Ionic currents activated via purinergic receptors in the cumulus cell-enclosed mouse oocyte

Abstract

Several chemical signals synthesized in the ovary, including neurotransmitters, have been proposed to serve as regulators of folliculogenesis, however, their mechanisms of action have not been completely elucidated. Here, electrophysiological and molecular biology techniques were used to study responses generated via purinergic stimulation in cultured mouse cumulus cell-enclosed oocytes (CEOs). Application of extracellular ATP elicited depolarizing responses in CEOs. Using the voltage clamp technique by impaling oocytes with two microelectrodes, we determined that these responses were mainly due to activation of two distinct ionic currents. The first corresponded to the opening of Ca2+-dependent Cl- channels (I(Cl(Ca))) and the second to the opening of Ca2+-independent channels that are permeable to Na+ (I(c+)). The potency order for different nucleotides (50 micro M) was UTP > ATP > 2meS-ATP > ADP, and alpha,betame-ATP and adenosine were found to be inactive. Suramin (100 micro M) blocked the response elicited by ATP or UTP. In addition, voltage dependent K+ currents activated by depolarization of CEOs were characterized. All CEO ionic currents recorded from the oocyte were completely inhibited by octanol (1 mM), a gap junction blocker. Thus, purinergic responses and K+ currents originate mainly in the membrane of cumulus cells. Transcripts of the purinergic receptor P2Y2 subtype were amplified by polymerase chain reaction from the cDNA of granulosa cells or cumulus cells. This study shows that P2Y2 receptors are expressed in CEOs, and that their stimulation opens at least two different types of ion channels. Both the ion channels and the receptors seemed to be located in the cumulus cells, which transmit their corresponding electrical signals to the oocyte via gap junction channels.