y5⫺/⫺ mice show late-onset obesity and increased food intake, have reduced sensitivity to NPY, and are seizure prone (Marsh et al., 1999). It was also found that Y5⫺/⫺ mice maintained on a pure 129/SvEv genetic background showed normal voluntary consumption of solutions containing 3, 6, 10, and 20% (v/v) ethanol but had increased sleep time after administration of ethanol 2.5 or 3.0 g/kg. However, the Y5⫺/⫺ mice also showed high plasma ethanol levels relative to wild-type mice after injection of a 3.0 g/kg dose. In summary, these data suggest that voluntary con-
PANDEY ET AL.
sumption of ethanol is modulated by Y1 and Y2 receptors and that ethanol-induced sedation is modulated by Y1 and, perhaps, Y5 receptors. NPY IN P AND NP RATS: POLYMORPHISM AND MRNA EXPRESSION
Lucinda G. Carr Several lines of evidence suggest that NPY may play a role in the drinking behavior of P rats (Badia-Elder et al., 2001; Carr et al., 1998; Hwang et al., 1999). To determine whether there were differences in NPY receptor levels between the P and NP rats, the ligands 125I-Leu31Pro34-Pyy and 125IPYY3-3 were used to detect Y1 and Y2 receptor binding, respectively. Dr. Carr presented data that demonstrated significantly higher Y1 receptor binding in the dentate gyrus, the CA1 (cornu ammonis 1) hippocampus molecular layer, and the lateral amygdaloid complex of the P rats compared with NP rats. Y2 receptor binding differences were observed in the lateral septal nucleus and paraventricular nucleus, with P rats having higher binding in the lateral septal nucleus and lower binding in the paraventricular nucleus than NP rats. Dr. Carr and her group also found that intracerebroventricular administration of NPY is able to block the increase in alcohol drinking exhibited by P rats after alcohol deprivation. NPY blocked the reacquisition of ethanol drinking at both 4 and 24 hr after a period of ethanol deprivation. Interestingly, alcohol drinking remained low for 5 days after alcohol reinstatement in those rats that received NPY. Quantitative real-time polymerase chain reaction was used to determine whether there were differences in NPY messenger RNA (mRNA) expression between the P and NP rats 1 hr after IP injection with saline or ethanol. Preliminary data indicated that there was not a significant difference in NPY mRNA levels in the frontal cortex, hippocampus, or amygdala between P and NP alcoholnaïve rats. After ethanol treatment, there was a significant decrease in NPY mRNA in the amygdala of P rats, but not NP rats, and no difference in mRNA levels was observed between the rat lines in the cortex or hippocampus. To determine whether the differences in NPY levels between P and NP rats were due to a polymorphism in the prepro-NPY gene, the four exons and portions of the noncoding regions of the gene were sequenced. Three polymorphisms were identified: two in introns and one in the 3' untranslated region. The luciferase gene is being used as a reporter to assess whether these alterations in the preproNPY gene are involved in differential NPY expression. THE CAMP-DEPENDENT PKA IN THE CENTRAL AMYGDALA REGULATES ALCOHOL INTAKE THROUGH NPY GENE
Subhash C. Pandey Pandey et al. (1999) reported earlier that the expression and phosphorylation of cAMP-responsive element-binding
151
NEUROPEPTIDE Y AND ALCOHOLISM
(CREB) protein was lower in the amygdala of P rats as compared with NP rats. Dr. Pandey and his group extended these findings and observed that CREB expression and phosphorylation, as well as expression of NPY, were lower in the central and medial, but not in the basolateral, amygdala of P rats as compared with NP rats. They also observed that P rats were anxious and consumed larger amounts of alcohol as compared with NP rats. These results suggest the possibility that decreased CREB activity and NPY expression in the central and medial amygdala could be associated with the innate anxiety and excessive alcohol-drinking behaviors of P rats. If this is the case, then decreasing the phosphorylation of CREB in the central amygdala by inhibiting protein kinase A (PKA) may provoke anxiety and promote alcohol intake even in an unselected stock of Sprague-Dawley rats (Pandey et al., 2003). To examine this notion, we implanted bilateral cannulae