Carbachol induces oscillations in membrane potential and intracellular calcium in a colonic tumor cell line, HT-29.
The patch-clamp technique was used to study the effects of carbachol (CCh) on HT-29 cells. During CCh exposure, the cells (n = 23) depolarized close to the equilibrium potential for Cl- (E(Cl-); -48 mV) and the membrane potential then started to oscillate (16/23 cells). In voltage-clamp experiments, similar oscillations in whole cell currents could be demonstrated. The whole cell conductance increased from 225 +/- 25 pS in control solution to 6,728 +/- 1,165 pS (means +/- SE, n = 17). In substitution experiments (22 mM Cl- in bath solution, E(Cl-) = 0 mV), the reversal potential changed from -41.6 +/- 2.2 mV (means +/- SE, n = 9) to -3.2 +/- 2.0 mV (means +/- SE, n = 7). When the cells were loaded with the calcium-sensitive fluorescent dye, fluo 3, and simultaneously patch clamped, CCh caused a synchronous oscillating pattern of fluorescence and membrane potential. In cell-attached patches, the CCh-activated currents reversed at a relative membrane potential of 1.9 +/- 3.7 mV (means +/- SE, n = 11) with control solution in the pipette and at 46.2 +/- 5.3 mV (means +/- SE, n = 10) with a 15 mM Cl- solution in the pipette. High K+ (144 mM) did not change the reversal potential significantly (P < or = 0.05, n = 8). In inside-out patches, calcium-dependent Cl- channels could be demonstrated with a conductance of 19 pS (n = 7). It is concluded that CCh causes oscillations in membrane potential that involve calcium-dependent Cl- channels and a K+ permeability.
Sand
P
P
Department of Physiology and Pharmacology, Karolinska Institutet, Stockholm, Sweden.
Svenberg
T
T
Rydqvist
B
B
D002118:Calcium / Q000378:metabolism*; D002217:Carbachol / Q000494:pharmacology*; D002462:Cell Membrane / Q000187:drug effects / Q000502:physiology*; D002712:Chlorides / Q000494:pharmacology; D003110:Colonic Neoplasms; D006801:Humans; D007700:Kinetics; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D013997:Time Factors; D014407:Tumor Cells, Cultured
D002712:Chlorides; D002217:Carbachol; D002118:Calcium