CFTR-mediated anion secretion in parathyroid hormone-treated Caco-2 cells is associated with PKA and PI3K phosphorylation but not intracellular pH changes or Na⁺/K⁺-ATPase abundance

Abstract

Parathyroid hormone (PTH) has previously been shown to enhance the transepithelial secretion of Cl− and HCO3− across the intestinal epithelia including Caco-2 monolayer, but the underlying cellular mechanisms are not completely understood. Herein, we identified the major signaling pathways that possibly mediated the PTH action to its known target anion channel, i.e., cystic fibrosis transmembrane conductance regulator anion channel (CFTR). Specifically, PTH was able to induce phosphorylation of protein kinase A and phosphoinositide 3-kinase. Since the apical HCO3− efflux through CFTR often required the intracellular H+/HCO3− production and/or the Na+-dependent basolateral HCO3− uptake, the intracellular pH (pHi) balance might be disturbed, especially as a consequence of increased endogenous H+ and HCO3− production. However, measurement of pHi by a pH-sensitive dye suggested that the PTH-exposed Caco-2 cells were able to maintain normal pH despite robust HCO3− transport. In addition, although the plasma membrane Na+/K+-ATPase (NKA) is normally essential for basolateral HCO3− uptake and other transporters (e.g., NHE1), PTH did not induce insertion of new NKA molecules into the basolateral membrane as determined by membrane protein biotinylation technique. Thus, together with our previous data, we concluded that the PTH action on Caco-2 cells is dependent on PKA and PI3K with no detectable change in pHi or NKA abundance on cell membrane.