Involvement of G protein and purines in Rhinella arenarum oocyte maturation.
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.
Instituto Superior de Investigaciones Biológicas (INSIBIO) Universidad Nacional de Tucumán Chacabuco 461 (4000) S.M. de Tucumán. Argentina.
L I
LI
Zelarayán
M T
MT
Ajmat
F
F
Bonilla
M I
MI
Bühler
D000262:Adenylyl Cyclases / Q000378:metabolism; D000818:Animals; D002025:Bufonidae / Q000378:metabolism*; D002467:Cell Nucleus / Q000235:genetics / Q000378:metabolism; D002478:Cells, Cultured; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects; D005260:Female; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000378:metabolism*; D059471:In Vitro Oocyte Maturation Techniques; D036341:Intercellular Signaling Peptides and Proteins; D007546:Isoquinolines; D016200:Maturation-Promoting Factor / Q000378:metabolism; D009865:Oocytes / Q000187:drug effects / Q000502:physiology*; D006080:Ovarian Follicle / Q000378:metabolism; D010455:Peptides / Q000494:pharmacology; D011374:Progesterone / Q000494:pharmacology; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011687:Purines / Q000378:metabolism*; D013449:Sulfonamides
D036341:Intercellular Signaling Peptides and Proteins; D007546:Isoquinolines; C084070:Mas7 protein, synthetic; D010455:Peptides; D047428:Protein Kinase Inhibitors; D011687:Purines; D013449:Sulfonamides; D005576:Colforsin; D011374:Progesterone; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D016200:Maturation-Promoting Factor; D019206:GTP-Binding Protein alpha Subunits, Gi-Go; D000262:Adenylyl Cyclases; C063509:N-(2-(4-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide