Gonocytes in testes of neonatal rats express thec-kit gene

Abstract

Information gathered from mutant mouse models and from studies on normal puberal and adult animals points to the product of the c‐kit gene, a tyrosine kinase surface receptor, and the kit‐ligand (KL) as important for gametogenesis in males. In fetuses, KL serves as a survival factor for primordial germ cells, at least in vitro, and in adults activity of the c‐kit gene has been indirectly related to survival and subsequent development of differentiating spermatogonia. However, because of the structural complexity of the seminiferous epithelium in adults, c‐kit mRNA has not yet been definitively localized to one or more types of spermatogenic cells. In addition, no information is currently available regarding the possible involvement of the c‐kit protein and its ligand in mediating germ cell development and/or Sertoli‐germ cell interactions immediately after birth when events critical for later onset of spermatogenesis are ongoing. Thus, the aims of the current study were (1) to determine whether the c‐kit gene is expressed in testes of neonatal and adult rats and, if so, by what specific cell types, and (2) to determine if those cells expressing the gene also produce the c‐kit receptor protein. For this, we isolated total RNA from testes of pups aged days 1–5 and from adult rat testes, and probed for the presence of c‐kit mRNA with Northern analysis. We identified the cells containing the c‐kit message by carrying out in situ hybridization with digoxigeninlabeled probes, thus allowing the colorimetric signal to be assigned beyond doubt to individual cells in sections of testes. We also utilized Western analysis and immunolocalization to confirm the presence of the c‐kit receptor protein in testes at these ages and to identify those cells types producing it. Our findings indicate that (1) neonatal gonocytes express the c‐kit gene and produce the receptor protein on postnatal days 1 through 5, spanning the time when they resume dividing and migrating, and (2) spermatogonia and, to a lesser extent, spermatocytes and spermatids of adults express the gene but c‐kit protein is present in detectable amounts only in spermatogonia and possibly a few early primary spermatocytes.