Effects of epoxyeicosatrienoic acids on the cardiac sodium channels in isolated rat ventricular myocytes

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Abstract
Whole-cell Na+ currents (holding potential, −80 mV; test potential, −30 mV) in rat myocytes were inhibited by 8,9-epoxyeicosatrienoic acid (8,9-EET) in a dose-dependent manner with 22 ± 4 % inhibition at 0.5 μM, 48 ± 5 % at 1 μM, and 73 ± 5 % at 5 μM (mean ± s.e.m., n = 10, P < 0.05 for each dose vs. control). Similar results were obtained with 5,6-, 11,12-, and 14,15-EETs, while 8,9-dihydroxyeicosatrienoic acid (DHET) was 3-fold less potent and arachidonic acid was 10- to 20-fold less potent. 8,9-EET produced a dose-dependent, hyperpolarized shift in the steady-state membrane potential at half-maximum inactivation (V½), without changing the slope factor. 8,9-EET had no effect on the steady-state activation of Na+ currents. Inhibition of Na+ currents by 8,9-EET was use dependent, and channel recovery was slowed. The effects of 8,9-EET were greater at depolarized potentials. Single channel recordings showed 8,9-EET did not change the conductance or the number of active Na+ channels, but markedly decreased the probability of Na+ channel opening. These results were associated with a decrease in the channel open time and an increase in the channel closed times. Incubation of cultured cardiac myocytes with 1 μM [3H]8,9-EET showed that 25 % of the radioactivity was taken up by the cells over a 2 h period, and most of the uptake was incorporated into phospholipids, principally phosphatidylcholine. Analysis of the medium after a 2 h incubation indicated that 86 % of the radioactivity remained as [3H]8,9-EET while 13 % was converted into [3H]8,9-DHET. After a 30 min incubation, 1–2 % of the [3H]8,9-EET uptake by cells remained as unesterified EET. These results demonstrate that cardiac cells have a high capacity to take up and metabolize 8,9-EET. 8,9-EET is a potent use- and voltage-dependent inhibitor of the cardiac Na+ channels through modulation of the channel gating behaviour.