We have described rapid increases in intracellular calcium and pH (pH_i) in human distal colon in response to the mineralocorticoid aldosterone at normal physiological levels (Maguire et al. 1999). In these experiments, we examine the effects of aldosterone on the sodium-proton exchanger (NHE) as measured by the pH_i recovery rate following acid loading using ammonium chloride (NH₄Cl).

Individual human distal colonic crypts, used with local ethics committee approval and informed patient consent, were isolated and loaded with the fluorescent probe 2Ì, 7Ì-bis(carboxyethyl)-5(6)-carboxyfluorescein (BCECF), and changes in pH_i were recorded using spectro-fluorescence microscopy.

A specific inhibitor of the NHE, 5-(N-ethyl-N-isopropyl) amiloride (EIPA, 100 µM) had little effect on control pH_i, suggesting activity of the NHE at resting pH is low. When subjected to a 20 mM NH₄Cl acid load, crypts in a bicarbonate-free Krebs solution underwent a basal recovery rate of 0.018 ± 0.007 pH units min^-1 (mean ± S.E.M., n = 9). Addition of aldosterone at concentrations of 0.1, 1 and 10 nM increased average crypt recovery rates to 0.037 ± 0.006, 0.064 ± 0.02 and 0.114 ± 0.02 pH units min^-1, respectively (all P < 0.005, Student’s unpaired t test for n = 9). Both basal and aldosterone-induced recovery was blocked by EIPA (100 µM). There was an observed heterogeneity along the crypt axis in that recovery was significantly more rapid in surface and middle regions when compared with base regions, although this was less marked at higher concentrations of aldosterone.

Aldosterone-induced recovery was not affected by incubation with spironolactone (25 µM), a competitive antagonist of the mineralocorticoid receptor, actinomycin D (20 µg ml^-1), an inhibitor of DNA transcription, or cycloheximide (20 µg ml^-1), an inhibitor of mRNA translation. Incubation with the PKC inhibitor chelerythrine chloride (1 µM) inhibited average crypt recovery by 39 %. These results, and the rapidly observed effects of aldosterone (< 90 s), indicate a non-genomic mechanism of NHE stimulation, which is at least partially PKC mediated.

These rapid effects of aldosterone on the NHE may represent a novel mechanism for enhancing transepithelial sodium absorption in the colonic crypt.

Maguire, D., MacNamara, B., Cuffe, J.E., Winter, D., Doolan, C.M., Urbach, V., O’Sullivan, G.C. & Harvey, B.J. (1999). Steroids 64, 51-63.