This study examined whether a reportedly functional polymorphism in the gene encoding the μ‐opioid receptor protein (A118G, which causes an Asn40Asp substitution in the receptor's extracellular domain), modifies the cortisol response to the opioid antagonist naloxone. The polymorphism occurs commonly in European Americans and some other population groups, underscoring its potential phenotypic significance. Methods: Using a balanced, within‐subject design involving two test sessions over a period of 3–7 days, we examined ACTH and cortisol responses to intravenous naloxone (125 μg/kg) or placebo in 30 healthy subjects (21 males, mean age = 24.4 years). Plasma ACTH and cortisol concentrations were measured over 120 min post infusion. DNA isolated from whole blood was PCR amplified and genotyped via restriction enzyme digestion, with genotypes assigned based on agarose gel size fractionation. Results: Subjects with one or more Asp40 alleles (n = 6; 5 heterozygotes and 1 homozygote) had significantly higher cortisol concentrations at baseline and at 15, 60, and 90 min after naloxone infusion than subjects homozygous for the Asn40 allele (n = 24). Subjects with the Asp40 allele also had a greater peak cortisol response and a greater area under the cortisol time curve than those homozygous for the Asn40 allele. There were no effects of the Asn40Asp polymorphism on plasma ACTH concentration or on self‐reported anxiety or distress. Conclusions: These findings are consistent with recent reports showing an enhanced cortisol response to naloxone and a reduced agonist effect of morphine‐6‐glucuronide among subjects with the Asp40 variant. Given evidence of its pharmacological significance, the clinical relevance of this polymorphism warrants further investigation. © 2003 Wiley‐Liss, Inc.