targeting the central or basolateral amygdaloid nuclei of rats, and after 1 week of recovery, we injected 0.5 l of artificial cerebrospinal fluid (aCSF) and 0.5 l of 40 nmol of PKA inhibitor (Rp-cAMPS; adenosine 3'5' cyclic monophosphorothioate, Rp isomer) bilaterally into the central or basolateral amygdala (BLA). One hour after infusion of aCSF or PKA inhibitor, open- and closed-arm activities were measured by using the elevated plus maze test. It was found that rats microinfused with PKA inhibitor into the central nucleus of the amygdala (CeA) showed a significant reduction in the percentage of time spent on the open arms and in the percentage of open-arm entries compared with aCSFmicroinfused rats. PKA inhibitor infusion into the BLA had no effects on open- or closed-arm activities of rats using the elevated plus maze test. The total number of entries (overall general activity) of Rp-cAMPS–infused rats (all groups) was similar to that of aCSF-infused rats. These results suggest that intracentral, but not basolateral, amygdaloid infusion of PKA inhibitor induces an anxiety state in rats. Immediately after measurements of anxiety, brains were used to examine the cellular expression of total CREB and phosphorylated CREB (p-CREB) in the brain of aCSF- or PKA inhibitor–infused (1 hr after infusion) rats as a measure of in vivo PKA activity. It was found that Rp-cAMPS infusion (1 hr after the infusion) into CeA also produced decreased CREB phosphorylation without modulation of total CREB protein. Injection of PKA inhibitor into the BLA also significantly decreased the protein levels of p-CREB, but not total CREB protein, as compared with aCSF-infused rats. To determine whether decreased PKAdependent CREB phosphorylation leads to the decreased expression of cAMP-inducible genes, the cellular expression of NPY in the CeA of aCSF- or PKA inhibitor–infused rats by using gold immunolabeling and in situ polymerase chain reaction was examined. It was found that infusion of PKA inhibitor into CeA significantly decreased the protein and mRNA levels of NPY. These results indicate that decreased CREB function in the CeA, but not in the BLA, may contribute to the development of anxiety.
Because decreased CREB phosphorylation in the CeA is correlated with the development of anxiety and because high anxiety levels promote alcohol preference, Dr. Pandey’s group also measured alcohol preference in rats by using the two-bottle free-choice paradigm. Bilateral infusions of a PKA inhibitor (0.5 l of 20 or 40 nmol of Rp-cAMPS) or aCSF (0.5 l) into the CeA were performed once daily for 3 days. Rats were provided with 7% ethanol solution in one of the bottles and water in the other bottle during these 3 days. The mean percentage of ethanol or water intake of their total fluid intake for 3 days was calculated. It was found that PKA inhibitor (both doses of Rp-cAMPS)–infused rats consumed significantly more ethanol solution and less water compared with aCSF-infused rats. The total fluid intake (ml/day) was similar in both groups. Interestingly, when the infusion of PKA inhibitor or aCSF was stopped, the alcohol intake in the PKAinhibitor group of rats on the third day after the postinfusion period was similar to that of the aCSF group of rats. These results indicate that in the central amygdala, the decreased function of PKA increases the preference to alcohol over water without modulating the total fluid intake of rats. This effect is specific to CeA, because infusion of PKA inhibitor (0.5 l of 40 nmol Rp-cAMPS) into the BLA has no effects on the alcohol intake of rats. The effects of an infusion of PKA activator (0.5 l of 80 nmol Sp-cAMPS; adenosine 3',5'cyclic monophosphorothioate, Sp isomer) into the CeA on alcohol intake were also examined. It was found that Sp-cAMPS (alone) infusion had no effects on alcohol intake, but when it was coinfused with PKA inhibitor (infused 15 min before 0.5 l of 40 nmol Rp-cAMPS infusion), it significantly antagonized the PKA inhibitor–induced increase in the alcohol intake of rats. This suggests that the increase in alcohol preference may be related to decreased CREB phosphorylation in the CeA. The possibility that decreases in PKA-dependent CREB phosphorylation may be involved in alcohol preference via the reduction of NPY expression in the CeA was also examined. It was found that NPY (0.5 l of 100 pmol) infusion (once daily for 3 days) alone into the CeA had no effects on alcohol intake, but coinfusing NPY with PKA inhibitor (0.5 l of 40 nmol Rp-cAMPS) significantly antagonized the PKA inhibitor–induced increase in the alcohol intake of rats. These results indicate that decreased CREB phosphorylation-induced decreased expression of NPY in the CeA may be associated with high alcohol intake. INVOLVEMENT OF NPY IN ALCOHOL DEPENDENCE: FROM ANIMAL MODELS TO HUMAN GENETICS
