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 micromol/l, basolateral), via an increase in cytosolic Ca2+ concentration ([Ca2+]i), activated a transient lumen-negative equivalent short-circuit current (Isc) [change (Delta) in Isc = -79.4 +/- 7.5 microA/cm2]. 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 micromol/l IBMX and 1 micromol/l forskolin) activated a sustained lumen-negative current (DeltaIsc = -42.4 +/- 7.2 microA/cm2) that was inhibited by basolateral trans-6-cyano-4-(N-ethylsulfonyl-N-methylamino)-3-hydroxy-2, 2-dimethyl&2-chromane (10 micromol/l), a blocker of KvLQT1 channels. In the presence of elevated cAMP, the CCH-activated currents were augmented (DeltaIsc = 167.7 +/- 32.7 microA/cm2), suggesting cooperativity of the Ca2+- and cAMP-mediated responses. Inhibition of endogenous cAMP production by indomethacin (10 micromol/l) significantly reduced CCH-activated currents and even reversed the polarity in 70% of the experiments. The transient lumen-positive Isc 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 micromol/l, basolateral), an increase of cAMP activated a sustained negative Isc. Under these conditions, CCH induced a large further increase in lumen-negative Isc (DeltaIsc = -100.0 +/- 21.0 microA/cm2). 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.