This study is first to show that chronic oral exposure of a known environmental toxicant and human carcinogen, arsenic, to adult male rats can alter reproductive functions by decreasing the paired testicular mass, inhibiting testicular androgenesis and decreasing concentrations of testosterone and gonadotrophins along with increasing adrenocortical activity. Paired testicular mass, a valuable index of reproductive toxicity in male animals  decreased in arsenic-treated animals and this decrease in testicular mass was consistent with elimination of germ cells . Similar effects were also observed after treatment with oestradiol alone or in combination with arsenic. This antigonadal activity of oestradiol is in agreement with the previous findings [18, 40–43].
Testicular steroidogenic events, Δ5, 3β-HSD and 17β-HSD play a key regulatory role, as these are the prime enzymes in testicular androgenesis [23, 44]. The diminution in these enzymes by sodium arsenite treatment in our study is in agreement with the findings of others, where arsenic treatment was associated with the inhibition of testicular androgenesis . A dose-dependent decrease in plasma and intratesticular concentrations of testosterone in arsenic-treated rats may occur due to the inhibition of these testicular androgenic enzymes activities, because these enzymes are responsible for the regulation of testosterone biosynthesis [23, 45]. Moreover, the inhibition of testicular androgenic enzymes in arsenic-treated rats may be a result of low plasma levels of LH as this is a prime regulator of testicular androgenic enzymes activities [46, 47]. In the present study, the spermatogenic disorder has been reflected by the diminution in the number of different generations of germ cells at stage VII of spermatogenic cycle. These findings also corroborated with the previous findings with arsenic in testis . The inhibition in spermatogenesis is may be due to low level of gonadotrophins i.e. LH and FSH . LH and FSH are required for quantitatively normal spermatogenesis in pubertal rats [22, 49]. The reduction of LH and FSH and consequent reduction in testosterone production may therefore, be held responsible for the arsenic induced changes in spermatogenesis. This prediction for low plasma concentration of LH, FSH and testosterone in arsenic-treated rats are consistent with our results when plasma concentrations of LH, FSH and testosterone have been decreased in treated rats in a dose-dependent manner. Moreover, the reduction in the number of Asg in arsenic-treated rats is possibly due to the low level of FSH, as FSH inhibits the normal degeneration of Asg and reduced FSH secretion may promotes Asg degeneration [15, 48]. Besides this, the germ cell degeneration by arsenic treatment is may be due to low intratesticular concentrations of testosterone, as high level of testosterone in testis is essential for normal spermatogenesis as well as for the maintenance of structural morphology and normal physiology of seminiferous tubule [50, 51]. However the maturation of mPSc and 7Sd are testosterone dependent . The reduction in the number of mPSc and 7Sd following sodium arsenite treatment is due to diminution in testosterone production. In experimental circumstances in which the intratesticular testosterone concentration is reduced a significant germ cell death is seen . Testosterone is required for the attachment of different generations of germ cells in seminiferous tubules and therefore low level of intratesticular testosterone may lead to detachment of germ cells from seminiferous epithelium and may initiate germ cell apoptosis . The decreases in plasma testosterone concentrations may be explained by the fact that the arsenic treatment causes a decrease in plasma LH concentrations. In male rats, circulating LH is responsible for maintaining normal plasma testosterone concentrations . The possibility of low plasma level of gonadotrophins in this experiment may be due to elevated secretion of glucocorticoids from adrenal gland as the high level of corticosterone also observed in arsenic-treated animals. Moreover, arsenic activates the stress signal pathway, the hypophysial- adrenocortical axis and increased the secretion of ACTH from pituitary . The elevation of plasma levels of corticosterone, which may suppress the sensitivity of gonadotroph cells to GnRH and therefore may, prevents gonadotrophin secretion . The high levels of ACTH and corticosterone also directly suppress testosterone production and secretion by decreasing the testicular LH receptor  resulting the reduction of spermatogenesis and epididymal sperm count. The diminution in the epididymal sperm count in arsenic-treated animals is due to the lower concentrations of testosterone as the sperm production in testis and maturation in epididymis is under the control of testosterone . Moreover, the increased activity of testicular acid phosphatase and alkaline phosphatase in arsenic-treated rats also reflects testicular degeneration, which may likely be a consequence of suppressed testosterone and indicative of lytic activity . The activities of SDH and LDH in testicular tissue are associated with the maturation of the germinal epithelial layer of seminiferous tubule [14, 59]. The activity of SDH increased markedly through out the maturation of germ cells and is reported to decrease during the depletion of germ cells . Moreover, the activity of this enzyme is regulated by testosterone and the diminution in this enzyme activity after arsenic exposure is due to the decreased production of testosterone . LDH is reported to be present in higher amounts in the testis of newborn rats and its activity declines with the development of the testis. The increase in activity of LDH and decrease in that of SDH observed in treated animals suggest that this chemical exposure causes deterioration of germinal epithelium [14, 59, 60]. Since the body weight gain was not altered significantly in arsenic-treated rats in comparison to controls. The deleterious effect of arsenic on the male reproductive system may be due to the toxic effect of arsenic itself on this particular system but not to the health of the animals. Sodium arsenite induced diminution in plasma gonadotrophins may be due to the low level of dopamine and elevated levels of noradrenaline and 5-HT in hypothalamus and pituitary as these catacholamines are the important regulators for gonadotrophins secretion and synthesis . However, the diminution in the level of dopamine by arsenic treatment caused the suppressed secretion of gonadotrophins as dopamine has the stimulatory effect on gonadotrophins secretion . Increases in noradrenaline and 5-HT and decrease in dopamine in hypothalamus and pituitary indicate that the effect of the arsenic can be potentiated in an environment in which steroid concentrations are high.