Markus Heilig An extensive line of research indicates that, in addition to other brain-mediated effects, central NPY modulates emotionality, stress responses, and experimental anxiety
152
(Heilig and Widerlov, 1995). In contrast to many other compounds, NPY’s antianxiety actions are found in all animal models in which they have been tested, indicating that NPY might act on a common core process of emotionality. Pharmacological antianxiety effects of NPY can be produced through activation of NPY Y1 receptors within the amygdala, but other brain areas, notably the hippocampus, are also likely to be involved (Heilig and Thorsell, 2002). Dr. Heilig and his group postulated that long-term NPY overexpression in relevant brain areas should render animals behaviorally insensitive to stress. This hypothesis has been supported by findings in transgenic rats with overexpression of NPY within the hippocampal CA1 region. These subjects showed normal behavior in a range of behavioral tests, including spontaneous locomotion and exploration, as well as spontaneous exploration of the elevated plus maze. However, when a stressor was introduced, the expected anxiogenic effects of NPY were absent in the transgenic subjects. Also, the fear-mediated behavioral suppression normally seen in the Vogel conflict test was entirely absent (Thorsell et al., 2000). Consumption of alcohol is thought to be driven in part by the anxiolytic-like properties of this compound, yet normal alcohol consumption was found in the NPY transgenic rats. A possible reason for this observation might be that NPY or NPY receptor expression and/or function in brain areas other than the hippocampus may be critical for controlling alcohol intake. Complex involvement of NPY in this mechanism is also indicated by a lack of effect of central NPY administration on alcohol intake in normal Wistar rats. Dr. Heilig’s group therefore analyzed the expression of NPY, Y1, and Y2 receptors in the AA (alcohol-accepting) rat line, as well as in alcohol-nonaccepting (ANA) and normal Wistar rats. Differential patterns of expression have been found for NPY and have been recently confirmed by a GeneChip (Affymetrix, Santa Clara, CA) analysis. Although differential hippocampal expression might contribute to the cognitive impairment seen in the ANA line, it did not seem to be related to alcohol intake. However, a differential expression of Y2 receptors within the medial amygdala has also been found and might contribute to the different degrees of alcohol preference in the three lines (Caberlotto et al., 2001). On the basis of these findings, it was hypothesized that, under normal, low-consumption conditions, NPY transmission may leave alcohol consumption unaffected or may even stimulate it through hypothalamic appetitestimulating effects, but it may suppress alcohol drinking in high-drinking states. This would make the NPY system an interesting target for pharmacological treatment of alcoholism. An additional hypothesis was that blockade of presynaptic Y2 receptors may potentiate endogenous NPY transmission and thereby suppress increased alcohol consumption, such as under limited-access conditions or in subjects with a history of dependence. In agreement with
PANDEY ET AL.
