During muscarinic activation of canine tracheal smooth muscle with carbachol, myosin phosphorylation is significantly more sensitive than stress to the external Ca2+ concentration ([Ca2+](o)) [W.T. Gerthoffer. Am. J. Physiol. 250 (Cell Physiol. 19): C597-C604, 1986]. To determine whether the intracellular Ca2+ concentration ([Ca2+](i)) correlated more closely with changes in phosphorylation or force, we measured isometric force and light emitted by the luminescent intracellular Ca2+ indicator aequorin as [Ca2+](o) was increased in the presence of 1 μM carbachol or 60 mM K+. Myosin phosphorylation was measured using an immunoblot assay in a second set of muscle strips treated identically. Stimulation with carbachol increased aequorin luminescence slightly in strips incubated in Ca2+-free solution. Active stress and aequorin luminescence subsequently increased in parallel as [Ca2+](o) was increased. Myosin phosphorylation at 0.05 mM [Ca2+](o) (0.30 ± 0.04 mol P(i)/mol light chain) was significantly higher than phosphorylation in Ca2+-free solution with no carbachol (0.12 ± 0.048 mol P(i)/mol light chain) and increased to a maximum of 0.56 ± 0.03 mol P(i)/mol light chain at 1.6 mM [Ca2+](o). In contrast, active stress and aequorin luminescence remained low at low [Ca2+](o) and reached a maximum at 2.4 mM [Ca2+](o). Stimulation with carbachol produced greater increases in myosin phosphorylation and active stress for a given change in aequorin luminescence than did K+ depolarization. Stimulation with carbachol also produced a different phosphorylation-stress relationship than did K+ depolarization. These observations are consistent with the possibility that carbachol induces increases in the Ca2+ sensitivity of contractile proteins in tracheal smooth muscle.
|Original language||English (US)|
|Journal||American Journal of Physiology - Cell Physiology|
|State||Published - Dec 1 1989|
ASJC Scopus subject areas
- Cell Biology