Role of intracellular cyclic adenosine 3',5'-monophosphate concentration and oocyte-cumulus cells communications on the acquisition of the developmental competence during in vitro maturation of bovine oocyte.
The present study was designed to address the physiological role played by cAMP on gap junction (GJ) mediated communications between oocyte and cumulus cells during in vitro maturation. Cyclic AMP was stimulated by different collection and maturation media known to induce different rates of nuclear maturation and developmental competence as well as different levels of cumulus expansion. Cumulus-oocyte complexes (COCs) were matured for 0, 3, 7, 12, 18, and 24 h in the absence of stimulation or in the presence of serum and gonadotropins (fetal bovine serum+human menopausal gonadotropins [FCS+hMG]) or 0.01 microg/ml of invasive adenylate cyclase (iAC). For each time point, intracellular cAMP concentration ([cAMP]i) was determined either in the whole COC or oocyte after cumulus cell removal. GJ functional status was analyzed by microinjection of Lucifer yellow fluorescent dye in cumulus-enclosed oocytes and by immunohistochemical localization of connexin 43 (Cx43). In the absence of stimulation, [cAMP]i in COC and oocyte was lower than in other groups, and communications declined after 3 h of culture. In the FCS+hMG group, [cAMP]i increased significantly in COC, with a peak between 3 and 7 h that was temporally correlated with the beginning of the cumulus expansion process, which occurred only in this group and with the termination of the communications. COC matured in the presence of iAC showed a moderate increase of [cAMP]i during all of the maturation times as well as a prolongation of oocyte-cumulus cell communications. The immunohistochemical localization of Cx43 confirmed the delay in connexons protein turnover in iAC-treated COCs. Our results show that cumulus expansion and oocyte developmental competence are induced by different levels of cAMP and that its intracellular concentration may affect cell coupling between oocyte and cumulus cells. We hypothesize that the higher developmental competence of COCs matured in the presence of iAC could be achieved through a moderate increase of intracellular cAMP, which in turn determines a prolongation of communications between the two cell types.
Luciano
Alberto M
AM
Institute of Anatomy of Domestic Animals, Histology and Embryology, Faculty of Veterinary Medicine, University of Milan, Milan, 20133, Italy. alberto.luciano@unimi.it
Modina
Silvia
S
Vassena
Rita
R
Milanesi
Elisabetta
E
Lauria
Antonio
A
Gandolfi
Fulvio
F
D000818:Animals; D002417:Cattle; D002450:Cell Communication / Q000502:physiology*; D004396:Coloring Agents / Q000493:pharmacokinetics; D018031:Connexin 43 / Q000378:metabolism; D000242:Cyclic AMP / Q000378:metabolism*; D005314:Embryonic and Fetal Development; D005260:Female; D005307:Fertilization in Vitro; D017629:Gap Junctions / Q000502:physiology; D007150:Immunohistochemistry; D066298:In Vitro Techniques; D009865:Oocytes / Q000166:cytology* / Q000502:physiology*
D004396:Coloring Agents; D018031:Connexin 43; D000242:Cyclic AMP