Elevations of intracellular calcium reflect normal voltage-dependent behavior, and not constitutive activity, of voltage-dependent calcium channels in gastrointestinal and vascular smooth muscle

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McCarron, J.G. and Olson, M. and Currie, S. and Wright, A. and Anderson, K.I. and Girkin, J.M. (2009) Elevations of intracellular calcium reflect normal voltage-dependent behavior, and not constitutive activity, of voltage-dependent calcium channels in gastrointestinal and vascular smooth muscle. Journal of General Physiology, 133 (4). pp. 439-457. ISSN 0022-1295 (https://doi.org/10.1085/jgp.200810189)

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Abstract

In smooth muscle, the gating of dihydropyridine-sensitive Ca2+ channels may either be stochastic and voltage dependent or coordinated among channels and constitutively active. Each form of gating has been proposed to be largely responsible for Ca2+ influx and determining the bulk average cytoplasmic Ca2+ concentration. Here, the contribution of voltage-dependent and constitutively active channel behavior to Ca2+ signaling has been studied in voltage-clamped single vascular and gastrointestinal smooth muscle cells using wide-field epifluorescence with near simultaneous total internal reflection fluorescence microscopy. Depolarization (-70 to +10 mV) activated a dihydropyridine-sensitive voltage-dependent Ca2+ current (ICa) and evoked a rise in [Ca2+] in each of the subplasma membrane space and bulk cytoplasm. In various regions of the bulk cytoplasm the [Ca2+] increase ([Ca2+]c) was approximately uniform, whereas that of the subplasma membrane space ([Ca2+]PM) had a wide range of amplitudes and time courses. The variations that occurred in the subplasma membrane space presumably reflected an uneven distribution of active Ca2+ channels (clusters) across the sarcolemma, and their activation appeared consistent with normal voltage-dependent behavior. Indeed, in the present study, dihydropyridine-sensitive Ca2+ channels were not normally constitutively active. The repetitive localized [Ca2+]PM rises ("persistent Ca2+ sparklets") that characterize constitutively active channels were observed rarely (2 of 306 cells). Neither did dihydropyridine-sensitive constitutively active Ca2+ channels regulate the bulk average [Ca2+]c. A dihydropyridine blocker of Ca2+ channels, nimodipine, which blocked ICa and accompanying [Ca2+]c rise, reduced neither the resting bulk average [Ca2+]c (at -70 mV) nor the rise in [Ca2+]c, which accompanied an increased electrochemical driving force on the ion by hyperpolarization (-130 mV). Activation of protein kinase C with indolactam-V did not induce constitutive channel activity. Thus, although voltage-dependent Ca2+ channels appear clustered in certain regions of the plasma membrane, constitutive activity is unlikely to play a major role in [Ca2+]c regulation. The stochastic, voltage-dependent activity of the channel provides the major mechanism to generate rises in [Ca2+].

ORCID iDs

McCarron, J.G. ORCID logoORCID: https://orcid.org/0000-0002-3302-3984, Olson, M., Currie, S. ORCID logoORCID: https://orcid.org/0000-0002-4237-4428, Wright, A., Anderson, K.I. and Girkin, J.M.;
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