these hypotheses, intracerebroventricular administration of the experimental Y2-receptor antagonist BIIE0246 dosedependently reduced operant alcohol self-administration under limited-access conditions (Thorsell et al., 2002). In preliminary experiments, they recently found that doses of the antagonist that are subthreshold in naïve rats are still markedly effective to reduce self-administration in animals with a history of alcohol dependence, implying a role for the NPY system in the process of developing dependence. The animal findings raise the question of whether genetically encoded and acquired dysfunction of the NPY system might be involved in the pathogenesis of alcoholism in humans. Dr. Heilig and his group therefore studied the possible association of single nucleotide polymorphisms within the human NPY gene by using carefully diagnosed alcoholism (DSM-IV) in a genetically homogenous sample of Swedish alcoholics, compared with Swedish health examination controls in whom dependence was ruled out through the use of a face-to-face structured diagnostic interview. In this work, they were unable to replicate the previously postulated association of the T1128C polymorphism. Other polymorphic sites were found within the promoter regions of the prepro-NPY gene; however, their association with alcoholism is unknown. ASSOCIATION OF NEUROPEPTIDE Y POLYMORPHISM WITH THE OCCURRENCE OF TYPE 1 AND TYPE 2 ALCOHOLISM
Erkki Ilveskoski A common polymorphism in the signal peptide of human prepro-NPY was identified by a Finnish research group in 1998 (Karvonen et al., 1998). This leucine to proline polymorphism (Leu7/Pro7) in the NPY protein is due to the substitution of thymidine to cytosine at site 1128 in the first exon of the NPY gene. In comparison with the wild-type Leu7/Leu7 genotype, the Pro7 allele has been reported to be associated with increased serum total and low-density lipoprotein cholesterol in obese subjects (Karvonen et al., 1998), with enhanced carotid atherosclerosis and even with increased blood pressure (Karvonen et al., 2001). The role of NPY as one potential factor in alcoholism emerged from animal studies. A recent population-based study in Finnish men indicated that the NPY Pro7 allele was associated with 34% higher average alcohol consumption (Kauhanen et al., 2000). To further investigate the role of NPY in alcohol dependence, Dr. Ilveskoski and his group performed a case-control study in Finnish men and women (Ilveskoski et al., 2001). The subjects of this study consisted of 122 alcoholics classified as type 1 and type 2 subtypes by psychiatric evaluation. A random sample of 59 social drinkers was used as a control group to compare the distribution of NPY genotypes with those of alcoholics. Both type 1 (p ⫽ 0.029) and type 2 (p ⫽ 0.049) alcoholics had significantly lower frequencies of the Leu7/Pro7 genotype as compared with the controls. In a logistic regression
153
NEUROPEPTIDE Y AND ALCOHOLISM
model adjusting for the effects of age and gender, there was a significantly lower frequency of the Leu7/Pro7 heterozygotes among type 2 alcoholics, compared with the social drinkers (10.8 vs. 24.1%, p ⫽ 0.028). The data provided by Dr. Ilveskoski and his group were opposite to previous findings of Kauhanen et al. (2000), because the Leu7/Pro7 genotype seemed to protect from alcoholism. Several reasons, such as selection bias and differences in study material and design, could have led to the discrepancy (Ilveskoski et al., 2001). However, one probable cause is a difference in traits. The Leu7/Leu7 carriers in the study of Kauhanen et al. (2000) drank an average of 12.3 g of ethanol per day, whereas Pro7 carriers drank 16.4 g/day. Neither of these amounts can be considered heavy consumption. However, the alcoholics in our study had a serious drinking problem that led to hospitalization and treatment. In other words, Kauhanen et al. (2000) studied the amount of alcohol consumption, and we studied alcoholism. In conclusion, the NPY gene may be a susceptibility locus for alcoholism and also for alcohol consumption. However, more functional and association studies are needed to draw any firm conclusions of the role of NPY polymorphism in the development of alcoholism. SIGNIFICANCE
