Endothelium-dependent frequency modulation of Ca2+ signalling in individual vascular smooth muscle cells of the rat

1. We visualized intracellular Ca2+ concentration ([Ca2+](i)) changes, using fluo-3 as an indicator, of individual vascular smooth muscle cells and endothelial cells within intact rat tail arteries by confocal microscopy. 2. Using a piezo-driven objective, we focused on endothelial and smooth muscle cell layers alternately to obtain Ca2+ images of their cells. In the presence of 1 mu M acetylcholine (ACh), individual endothelial cells responded with intermittent increases in the [Ca2+](i) (Ca2+ oscillations). At the same time, the frequency of Ca2+ oscillations in smooth muscle cells induced by electrical stimulation of the perivascular sympathetic nerve was greatly decreased. 3. A [Ca2+](i) rise during the oscillations in the endothelial cells propagated in the form of a wave along the long axis of the cells. 4. In the presence of a NO synthase inhibitor, no significant inhibitory effect of ACh on the Ca2+ signalling in the vascular smooth muscle cells was detected, although the Ca2+ oscillations in the endothelial cells persisted. 5. The inhibitory effect of ACh on the frequency of Ca2+ oscillations in the vascular smooth muscle cells was mimicked by 1 mu M Sodium nitroprusside, a NO donor. 6. These results indicate that Ca2+ waves and oscillations in vascular endothelial cells regulate NO production, which modulates vascular tone by decreasing the frequency of Ca2+ oscillations in smooth muscle cells activated by sympathetic agonists.