In previous studies of testicular biopsy tissue from healthy men, intratesticular testosterone (ITT) has been shown to be much higher than serum testosterone (T), suggesting that high ITT is needed relative to serum T for normal spermatogenesis in men. However, the quantitative relationship between ITT and spermatogenesis is not known. To begin to address this issue experimentally, we determined the dose-response relationship between human chorionic gonadotropin (hCG) and ITT to ascertain the minimum dose needed to maintain ITT in the normal range. Twenty-nine men with normal reproductive physiology were randomized to receive 200 mg T enanthate weekly in combination with either saline placebo or 125, 250, or 500 IU hCG every other day for 3 wk. ITT was assessed in testicular fluid obtained by percutaneous fine needle aspiration at baseline and at the end of treatment. Baseline serum T (14.1 nmol/liter) was 1.2% of ITT (1174 nmol/liter). LH and FSH were profoundly suppressed to 5% and 3% of baseline, respectively, and ITT was suppressed by 94% (1234 to 72 nmol/liter) in the T enanthate/placebo group. ITT increased linearly with increasing hCG dose (P < 0.001). Posttreatment ITT was 25% less than baseline in the 125 IU hCG group, 7% less than baseline in the 250 IU hCG group, and 26% greater than baseline in the 500 IU hCG group. These results demonstrate that relatively low dose hCG maintains ITT within the normal range in healthy men with gonadotropin suppression. Extensions of this study will allow determination of the ITT concentration threshold required to maintain spermatogenesis in man.