Protein phosphatase 2A and Ca 2+-activated K + channels contribute to 11,12-epoxyeicosatrienoic acid analog mediated mesenteric arterial relaxation

Abstract

Epoxyeicosatrienoic acids (EETs) are considered to be endothelium-derived hyperpolarizing factors, and are potent activators of the large-conductance, Ca 2+-activated K + (BK Ca) channel in vascular smooth muscle. Here, we investigate the signal transduction pathway involved in the activation of BK Ca channels by 11,12-EET and 11,12-EET stable analogs in rat mesenteric vascular smooth muscle cells. 11,12-EET and the 11,12-EET analogs, 11-nonyloxy-undec-8( Z)-enoic acid (11,12-ether-EET-8-ZE), 11-(9-hydroxy-nonyloxy)-undec-8( Z)-enoic acid (11,12-ether-EET-8-ZE-OH) and 11,12- trans-oxidoeicosa-8( Z)-enoic acid (11,12-tetra-EET-8-ZE), caused vasorelaxation of mesenteric resistance arteries. Mesenteric myocyte whole-cell (perforated-patch) currents were substantially (∼150%) increased by 11,12-EET and 11,12-EET analogs. Single-channel recordings were conducted to identify the target for 11,12-EET. 11,12-EET and 11,12-EET analogs also increased mesenteric myocyte BK Ca channel activity in cell-attached patches. Similar results were obtained in cell-free patches. Baseline mesenteric myocyte BK Ca channel activity (NPo) in cell-free patches averaged less than 0.001 at +50 mV and 11,12-EET (1 μmol/L) increased NPo to 0.03 ± 0.02 and 11,12-EET analogs (1 μmol/L) increased NPo to 0.09 ± 0.006. Inhibition of protein phosphatase 2A (PP2A) activity with okadaic acid (10 nmol/L) completely reversed 11,12-EET stimulated BK Ca channel activity and greatly attenuated 11,12-ether-EET-8-ZE mesenteric resistance artery vasorelaxation. 11,12-EET and 11,12-EET analogs increased mesenteric myocyte PP2A activity by 3.5-fold. Okadaic acid and the EET inhibitor, 14,15-epoxyeicosa-5( Z)-enoic acid (14,15-EEZE) inhibited the 11,12-EET mediated increase in PP2A activity. These findings provide initial evidence that PP2A activity contributes to 11,12-EET and 11,12-EET analog activation of mesenteric resistant artery BK Ca channels and vasorelaxation.