Adrenal sexual dimorphism is thought to be important in explaining gender-related differences regarding prevalent diseases and the responses to stress and drugs. We report here that in CD1 mice there is marked sexual dimorphism affecting not only gland size and corticoid hormone secretion but also adrenal ornithine decarboxylase (ODC), polyamine and catecholamine levels, in which testosterone appears to be a major determinant. Our results show that adrenal weight, ODC activity and corticosterone and aldosterone secretion were higher in female than in male mice, and that orchidectomy brought these male parameters closer to the values found in females. mRNA levels of steroidogenic proteins SF-1, Dax-1, steroid 21-hydroxylase (CYP21) and aldosterone synthase (CYP11B2) appeared to be slightly higher in female than in male adrenals. Immunocytochemical analysis of adrenal ODC revealed that immunoreactivity was higher in females than in males, and was mainly located in the cortical cells, and especially in zona glomerulosa, whereas no gender differences in ODC mRNA levels were observed. These results suggest that sex-associated differences in the expression of ODC in the mouse adrenal gland appear to be mainly related to post-transcriptional mechanisms. Combination treatment of mice with alfa-difluoromethylornithine (a suicide inhibitor of ODC), and a polyamine-deficient diet, produced a marked decrease in adrenal polyamine and catecholamine levels and a significant reduction in plasma corticosterone and aldosterone concentrations that were not associated with a decrease in the mRNA levels of steroidogenic proteins. All these data suggest a relevant role for testosterone, ODC and polyamines in the mouse adrenal function.