The data presented in this symposium clearly suggest a role for the NPYergic system in anxiety and alcoholdrinking behaviors. The knock-out studies related to NPY Y1, Y2, and Y5 receptors suggest the direct involvement of NPY Y1 receptors in high–alcohol-drinking behaviors, whereas NPY Y2 receptors are inhibitory to alcohol intake. The observation made with selectively bred models, such as P and NP rats, as well as AA and ANA rats, also suggests a role for NPY and NPY Y1 and Y2 receptors in alcoholdrinking behaviors. Pharmacological studies conducted in an unselected stock of rats suggest that PKA 3 CREB 3 NPY signaling in the central amygdala modulates anxiety and alcohol-drinking behaviors. These data also support the notion that decreased CREB and NPY in the central amygdala in P rats may be involved in the genetic predisposition to anxiety and alcohol-drinking behaviors. Future studies are needed to investigate the changes in the expression of NPY receptor subtypes in various brain structures during ethanol exposure and withdrawal. It is also important to know how the signaling related to NPY is modified in various NPY-related knock-out mice as a result of compensatory changes due to specific gene deletion. The NPY gene polymorphism studies in Swedish alcoholics indicate that there is no T1128C polymorphism. However, it would be interesting to examine whether other polymorphic sites within the NPY gene of Swedish alcoholics are associated with an alcoholism phenotype. On the other hand, Finnish alcoholic studies suggest that the Leu7/ Pro7 genotype seems to protect against alcoholism. There is also some evidence of functional consequences of the
Leu7/Pro7 substitution. Kallio et al. (2001) showed that the Pro7 allele is associated with ⬎40% higher exerciseinduced NPY concentrations in blood and with lower postexercise free fatty acid concentrations. Considering the fact that the polymorphism probably modifies the processing of prepro-NPY in the endoplasmic reticulum, this finding suggests that the Pro7 allele might be related to enhancing the intracellular processing of NPY and to increased NPY concentrations in blood and tissue. It is possible that Pro7 alleles may lead to increased NPY signaling, thereby protecting the development of alcohol-drinking behaviors. More functional studies are needed to clarify the association of polymorphic sites within the NPY gene and alcoholism. In summary, the studies related to the NPYergic system accumulated so far suggest the possibility that this system represents a novel therapeutic target for the treatment of anxiety and alcohol-abuse disorders. REFERENCES Badia-Elder NE, Stewart RB, Powrozek TA, Roy KF, Murphy JM, Li T-K (2001) Effect of neuropeptide Y (NPY) on oral ethanol intake in Wistar, alcohol-preferring (P), and -nonpreferring (NP) rats. Alcohol Clin Exp Res 25:386 –390. Bice P, Foroud T, Bo R, Castelluccio P, Lumeng L, Li T-K, Carr LG (1998) Genomic screen for QTL underlying alcohol consumption in the P and NP rat lines. Mamm Genome 9:949 –955. Caberlotto L, Thorsell A, Rimondini R, Sommer W, Hyytia P, Heilig M (2001) Differential expression of NPY and its receptors in alcoholpreferring AA and alcohol-avoiding ANA rats. Alcohol Clin Exp Res 25:1564 –1569. Carr L, Foroud T, Bice P, Gobbett T, Ivashina J, Edemberg H, Lumeng L, Li T-K (1998) A quantitative trait locus for alcohol consumption in selectively bred rat lines. Alcohol Clin Exp Res 22:884 – 887. Ehlers C, Li T-K, Lumeng L, Somes C, Himinez P, Mathe A (1998) Neuropeptide Y (NPY) levels in ethanol-naïve alcohol-preferring and nonpreferring rats and in Wistar rats after ethanol exposure. Alcohol Clin Exp Res 22:1778 –1782. Heilig M, Thorsell A (2002) Brain neuropeptide Y (NPY) in stress and alcohol dependence. Rev Neurosci 13:85–94. Heilig M, Widerlov E (1995) Neurobiology and clinical aspects of neuropeptide Y. Crit Rev Neurobiol 9:115–136. Hwang BH, Zhang JK, Ehlers CL, Lumeng L, Li T-K (1999) Innate differences of neuropeptide Y (NPY) in hypothalamic nuclei and central nucleus of the amygdala between selectively bred rats with high and low alcohol preference. Alcohol Clin Exp Res 23:1023–1030. Ilveskoski E, Kajander OA, Lehtimäki T, Kunnas T, Karhunen PJ, Heinälä P, Virkkunen M, Alho H (2001) Association of neuropeptide Y polymorphism with the occurrence of type 1 and type 2 alcoholism. Alcohol Clin Exp Res 10:1420 –1422. Kallio J, Pesonen U, Kaipio K, Karvonen MK, Jaakkola U, Heinonen OJ, Uusitupa MI, Koulu M (2001) Altered intracellular processing and release of neuropeptide Y due to leucine 7 to proline 7 polymorphism in the signal peptide of preproneuropeptide Y in humans. FASEB J 7:1242–1244. Karvonen MK, Pesonen U, Koulu M, Niskanen L, Laakso M, Rissanen A, Dekker JM, Hart LM, Valve R, Uusitupa MI (1998) Association of a leucine(7)-to-proline(7) polymorphism in the signal peptide of neuropeptide Y with high serum cholesterol and LDL cholesterol levels. Nat Med 12:1434 –1437. Karvonen MK, Valkonen VP, Lakka TA, Salonen R, Koulu M, Pesonen U, Tuomainen TP, Kauhanen J, Nyyssönen K, Lakka HM, Uusitupa MI, Salonen JT (2001) Leucine7 to proline7 polymorphism in the
