Calcium influx factor, further evidence it is 5,6-epoxyeicosatrienoic acid

We present evidence in astrocytes that 5,6-epoxyeicosatrienoic acid, a cytochrome P450 epoxygenase metabolite of arachidonic acid, may be a component of calcium influx factor, the elusive link between release of Ca²⁺ from intracellular stores and capacitative Ca²⁺ influx. Capacitative influx of extracellular Ca²⁺ was inhibited by blockade of the two critical steps in epoxyeicosatrienoic acid synthesis: release of arachidonic acid from phospholipid stores by cytosolic phospholipase A₂ and cytochrome P450 metabolism of arachidonic acid. AAOCF₃, which inhibits cytosolic phospholipase A₂, blocked thapsigargin-stimulated release of arachidonic acid as well as thapsigargin-stimulated elevation of intracellular free calcium. Inhibition of P450 arachidonic acid metabolism with SKF525A, econazole, or N-methylsulfonyl-6-(2-propargyloxyphenyl)hexanamide, a substrate inhibitor of P450 arachidonic acid metabolism, also blocked thapsigargin-stimulated Ca²⁺ influx. Nanoto picomolar 5,6- epoxyeicosatrienoic acid induced [Ca²⁺](i) elevation consistent with capacitative Ca²⁺ influx. We have previously shown that 5,6- epoxyeicosatrienoic acid is synthesized and released by astrocytes. When 5,6- epoxyeicosatrienoic acid was applied to the rat brain surface, it induced vasodilation, suggesting that calcium influx factor may also serve a paracrine function. In summary, our results suggest that 5,6- epoxyeicosatrienoic acid may be a component of calcium influx factor and may participate in regulation of cerebral vascular tone.