Molecular Psychiatry (2006) 11, 312–322 & 2006 Nature Publishing Group All rights reserved 1359-4184/06 $30.00 www.nature.com/mp ORIGINAL ARTICLE Why do young women smoke? I. Direct and interactive effects of environment, psychological characteristics and nicotinic cholinergic receptor genes L Greenbaum1,3, K Kanyas1,3, O Karni1, Y Merbl2, T Olender2, A Horowitz2, A Yakir2, D Lancet2, E Ben-Asher2 and B Lerer1 1Biological Psychiatry Laboratory, Department of Psychiatry, Hadassah-Hebrew University Medical Center, Jerusalem, Israel and 2Department of Molecular Genetics, Weizmann Institute of Science, Rehovot, Israel Despite the health hazards, cigarette smoking is disproportionately frequent among young women. A significant contribution of genetic factors to smoking phenotypes is well established. Efforts to identify susceptibility genes do not generally take into account possible interaction with environment, life experience and psychological characteristics. We recruited 501 female Israeli students aged 20–30 years, obtained comprehensive background data and details of cigarette smoking and administered a battery of psychological instruments. Smoking initiators (n = 242) were divided into subgroups with high (n = 127) and low (n = 115) levels of nicotine dependence based on their scores on the Fagerstrom Tolerance Questionnaire and genotyped with noninitiators (n = 142) for single nucleotide polymorphisms (SNPs) in 11 nicotinic cholinergic receptor genes. We found nominally significant (P < 0.05) allelic and genotypic association with smoking initiation of SNP rs2072660 and multilocus haplotypes (P < 0.007–0.05) in CHRNB2 and nominal (P < 0.05) allelic or genotypic association of SNPs in CHRNA7 (rs1909884), CHRNA9 (rs4861065) and CHRNB3 (rs9298629) with nicotine dependence. Employing logistic regression and controlling for known risk factors, the best- fitting model for smoking initiation encompassed a 5 SNP haplotype in CHRNB2, neuroticism and novelty seeking (P = 5.9 Â 10À14, Nagelkerke r2 = 0.30). For severity of nicotine dependence, two SNPs in CHRNA7 (rs1909884 and rs883473), one SNP in CHRNA5 (rs680244) and the interaction of a SNP in CHRNA7 (rs2337980) with neuroticism, were included in the model (P = 2.24 Â 10À7, Nagelkerke r2 = 0.40). These findings indicate that background factors, psychological characteristics and genetic variation in nicotinic cholinergic receptors contribute independently or interactively to smoking initiation and to severity of nicotine dependence in young women. Molecular Psychiatry (2006) 11, 312–322. doi:10.1038/sj.mp.4001774; published online 13 December 2005 Keywords: smoking initiation; nicotine dependence; nicotinic cholinergic receptor genes; environment; life experience; personality Introduction women typically serve in the army for 18–24 months, representing the age group of 18–21 years. Since Despite global trends indicating an overall decline, smoking behavior is considered to be fixed by 25 cigarette smoking is increasing among women in years,4 these figures are alarming. high-income countries and is highest among women Research on smoking behavior has focused on risk of reproductive age.1,2 In the US, nearly 30% of female factors for both smoking initiation (SI) and nicotine high school seniors were smoking in 2000.2 In Israel, dependence (ND). Especially among adolescents, SI 21.9% of Jewish women aged 21–34 years smoked in has been associated with family risk factors, such as 2003.3 The Israel Defense Force (IDF) reports an parental smoking5,6 and family protective factors, increase of 35% in smoking over the last decade such as a strong parent–child relationship.7,8 Psycho- among women at induction and demobilization with logical factors, such as extraversion,9 novelty seek- 39.1% of female soldiers smoking regularly.3 In Israel, ing,10,11 neuroticism12 and external locus of control13 have also been associated with SI. Having two parents Correspondence: Professor B Lerer, Biological Psychiatry Labora- who smoked has also been related to adult ND7,14 tory, Department of Psychiatry, Hadassah-Hebrew University as has harm avoidance.10 High harm avoidance and Medical Center, Ein Karem, Jerusalem 91120, Israel. neuroticism were related to severity of withdrawal E-mail: [email protected] 15 3These authors contributed equally to this work. symptoms from nicotine. Received 5 August 2005; revised 6 October 2005; accepted 11 A substantial body of literature indicates that October 2005; published online 13 December 2005 nicotine is the major component in cigarette smoke Environment, personality, genes and smoking L Greenbaum et al 313 that leads to addiction.16 The development of ND in for the beta4 nicotinic receptor subunit were demon- vulnerable individuals requires exposure