The Presence of NHE1 and NHE3 Na⁺‐H⁺ Exchangers and an Apical cAMP‐Independent Cl⁻ Channel Indicate that Both Absorptive and Secretory Functions are Present in Calf Gall Bladder Epithelium

Abstract

We investigated the transport systems that can sustain Na+ and Cl− movements across bovine gall bladder epithelium, focusing on the Na+‐H+ exchanger (NHE) family and chloride conductive pathways. Experiments conducted using the fluorescent probe acridine orange (AO) with brush‐border membrane vesicles (BBMV) or vesicles obtained from the total epithelium (EMV) demonstrated the presence of a Na+‐H+ exchange in both preparations. The use of specific inhibitors indicated the presence of an apical NHE3 exchanger and a NHE1 isoform which should reside in the basolateral membrane. Using reverse transcriptase (RT) PCR, we identified cDNA fragments corresponding to the NHE1, NHE3, Cl−‐HCO3− (AE2a) transporters and to the CFTR channel. Using the patch‐clamp technique, we investigated Cl− conductances on cultured epithelial cells. We found a 5 pS Cl− channel with a voltage‐independent open probability, insensitive to stilbenes (SITS), Zn2+ and cAMP. The results suggest that absorption and secretion coexist in calf gall bladder epithelium. A Na+‐H+‐Cl−‐HCO3− double exchange may, at least partially, sustain the absorptive function, and a Cl− apical conductive pathway may be involved in secretion. The conductance we observed does not seem to be cAMP‐regulated, unlike other mammalian gall bladders.

Ion Transport Across the Gallbladder Epithelium

“…Furosemide (10 -4 M) and SITS (10 -4 M), considered typical inhibitors of AE at these concentrations, have an effect similar to that of removing bicarbonate from bathing salines or adding acetazolamide (10 -4 M, inhibitor of carbonic anidrase) [29]. All these data suggest the presence of a Cl -/HCO 3 -antiport which seems to be due to an AE2 isoform, at least in rabbit [31] and calf [20].…”

“…Concerning this function, a potential cooperation between these two isoforms has been postulated [26]. NHE1, possibly responsible of intracellular pH homeostasis, has been detected in rabbit, human and calf gallbladder [20,24,25]. Extracellular HCO 3 -has a stimulatory effect on transepithelial NaCl transport with an apical primary action [27,28] but using a bicarbonate free luminal medium the Cl -influx through the apical membrane is strongly reduced but not completely abolished, at least in rabbit gallbladder [29,30].…”

“…H + secretion is dependent on luminal Na + concentration, following a saturation kinetics, which is compatible with an antiport mechanism. The transport is sensitive to amiloride and its analogs with an IC 50 of 10 -4 -10 -5 M [20,21]. Intracellular pH can regulate salt absorption via changes in Fig.…”

“…Na + /H + , probably also by an allosteric modulation at the cytosolic site [22,23]. NHE3 [20,21,24,25] and NHE2 [21] seem to be the main apical isoforms of the Na + /H + exchanger, involved in transepithelial salt absorption. Concerning this function, a potential cooperation between these two isoforms has been postulated [26].…”

Cholesterol gallstone disease: focusing on the role of gallbladder

“…On the other hand, the gallbladder also has secretive function that is mainly mediated by cystic fibrosis transmembrane conductance regulator (CFTR). 56 Decrease in the secretive function of the epithelium quite possibly translates into enhanced absorption. Thus, the existence of other transporters in GBECs, especially CFTR, may also have a role in the pathogenesis of CGD.…”