AJP - Cell Physiology, Vol 265, Issue 4 C877-C886, Copyright © 1993 by American Physiological Society

ARTICLES

Emptying and refilling of Ca2+ store in tracheal myocytes as indicated by ACh-evoked currents and contraction

L. J. Janssen and S. M. Sims
Department of Physiology, University of Western Ontario, London, Canada.

Membrane currents and contractions evoked by acetylcholine (ACh) in freshly dissociated canine tracheal myocytes were investigated using the nystatin perforated-patch recording technique. In cells held at -60 mV in the presence of nifedipine, ACh evoked inward current (IACh) and contraction. Caffeine mimicked the effects of ACh. IACh and contractions could be evoked 3-4 min after removing external Ca2+ but were abolished by prolonged exposure to Ca(2+)-free media. Both responses were restored within minutes of reintroduction of Ca2+, even though the cells were held at -60 mV in the presence of nifedipine. IACh and ACh-evoked contractions were also reversibly abolished by continued exposure to caffeine. Cyclopiazonic acid (CPA), a blocker of the sarcoplasmic reticulum (SR) Ca(2+)-ATPase, reduced IACh by > 95% within 15 min but had little or no effect on the contractile responses evoked by ACh. IACh was restored after washout of CPA even though cells were held at -60 mV. After depleting the Ca2+ store with the use of CPA, depolarization of the membrane to +10 mV immediately before application of ACh led to a partial restoration of IACh. This restorative effect of depolarization was potentiated by Bay K 8644 and antagonized by nifedipine. In conclusion, IACh and contractions in canine tracheal myocytes are mediated by Ca2+ released from an internal store that can be depleted by prolonged removal of extracellular Ca2+, prolonged exposure to caffeine, or by blockade of the SR Ca(2+)-ATPase. At least two Ca2+ influx pathways appear to contribute to refilling of the internal store: one pathway that is not activated by depolarization or ACh and a second involving dihydropyridine-sensitive voltage-activated Ca2+ channels that may be in direct contact with the SR (i.e., conduct extracellular Ca2+ directly into the SR, bypassing the cytosol).

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