The effect of T₃ on basal and LH mediated synthesis and secretion of testosterone and oestradiol by puberal rat Leydig cells was studied in vitro. Percoll gradient purified Leydig cells (1×10³) were cultured for 48 hours at 34°C in a medium containing a range of 5-400ng/mL concentration of T₃ or ovine LH after 24 hours initial culture at 37°C. T₃ increased testosterone and oestradiol secretions in a dose dependent manner which reached the saturation point with 50ng dose. While the minimum effective dose of T₃ (25ng) potentiated the stimulatory effect of the minimum effective dose of LH (25ng) on testosterone secretion, it suppressed the effect of the saturation dose of LH (100ng). Fifty ng T₃ quelled the stimulatory effect of either dose of LH. Both doses of T₃ increased oestradiol secretion, irrespective of the dose of LH. Addition of androstenedione (500ng/mL) to the culture medium enhanced 25ng T₃ induced testosterone and oestradiol secretions. While androstenedione potentiated the stimulatory effect of T₃ (25ng) on LH (25ng) induced testosterone and oestradiol secretions, it reversed the inhibitory effect of 50ng T₃ on LH mediated testosterone secretion which was accompanied by a decrease in oestradiol secretion. Puromycin (35μg/mL) suppressed the stimulatory effect of T₃ on basal and LH mediated testosterone and oestradiol production. Taken together, the present results indicate a direct stimulatory effects of T₃ on basal production of testosterone and oestradiol by Leydig cells and its modulatory effect on LH mediated steroidogenic activity varies depending upon the intensity of LH stimuli.