AJP - Lung Cellular and Molecular Physiology, Vol 264, Issue 5 496-L503, Copyright © 1993 by American Physiological Society

ARTICLES

Calcium mobilization by muscarinic cholinergic stimulation in bovine single airway smooth muscle

J. Kajita and H. Yamaguchi
Department of Physiology, University of Massachusetts Medical School, Worcester 01655.

Cellular mechanisms for Ca2+ mobilization by muscarinic cholinergic stimulation were investigated in dispersed bovine airway smooth muscle cells by the fura-2 method. Because the dissociation constant (Kd) of the intracellular fura-2 was not known, we first determined the Kd as 386 +/- 6 nM, which ranged from 368 to 394 nM. By using the Kd, resting intracellular Ca2+ concentration ([Ca2+]i) was calculated to be 177 +/- 11 nM (n = 22). Carbachol (CCh) at 10(-7) - 10(-5) M caused two distinct increases in [Ca2+]i; an initial transient which reached 1.3-2.7 microM followed by a sustained increase in [Ca2+]i of 250-400 nM. The sustained Ca2+ elevation was not observed in the absence of extracellular Ca2+. D 600 (10(-5) - 3 x 10(-5) M) also eliminated this response. An analogue of diacylglycerol (DAG), 1,2-dioctanoyl-sn-glycerol, caused a gradual increase in [Ca2+]i to 303 +/- 24 nM (n = 13), which was equivalent to the sustained response to 10(-7) and 10(-6) M CCh and was again blocked by D 600. These findings are consistent with the idea that inositol 1,4,5-trisphosphate releases stored Ca2+ as the main cause for the initial transient and that activation of DAG by CCh may result in the sustained increase in [Ca2+]i through voltage-dependent Ca2+ channels.

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