Co- administration of hCG in arsenic-treated rats resulted a significant protection in testicular Δ5, 3β-HSD and 17β-HSD activities, plasma and intratesticular testosterone to the control along with increased total sperm count. The protection of steroidogenesis in arsenic-treated animals by hCG is due to the stimulatory effects of hCG on steroidogenesis [62, 63]. However, in this experiment, administration of hCG along with arsenic partially prevented the degeneration of 7Sd but failed to restored spermatogenesis quantitatively. Although hCG failed to restore the number of spermatocytes and spermatids, but the numbers of Asg were significantly increased compared with that of vehicle control. Though the effect of hCG on Asg at stage VII of the seminiferous cycle in adult rats is not clear from this experiment but the initiation and maintenance of spermatogenesis require LH and FSH in pre-pubertal and pubertal rats  and hCG has a stimulatory effect on the animals own pituitary to secrete FSH, while possessing LH like activity itself [16, 64]. LH stimulates Leydig cells to produce testosterone within the testis. Intratesticular testosterone is an absolute prerequisite for normal spermatogenesis . FSH is also vital for normal testicular function and is necessary for quantitatively normal spermatogenesis . Moreover, hCG maintains the baseline levels of intratesticular testosterone in gonadotrophins withdrawal from exogeneous testosterone administration . The hCG acts as a LH and stimulated the Leydig cells to produce testosterone and that maintains the normal spermatogenesis after gonadotrophin suppression [62, 63]. The restoration of the activities of SDH, LDH, ACP, and ALP in hCG co- administration may be due to the significant protection of germ cells from arsenic induced degeneration. However, the direct detrimental action of arsenic on germ cells can't be ruled out since administration of hCG along with arsenic treatment failed to restore quantitatively the number of all varieties of germ cells at stage VII of the seminiferous cycle.
The spermatogenic degeneration with arsenic treatment in the present work could also be consistent with an estrogen related mechanism. It is quite clear from this experiment that chronic oestradiol treatment in adult rat can induce the same spermatogenic degeneration as induced by arsenic treatment. Moreover, the antispermatogenic effects of arsenic could be enhanced by oestradiol co- administration in arsenic-treated animals. It is widely accepted that oestradiol treatment mimics the effects of hypophysectomy in the testis, due to gonadotrophins and testosterone suppression [17, 18, 67]. Oestradiol also suppressed the transcription of gonadotrophin genes . In our study, oestradiol treatment resulted more than 75% suppression of LH, FSH and testosterone concentrations, respectively; that is its effect on germ cells degeneration should be equivalent to other gonadotrophin suppressing treatments [17, 42, 43]. Moreover, oestradiol exposure has been shown to decrease Leydig cell volume and impair the levels of mRNA and protein of important enzymes of steroidogenesis [69, 70] that is consistent with our result where the key steroidogenic enzymes were diminished after oestradiol treatment. Oestradiol treatment resulted severe degeneration of germ cells at stage VII of seminiferous epithelium cycle. Earlier morphological studies showed that withdrawal of either gonadotrophins  or testosterone  leads to a marked increase in this type of cell death mainly at stage VII. It has been reported that germ cell apoptosis occurs at several stages of the cycle after either gonadotrophin  or testosterone  deprivation. Blanco- Rodriguez and Martinez-Garcia  also showed that oestradiol treatment caused germ cell apoptosis along with suppressed the gonadotrophin and testosterone concentrations in rat.
Several arsenic-treated animals showed reproductive tract abnormalities including gonadotrophins and androgen suppression along with germ cell degeneration in testes, effects similar to those induced by estrogenic agonists, such as di-ethylstilbestrol, methoxychlor and tamoxifen . Further study, demonstrated that the enhanced expression of estrogen receptors (ERs) and various ER- associated gene products were evident in arsenic induced testicular lesions  suggesting that arsenate may exhibit the estrogenic properties.