to nicotine, strated, but not in mice null for the beta2 gene.32 A which occurs by voluntary initiation of cigarette role of CHRNB2 in nicotine reinforcement was smoking. Thus, consideration of genetic factors that recently demonstrated by specifically re-expressing influence cigarette smoking should take into account this gene in the ventral-tegmental area of mutated factors that influence the likelihood of smoking the mice.33 A few studies have shown association of first cigarette, the development of a regular smoking nAChR genes with smoking in schizophrenic patients habit and, once smoking behavior is established, the and have raised the possibility of common neuro- degree to which the smoker is dependent on nicotine. biological mechanisms. CHRNA7 was found to be Extensive evidence from twin and adoption studies associated with smoking34 and linkage of CHRNA2 supports a significant contribution of genetic factors and CHRNB2 was reported,35 based on the results of a to SI as well as progression to ND, with heritability of genome scan of schizophrenia patients. CHRNA7 was the order of 50–60%.17–19 shown to be involved in auditory sensory processing The physiological and behavioral effects of nicotine deficits in schizophrenia patients.36,37 Nicotine may are primarily mediated by neuronal nicotinic choli- ameliorate some of these deficits.38 nergic receptors (nAChRs), which modulate the In this study, we examined the association of 11 release of dopamine in the mesolimbic system. Other nAChR genes, including CHRNA4 and CHRNAB2, addictive drugs such as cocaine and amphetamine are with SI and ND in a case–control sample of young thought to act through this central reward path- female students, taking into account the contribution way.20,21 Neuronal nAChRs are ligand gated, penta- of background, life experience and psychological meric ion channels. A number of different subtypes of characteristics of the subjects and the possible nAChR exist, each with individual pharmacological interaction of these variables with nAChR gene and physiological profiles and distinct anatomical variants. distribution in the brain.22 Each nAChR is composed of five subunits arranged in either homomeric or Materials and methods heteromeric complexes of alpha or beta subunits. The different combination of subunits is responsible for Subjects the unique properties of the receptor.23 In all, 12 genes A total of 501 female subjects were recruited from the coding for neuronal nAChR subunits have been Jerusalem area between September 2002 and May cloned: nine neuronal alpha-subunits (alpha2– 2004. Subjects were recruited through advertisements alpha10) and three neuronal beta-subunits (beta2– at institutions of higher education reflecting both beta4). They encode peptides that have a relatively secular and religious environments and various fields hydrophilic extracellular amino terminal portion, of study. Inclusion criteria were age (20–30 years), followed by three hydrophobic trans-membrane doma- enrollment in an institute of higher learning, Jewish ins (M–M3), a large intracellular loop, and then a origin with both parents Ashkenazi or non-Ashkenazi fourth hydrophobic trans-membrane domain (M4).22 and no history of psychiatric treatment of any kind. The two binding sites for acetylcholine (both of which Subjects who were not born and raised in Israel were need to be occupied to cause the channel to open) excluded in order to prevent cultural bias. reside at the interface between the extracellular After screening by telephone, each subject was domain of each of the alpha-subunits and its invited to visit the laboratory where she received an neighbor. Acetylcholine acts on the postsynaptic explanation of the project and was asked to provide receptor to cause a large increase in its permeability signed informed consent as approved by the Helsinki to cations and depolarization of the membrane.24 The Committee (Internal Review Board) of the Hadassah primary brain nAChR subtype with high affinity for Medical Organisation. All consenting subjects com- nicotine contains alpha4 and beta2 subunits.25 pleted a booklet containing psychological and back- Studies to date on the association of nAChRs with ground measures and provided 30–50 ml of fresh cigarette smoking and ND have focused on the genes blood for DNA extraction and lymphoblast transfor- encoding the alpha4 and beta2 subunits. A recent mation. All subjects received monetary compensation study26 reported significant protective effects against for their time and travel expenses. ND in Chinese men of two single nucleotide poly- Subjects were coded as smoking initiators (SI) if morphisms (SNPs) and a multilocus haplotype in the they had smoked at some time during their lifetime CHRNA4 gene.