Purinergic receptor-induced changes in paracellular resistance across cultures of human cervical cells are mediated by two distinct cytosolic calcium-related mechanisms

Abstract

In human cervical (CaSki) cells, extracellular adenosine triphosphate (ATP) induces an acute decrease in the resistance of the lateral intercellular space (RLIS), phase I response, followed by an increase in tight junctional resistance (RTJ), phase II response. ATP also stimulates release of calcium from intracellular stores, followed by augmented calcium influx, and both effects have similar sensitivities to ATP (EC50 of 6 microM). The objective of the study was to determine the degree to which the changes in [Ca2+]i mediate the responses to ATP. 1,2-bis (2-aminophenoxy) ethane-N,N,N1,N1-tetraacetic acid (BAPTA) abrogated calcium mobilization and phase I response; in contrast, nifedipine and verapamil inhibited calcium influx and attenuated phase II response. Barium, La3+, and Mn2+ attenuated phase I response and attenuated and shortened the ionomycin-induced phase I-like decrease in RLIS, suggesting that store depletion-activated calcium entry was inhibited. Barium and La3+ also inhibited the ATP-induced phase II response, but Mn2+ had no effect on phase II response, and in the presence of low extracellular calcium it partly restored the increase in RTJ. KCl-induced membrane depolarization stimulated an acute decrease in RLIS and a late increase in RTJ similar to ATP, but only the latter was inhibited by nifedipine. KCl also induced a nifedipine-sensitive calcium influx, suggesting that acute increases in [Ca2+]i, regardless of mobilization or influx, mediate phase I response. Phase II-like increases in RTJ could be induced by treatment with diC8, and were not affected by nifedipine. Biphasic, ATP-like changes in RTE could be induced by treating the cells with ionomycin plus diC8. We conclude that calcium mobilization mediates the early decrease in RLIS, and calcium influx via calcium channels activates protein kinase C and mediates the late increase in RTJ.

P2X7 receptor expression is decreased in epithelial cancer cells of ectodermal, uro-genital sinus, and distal paramesonephric duct origin

“…1g-j). Previous studies also reported preferential sorting of the P2X 7 receptor into apical domains of the plasma membrane in cervical epithelial cells [40], as well as of other purinergic receptors (P2Y 2 and P2X 4 [47][48][49][50]). However, the biological significance of these findings is at present unclear.…”

Changes in Tight Junctional Resistance of the Cervical Epithelium Are Associated with Modulation of Content and Phosphorylation of Occludin 65-Kilodalton and 50-Kilodalton Forms

“…1A). ATP phase-I and phase-II responses were previously described and characterized (25)(26)(27). However, until recently, less was known about the phase-III response, and one of our objectives was to better understand the mechanisms that mediate the ATP-induced late decrease in R TE .…”

Purinergic Signalling in the Reproductive System

“…The P2X 4 effect is mediated by increased Ca 2+ influx, Ca 2+ i -dependent activation of phospholipase (PL)-D, upregulation of diacylglycerol and PKC-mediated threonine dephosphorylation of the tight-junctional protein occludin (Gorodeski, 2002a;Zhu et al, 2006). The data suggest that fluid and solutes secretion in vivo is determined by the combined effects of P2Y 2 and P2X 4 receptors signalling (Gorodeski et al, 1998a). The increases in P2X 7 A immunoreactivity in apical regions of the plasma membranes (facing the lumen) represent P2X 7 A aggregates composing P2X 7 A pores.…”

“…HeLa cervical epithelial cells express P2Y 1 , P2Y 2 , P2Y 6 and P2X 7 receptors (Welter-Stahl et al, 2009). Human normal endocervical and ectocervical epithelial cells express P2Y 2 , P2X 4 , and P2X 7 receptors (Meldrum and Burnstock, 1985;Wali and Greenidge, 1989;Pinna et al, 1997;Gorodeski, 1996aGorodeski, , 2008Gorodeski et al, 1994a,b;Gorodeski et al, , 1996Gorodeski and Goldfarb, 1997;Gorodeski et al, 1998a;Gorodeski, 2002b). The P2Y 2 and P2X 4 receptors are located on the apical surface of the epithelial cells (Gorodeski, 2000a,b), and are involved in the regulation of epithelial transport (Gorodeski, 2008).…”

“…In the human uterus and cervix, extracellular ATP regulates secretion of fluid and solutes into the lumen via P2Y 2 and P2X 4 receptors mechanisms (Gorodeski, 2002a;Gorodeski et al, 1994a,b;Gorodeski et al, , 1996Gorodeski and Goldfarb, 1997;Gorodeski et al, 1998a;Gorodeski, 2002b). P2Y 2 and P2X 4 receptors are expressed on the apical membrane of endometrial epithelial cells (Gorodeski, 2000a,b), and their expression is stable throughout the menstrual cycle.…”