The absorption of water and electrolytes by the gallbladder seems to be largely dependent upon a Na+/H+ exchange at the apical membrane of the gallbladder epithelium. To find out if the exchanger involved is the NHE3 isoform, as in other absorbing epithelia, two studies were performed using the rabbit gallbladder. First, we studied 22Na absorption in Ussing chambers with Krebs buffer as a control solution, and in the presence of amiloride (100, 200 or 1000 microM), ethyl-isopropyl-amiloride (EIPA, 1 or 5 microM), or the phorbol ester, phorbol 12-myristate 13-acetate (PMA, 1 microM). A net mucosal-to-serosal Na+ flux was observed with control buffer. No inhibition of this net flux was observed with 5 microM EIPA, and the IC50 for amiloride was found to be 200 microM. PMA induced a reduction of absorption by 30% that was prevented by incubation with calphostin C. Resistance to amiloride and EIPA, and inhibition by PMA are consistent with the involvement of the NHE3 isoform. The second study involved reverse-transcriptase polymerase chain reaction (RT-PCR) of total gallbladder RNA, with two primers designed to amplify a 645-base-pair fragment from NHE3 mRNA. A cDNA fragment of the expected size was actually obtained from gallbladder RNA, while RT-PCR of RNA from the liver, which does not contain NHE3, gave negative results. A sequence of 492 nucleotides of the amplified product was determined, which was almost superimposable onto the known sequence of the corresponding fragment of rabbit NHE3. It is concluded that, in rabbit gallbladder, neutral NaCl absorption is, at least in part, dependent on the NHE3 isoform of the Na+/H+ exchanger.

The finding of a high PCO2 in basally secreted pancreatic juice of man and dog raises the hypothesis of proton secretion from ductal epithelial cells presumably through a Na+/H+ exchanger. To test this possibility, H+ luminal secretion and Na+ movements were measured in vitro on samples of bovine pancreatic ducts mounted in Ussing-type chambers. The rate of luminal acidification measured by the pH stat method, using bicarbonate-free media gassed with 100% O2, reached 2.75 muEq/cm2/hr. Proton secretion was blocked in the presence of 1 nM amiloride or in the absence of Na+ (replaced by choline) in the mucosal solution. Study of transepithelial 22Na fluxes in short-circuited tissue, bathed on both sides by control Ringer solution, gassed by 95% O2-5% CO2 demonstrated a net sodium transport from the mucosal to the interstitial side of the duct (net 22Na flux = 3.23 +/- 0.8 muEq/cm2/hr). This net sodium transport was electroneutral and blocked by mucosal amiloride (0.5-1 mM/liter) or by interstitial ouabain (1 mM/liter). These results are consistent with the existence of a Na+/H+ exchanger on the luminal side of the bovine main pancreatic duct.

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1. In man and in various animal species, absorption of NaCl from bile by the gallbladder mucosa is associated with luminal proton secretion. A similar absorption of NaCl in small intestine, colon and renal tubule is related, at least in part, to the presence of the NHE3 isoform of the Na+/H+ exchanger. This work was designed to find out whether NHE3 is also present in human gallbladder. 2. At surgery, 100-200 mg of the gallbladder wall was obtained from patients treated by cholecystectomy for gallstones. After isolation of the mucosa, total RNA was extracted and submitted to reverse transcription-polymerase chain reaction with two primers: 5'-AAGCCICTGGTGCAGTGGCTGAAGG-3' and 5'-GGAGTCCTTIAAGTCGGCIAAGCTGGGC-3', designed to amplify a sequence of 645 bp of rabbit NHE3 mRNA (642 bp in man). RNA from human liver and from rabbit heart, neither of which contain NHE3, and human ileal RNA, which does contain NHE3, were used as controls. 3. RNA extracted from the mucosal moiety of the gallbladder wall gave an amplification product of about 645 nucleotides. Controls gave the expected negative or positive results. Sequencing of the amplified RNA showed it was almost identical to previously determined sequences of NHE3 in other human tissues. 4. It is concluded that the mucosa of human gallbladder contains the mRNA of NHE3 isoform. This isoform could therefore play a role in sodium absorption from bile.