SOURCES OF CALCIUM UTILIZED IN CHOLINERGIC RESPONSES IN CANINE COLONIC SMOOTH-MUSCLE

Ratiometric fura 2 fluorescence techniques were used to investigate th e sources of Ca2+ that lead to an increase in cytosolic Ca2+ concentra tion ([Ca2+](i)) and the generation of force during cholinergic stimul ation of canine colonic circular smooth muscle tissues. Acetylcholine (ACh; 1 mu M) caused a biphasic increase in [Ca2+](i) and force. The i nitial upstroke phase was characterized by an increase in [Ca2+](i) an d a pronounced increase in force. The sustained phase was characterize d by concurrent oscillations in [Ca2+](i) and force (2-3/ min) that pe rsisted as long as ACh was present. The increase in [Ca2+](i) in respo nse to ACh was reduced to similar to 30% in the presence of nicardipin e (1 mu M), suggesting that L-type Ca2+ channels contribute to the ris e in [Ca2+](i) but that other sources also contribute. Preincubation i n caffeine (10 mM) and ryanodine (10 mu M) reduced the upstroke phase of the increase in [Ca2+](i) and contractile responses to ACh, indicat ing that release of Ca2+ from intracellular stores contributes only to the initial cholinergic response. Responses to ACh persisted when nic ardipine (1 mu M) was included after emptying of caffeine-ryanodine-se nsitive stores, suggesting the presence of additional sources of Ca2+. Data suggest that cholinergic regulation of [Ca2+](i) in colonic smoo th muscle occurs by a number of parallel pathways. Influx through volt age-dependent Ca2+ channels, release of Ca2+ from intracellular stores , and possibly Ca2+ entry through additional conductances activated by ACh all contribute to the regulation of [Ca2+](i).