Two-photon imaging of endothelin-1-mediated intracellular Ca^2 + handling in smooth muscle cells of rat renal resistance arteries

Aims Endothelin-1 (ET-1) is a potent vasoconstrictor which regulates the physiology of cardiorenal system. The aim of this study was to evaluate ET-1-mediated elevation of intracellular Ca^2 + in smooth muscle cells (SMC) of renal resistance arteries. Main methods In in vitro studies of primary SMC, which were isolated from rat renal microvessels, the levels of intracellular Ca^2 + were calculated from the ratio of emissions at 340 and 380 nm after loading cells with Fura 2-AM dye. In ex vivo studies we used two-photon imaging of renal resistance arteries excised from rat kidneys and loaded with fluorescent Ca^2 + indicator Fluo-4 AM. Key findings The two-photon imaging demonstrates that treatment of isolated rat renal resistance arteries with ET-1 causes a rapid increase of intracellular Ca^2 + concentration in smooth muscle vasculature of these vessels. These ex vivo observations are in accordance with in vitro findings indicating that ET-1 mediates activation of TRPC channels and increases the level of intracellular Ca^2 + in cultured SMC to 510 ± 83 nM. Significance ET-1-mediated elevation of intracellular Ca^2 + is strongly linked to renal microvascular contraction and is crucial for ET-1-induced contraction of SMC. The two-photon imaging of intracellular Ca^2 + in intact SMC of rat renal resistance arteries is a powerful technique which allows the detailed ex vivo analysis of intracellular Ca^2 + handling by ET-1, an important player in hypertension-related kidney diseases.