Junctional contacts between Sertoli cells in normal and aspermatogenic rat seminiferous epithelium contain alpha6beta1 integrins, and their formation is controlled by follicle-stimulating hormone.

The distribution of alpha6beta1 integrins at the level of cell-to-cell contacts within the rat seminiferous epithelium was investigated. Double fluorescence experiments using phalloidin staining of actin filaments and anti-integrin subunit antibodies showed that the receptor belongs to the Sertoli cell lateral domains engaged in the characteristic junctional structures known as ectoplasmic specializations (ES), at the level both of inter-Sertoli junctions and of the contacts between Sertoli cells and elongating spermatids. In the seminiferous epithelium of aspermatogenic testes, obtained through X-irradiation in utero (Sertoli-cell-only testes), at the level of inter-Sertoli junctions both ES and alpha6beta1 integrins are present. In order to study the dependence of alpha6beta1 receptors and ES formation upon FSH stimulation during development, 9-day-old testes were grown in organ culture in basal as well as FSH-supplemented conditions. FSH stimulation, which is necessary for the progression of spermatogenesis to early meiotic stages, appears to be required for the development of inter-Sertoli junctional structures containing ES and alpha6beta1 integrins. These observations indicate that the receptor belongs to the inter-Sertoli junctional machinery and that its expression at that level is not dependent on active spermatogenesis but requires FSH stimulation.

Stage and cell-specific expression of the adenosine 3',5' monophosphate-phosphodiesterase genes in the rat seminiferous epithelium.

Four genes (ratPDE1/IVc, ratPDE2/IVa, ratPDE3/IVd, and ratPDE4/IVb) encoding different isoforms of phosphodiesterase that specifically hydrolyze the second messenger cAMP (cAMP-PDEs) are present in the rat. Previous data from our laboratory indicated that these genes are differentially expressed in the somatic and germ cells of the seminiferous epithelium of the testis. To further characterize their spatial and temporal expression in the seminiferous tubules, in situ hybridization was used to monitor the expression of the four cAMP-PDE messenger RNAs (mRNAs). The signals corresponding to ratPDE1/IVc and ratPDE2IVa mRNAs were localized in two restricted layers of the seminiferous epithelium. The ratPDE1/IVc mRNA was present in a region of the epithelium corresponding to the location of middle-late pachytene spermatocytes. Conversely, the ratPDE2/IVa signal was confined to a more adluminal area corresponding to the location of maturing round spermatids. The ratPDE3/IVd and ratPDE4/IVb mRNAs were distributed throughout the span of the seminiferous epithelium, indicating a localization in the Sertoli cell cytoplasm. Although the intensity of the signal corresponding to ratPDE4/IVb was similar in all seminiferous tubule stages, the ratPDE3/IVd signal varied in intensity in tubules at different stages of the seminiferous cycle. Maximal expression was present in tubules at stages I-V and XI-XIII of the cycle and minimal at stages VIII-IX of the cycle. The expression of the ratPDE3/IVd mRNA positively correlated with the ability of specific inhibitors of the cAMP-PDEs to potentiate the FSH-dependent cAMP accumulation in tubules at different stages of the seminiferous cycle, with maximal potentiation observed at stages II-VI of the cycle. These data demonstrate that different cAMP-PDE genes are active in different cells of the seminiferous tubules and that the ratPDE3/IVd gene is expressed in the Sertoli cell in a cyclical fashion during the seminiferous cycle.

Development and cytodifferentiation of peritubular myoid cells in the rat testis.

The cytodifferentiation of peritubular myoid cells was studied in developing rats from fetal day 18 through approachment of puberty. The parameters taken into consideration were 1) the presence of desmin, a component of intermediate filaments in contractile cells; 2) the expression of alkaline phosphatase, a cell surface enzyme present in no other cell type of the seminiferous tubule; 3) the expression of the smooth muscle specific isoform of alpha-actin, a marker of terminal differentiation in smooth muscle cells; 4) cell proliferation rate, evaluated in radioautography as labeling index after incorporation of 3H-thymidine in short-term organ culture; and 5) cytoarchitectural changes detected with scanning electron microscopy. By means of immunofluorescence and cytochemistry it was observed that the three markers are expressed early during life, long before the onset of the first spermatogenic wave; in particular desmin is already present in fetal samples and alkaline phosphatase activity appears a few days after birth, whereas alpha-smooth muscle isoactin is first detected around birth. As for myoid cell replication, the high prenatal labeling index was found to drop soon after birth and to further slow down during the first month of postnatal life, suggesting that myoid cell proliferation is not a major factor in peritubular expansion. SEM examination of developing peritubulum has shown that, when approaching puberty, the myoid cell undergoes a dramatic change in cytoarchitecture, consisting in extreme flattening and cytoplasmic expansion resulting in an apparent increase in peritubular surface.