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

C. Wu*, G. Sui and C. H. Fry

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 clamp. 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.

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