Sertoli cell number is considered to be stable and unmodifiable by hormones after puberty in mammals, although recent data using the seasonal breeding adult Djungarian hamster (Phodopus sungorus) model challenged this assertion by demonstrating a decrease in Sertoli cell number after gonadotropin depletion and a return to control levels following 7 days of FSH replacement. The present study aimed to determine whether adult Sertoli cells are terminally differentiated using known characteristics of cellular differentiation, including proliferation, junction protein localization, and expression of particular maturational markers, in the Djungarian hamster model. Adult long-day (LD) photoperiod (16L:8D) hamsters were exposed to short-day (SD) photoperiod (8L:16D) for 11 wk to suppress gonadotropins and then received exogenous FSH for up to 10 days. Sertoli cell proliferation was assessed by immunofluorescence by the colocalization of GATA4 and proliferating cell nuclear antigen and quantified by stereology. Markers of Sertoli cell maturation (immature, cytokeratin 18 [KRT18]; mature, GATA1) and junction proteins (actin, espin, claudin 11 [CLDN11], and tight junction protein 1 [TJP1, also known as ZO-1]) also were localized using confocal immunofluorescence. In response to FSH treatment, proliferation was upregulated within 2 days compared with SD controls (90% vs. 0.2%, P < 0.001) and declined gradually thereafter. In LD hamsters, junction proteins colocalized at the basal aspect of Sertoli cells, consistent with inter-Sertoli cell junctions, and were disordered within the Sertoli cell cytoplasm in SD animals. Exogenous FSH treatment promptly restored localization of these junction markers to the LD phenotype. Protein markers of maturity remain consistent with those of adult Sertoli cells. It is concluded that adult Sertoli cells are not terminally differentiated in the Djungarian hamster and that FSH plays an important role in governing the differentiation process. It is proposed that Sertoli cells can enter a transitional state, exhibiting features common to both undifferentiated and differentiated Sertoli cells.