Calcium, Ca2+-ATPase, Calmodulin, and Calbindin D-28KD Localization in Testis of Leptodactylus chaquensis (Anura: Leptodactylidae).

The intracellular localization of Ca2+, Ca2+-ATPase, Calmodulin, and Calbindin D-28KD have been studied in testes of the toad Leptodactylus chaquensis, using ultracytochemical and immunohistochemical techniques. The Ca2+ presences in the nucleus and into the mitochondria of the germ cells, together with the activity of Ca2+-ATPase detected in the nuclear envelope and mitochondrial crests, suggest the participation of this transporter in the storage of Ca2+. In Sertoli cells, Ca2+ deposits were also found in vesicles and lamellar bodies. Calmodulin and Calbindin D-28KD were revealed in the cytoplasm of both cell types. At the spermatozoon level, the cation deposits were located in the subacrosomal space and in the acrosomal vesicle. Ca2+-ATPase activity was observed in the acrosomal and plasma membranes of the gamete that suggests the existence of a transport system responsible for maintaining low cytoplasmic Ca2+ levels. The activity of Ca2+-ATPase and the location of Ca2+ deposits in gamete tail would be related to flagellar movement. The colocalization of Ca2+ deposits and their binding proteins in efferent duct cells would probably be associated with secretory activity. Considering that intracellular Ca2+ is present in different gonadal cells, this work would provide a better understanding of the cation importance in the testicular functions of this species.

Effect of nervous stimulation on ovarian steroid secretion in amphibians.

In the present study, the effect of nerve stimulation on the secretory activity of the ovary of adult females was analyzed for the first time in amphibians. Results revealed that in Rhinella arenarum the stimulation of nerves that supply the gonad induced an increase in estradiol and progesterone secretion, this response showing differences during the reproductive cycle of the species. During the postreproductive period, an increase in estradiol secretion was observed while, in the reproductive period, progesterone secretion increased. Our results suggest that the sympathetic division of the autonomic nervous system would be responsible for this increase, taking into account that, under our experimental conditions, acetylcholine did not affect the endocrine activity of the gonad, while adrenaline (epinephrine) was effective in inducing steroid secretion an effect that could be due to interaction with ß receptors. On the other hand, our data show that the association of adrenaline with follicle-stimulating hormone increased estradiol secretion during the postreproductive period, while the association of catecholamine with LH or hCG increased progesterone secretion during the reproductive period. Our results would suggest that nerve stimulation, mediated by the release of adrenaline, would act synergistically with gonadotrophins to stimulate steroid secretion.

Innervation of amphibian reproductive system. Histological and ultrastructural studies.

In the present study we describe for the first time in anuran amphibians the histological and ultrastructural characteristics of innervation in the female reproductive organs. The observations in Rhinella arenarum revealed the presence of nerve fibers located predominantly in the ovarian hilium and in the oviduct wall. In both organs the nerves fibers are placed near blood vessels and smooth muscles fibers. In the present study the histological observations were confirmed using antibodies against peripherin and neurofilament 200 proteins. Ultrastructural analyses demonstrated that the innervation of the reproductive organs is constituted by unmyelinated nerve fibers surrounded by Schwann cells. Axon terminals contain a population of small, clear, translucent vesicles that coexist with a few dense cored vesicles. The ultrastructural characteristics together with the immunopositive reaction to tyrosine hydroxylase of the nerve fibers and the type of synaptic vesicles present in the axon terminal would indicate that the reproductive organs of R. arenarum females are innervated by the sympathetic division of the autonomic nervous system.