Effects of chronic ethanol administration on the NOS-related NADPH-diaphorase activity in the mouse Leydig cells

INTRODUCTION - Nitric oxide (NO) is a free radical involved in several physiological and pathological processes (1). NO appears to be involved in crucial aspects of male genital physiology, including relaxation of corpora cavernosa and inhibition of sperm mobility and testosterone secretion (2). The gonadotoxic effects of the alcohol were well documented in humans and in animal models (2, 3). In-deed, ethanol administration has been shown to cause morphological alterations on seminiferous tubules (2) and Leydig cells (3) and to decrease testosterone and LH plasmatic levels (4). We examined by TEM the effects of chronic ethanol treatment on the NADPH-diaphorase (NADPH-d) activity in the mouse Leydig cells. MATERIAL AND METHODS - Ten adult Wistar mice were treated with ethanol 0.5g/Kg/die intragastrically for two weeks. The animals were anaesthetised, perfused by aldehydes and treated for NADPH-d histochemistry (5). Specimens incubated with NADPH-free medium were utilised as controls. In order to test the specificity of NADPH-d staining for NOS activity, some specimens were immersed in the medium containing 0.3 mM iodonium diphenyl (6). The specimens were post-fixed in osmium tetroxide, and embedded in Epon 812. RESULTS AND DISCUSSION - About 20% of Leydig cells of the ethanol-treated mice showed morphological alterations. The cells were characterised by irregular protrusions of the plasmatic membrane, rarefaction of the cytoplasmic matrix and increased number of lipid droplets. Irregular mitochondria were also observed. Some Leydig cells (about 10%) showed signs of degeneration. As regards the enzymatic study, the controls animals exhibited the NADPH-d activity in the nuclear cistern, mitochondria and SER. In the ethanol treated-mice the enzymatic reaction was strongly reduced both in apparently normal and injured Leydig cells. A moderate reactivity was detected only in the SER. These findings suggest that chronic ethanol treatment inhibits the NOS-related NADPH-d activity in the Leydig cells of mouse. This effect could be a consequence of the impaired synthesis of the testosterone, inducing an inhibition of NO production through a negative feedback mechanism. However it can not be excluded a direct action of the alcohol on the NOS/cGMP enzymatic pathway.