The Pharmacogenomics Journal (2006) 6, 34–41 & 2006 Nature Publishing Group All rights reserved 1470-269X/06 $30.00 www.nature.com/tpj ORIGINAL ARTICLE Ionotropic glutamate receptor gene GRIK3 SER310ALA functional polymorphism is related to delirium tremens in alcoholics UW Preuss1,2, P Zill1, G Koller1, Upregulation of glutamatergic neurotransmission resulting from chronic 1 3 ethanol intoxication may cause a hyperexcitable state during alcohol B Bondy , V Hesselbrock and withdrawal, which may lead to seizures and delirium tremens. The aim of 1 M Soyka our study was to evaluate the association between a history of alcohol withdrawal-induced seizures and delirium tremens, and a functional 1Department of Psychiatry, Ludwig-Maximilians Universita¨t Mu¨nchen, Mu¨nchen, Germany; polymorphism (Ser310Ala) of the GRIK3 gene coding for the glutamatergic 2Johanna-Odebrecht-Stiftung, Greifswald, kainate receptor subunit GlurR7 in a sample of well-characterized alcoholics Greifswald, Germany and 3Department of compared to controls. In total, 233 patients meeting DSM-IV alcohol Psychiatry, University of Connecticut School of dependence criteria and 309 controls, all of German descent, were Medicine, Farmington, CT, USA investigated. GRIK3 functional polymorphism was determined using PCR Correspondence: (polymerase chain reaction) of lymphocyte DNA. History of alcohol PD Dr med. U.W. Preuss, Johanna-Odebrecht- withdrawal-induced delirium tremens and seizures were obtained using the Foundation, Gu¨tzkower Landstrasse 69, 17489 SSAGA (Semi-Structured Assessment for the Genetics of Alcoholism). Data Greifswald, Germany. were cross-checked with in-patients’ clinical files. While a significant E-mail: [email protected] relationship between history of delirium tremens and the Ser310 allele was detected, no significant results were obtained for alcohol withdrawal-related seizures. Although this result is suggestive for a significant role of this polymorphism in the pathogenesis of delirium tremens in alcohol-dependent individuals, further investigation and confirmation are warranted. The Pharmacogenomics Journal (2006) 6, 34–41. doi:10.1038/sj.tpj.6500343; published online 29 November 2005 Keywords: GRIK3; glutamate system; alcohol dependence; withdrawal; delirium tremens Introduction Ethanol withdrawal following chronic alcohol consumption is reported to produce dramatic changes in the content of brain excitatory neurotransmitters as well as their associated receptor sensitivity and regulation.1 Upregulation of glutamate neurotransmission as a consequence of chronic alcohol intake may contribute not only to neurotoxicity but also to neuronal death observed in brains of alcohol-dependent individuals (e.g.Davis and Wu,2 and Bleich3). This upregulation is related to the neuroadaptation, which results from chronic ethanol intoxication, causing a hyperexitable state that may also lead to seizures and delirium tremens during alcohol withdrawal.1,4,5 However, while most research focuses on the effects and consequences of acute and chronic alcohol Received 16 November 2004; revised 31 August 2005; accepted 1 September 2005; consumption on glutamatergic ionotropic NMDA (N-methyl-D-aspartate) recep- published online 29 November 2005 tors, several other ligand-gated channels activated by glutamate might be Kainate receptor functional polymorphism and delirium tremens UW Preuss et al 35 involved in the pathophysiology of alcoholism, including confirmed the functional relevance of this polymorphism. alcohol withdrawal-related complications such as delirium First, a lower KA-receptor expression of T-allele carriers tremens and seizures. (Ser310) was detected. In seven of nine human brain tissue Besides NMDA and AMPA (alpha-amino-3-hydroxy- samples under investigation, a 1.4- to 4.7-fold reduction of 5-methylisoxazole-4-propionate) receptors, kainate (KA) this allele expression was found. Second, while the Ser310 receptors are the third class in members of the glutamate- allele expression levels in the occipital cortex, mesencepha- gated family of ionotropic receptors. It is assumed that lon, cerebellum basal ganglia and thalamus was significantly KA-receptors channels are formed by tetramers or pentamers lower, the frontal brain had a two-fold higher expression of GluR5-GluR7, KA1, KA2 subunits. While GluR5-7 repre- level of this allele.28 sent low-affinity subunits and KA1-2 are considered high- The aim of this analysis is to investigate the influence of a affinity subunits,6 it is not clear as to which combination GluR7 functional polymorphism on a history of epileptic actually exist in native KA receptors in mammalian brains. seizures or delirium tremens during alcohol withdrawal in Some pathways involved in physiological activation of