In the current study, localization of D-aspartic acid (D-Asp) in rat testis was studied by immunohistochemical and biochemical techniques. Immunohistochemical staining of this tissue using specific polyclonal antibody to D-Asp revealed D-Asp immunoreactivity (IR) in the cytoplasm of germ cells, especially around the region rich in elongate spermatids, the most mature of the germ cells. Weak IR was also noted in cytoplasm of spermatocytes and round spermatids; however, it was negligible in interstitial cells and Sertoli cells. The intensity of immunostaining in each seminiferous tubule differed according to its distinct germ cell composition. In testis of young rats, seminiferous tubules lack elongated spermatids, and D-Asp was found to be localized in spermatocytes, the most mature population of germ cells at that age. We used various toxicants to destroy specific testicular cell populations and to confirm the localization of D-Asp in rat testis. Administration of ethane dimethane sulfonate induced a selective destruction of all Leydig cells in this tissue. This resulted in a significant decrease in the D-Asp level, which was probably due to a drop in testosterone brought about by this treatment, and this was followed by a modulation of spermatogenesis. Three days after treatment with methoxyacetic acid (MAA), many seminiferous tubules were found to lack or to have severe depletions of pachytene spermatocytes, but not of elongate spermatids. This caused reductions in protein content and in the total amount of L-Asp, but not that of D-Asp. Twenty days after treatment with MAA, the depleted population of germ cells progressed through the spermatogenic cycle from pachytene spermatocytes to elongate spermatids. At this time, the level of D- Asp decreased significantly, as did that of L-Asp and protein, consistent with D-Asp localization in elongate spermatids. This decrease in the D-Asp level was also seen with immunostaining.
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