Ghrelin signalling in guinea-pig femoral artery smooth muscle cells

Our aim was to study the new signalling pathway of ghrelin in the guinea-pig femoral artery using the outward IK as a sensor. Whole-cell patch-clamp experiments were performed on single smooth muscle cells, freshly isolated from the guinea-pig femoral artery. The contractile force of isometric preparations of the same artery was measured using a wire-myograph. In a Ca²⁺- and nicardipine-containing external solution, 1 mmol L⁻¹ tetraethylammonium reduced the net IK by 49 ± 7%. This effect was similar and not additive to the effect of the specific BKCa channel inhibitor iberiotoxin. Ghrelin (10⁻⁷ mol L⁻¹) quickly and significantly reduced the amplitudes of tetraethylammonium- and iberiotoxin-sensitive currents through BKCa channels. The application of 5 x 10⁻⁶ mol L⁻¹ desacyl ghrelin did not affect the amplitude of the control IK but it successfully prevented the ghrelin-induced IK decrease. The effect of ghrelin on IK was insensitive to selective inhibitors of cAMP-dependent protein kinase, soluble guanylyl cyclase, cGMP-dependent protein kinase or a calmodulin antagonist, but was effectively antagonized by blockers of BKCa channels, phosphatidylinositol-phospholipase C, phosphatidylcholine-phospholipase C, protein kinase C, SERCA, IP₃-induced Ca²⁺ release and by pertussis toxin. The ghrelin-induced increase in the force of contractions was blocked when iberiotoxin (10⁻⁷ mol L⁻¹) was present in the bath solution. Ghrelin reduces IK₍Ca₎ in femoral artery myocytes by a mechanism that requires activation of Gαi/o-proteins, phosphatidylinositol phospholipase C, phosphatidylcholine phospholipase C, protein kinase C and IP₃-induced Ca²⁺ release.