Abstract-Sudden cardiac death attributable to ventricular tachycardia/fibrillation (VF) remains a catastrophic outcome of myocardial ischemia and infarction. At the same time, conventional antagonist drugs targeting ion channels have yielded poor survival benefits. Although pharmacological and genetic models suggest an association between sodium (Na ϩ ) channel loss-of-function and sudden cardiac death, molecular mechanisms have not been identified that convincingly link ischemia to Na ϩ channel dysfunction and ventricular arrhythmias. Because ischemia can evoke the generation of reactive oxygen species, we explored the effect of oxidative stress on Na ϩ channel function. We show here that oxidative stress reduces Na ϩ channel availability. Both the general oxidant tert-butyl-hydroperoxide and a specific, highly reactive product of the isoprostane pathway of lipid peroxidation, E 2 -isoketal, potentiate inactivation of cardiac Na ϩ channels in human embryonic kidney (HEK)-293 cells and cultured atrial (HL-1) myocytes. Furthermore, E 2 -isoketals were generated in the epicardial border zone of the canine healing infarct, an arrhythmogenic focus where Na ϩ channels exhibit similar inactivation defects. In addition, we show synergistic functional effects of flecainide, a proarrhythmic Na ϩ channel blocker, and oxidative stress. These data suggest Na ϩ channel dysfunction evoked by lipid peroxidation is a candidate mechanism for ischemia-related conduction abnormalities and arrhythmias.

We showed that muscarinic acetylcholine (ACh)-stimulation increased the cellular content of cADPR in the pancreatic acinar cells from normal mice but not in those from CD38 knockout mice.

Local anaesthetics such as lidocaine (lignocaine) interact with sodium channels in a manner that is exquisitely sensitive to the voltage‐dependent conformational state of the ion channel. When depolarized in the presence of lidocaine, sodium channels assume a long‐lived quiescent state. Although studies over the last decade have localized the lidocaine receptor to the inner aspect of the aqueous pore, the mechanistic basis of depolarization‐induced ‘use‐dependent’ lidocaine block remains uncertain. Recent studies have shown that lowering the extracellular Na+ concentration ([Na+]o) and mutations in the sodium channel outer P‐loop modulate occupancy of a quiescent ‘slow’ inactivated state with intermediate kinetics (termed IM) that involves structural rearrangements in the outer pore. Site‐directed mutagenesis and ion‐replacement experiments were performed using voltage‐clamped Xenopus oocytes and cultured (HEK‐293) cells expressing wild‐type and mutant rat skeletal muscle (μ1) sodium channels. Our results show that lowering [Na+]o potentiates use‐dependent lidocaine block. The effect of [Na+]o is maintained despite a III‐IV linker mutation that partially disrupts fast inactivation (F1304Q). In contrast, the effect of lowering [Na+]o on lidocaine block is reduced by a P‐loop mutation (W402A) that limits occupancy of IM. Our findings are consistent with a simple allosteric model where lidocaine binding induces channels to occupy a native slow inactivated state that is inhibited by [Na+]o.

. Cytochrome P-450 metabolites but not NO, PGI2, and H2O2 contribute to ACh-induced hyperpolarization of pressurized canine coronary microvessels. Am J Physiol Heart Circ Physiol 285: H1939-H1948, 2003. First published July 24, 2003 10.1152/ajpheart.00190.2003.-The endothelium-dependent hyperpolarization of cells has a crucial role in regulating vascular tone, especially in microvessels. Nitric oxide (NO) and prostacyclin (PGI 2), in addition to endothelium-derived hyperpolarizing factor (EDHF), have been reported to hyperpolarize vascular smooth muscle in several organs. Studies have reported the hyperpolarizing effects of these factors are increased by a stretch in large coronary arteries. EDHF has not yet been identified and cytochrome P-450 metabolites and H2O2 are candidates for EDHF. With the use of the membrane potential-sensitive fluorescent dye bis-(1,3-dibutylbarbituric acid)trimethione oxonol [DiBAC4(3)], we examined whether NO, PGI2, cytochrome P-450 metabolites, and H2O2 contribute to AChinduced hyperpolarization in pressurized coronary microvessels. Canine coronary arterial microvessels (60-356 m internal diameter) were cannulated and pressurized at 60 cmH2O in a vessel chamber perfused with physiological salt solution containing DiBAC4(3). Fluorescence intensity and diameter were measured on a computer. There was a linear correlation between changes in the fluorescence intensity and membrane potential. ACh significantly decreased the fluorescence intensity (hyperpolarization) of the microvessels without any inhibitors. Endothelial damage caused by air perfusion abolished the ACh-induced decrease in fluorescence intensity. The inhibitors of NO synthase and cyclooxygenase did not affect the ACh-induced decreases in the fluorescence intensity. The addition of 17-octadecynoic acid, a cytochrome P-450 monooxygenase inhibitor, to those inhibitors significantly attenuated the ACh-induced decreases in fluorescence intensity, whereas catalase, an enzyme that dismutates H 2O2 to form water and oxygen, did not. Furthermore, catalase did not affect the vasodilation produced by ACh. These results indicate that NO and PGI 2 do not contribute to the AChinduced hyperpolarization and that the cytochrome P-450 metabolites but not H 2O2 are involved in EDHF-mediated hyperpolarization in canine coronary arterial microvessels. microcirculation; endothelium-derived hyperpolarizing factor; endothelium THE ENDOTHELIUM PLAYS an important role in regulating the vascular tone by releasing vasodilator substances such as prostacyclin (PGI 2 ), nitric oxide (NO), and endothelium-derived hyperpolarizing factor (EDHF) (37, 50). Part of the endothelium-dependent relaxation is accompanied by endothelium-dependent hyperpolarization of the vascular smooth muscle cells (10) and the contribution of the hyperpolarizing mechanism to endothelium-dependent relaxation increases as the vessel size decreases, whereas the contribution of NO decreases (24,29,36,48).It has been shown that NO and PGI 2 , in addition to EDHF, hyperpolarize the m...