A “Living Cell Method” for Testing the Early Effects of Antispermatogenic Compounds: Model Experiments with Two Alkylating Agents ThioTepa and Nitrogen Mustard

Abstract

An accurate identification of stages of spermatogenesis in freshly isolated unstained seminiferous tubules is possible by a simple transillumination. Physiologically, the acrosome and maturation phase spermatids are arranged in bundles or as a continous layer and give rise to light absorption patterns characteristic for different cell associations. The damaged and dying cells cause alterations in the light absorption pattern which helps their specific selection for further analyses by morphological and biochemical methods.In model experiments, we have analyzed the early stage‐specific effects of two alkylating anticancer agents, nitrogen mustard and ThioTepa. Increased light absorption was seen in segments V–VI caused by degenerating early spermatids and mid‐pachytene spermatocytes, and in segments XII–I caused mainly by pachytene spermatocytes. Phase contrast microscopy proved to be valuable in analysis of the phagocytosing function of the Sertoli cells. Electron microscopy revealed that probably the earliest observable alterations were disintegrations of the developing acrosomic system, of the chromatoid body in early spermatids and of the spermatid nuclei during the late acrosome phase. Biochemical analyses revealed an inhibition of physiological DNA and RNA syntheses and an activation of unscheduled repair‐DNA synthesis.The “living cell method” gives new possibilities for combined morphological and biochemical analyses of specific effects of antifertility and anticancer drugs on different stages of mitotic and meiotic cell cycles during spermatogenesis.