Inhibitory effect of nitrovasodilators and cyclic GMP on ET‐1‐activated Ca²⁺‐permeable nonselective cation channel in rat aortic smooth muscle cells

Abstract

1 In single vascular smooth muscle cells (VSMCs) isolated from the aortae of male Wistar rats, we examined the e ects of nitric oxide (NO) donors such as sodium nitroprusside (SNP) and S-nitroso-Nacetyl-DL-penicillamine (SNAP), and 8-bromo-guanosine-3':5'-cyclic monophosphate (8-bromo-cyclic GMP) on endothelin-1 (ET-1)-activated Ca 2+ -permeable nonselective cation channel by use of whole-cell recordings of patch-clamp technique and monitoring of intracellular free Ca 2+ concentration ([Ca 2+ ] i ) with fura-2 real-time digital micro¯uorometry. 2 ET-1 evoked an initial transient peak and a subsequent sustained elevation in [Ca 2+ ] i . After removal of extracellular Ca 2+ , ET-1 evoked only an initial transient peak without a sustained phase. Nifedipine (1 mM), a speci®c blocker of the L-type voltage-operated Ca 2+ channel (VOC), reduced the sustained phase to about 40% of the control level. The remaining part of the sustained phase was abolished by 30 mM SK&F 96365, a blocker of nonselective cation channels. 3 The nifedipine-resistant sustained elevation in [Ca 2+ ] i was abolished by 100 mM SNP, 10 mM SNAP and 300 mM 8-bromo-cyclic GMP. Neither SNP, SNAP nor 8-bromo-cyclic GMP signi®cantly a ected the basal level of [Ca 2+ ] i . 4 In a VSMC clamped at a holding potential of 760 mV with K + in the pipette solution replaced by Cs + , application of 10 78 M ET-1 induced an inward current with an increase in baseline¯uctuation. With¯uctuation analysis, unit conductance of the ET-1-induced current was calculated to be about 21 pS. The ET-1-induced current was linearly related to the membrane potentials with its reversal potential of 75.5 mV. 5 The ET-1-induced current was reversibly and completely inhibited by 30 mM SK&F 96365 or 500 mM Cd 2+ . The current inhibited by SK&F 96365 or Cd 2+ was linearly related to membrane potential with a reversal potential of about 75 mV. 6 The ET-1-induced current was reversibly and completely inhibited by 100 mM SNP, 10 mM SNAP and 300 mM 8-bromo-cyclic GMP. The current inhibited by SNP, SNAP or 8-bromo-cyclic GMP showed linear voltage-dependence and reversed at about 75 mV. 7 In a bath solution in which all cations were replaced by 30 mM Ca 2+ and 100 mM nonpermeant cation N-methyl-D-glucamine (NMDG), ET-1 evoked a current with a reversal potential of 711 mV, from which P Ca 2+ /P Cs + was calculated to be 2.1. This Ca 2+ current was also abolished by 100 mM SNP, 10 mM SNAP and 300 mM 8-bromo-cyclic GMP. The current inhibited by SNP, SNAP or 8-bromo-cyclic GMP showed linear voltage-dependence and reversed at about 711 mV. 8 These results taken together indicate that NO through a cyclic GMP signalling pathway inhibits ET-1-activated Ca 2+ -permeable nonselective cation channels, thereby suppressing the sustained increase in [Ca 2+ ] i . Thus, the present study indicates that this Ca 2+ -permeable nonselective cation channel is an important target for nitrovasodilators.

Activation of hypoxia-inducible factor-1 in pulmonary arterial smooth muscle cells by endothelin-1

“…The activation of ERK1/2 depended on increased ROS levels, since antioxidants blocked ET-1-dependent ERK1/2 activation. ET-1 also increased [Ca 2ϩ ] i and ROS levels in AoSMCs, consistent with previous reports (21,31,41), but did not activate ERK1/2 or induce HIF-1␣ protein accumulation. The mechanism by which ROS leads to ERK1/2 activation in PASMCs is unclear, as is the reason why ET-1-induced increases in ROS do not activate ERK1/2 in AoSMCs.…”

In vitroeffect of endothelin-1 and nifedipine on pulmonary vascular contraction of pulmonary hypertensive and non-pulmonary hypertensive chickens

“…Thus, concentrations above 10 −9 M lead to an increase in [(Ca 2+ ) i ] through the liberation of Ca 2+ from intracellular deposits, which induces a short and transient contraction phase, and also through the activation of VOCC, which induces a higher, sustained increase in [(Ca 2+ ) i ]. Thus, a contraction phase with a longer duration and higher magnitude is produced (Takuwa et al 1990;Komuro et al 1997;Minowa et al 1997;Zhang et al 1998). In contrast, lower ET-1 concentrations only induce the sustained phase; that is, the contraction in this case is mainly dependent on extracellular Ca 2+ Kawanabe et al 2002).…”

Pharmacological characterization of receptor-mediated Ca2+ entry in endothelin-1-induced catecholamine release from cultured bovine adrenal chromaffin cells

“…Several researchers have shown that Ca 2+ -permeable nonselective cation channels (NSCC) are activated by stimulation of native ET A receptor in vascular smooth muscle cells (van Renterghem et al 1988;Chen and Wagoner 1991;Enoki et al 1995;Minowa et al 1997) and of recombinant human ET A receptors expressed in Ltk -cells (Enoki et al 1995). We have recently found that in A7r5 cells derived from rat thoracic aortic smooth muscle cells, ET-1 can activate three types of voltage-independent Ca 2+ entry channels: type-1 Ca 2+ -permeable NSCC (NSCC-1), type-2 Ca 2+ -permeable NSCC (NSCC-2) and store-operated Ca 2+ channel (SOCC), in addition to VOCC (Iwamuro et al 1998(Iwamuro et al , 1999.…”