KA an in patient sample of alcohol-dependent subjects. Delir- receptors are still unknown,7 although physiological studies ium tremens and epileptic seizure may have different and have identified both post- and presynaptic roles for KA independent neurobiological mechanisms, they often co- receptors. This class of receptors has also been suggested to occur during withdrawal. For a subsequent analysis, there- play an important role in short- and long-term synaptic fore, three groups of alcohol-dependent subjects with a plasticity.8 KA receptors have been reported to generate a lifetime history of delirium tremens only, epileptic seizures slow excitatory activity-dependent synaptic current, which only and co-occuring delirium tremens and seizures are greatly augments excitatory currents of hippocampal CA3 associated with this GluR7 functional polymorphism. pyramidal cells.9 While located both pre- and postsynap- Furthermore, the potential influence of severity of alcohol ticlly, KA receptors play significant roles in the regulation of dependence, reflected by illness duration, severity of alcohol hippocampal excitability. They inhibit glutamate release dependence according to number of DSM IV criteria presynaptially10–12 and also regulate action potential-depen- endorsed, poor somatic condition, early age of onset and dent GABA release from interneurons. KA receptor mediated daily alcohol intake as well as subtypes of alcohol-depen- increase of interneuronal GABA release may function as an dent subjects (Babor’s Type A and B) are evaluated. All these important homeostatic mechanism that prevents overexci- characteristics of alcohol dependence might increase the tation of hippocampal CA1 pyramidal neurons.13,14 risk for both epileptic seizures and delirium tremens during While little was known about adaptations in KA receptors alcohol withdrawal and mediate the influence of genetic after chronic ethanol intake, in recent years KA receptors polymorphism on alcohol withdrawal complications. have emerged as important targets of alcohol’s action in the central nervous system.15 Exposure to ethanol for 1–5 days selectively decreased the sensitivity of KA receptor responses Results to low KA concentrations.16 Furthermore, an upregulation of these receptors under chronic administration of alcohol Two hundred and thirty-three alcohol-dependent indivi- in hippocampal regions has been reported. This upregula- duals (189 males and 44 females, aged from 18 to 74 years, 7 tion was suggested to eventually result in an increased mean age 42.02 9.8 years, see Table 1), all meeting ICD10 inhibition of GABAergic interneurons in the hippocampus and DSM-IV criteria for alcohol dependence were recruited. during alcohol withdrawal.17 This makes KA receptors As controls, 309 persons (161 males, 148 females, aged from 7 prime candidates in hippocampal kindling, which is 19 to 76 years; mean 46.8 12.1 years) were enrolled into suggested to be an important pathophysiological factor the study. History of alcohol withdrawal-induced delirium generating alcohol withdrawal-induced seizures and tremens was noted in 45 (19.3%), while a history of alcohol delirium tremens.18 withdrawal-induced epileptic seizures was reported from 54 The GluR7 subunit of KA receptors has been mapped to (23.2%) of the 233 alcohol-dependent subjects. Also pre- chromosome 1 at 1p34–p33,19 close to a region reported to sented in Table 1 are the duration of alcohol dependence, be associated with alcoholism and alcoholism-related phe- number of DSMIV alcohol-dependence criteria endorsed and notypes from linkage studies.20–22 GluR7 receptor subunits the number of subjects meeting Babor’s type A and B criteria. form a functional homomeric receptor channel with low sensitivity to glutamate.23 The GluR7 contains an exonic Genotype results thymine/guanine nucleotide variation that determines a The genotype frequencies of alcohol dependent subjects and serine or alanine at position 310 in the N-terminal controls are presented in Table 2. Data for both alcohol- extracellular domain of the receptor.24 This domain has dependent individuals and controls followed the Hardy– been reported to affect receptor desensitization25 and to Weinberg equilibrium (w2: 2.18, df: 1, P: 0.34 and w2: 0.17, participate in ligand binding.26,27 Prior electrophysiological df: 1, P: 0.92). investigations of both alleles in HEK 293 cells did not To assess the influence of GRIK3 polymorphism on indicate that the replacement of serine 310 by alanine alcohol dependence, GRIK3 genotype and allele frequencies affects receptor responses to glutamate.28 However, expres- were compared between alcoholic subjects and controls. No sion studies and characterization of putative consequences significant differences