STUDIES OF SPERMATOGENESIS

II. Effect of Testosterone and FSH on Spermatogenesis in Organ Culture (Studies on Testicular Function, Report 8)

Abstract

The effect of glucose, testosterone and FSH on spermatogenesis was studied in organ cultures of 21-day-old rat testis. Since previous experiments on the determinaiton of appropriate experimental conditions for rat testicular culture suggested that the best maintenance of primary spermatocytes was 2mg/ml of glucose, 50μg/ml of testosterone and 200μ/ml of FSH, the present investigation was undertaken to define the actions of these additives in the aforementioned concentrations. Data from 7 days and 14 days of cultures were statistically analyzed by analysis of variance.
As the interpretation of the mode of action of these factors on spermatogenesis is very intricte, data obtained in this series of experiments may be summarized by opinion. The results are summarized as follows.
1) In cultures of 2mg/ml of glucose medium containing a concentration of glucose double that normally present in a control medium, it was demonstrated that differentiation of germ cells was not induced by addition of glucose, while good maintenance of primary spermatocytes was observed for at least 7 days in culture. This suggests that glucose, as the energy source, acts on Sertoli cells to give nutrients to germ cells.
2) In cultures of 50μg/ml concentration of testosterone medium, it was demonstrated that the number of resting, leptotene and zygotene spermatocytes, i. e. early stages of primary spermatocytes, which have been associated with differentiation of spermatogonia increased. A reduction in the number of pachytene spermatocytes, a late stage of primary spermatocytes, and an increase in the number of pyknotic cells were noticiable, since the supplying capacity of Sertoli cells could not be accompanied by an increase in the number of early stages of primary spermatocytes (an explanation is given in subsection 3, concerned with the action of FSH). The primary spermatocytes are suggested to be the source of these pyknotic cells because of the anatomical arrangement of the intratubular cells. Therefore, testosterone may affect the mitotic division of spermatogonia.
Since the duration of spermatogenesis always progresses at a constant speed, resting, leptotene and zygotene spermatocytes observed in preculture must develop into pachytene spermatocytes, the next stage of zygotene primary spermatocytes, in 7 days and 14 days in culture, and resting, leptotene and zygotene spermatocytes ater 7 days in culture must be differentiated from spermatogonia in precultures.
Our interpretation of pyknotic cells is as follows: It has been demonstrated that degenerative processes of germ cells, such as degeneration, pyknosis (necrosis) and disappearance, should be considered as a constant and regular feature of in vivo normal testis, and that the number of spermatozoa produced per spermatogonia should be, in the rat, only 52% of theoretical yield. The pyknotic cells, the degenerated germ cells, are phagocytosed by Sertoli cells. The increase in the number of pyknotic cells and the reduction in the number of primary spermatocytes indicate a decrease in the supplying capacities as well as the phagocytic properties of Sertolic cells. Therefore, the presence of pyknotic cells may reflect the Sertoli cell function.
3) Data from experiments containing 200μg/ml of FSH produced the following two effects.
3)-a) The effect of FSH on the increase in the number of resting, leptotene and zygotene spermatocytes was observed, and was greater than that of testosterone. This indicates that mitotic division of spermatogonia is affected by some action of FSH as well as of testosterone.
3)-b) The other effect of FSH was maintenance of pachytene spermatocytes, and a significant observed increase in the number of primay spermatocytes on the whole. However, no increase in the number of pyknotic cells was observed. The relationship between testosterone and FSH in the number of pyknotic cells of 14-day cultivated explants statistically exhibited