154
preproneuropeptide Y is associated with the progression of carotid atherosclerosis, blood pressure and serum lipids in Finnish men. Atherosclerosis 1:145–151. Kauhanen J, Karvonen MK, Pesonen U, Koulu M, Tuomainen TP, Uusitupa MI, Salonen JT (2000) Neuropeptide Y polymorphism and alcohol consumption in middle-aged men. Am J Med Genet 93:117–121. Kelley SP, Nannini MA, Bratt AM, Hodge CW (2001) Neuropeptide-Y in the paraventricular nucleus increases ethanol self-administration. Peptides 22:515–522. Marsh DJ, Baraban SC, Hollopeter G, Palmiter RD (1999) Role of the Y5 neuropeptide Y receptor in limbic seizures. Proc Natl Acad Sci USA 96:13518 –13523. Mayfield RD, Lewohl JM, Dodd PR, Herlihy A, Liu J, Harris RA (2002) Patterns of gene expression are altered in the frontal and motor cortices of human alcoholics. J Neurochem 81:802– 813. Naveilhan P, Hassani H, Canals JM, Ekstrand AJ, Larefalk A, Chhajlani V, Arenas E, Gedda K, Svensson L, Thoren P, Ernfors P (1999) Normal feeding behavior, body weight and leptin response require the neuropeptide Y Y2 receptor. Nat Med 5:1188 –1193. Naveilhan P, Neveu I, Arenas E, Ernfors P (1998) Complementary and overlapping expression of Y1, Y2 and Y5 receptors in the developing and adult mouse nervous system. Neuroscience 87:289 –302. Palmiter RD, Erickson JC, Hollopeter G, Barban SC, Schwartz MW (1998) Life without neuropeptide Y. Recent Prog Horm Res 53:163– 199. Pandey SC, Mittal N, Lumeng L, Li T-K (1999) Involvement of the cyclic AMP-responsive element binding protein gene transcription factor in genetic preference for alcohol drinking behavior. Alcohol Clin Exp Res 23:1425–1434. Pandey SC, Roy A, Zhang H (2003) The decreased phosphorylation of cyclic AMP response element-binding (CREB) protein in the central amygdala act as a molecular substrate for anxiety related to ethanol withdrawal in rats. Alcohol Clin Exp Res (in press).
PANDEY ET AL.
Pedrazzini T, Seydoux J, Kunstner P, Aubert JF, Grouzmann E, Beermann F, Brunner HR (1998) Cardiovascular response, feeding behavior and locomotor activity in mice lacking the NPY Y1 receptor. Nat Med 4:722–726. Roy A, Pandey SC (2002) The decreased cellular expression of neuropeptide Y protein in rat brain structures during ethanol withdrawal after chronic ethanol exposure. Alcohol Clin Exp Res 26:796 – 803. Slawecki CJ, Betancourt M, Walpole T, Ehlers CL (2000) Increases in sucrose consumption, but not ethanol consumption, following ICV NPY administration. Pharmacol Biochem Behav 66:591–594. Thiele TE, Marsh DJ, Ste Marie LS, Bernstein IL, Palmiter RD (1998) Ethanol consumption and resistance are inversely related to neuropeptide Y levels. Nature 396:366 –369. Thiele TE, Miura GI, Marsh DJ, Bernstein IL, Palmiter RD (2000) Neurobiological responses to ethanol in mutant mice lacking neuropeptide Y or the Y5 receptor. Pharmacol Biochem Behav 67:683– 691. Thiele TE, Koh MT, Pedrazzini T (2002) Voluntary alcohol consumption is controlled via the neuropeptide Y Y1 receptor. J Neurosci 22:RC208. Thorsell A, Michalkiewicz M, Dumont Y, Quirion R, Caberlotto L, Rimondini R, Mathe AA, Heilig M (2000) Behavioral insensitivity to restraint stress, absent fear suppression of behavior and impaired spatial learning in transgenic rats with hippocampal neuropeptide Y overexpression. Proc Natl Acad Sci USA 97:12852–12857. Thorsell A, Rimondini R, Heilig M (2002) Blockade of central neuropeptide Y (NPY) Y2 receptor reduces ethanol self-administration in rats. Neurosci Lett 332:1– 4. Wettstein JG, Earley B, Junien JL (1995) Central nervous system pharmacology of neuropeptide Y. Pharmacol Ther 65:397– 414. Woldbye DPD, Ulrichsen J, Haugbol S, Bolwig TG (2002) Ethanol withdrawal in rats is attenuated by intracerebroventricular administration of neuropeptide Y. Alcohol Alcohol 37:318 –321.
