Acetylcholine-activated inward current induces cytosolic Ca2+ mobilization in mouse C2C12 myotubes

A. Giovannelli, F. Grassi, C. Limatola, E. Mattei, D. Ragozzino, F. Eusebi

Research output: Contribution to journalArticle

Abstract

We examined the spatiotemporal pattern of intracellular Ca2+, liberation in mouse myotubes by means of fluorescence imaging of cytosolic free Ca2+, together with the simultaneous recording of membrane whole-cell currents. Acetylcholine (ACh) applications to C2C12 myotubes equilibrated in Ca2+-free medium and voltage clamped at -50 mV evoked localized fluorescence transients of variable amplitude with less than 0.5 s delay. Under the same experimental conditions, fluorescence transients were elicited by ACh also in mouse primary myotubes. Ca2+ transients were inhibited in myotubes clamped at depolarized potentials (-10 mV to +50 mV), or equilibrated in a Na+,Ca2+-free medium, as well as in cells loaded with heparin, or with inositol (1,4,5 trisphosphate (InsP3). To investigate whether InsP3 could induce Ca2+ mobilization, [Ca2+]i determinations were carried out in myotubes loaded with InsP3 through the whole-cell patch-clamp recording pipette or by extracellular application in permeabilized cells. InsP3 diffusion into the myoplasm caused Ca2+ spikes with 5 ±1 s (mean ± SEM) delay from the rupture of the membrane patch. Spikes were followed by sustained increases in fluorescence or by damped oscillations. In permeabilized myotubes, InsP3 induced the release of sequestered 45Ca2+ with a half-maximally effective concentration (EC50) of 0.28 ± 0.05 μM, and Hill coefficient of 0.79 ± 0.09. It is concluded that the ACh-activated inward current in mouse myotubes is coupled to cytosolic Ca2+ mobilization from internal InsP3-sensitive pools.

Original languageEnglish
Pages (from-to)41-50
Number of pages10
JournalCell Calcium
Volume18
Issue number1
DOIs
Publication statusPublished - 1995

ASJC Scopus subject areas

  • Cell Biology
  • Molecular Biology
  • Physiology
  • Endocrinology

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