NHE3 inhibition activates duodenal bicarbonate secretion in the rat

Abstract

We examined the effect of inhibition of Na ϩ /H ϩ exchange (NHE) on duodenal bicarbonate secretion (DBS) in rats to further understand DBS regulation. DBS was measured by using the pH-stat method and by using CO2-sensitive electrodes. 5-(N,N-dimethyl)-amiloride (50 M; DMA), a concentration that selectively inhibits the NHE isoforms NHE1 and NHE2, but not NHE3, did not affect DBS. Nevertheless, 3 mM DMA, a higher concentration that inhibits NHE1, NHE2, and NHE3, significantly increased DBS. Moreover, S1611 and S3226, both specific inhibitors of NHE3 only, or perfusion with Na ϩ -free solutions, dose dependently increased DBS, as measured by pH-stat and CO2-sensitive electrode, without affecting intracellular pH. Coperfusion with 0.1 M indomethacin, 0.5 mM DIDS, or 1 mM methazolamide did not affect S3226-induced DBS. Nevertheless, coperfusion with 0.1 and 0.3 mM 5-nitro-2-(3-phenylpropylamino) benzoic acid, which inhibits the cystic fibrosis transmembrane conductor regulator (CFTR), dose dependently inhibited S3226-induced DBS. In conclusion, only specific apical NHE3 inhibition increased DBS, whereas prostaglandin synthesis, Na ϩ -HCO 3 Ϫ cotransporter activation, or intracellular HCO 3 Ϫ formation by carbonic anhydrase was not involved. Because NHE3 inhibition-increased DBS was inhibited by an anion channel inhibitor and because reciprocal CFTR regulation has been previously shown between NHE3 and apical membrane anion transporters, we speculate that NHE3 inhibition increased DBS by altering anion transporter function. epithelial cells; cystic fibrosis transmembrane conductance regulator; back titration; S3226 DUODENAL EPITHELIAL BICARBONATE secretion (DBS) is one of the most important mechanisms by which the duodenum is protected from the injurious effects of secreted gastric acid (6, 16). DBS is regulated by humoral factors such as PGE 2 , VIP, glucagon, gastric inhibitory peptide, and the enteric nervous system (6). These factors promote cAMP production, which stimulates the cystic fibrosis transmembrane conductor regulator (CFTR), an apical anion channel (19,20), and the basolateral Na ϩ -HCO 3 Ϫ cotransporter (NBC1), an HCO 3 Ϫ uptake pathway (4,22,35). Recently, six Na ϩ /H ϩ exchanger (NHE) isoforms have been cloned. Of these isoforms, NHE1, 2, and 3 are expressed in the intestine of humans, rabbits, and rats (10, 21). In the small intestine, particularly in the duodenum, apical NHE2 and NHE3 are expressed in human, rabbits, rats, and mice (10,21,34). In the colonic surface mucosa, an apical NHE3 plays a key absorptive role for Na ϩ , concomitant with H ϩ excretion (31). In contrast, NHE2 may promote Na ϩ absorption from colonic crypts (12), whereas playing only a minor role in overall small intestinal electrolyte transport (18,29).In a recently published clinical study (36), inhibition of NHE2 and NHE3 by amiloride increased DBS. This increased DBS was thought to result from decreased NHE2-and NHE3-mediated H ϩ secretion into the lumen, increasing the amount of measured titratable alkalinity. Although it...

CFTR inhibition augments NHE3 activity during luminal high CO2exposure in rat duodenal mucosa

“…In contrast, S3226, which had no effect on the acid-induced H ϩ loss or PV acidification (Fig. 5, B-D), increased total CO 2 output, similar to the previously observed augmented HCO 3 Ϫ secretion during NHE inhibition (15). These results suggest that, during luminal acid perfusion, HCO 3 Ϫ secretion and transmucosal CO 2 and H ϩ movement are mediated by CFTR and DIDS-sensitive transporters but not by NHE3.…”

“…Another possibility is that DIDS inhibits basolateral NBC activity, lowering intracellular [HCO 3 Ϫ ], increasing the gradient for HCO 3 Ϫ uptake from lumen to cell (2). Furthermore, although S3226 increased HCO 3 Ϫ secretion via CFTR activation (15), S3226 has no effect on CO 2 /HCO 3 Ϫ movement during CO 2 exposure (27), suggesting that the presence of luminal HCO 3 Ϫ or CO 2 may mask S3226-induced HCO 3 Ϫ secretion. Therefore, DIDS most likely impairs basolateral HCO 3 Ϫ uptake due to NBC inhibition, diminishing the effect of S3226-induced CFTR activation on HCO 3 Ϫ secretion.…”

“…In the duodenum, where SLC26A is implicated in HCO 3 Ϫ secretion rather than in Cl Ϫ absorption, this mechanism is less clear. We have shown that the selective NHE3 inhibitor, S3226, increases duodenal HCO 3 Ϫ secretion via CFTR activation rather than by decreasing the neutralization of HCO 3 Ϫ by NHE3-secreted H ϩ (15).…”

“…The transport function of NHE3, which plays an important role in lower intestinal Na ϩ absorption, (33) is unclear in the duodenum. We have previously shown that NHE3 is not involved in CO 2 /H ϩ movement between the lumen and cell during high CO 2 exposure (27), nor is it involved in the regulation of enterocyte intracellular pH (pH i ) at neutral luminal pH (15).…”

“…CFTR inh-172 inhibition can mimic CFTR dysfunction in human tracheal epithelia (29). To examine the effect of the inhibition of NHE3 or anion exchanger/Na ϩ :HCO 3 Ϫ cotransporter (NBC), S3226 (10 M) (15,27) or DIDS (0.1 mM) (14), respectively, were perfused with the high CO 2 solution or acid solution. We confirmed that S3226 or DIDS in these concentrations had no effect on solution pH or [CO 2].…”