Spermatogenesis is a complex cellular event during which the diploid germ cells differentiate and divide by mitosis and meiosis at specific time points along the spermatogenic cycle to generate the haploid spermatozoa. For this complex event to go in an orderly manner, cell differentiation and division must be precisely controlled by signals arising from within and outside the seminiferous tubules. Changes in the membrane potential of the germ cells are likely to be an important part of the signaling mechanism. We have applied the whole-cell patch clamp technique to identify and characterize ion channels in different spermatogenic cells from immature and mature rat testes fractionated by discontinuous Percoll gradient. A voltage- and Ca² - dependent, outwardly rectifying current with gating and pharmacologic properties resembling the large conductance K channels (BK_Ca) was recorded from the spermatogonia and primary spermatocytes. Another voltage-dependent, outwardly rectifying current that was sensitive to 4-aminopyridine, a K_v channel blocker, was detected in spermatocytes and early spermatids. This current is likely caused by the smaller conductance, voltage-sensitive K channels (K_v). In some spermatogonia, both the BK_Ca channels and the K_v channels could be simultaneously detected in the same cell. It appears that during the course of spermatogenesis, there is up-regulation of K_v but down-regulation of BK_Ca. Reverse transcription-polymerase chain reaction, Western blot analysis, and immunohistochemistry further confirmed the differential expression of the ion channels in different spermatogenic cells. We conclude that these ion channels may play an important role in the control of spermatogenesis.