Potential parameters of male reproductive toxicity: reproductive performance, histopathology and sperm evaluation in SD rats given nitrazepam.
The present study was designed to elucidate the correlation between findings from reproductive performance testing and those from histopathological examination of the testis and sperm analysis in rats given a benzodiazepine derivative, nitrazepam, for 2 and 4 weeks. The mechanisms of toxicological action of nitrazepam on the male reproductive organs were also investigated. Nitrazepam was given orally to Sprague-Dawley male rats (6-week-old) at a daily dose of 80 mg/kg for 2 weeks or at daily doses of 20, 40 or 80 mg/kg for 4 weeks. Treated males were mated to examine reproductive performance with untreated females after each dosing period, and after 4 and 9 week of recovery periods. Necropsy was performed for histopathological examination of the testis and epididymis and for sperm analysis after each dosing period and the final mating trial (total of 11 weeks recovery). In the findings from reproductive performance testing, significant decrease in the fertility index was observed in the 80 mg/kg group even after 2 weeks dosing and thereafter until 4 weeks recovery, though the mating index did not significantly differ from that of controls through the experiment. In the histopathological examination and sperm analysis, testicular signs of toxicity, decrease in number of sperm heads in the testis and increase in number of sperm with abnormal heads in the seminiferous tubules were noted in the 80 mg/kg group after 2 weeks dosing and in the 40 and 80 mg/kg groups after 4 weeks dosing. Concentrations of plasma testosterone and content of testis testosterone in nitrazepam-treated groups were not significantly different from those of controls. Plasma FSH concentration was significantly elevated in the 80 mg/kg group through the experiment, although significant elevation of plasma LH was observed only after 2 weeks dosing. These results indicate that histopathological examination is the most reliable approach to detect male reproductive adverse effects induced by nitrazepam rather than using parameters from mating trials. The four-week-dosing period is appropriate for their detection. Hypospermatogenesis induced by nitrazepam is suggested to be caused by direct action of nitrazepam on germ cells and/or Sertoli cells rather than by indirect action through inhibition of testosterone secretion.