The present work aimed to compare some features of the meiotic process which develops in the testis of pubertal rats, in vivo and in vitro, paying special attention to the time-course of the phenomenon. The differentiation of spermatocytes was assessed in testes of 20- to 46-day-old rats and in tubule segments of 20- or 28-day-old rats cultured over a 4-week period. Very similar results were obtained in vivo and in vitro, during the first week of culture, when considering the changes in the cell populations of different ploidy, the gene expression of germ cells, the kinetics of differentiation of BrdU-labeled early or middle pachytene spermatocytes and the levels of apoptosis in the different cell populations. However, during the second week of culture, the decrease in the proportion of the 4C cell population which was only slightly more marked than that observed in vivo between 27 and 34 days, was not associated with an increase in the 1C cell population as large as in vivo. This result could be explained partly by a high proportion of apoptotic 1C cells beyond one week of culture. Concomitantly, the rate of in vitro differentiation of BrdU-labeled spermatocytes slowed down when reaching the stage of middle pachytene spermatocytes and BrdU-labeled round spermatids were observed 6-11 days later than when BrdU-labeled spermatocytes differentiated in vivo. Taken together, our results indicate that the bottleneck for the development of the meiotic cells in vitro is at the transition from middle to late pachytene spermatocytes. Hence, comparing the expression of locally produced regulatory molecules in vivo and in vitro at different days of culture should allow to identify key regulators of the meiotic step of spermatogenesis.
Copyright 2003 Wiley-Liss, Inc.