The role of the L-type Ca2+ channel in refilling functional intracellular Ca2+ stores in guinea-pig detrusor smooth muscle

Abstract

The transient rise of intracellular Ca2+ in detrusor smooth muscle cells is due to the release of Ca2+ from intracellular stores. However, it is not known how store refilling is maintained at a constant level to ensure constancy of the contractile response. The aim of these experiments was to characterise the role of L-type Ca2+ channels in refilling. Experiments used isolated guinea-pig detrusor myocytes and store Ca2+ content was estimated by measuring the magnitude of change to the intracellular [Ca2+] ([Ca2+]i) after application of caffeine or carbachol. using epifluorescence microscopy. Membrane potential was controlled when necessary by voltage damp. After Ca2+ stores were emptied they refilled with an exponential time course, with a time constant of 88 s. The value of the time constant was similar to that of the undershoot of [Ca2+]i following store Ca2+ release. The degree of store filling was enhanced by maintained depolarisation, or by transient depolarising pulses, and attenuated by L-type Ca2+ channel antagonists. Inhibition of the sarcoplasmic reticular Ca2+-ATPase prevented refilling. Reduction of the resting [Ca2+]i was accompanied by membrane depolarisation; under voltage clamp reduction of [Ca2+]i decreased the number and magnitude of spontaneous transient outward currents. Ca2+ release from intracellular stores, elicited by caffeine or carbachol, is independent of membrane potential under physiological conditions. However, store refilling occurs via Ca2+ influx through L-type Ca2+ channels. Ca2+influx is regulated by a feedback mechanism whereby a fall of [Ca2+]i reduces the activity of Ca2+-activated K+ channels, causing cell depolarisation and an enhancement of L-type Ca2+ channel conductance.