Cholinergic ion secretion in human colon requires coactivation by cAMP

Cl- secretion in the colon can be activated by an increase of either intracellular Ca2+ or cAMP. In this study we examined a possible interdependence of the two second-messenger pathways in human colonic epithelium. When measured in a modified Ussing chamber, carbachol (CCH; 100 mu mol/l, basolateral), via an increase in cytosolic Ca2+ concentration ([Ca2+](i)), activated a transient lumen-negative equivalent short-circuit current (I-sc) [change (Delta) in I-sc = -79.4 +/- 7.5 mu A/cm(2)]. Previous studies indicated that intracellular Ca2+ directly acts on basolateral K+ channels, thus enhancing driving force for luminal Cl- exit. Increased intracellular cAMP (by basolateral addition of 100 mu mol/l IBMX and 1 mu mol/l forskolin) activated a sustained lumen-negative current (Delta I-sc = -42.4 +/- 7.2 mu A/cm(2)) that was inhibited by basolateral trans-6-cyano-4-(N-ethylsulfonyl-N-methylamino)-3-hydroxy-2,2-dimethyl&2-chromane (10 mu mol/l), a blocker of KvLQT1 channels. In the presence of elevated cAMP, the CCH-activated currents were augmented (Delta I-sc = 167.7 +/- 32.7 mu A/cm(2)), suggesting cooperativity of the Ca2+- and cAMP-mediated responses. Inhibition of endogenous cAMP production by indomethacin (10 mu mol/l) significantly reduced CCH-activated currents and even reversed the polarity in 70% of the experiments. The transient lumen-positive I-sc was probably due to activation of apical K+ channels because it was blocked by luminal Ba2+ (5 mmol/l) and tetraethylammonium (10 mmol/l). In the presence of indomethacin (10 mu mol/l, basolateral), an increase of cAMP activated a sustained negative I,,. Under these conditions, CCH induced a large further increase in lumen-negative I-sc (Delta I-sc = -100.0 +/- 21.0 mu A/cm(2)). We conclude that CCH acting via [Ca2+](i) can induce Cl- secretion only in the presence of cAMP, i.e., when luminal Cl- channels are already activated. The activation of a luminal and basolateral K+ conductance by CCH may be essential for transepithelial KCl secretion in human colon.