Specific mRNAs in sertoli and germinal cells of testes from stage synchronized rats

Abstract

A treatment which used vitamin A depletion followed by vitamin A repletion was used to synchronize seminiferous tubules to a few related stages of the cycle of the seminiferous epithelium. The success of the synchronization procedure was dependent on the age and size of the rat at the initiation of the experiment (20 days of age and 35-40 g) and the extent to which the vitamin A deficiency had progressed. Administration of retinol was done when the only viable germinal cells in the testis were preleptotene spermatocytes and type A spermatogonia but if the deficiency was prolonged spermatogenesis did not recover. Once established synchrony appeared to be sustained at least through several consecutive cycles. A combination of molecular probes was used to determine if the synchronized testes displayed stage specific variations in Sertoli cell and germinal cell mRNA levels as has been reported for normal asynchronized rats. Sertoli cells in the synchronized testes were shown by quantitative in situ hybridization and by Northern blot analysis to have stage specific variations in the levels of mRNA for transferrin, sulfated glycoprotein-1, and sulfated glycoprotein-2. The mRNA levels in the different stages were qualitatively similar to those in equivalent stages previously reported for testes from asynchronous rats. The germinal cell content of the synchronized testes were examined with Northern blots probed with nick-translated protamine 1 and transition protein 1 cDNAs. The results showed that testes synchronized in stages l-VI contained spermatid steps 1-6 and 15-18 and had little or no protamine 1 or transition protein 1 mRNA but that testes synchronized to later stages VIII to XIV had both high levels and the predicted nonpolysomal and polysomal sizes of these two mRNAs. Since these germinal cell mRNAs are found in step 7 to 14 spermatids in normal rats, these results confirm the extent of synchrony in these testes. Thus, the present investigation confirms the relationship of the synchronized testes to the corresponding stages in normal rats and establishes the value of this model in future studies on the nature of the cycle of the seminiferous epithelium. © 1989 by The Endocrine Society.