Changes in cytosolic calcium level in vascular smooth muscle strip measured simultaneously with contraction using fluorescent calcium indicator fura 2.

Abstract

In rat aortic strips, muscle contraction was recorded simultaneously with cytosolic Ca++ level, which was indicated by the 500 nm fluorescence of Ca++ indicator, fura 2, due to excitation at either 340 nm (F340) or 380 nm (F380) and the ratio of F340 to F380 (R340/380). On the addition of 72.7 nM K+ or 1 microM norepinephrine, muscle contraction followed the increase in R340/380 (resulted from the increased F340 and the decreased F380). Cytosolic Ca++ concentrations of resting, 72.7 mM K+-stimulated and 1 microM norepinephrine-stimulated aortas were tentatively calculated as 228 +/- 25, 1784 +/- 154 and 1528 +/- 180 nM, respectively. Cumulative addition of K+ or norepinephrine induced concentration-dependent increase in both muscle tension and R340/380. However, norepinephrine induced greater contraction than K+ when both of these stimulants induced similar increase in R340/380. Addition of 10 mM tetraethylammonium and 1 microM Bay k8644 caused rhythmic contractions which followed the rhythmic changes in R340/380. EGTA decreased the muscle contraction and decreased R340/380. In Ca++-free solution, addition of 10 microM norepinephrine or 20 mM caffeine induced transient increase in both muscle tension and R340/380. Tension changes always were preceded by the fluorescent changes. Verapamil (10 microM) decreased both tension development and R340/380 in high K+- and norepinephrine-stimulated tissues. Sodium nitroprusside (1 microM) also decreased both tension and R340/380 in norepinephrine-stimulated tissues, whereas it decreased tension more strongly than R340/380 in high K+-stimulated tissues. These results indicate that vasoconstrictors and vasodilators may modulate smooth muscle contraction by changing the cytosolic Ca++ concentrations and also by changing the sensitivity of contractile elements to Ca++.