Background: Preclinical studies have implicated GABAergic neurosteroids in behavioral responses to alcohol. Naltrexone is thought to blunt the reinforcing effects of alcohol, and a few studies have found that the effects of naltrexone are moderated by the Asn40Asp polymorphisms of theOPRM1gene. The present study seeks to integrate these lines of research by testing (i) the moderating role of the functional Asn40Asp polymorphism of theOPRM1gene on naltrexone‐induced alternations in GABAergic neurosteroid levels, namely (3α,5α)‐3‐hydroxypregnan‐20‐one (allopregnanolone, ALLO); and (ii) the combined effects of naltrexone or genotype with alcohol administration on neurosteroid levels in a sample of at‐risk drinkers.Methods: Participants were 32 (9 females) nontreatment‐seeking heavy drinkers who completed a placebo‐controlled laboratory study of naltrexone (50 mg/d for 3 days) and provided complete sets of serum samples for ALLO assays before and after alcohol administration under both naltrexone and placebo conditions.Results: Naltrexone treatment raised ALLO levels among carriers of the Asp40 allele, but not homozygotes for the Asn40 allele. The Asn40Asp polymorphism did not moderate effects of naltrexone on cortisol levels. Ethanol infusion modestly reduced ALLO levels in all subjects, independent of genotype or naltrexone exposure.Conclusions: Naltrexone increased ALLO levels among individuals with the Asn40Asp allele suggesting a potential neurosteroid contribution to the neuropharmacological effects of naltrexone among Asp40 carriers.