Intracellular Cl−fluxes play a novel role in Ca2+handling in airway smooth muscle
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Intracellular Ca(2+) is actively sequestered into the sarcoplasmic reticulum (SR), whereas the release of Ca(2+) from the SR can be triggered by activation of the inositol 1,4,5-trisphosphate and ryanodine receptors. Uptake and release of Ca(2+) across the SR membrane are electrogenic processes; accumulation of positive or negative charge across the SR membrane could electrostatically hinder the movement of Ca(2+) into or out of the SR, respectively. We hypothesized that the movement of intracellular Cl(-) (Cl(i)(-)) across the SR membrane neutralizes the accumulation of charge that accompanies uptake and release of Ca(2+). Thus inhibition of SR Cl(-) fluxes will reduce Ca(2+) sequestration and agonist-induced release. The Cl(-) channel blocker 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB; 10(-4) M), previously shown to inhibit SR Cl(-) channels, significantly reduced the magnitude of successive acetylcholine-induced contractions of airway smooth muscle (ASM), suggesting a "run down" of sequestered Ca(2+) within the SR. Niflumic acid (10(-4) M), a structurally different Cl(-) channel blocker, had no such effect. Furthermore, NPPB significantly reduced caffeine-induced contraction and increases in intracellular Ca(2+) concentration ([Ca(2+)](i)). Depletion of Cl(i)(-), accomplished by bathing ASM strips in Cl(-)-free buffer, significantly reduced the magnitude of successive acetylcholine-induced contractions. In addition, Cl(-) depletion significantly reduced caffeine-induced increases in [Ca(2+)](i). Together these data suggest a novel role for Cl(i)(-) fluxes in Ca(2+) handling in smooth muscle. Because the release of sequestered Ca(2+) is the predominate source of Ca(2+) for contraction of ASM, targeting Cl(i)(-) fluxes may prove useful in the control of ASM hyperresponsiveness associated with asthma.
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