Endothelial and inducible nitric oxide synthase (NOS) immunoreactivity and NOS-associated NADPH-diaphorase...

In this study, the cellular localization of nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d) and the endothelial (eNOS) and inducible (iNOS) forms of nitric oxide (NO) synthase in the cat testis were studied using enzyme histochemical and immunohistochemical techniques. Stage-dependent nuclear and cytoplasmic eNOS/iNOS immunoreactivity and cytoplasmic NADPH-d reactivity were found in all germ cells, including spermatogonia, primary spermatocytes (preleptotene, zygotene, and pachytene spermatocytes), and round (Sa, Sb1) and elongating spermatids (Sb2, Sc) of the seminiferous epithelium. The pachytene spermatocytes exhibited strong positive reactions at all spermatogenic stage. Interestingly, in elongated spermatids (Sd1) at stages VI to VII, eNOS and iNOS immunostainings was observed only in the cytoplasm but not in the nuclei. eNOS and iNOS immunolabeling was observed in the acrosomal vesicle of some round spermatids (Sb1) at stages I, VII, and VIII, and in the acrosomal cap of elongating spermatids (Sb2) at stage II. Furthermore, eNOS, iNOS, and NADPH-d reactions in elongated spermatids (Sd2) just before spermiation at stage VIII were restricted only to the middle and principal pieces of the tail. Positive reactions were also observed in the Sertoli and Leydig cells as well as in other tissues including vascular endothelial and smooth muscle cells and peritubular myoid cells. These results suggest that NO may play an important role in chromatin condensation, spermatid shaping, and the final release of sperm from the spermatogenic epithelium. Furthermore, NO may also be involved in spermiogenesis, steroidogenesis, and apoptotic cell death.