Non-ionic diffusion and carrier-mediated transport drive extracellular pH regulation of mouse colonic crypts

1. Extracellular pH (pH(o)) regulation within mouse colonic crypt lumens is stimulated by transepithelial gradients of short-chain fatty acids (SCFAs). Current work assesses underlying mechanisms contributing to pH(o) regulation. 2. Crypt luminal alkalinization was saturable by apical SCFA (substrate concentration activating half-maximal transport (K-T) of isobutyrate = 45 mM). However, saturation was consistent with either carrier-mediated SCFA flux or non-ionic diffusion, because the nonionized form was titrated by luminal alkalinization. Direct acidification of apical perfusates increased the magnitude of SCFA-induced luminal alkalinization, roughly in the same proportion to the increased concentration of non-ionized SCFA in the crypt lumen. 3. Transepithelial gradients of an alternative weak acid (CO2) produce pH(o) changes similar to SCFA. In contrast, a weak base (NH3) changes pH(o) with reverse dependence on the orientation of the transepithelial gradient compared with SCFA. Results implicate non-ionic diffusion in pH(o) regulation, and suggest that pH(o) changes may underly SCFA-stimulated bicarbonate secretion and ammonium absorption. 4. SCFA metabolism plays a minor role in extracellular pH regulation. An avidly metabolized SCFA (N-butyrate) augments crypt luminal alkalinization only slightly (0.08 pH units) versus a poorly metabolized SCFA (isobutyrate). 5. Apical addition of 1 mM 4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid (DIDS) partially inhibits luminal alkalinization caused by apical SCFA. DIDS has no effect on luminal alkalinization caused by transepithelial CO2 gradients. Probenecid (1 mM), alpha-cyano-4-hydroxycinnamic acid (4 mM) or basolateral DIDS (1 mM) do not affect pH(o) regulation. Results suggest that DIDS-sensitive, SCPB-dependent transport in the colonocyte apical membrane contributes to pH(o) regulation.