Regulation of chloride secretion across porcine endometrial epithelial cells by prostaglandin E2

1. The objective of this study was to investigate the mechanism of PGE(2) regulation of Cl- transport across glandular endometrial cells grown in primary culture. 2. Most of the basal short circuit current (I-sc) was inhibited by luminal addition of 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB) or glibenclamide, suggesting the presence of a basally active Cl- conductance in the apical membrane. 3. Basolateral addition of 10 mu M PGE(2) increased I-sc by 41 +/- 3 mu A. A similar response was observed when cells were treated with 8-(4-chlorophenylthio) adenosine 3',5'-cyclic monophosphate (CPT-cAMP). Pretreatment of monolayers with NPPB and glibenclamide blocked the PGE(2) and cAMP-mediated increase in I-sc, suggesting that the effects of PGE(2) and cAMP were dependent on the activity of an apical NPPB- and glibenclamide-sensitive conductance. 4. Addition of 50 nM antiPGE(2) antibody to the basolateral bathing solution decreased basal I-sc by 20% and shifted the threshold response to exogenous PGE(2). This result suggests autocrine regulation of electrogenic Cl- transport by PGE(2). 5. Experiments with amphotericin B-permeabilized monolayers revealed that the apical PGE(2)-activated, NPPB- and glibenclamide-sensitive conductance was Cl- dependent and that the current-voltage relationship and anion permeation properties (SCN->Br->Cl->I-) were characteristic of the cystic fibrosis transmembrane conductance regulator (CFTR). 6. Cultured porcine endometrial epithelial cells were specifically labelled with an antibody to a peptide sequence within the regulatory domain of CFTR. 7. The effect of PGE(2) was blocked by basolateral addition of bumetanide and furosemide at concentrations that are selective for inhibition of Na+-K+-2Cl(-) cotransport activity. The effect of bumetanide on I-sc was Cl- dependent, suggesting a role for the bumetanide-sensitive transport pathway in Cl- secretion. 8. PGE(2) and cAMP also activated an outwardly rectifying basolateral K+ channel which presumably sustains the driving force for electrogenic Cl- efflux across the apical membrane. 9. The concentration-conductance and concentration-I-sc response relationships for PGE(2) showed that basolateral K+ permeability was rate limiting with respect to transepithelial anion secretion and that activation of a basolateral K+ channel by PGE(2) was necessary to achieve maximum rates of Cl- secretion.