Reinitiation of Spermatogenesis by Hormones as Assessed by Cellular Parameters in vitro

Gonadotropic hormone stimulation of the reinitiation of meiosis and DNA synthesis following estradiol-induced suppression of spermatogenesis was measured in cultured testicular tissue and dissociated cells. Male rats were given estradiol dipropionate every 5 days starting at Day 1 after birth. At 40 days of age, a group of estradiol treated animals were given daily injections of PMSG. At 41, 43, 46 and 49 days of age, normal, estradiol, and estradiol + PMSG treated animals were sacrificed. Testes were dissociated, cultured in a medium containing [ 3 H]-thymidine and compared. One day after PMSG administration in vivo, significant stimulation of DNA synthesis in cultured testicular cells, as measured by scintillation counting and autoradiography, was observed. Increase in [ 3 H]-thymidine incorporation was associated with the entry of new cells into the S-phase, as determined by an increased labeling index. Autoradiography revealed that cells active in DNA synthesis included both mitotic and meiotic cells. Cells that subsequently developed were primary spermatocytes as indicated by a corresponding increase in the tetraploid peak in cellular DNA profiles measured by flow cytometry and in the appearance of pachytene spermatocytes in sectioned testicular tissue. These results indicate that PMSG is effective in inducing, within 1 day, an immediate and dramatic stimulatory response on meiosis and that the different in vitro techniques used allow an accurate and comprehensive assessment of the status of testicular tissue and of meiotic activity.