Involvement of G protein and purines inRhinella arenarumoocyte maturation

Abstract

We investigated the participation of G(αi) protein and of intracellular cAMP levels on spontaneous and progesterone-mediated maturation in Rhinella arenarum fully grown follicles and denuded oocytes. Although progesterone is the established maturation inducer in amphibians, Rhinella arenarum oocytes obtained during the reproductive period (competent oocytes) resume meiosis with no need for an exogenous hormonal stimulus if deprived of their enveloping follicular cells, a phenomenon called spontaneous maturation. In amphibian oocytes, numerous signalling mechanisms have been involved in the rapid, non-genomic, membrane effects of progesterone, but most of these are not fully understood. The data presented here demonstrate that activation of the G(αi) protein by Mas-7 induced maturation in non-competent oocytes and also an increase in GVBD (germinal vesicle breakdown) in competent oocytes. Similar results were obtained with intact follicles independent of the season. The activation of adenylyl cyclase (AC) by forskolin seems to inhibit both spontaneous and progesterone-induced GVBD. In addition, the high intracellular levels of cAMP caused by activation of AC by forskolin treatment or addition of db-cAMP inhibited maturation that had been induced by Mas-7 and in a dose-dependent manner. Treatment with H-89, a protein kinase A (PKA) inhibitor, was able to trigger GVBD in a dose-dependent manner in non-competent oocytes and increased the percentages of GVBD in oocytes competent to mature spontaneously. The results obtained with whole follicles and denuded oocytes were similar, which suggested that effects on AC and PKA were not mediated by follicle cells. The fact that Mas-7 was able to induce maturation in non-competent oocytes in a similar manner to progesterone and to increase spontaneous maturation suggests that G(αi) activation could be an important step in meiosis resumption. Thus, the decrease in cAMP as a result of the regulation of the G proteins on AC and the inactivation of PKA by H-89 could contribute to the activation of MPF (maturation promoting factor) and induce maturation of the oocytes of Rhinella arenarum.

Involvement of PLA2, COX and LOX inRhinella arenarumoocyte maturation

“…Two COX inhibitors were assayed: indomethacin (5-100 M), which inhibits both isoforms (COX 1 and COX 2 ), and rofecoxib (50-400 M), a selective COX 2 inhibitor. Although in Rhinella arenarum oocytes P 4 -induced maturation takes place throughout the year, oocytes showed a different P 4 response capacity dependent upon the period in which animals had been captured (Zelarayán et al, 2009). 4).…”

“…P 4 interacts with the oocyte surface and starts a cascade of events that leads to meiosis resumption or germinal vesicle breakdown (GVBD) (Sánchez Toranzo et al, 2006;Zelarayán et al, 2012). At this time, follicular cells release progesterone (P 4 ) to induce meiosis resumption, however mechanisms involved in this process have not been fully elucidated.…”

Validation of Electrochemiluminiscence Immunoassay for Ovarian Steroid Determination in Rhinella arenarum

“…However, we demonstrated that in experiments conducted with R. arenarum follicles incubated in vitro, exogenous P 4 was the most efficient maturation inducer throughout the year, especially during the reproductive season (Arias Torres et al., ). Numerous publications show that exogenous P 4 induces maturation (Zelarayán et al., , Arias Torres et al., ) and ovulation of amphibian oocytes in the absence of gonadotropins (Schuetz, ). In X. laevis and in B. vulgaris , hCG induces the maturation and ovulation of the oocyte by stimulating an increase in endogenous P 4 (Fortune et al., ).…”

“…In amphibians, progesterone (P 4 ) has long been considered the physiological inducer of maturation (Maller and Krebs, ; Zelarayán et al., , ). Nowadays it has been suggested that other steroids such as androgens might be involved in this process in Xenopus laevis (White et al., ) and Rhinella arenarum oocytes (Arias Torres et al., ).…”

Dynamics of steroid‐induced oocyte maturation in three amphibian species: Mathematical modeling and simulation

“…Sex steroid production in amphibians is essential for follicle growth and oocyte maturation. Numerous published works have suggested that progesterone (P 4 ) synthesized by ovarian follicles in response to Gns interacts with the oocyte surface to trigger a complex chain of morphological and biochemistry events that induce GVBD in vitro (Maller, ; Zelarayán, Ajmat, Bonilla, & Bühler, ). For several decades, P 4 was the MIS in amphibian oocytes (Smith, ).…”