1. The effect of FSH on the membrane potential of Sertoli cells has been investigated by intracellular microelectrode recording from monolayer cultures of cells, isolated from immature rat testes by enzymatic treatment. 2. In standard Earle's solution, the membrane potential of unstimulated cells in a 3-day-old monolayer was -21.6 +/- 0.2 mV (n = 300). The recorded potentials were unchanged on varying the culture period from 3 to 7 days or by removal of chloride or calcium from the media. However, they were decreased in a low-sodium or potassium-rich medium. 3. Ovine FSH caused hyperpolarization of the cell in a dose-dependent manner. Maximal values were obtained with concentrations ranging between 2.9 and 5.9 micrograms/ml (-37.0 +/- 0.2 mV; n = 310). Dibutyryl cyclic AMP (1 mM) produced a similar effect to FSH. Under similar conditions human chorionic gonadotrophin (hCG) had no effect on the membrane potential. 4. The FSH-induced hyperpolarization was unchanged by chloride or bicarbonate replacement. It was decreased by low-sodium media (9 mM) (-29.6 +/- 0.3 mV; n = 110), by removal of calcium (-32.4 +/- 0.6 mV; n = 128) and by an increase in the potassium concentration of the bathing medium. A tenfold increase in potassium concentration depolarized the cell by 29.5 mV against 16 mV for unstimulated cells. 5. FSH-induced hyperpolarization was decreased by application of ouabain (10(-4) M), quinidine (1.4 x 10(-4) M) and cobalt (10(-3) M) (respectively: -28.7 +/- 0.3 mV, n = 124; -26.6 +/- 0.3 mV, n = 52 and -24.8 +/- 0.3 mV, n = 68) but was unchanged by amiloride (10(-4) M) and TTX (3 x 10(-6) M). None of these drugs modified the membrane potential of unstimulated cells. 6. All these results suggest that FSH stimulation of Sertoli cells in monolayer culture involves the modification of their membrane permeabilities.