The continuous production of male gametes requires the precise regulation of the differentiation and replication of stem spermatogonia. These critical processes occur in a niche that is a dynamic environment created in part by Sertoli cells, the somatic cells with which all spermatogenic cells interact. As yet, the in vivo mechanisms by which Sertoli cells regulate stem spermatogonial differentiation and replication have not been elucidated. Neither is it understood how those regulatory functions of Sertoli cells are themselves controlled. The central hypothesis of this application is that differentiated spermatogenic cells control the expression by Sertoli cells of growth factors that regulate the differentiation and replication of stem spermatogonia. We propose that by regulating expression of those growth factors, differentiated spermatogenic cells control the functioning of the stem spermatogonia niche. Using mice as a model, this proposal focuses on three growth factors that been shown to stimulate the growth of stem spermatogonia either in vivo or in vitro. These growth factors are: glial cell line derived neurotrophic factor (GDNF), fibroblast growth factor 2 (FGF-2) and insulin like growth factor 1 (IGF-1). Specific Aim I tests the hypothesis that in vivo the stage-specific expression of GDNF by Sertoli cells regulates replication and differentiation of stem spermatogonia and that FGF-2 and IGF-1 act in consort with GDNF in the regulation of these processes. Specific Aim II tests the hypothesis that Type A1 spermatogonia or more advanced spermatogenic cells regulate expression of GDNF, FGF-2 and IGF-1 by Sertoli cells. To begin to define the molecular basis of this regulation, we also propose an unbiased search for transcription factors and components of signal transduction pathways that mediate the effects of spermatogenic cells on GDNF expression by Sertoli cells. Specific Aim III will test the hypothesis that differentiated spermatogenic cells by their feedback on Sertoli cells control stem spermatogonia renewal and differentiation. Stage-specific gene expression by Sertoli cells is a well-established phenomenon in many species, including humans and GDNF, IGF-1 and FGF-2 are all expressed in human testes. Therefore, results from our proposed experiments hold promise for understanding the biological basis of the infertility of some men.