CRH haplotype as a factor influencing cerebrospinal fluid levels of corticotropin-releasing hormone, hypothalamic-pituitary-adrenal axis activity, temperament, and alcohol consumption in rhesus macaques.

CONTEXT: Both highly stress-reactive and novelty-seeking individuals are susceptible to alcohol use disorders. Variation in stress reactivity, exploration, and response to novelty have been attributed to differences in corticotropin-releasing hormone (CRH) system function. As such, CRH gene variation may influence risk for alcohol use and dependence. OBJECTIVE: To determine whether CRH variation influences relevant intermediate phenotypes, behavior, and alcohol consumption in rhesus macaques. DESIGN: We sequenced the rhesus macaque CRH locus (rhCRH) and performed cladistic clustering of haplotypes. In silico analysis, gel shift, and in vitro reporter assays were performed to identify functional variants. Cerebrospinal fluid (CSF) and blood samples were obtained, and levels of CRH and corticotropin (ACTH) were measured by radioimmunoassay. Behavioral data were collected from macaques during infancy. Among adolescent/adult animals, we recorded responses to an unfamiliar conspecific and measured levels of ethanol consumption. SETTING: National Institutes of Health Animal Center. PARTICIPANTS: Rhesus macaques. MAIN OUTCOME MEASURES: Animals were genotyped for a single-nucleotide polymorphism disrupting a glucocorticoid response element, rhCRH -2232 C>G, and the effects of this allele on CSF levels of CRH, plasma levels of ACTH, behavior, and ethanol consumption were assessed by analysis of variance. RESULTS: We show that -2232C>G alters DNA x protein interactions and confers decreased sensitivity of the CRH promoter to glucocorticoids in vitro. Consistent with the known effects of glucocorticoids on CRH expression in the brain, carriers of the G allele had lower CSF levels of CRH but higher levels of ACTH. Infants carrying the G allele were more exploratory and bold, and among adolescent and adult male macaques, the G allele was associated with exploratory/bold responding to an unfamiliar male. Adults with the C/G genotype also exhibited increased alcohol consumption in the social group, a model for high-risk alcohol-seeking behavior. CONCLUSION: Haplotypes that differ in terms of corticosteroid sensitivity have been identified in humans. Our data may suggest that functionally similar CRH variants could influence risk for externalizing disorders in human subjects.

Rearing condition and rh5-HTTLPR interact to influence limbic-hypothalamic-pituitary-adrenal axis response to stress in infant macaques.

BACKGROUND: In humans and macaques, a promoter polymorphism that decreases transcription of the serotonin transporter gene is associated with anxiety. Serotonin transporter gene disruption in rodents produces anxious animals with exaggerated limbic-hypothalamic-pituitary-adrenal (LHPA) responses to stress. We wanted to determine whether serotonin transporter gene promoter variation (rh-5HTTLPR) and rearing condition would interact to influence endocrine responses to stress in infant rhesus macaques. METHODS: Animals were reared with their mothers (MR, n = 141) or in peer-only groups (PR, n = 67). At 6 months of age, adrenocorticotropic hormone (ACTH) and cortisol levels were determined at baseline and during separation stress. Serotonin transporter genotype (l/l and l/s) was determined with polymerase chain reaction followed by gel electrophoresis. RESULTS: Cortisol levels increased during separation, and there was a main effect of rearing condition, with decreased cortisol levels among PR macaques. Animals with l/s rh5-HTTLPR genotypes had higher ACTH levels than did l/l animals. Adrenocorticotropic hormone levels increased during separation, and there was a separation x rearing x rh5-HTTLPR interaction, such that PR-l/s animals had higher ACTH levels during separation than did other animals studied. CONCLUSIONS: These data demonstrate that serotonin transporter gene variation affects LHPA axis activity and that the influence of rh5-HTTLPR on hormonal responses during stress is modulated by early experience.

Serotonin transporter gene variation is associated with alcohol sensitivity in rhesus macaques exposed to early-life stress.

BACKGROUND: Decreased sensitivity to alcohol has been demonstrated to be a predictor of alcoholism in humans, and variation in the gene-linked polymorphic region of the serotonin transporter (5-HTTLPR) is associated with the response to the motor-impairing effects of alcohol. In a nonhuman primate model of excessive alcohol intake, we have shown that decreased serotonin turnover is associated with both lower initial sensitivity to alcohol and higher prospective alcohol consumption using rhesus macaques. In addition, we have demonstrated that macaques separated from their mothers and reared in peer-only groups are more likely to consume alcohol as adults. METHOD: To examine the relationship between serotonin transporter genotype, early rearing experience, and initial sensitivity to alcohol, peer- and mother-reared, adolescent, alcohol-naive rhesus macaques (n = 123) were rated for intoxication after intravenous administration of ethanol (2.2 g/kg and 2.0 g/kg for males and females, respectively) during two testing periods. Serotonin transporter (rh5-HTTLPR) genotype was determined using polymerase chain reaction followed by gel electrophoresis, and data were analyzed using ANOVA and the Mann-Whitney U test. RESULTS: Our analyses demonstrate an effect of serotonin transporter gene variation on ethanol sensitivity, such that animals homozygous for the l allele exhibited decreased sensitivity to the ataxic and sedating effects of alcohol. This effect remained after correction for blood ethanol concentrations and birth cohort. When animals were segregated according to rearing condition, serotonin transporter gene variation predicted intoxication scores among peer-reared animals. CONCLUSIONS: As in some human reports, this study demonstrates a diminution in the response to alcohol in animals homozygous for the l rh5-HTTLPR allele. The phenotypic expression of this genotype in l/s animals, however, is environmentally dependent.