P2Y4-mediated regulation of Na+ absorption in the Reissner's membrane of the cochlea

Abstract

The epithelial cells of Reissner's membrane (RM) are capable of transporting Na+ out of endolymph via epithelial Na+ channel (ENaC). However,muchremains to beknownas to mechanism of regulation ofNa+ absorption inRM.WeinvestigatedP2Ysignaling as a possible regulatory mechanism of ENaC in gerbil RM using voltage-sensitive vibrating probe technique and immunohistochemistry. Results showed thatUTPinduced partial inhibition of the amiloride-sensitive short-circuit current but did not change short-circuit current when applied in the presence of amiloride. The inhibitory effect of UTP was not completely reversible in minutes. The response to UTP was inhibited by reactive blue-2 and 2′,3′-O-(4-benzoylbenzoyl)adenosine 5′-triphosphate but not by suramin or pyridoxalphosphate-6-azophenyl- 2′, 4′-disulfonic acid, which indicates this P2Y receptor as the P2Y4 subtype. The phospholipase C (PLC) inhibitors 1-[6[[(17β)-3-methoxyestra-1,3,5(10)-trien-17-yl]amino]hexyl]-1H-pyrrole-2, 5-dione and 1-O-octadecyl-2-O-methyl-rac-glycero-3-phosphocholine markedly inhibited the effect of UTP on ENaC. In contrast, neither modulation of protein kinase C nor application of 2-aminoehoxydiphenyl borate affected P2Y 4-mediated inhibition of ENaC. Immunoreactive staining for P2Y 4 was observed in the RM, apical membrane of stria vascularis, spiral ligament, and organ of Corti, including outer hair cell, inner hair cell, outer pillar cell, Deiters' cell, and Hensen cell. These results suggest that the physiological role of P2Y4 receptor inRMis likely to regulate Na + homeostasis in the endolymph. The acute inhibition of ENaC activity by activation of P2Y4 receptor is possibly mediated by decrease of phosphatidylinositol 4,5-biphosphate in the plasma membrane through PLC activation. Copyright ©2010 the authors.