TNF-α dilates cerebral arteries via NAD(P)H oxidase-dependent Ca2+ spark activation

Expression of TNF-alpha, a pleiotropic cytokine, is elevated during stroke and cerebral ischemia. TNF-alpha regulates arterial diameter, although mechanisms mediating this effect are unclear. In the present study, we tested the hypothesis that TNF-alpha regulates the diameter of resistance-sized ( similar to 150-mu m diameter) cerebral arteries by modulating local and global intracellular Ca2+ signals in smooth muscle cells. Laser-scanning confocal imaging revealed that TNF-alpha increased Ca2+ spark and Ca2+ wave frequency but reduced global intracellular Ca2+ concentration ([ Ca2+](i)) in smooth muscle cells of intact arteries. TNF-alpha elevated reactive oxygen species ( ROS) in smooth muscle cells of intact arteries, and this increase was prevented by apocynin or diphenyleneiodonium ( DPI), both of which are NAD( P) H oxidase blockers, but was unaffected by inhibitors of other ROS-generating enzymes. In voltage-clamped ( -40 mV) cells, TNF-alpha increased the frequency and amplitude of Ca2+ spark-induced, large-conductance, Ca2+-activated K+ ( KCa) channel transients similar to 1.7- and similar to 1.4-fold, respectively. TNF-alpha-induced transient KCa current activation was reversed by apocynin or by Mn( III) tetrakis( 1-methyl-4-pyridyl) porphyrin ( MnTMPyP), a membrane-permeant antioxidant, and was prevented by intracellular dialysis of catalase. TNF-alpha induced reversible and similar amplitude dilations in either endothelium-intact or endothelium-denuded pressurized ( 60 mmHg) cerebral arteries. MnTMPyP, thapsigargin, a sarcoplasmic reticulum Ca2 alpha-ATPase blocker that inhibits Ca2+ sparks, and iberiotoxin, a KCa channel blocker, reduced TNF-alpha-induced vasodilations to between 15 and 33% of control. In summary, our data indicate that TNF-alpha activates NAD( P) H oxidase, resulting in an increase in intracellular H2O2 that stimulates Ca2+ sparks and transient KCa currents, leading to a reduction in global [ Ca2+](i), and vasodilation.