Role of gap junction-mediated communications in regulating large-scale chromatin configuration remodeling and embryonic developmental competence acquisition in fully grown bovine oocyte

Abstract

Purpose This study was aimed to test the hypothesis that gap junction mediated communications (GJC) are required to allow the progressive chromatin configuration remodeling (from GV1 to GV3) process to occur in fully grown oocytes in order to gain the final step of developmental competence acquisition, and that a premature disruption of GJC can alter this process. Methods Bovine cumulus-oocytes complexes collected from medium antral follicles were cultured for 2, 4, 6 and 8 h in the presence of 10 −4 IU/ml of r-hFSH and with 2 mM of the nonselective PDE inhibitor 3-isobutyl-1-methyl-xanthine (IBMX) to prevent meiotic resumption. GJC functionality and chromatin configuration were monitored during the culture period. After meiotic arrest, the developmental capability of oocytes was assessed after IVM and IVF. Results IBMX was effective in significantly sustaining GJC up to 6 h and maintaining meiotic arrest, when compared to control group. Moreover, the percentage of oocytes with less condensed chromatin (GV1) decreased within 4 h of culture, while the proportion of GV2 oocytes gradually increased up to 6 h.Interestingly, a decline in the proportion of GV2 oocytes and an increase in the proportion of GV3 oocytes were observed after 6 h of culture, when the major drop of GJC occurred. On the contrary, when GJC were uncoupled by adding 3 mM of 1-heptanol or through cumulus cells removal, chromatin condensation occurred rapidly throughout the culture period, more promptly in denuded oocytes. Moreover, the maintenance of GJC during meiotic arrest was accompanied by a significant increase of developmental competence compared to the control, as indicated by a higher percentage of hatched blastocysts and blastocyst cell number. Conclusions Altogether, our data indicate that both paracrine and junctional mechanisms are involved in modulating largescale chromatin structure during the final phase of oocyte differentiation.

The Effect of Cilostamide on Gap Junction Communication Dynamics, Chromatin Remodeling, and Competence Acquisition in Pig Oocytes Following Parthenogenetic Activation and Nuclear Transfer1

“…Many of these strategies, often referred to as 'prematuration culture', are based on the temporary blockage of oocyte spontaneous meiotic resumption, by modulating intra-oocyte cAMP content with different agents such as cAMP analogues or PDE inhibitors [12,16,[20][21][22][55][56][57][58][59][60][61][62][63][64][65][66][67]. Our results on the effect of the use of PDE inhibitors on meiotic resumption are largely supportive of previous data in pre-pubertal gilts in which cilostamide or other PDE3 inhibitors, such as milrinone, have been used [17,67,68].…”

“…Temporary blockage of spontaneous meiotic resumption can be achieved by preventing the intra-oocyte drop of cyclic adenosine monophosphate (cAMP) levels, that normally occurs after removal of the COC from the follicle, through the use of inhibitors of phosphodiesterases (PDEs), the enzymes that hydrolyze cAMP ( [15,16]. One of these compounds is cilostamide that specifically inhibits the type 3 PDE (PDE3), which is the major active cAMP-PDE in porcine oocytes [17].…”

PGRMC1 participates in late events of bovine granulosa cells mitosis and oocyte meiosis

“…47 Groups of 20-30 COCs were maintained meiotically arrested by adding 10 mM cilostamide as before described 48,49 until microinjection (minimum 30 minutes). A microinjection apparatus (Narishige Co. Ltd.) mounted on an inverted microscope (Nikon Diaphot; Nikon Corp.) and a Femtojet microinjector (Eppendorf, Hamburg, Germany) were used to inject 10 pl of 20 mM PGRCM1 RNAi or CTRL RNAi into the ooplasm of COC.…”

Cilostazol Improves Developmental Competence of Pig Oocytes by Increasing Intraoocyte Cyclic Adenosine Monophosphate Level and Delaying Meiotic Resumption

“…; Lodde et al. ). PDE3A is an enzyme that has negative effects on oocyte maturation and basically influences spontaneous meiotic resumption of immature oocytes.…”

“…Several culture techniques have been suggested to regain oocyte quality in mammals (Gilchrist and Thompson 2007). Many of these protocols, often indicated to as pre-IVM culture, are followed by the temporary halting of spontaneous meiotic resumption of oocytes by modulating intraoocyte cAMP content with various types of chemicals such as cAMP analogues or PDE inhibitors (Vanhoutte et al 2009;Lodde et al 2013).…”