In vitro formation of haploid spermatids has not been convincingly demonstrated in mammals. To investigate this problem we selected defined segments of rat seminiferous tubules containing late pachytene and diakinetic primary spermatocytes (Stages XII and XIII of the cycle) for culture in a chemically defined medium. After 2 days, most spermatocytes completed both meiotic divisions, and by 6 days the tubular epithelium developed morphologic characteristics of Stage V in which the newly formed spermatids had acrosanic systems characteristic of step 5 spermiogenesis. The seminiferous tubules also differentiated biochemically as evidenced by increased production of proteins characteristically secreted by Stage V. Since this in vitro differentiation of the germinal epithelium occurred in the absence of testosterone and FSH, we conclude that late pachytene spermatocytes and their associated Sertoli cells have all the information required for both meiotic divisions and early spermiogenesis.
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