Apelin-13 Inhibits Large-Conductance Ca2+-Activated K+ Channels in Cerebral Artery Smooth Muscle Cells via a PI3-Kinase Dependent Mechanism

Amit Modgil,Lirong Guo,Chengwen Sun,Stephen T O’Rourke,Zhe Zhang

doi:10.1371/journal.pone.0083051

Amit Modgil, Lirong Guo + Show 3 more

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https://doi.org/10.1371/journal.pone.0083051

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Abstract

Apelin-13 causes vasoconstriction by acting directly on APJ receptors in vascular smooth muscle (VSM) cells; however, the ionic mechanisms underlying this action at the cellular level remain unclear. Large-conductance Ca2+-activated K+ (BKCa) channels in VSM cells are critical regulators of membrane potential and vascular tone. In the present study, we examined the effect of apelin-13 on BKCa channel activity in VSM cells, freshly isolated from rat middle cerebral arteries. In whole-cell patch clamp mode, apelin-13 (0.001-1 μM) caused concentration-dependent inhibition of BKCa in VSM cells. Apelin-13 (0.1 µM) significantly decreased BKCa current density from 71.25±8.14 pA/pF to 44.52±7.10 pA/pF (n=14 cells, P<0.05). This inhibitory effect of apelin-13 was confirmed by single channel recording in cell-attached patches, in which extracellular application of apelin-13 (0.1 µM) decreased the open-state probability (NPo) of BKCa channels in freshly isolated VSM cells. However, in inside-out patches, extracellular application of apelin-13 (0.1µM) did not alter the NPo of BKCa channels, suggesting that the inhibitory effect of apelin-13 on BKCa is not mediated by a direct action on BKCa. In whole cell patches, pretreatment of VSM cells with LY-294002, a PI3-kinase inhibitor, markedly attenuated the apelin-13-induced decrease in BKCa current density. In addition, treatment of arteries with apelin-13 (0.1 µM) significantly increased the ratio of phosphorylated-Akt/total Akt, indicating that apelin-13 significantly increases PI3-kinase activity. Taken together, the data suggest that apelin-13 inhibits BKCa channel via a PI3-kinase-dependent signaling pathway in cerebral artery VSM cells, which may contribute to its regulatory action in the control of vascular tone.

Highlights

Apelin is a peptide isolated from bovine stomach extracts and identified as an endogenous ligand for the orphan Gprotein coupled receptor, APJ, which has seven transmembrane-spanning domains [1,2]
The present study was undertaken to examine the effect of apelin-13 on BKca channel activity in vascular smooth muscle (VSM) cells freshly isolated from rat middle cerebral arteries
The results demonstrate that apelin-13 inhibits BKCa channel activity in a concentrationdependent manner through G-protein and PI3-kinase dependent signaling pathways, which may contribute to its regulatory action in controlling vascular tone

Summary

Introduction

Apelin is a peptide isolated from bovine stomach extracts and identified as an endogenous ligand for the orphan Gprotein coupled receptor, APJ, which has seven transmembrane-spanning domains [1,2]. Despite sharing 31% of amino acid sequence homology with angiotensin II type 1 receptors (AT1-R), angiotensin II does not bind to these receptors [3] and apelin is the only known ligand for APJ receptors. The apelin gene, located on the long arm of the human X chromosome, codes for a preproprotein of 77-amino acid residues that is further cleaved to shorter active peptide fragments including apelin-36 (42-77), apelin-17 (61-77) and apelin-13 (65-77) [1,4,5]. Apelin-13 exhibits the greatest binding affinity and biological potency as compared to other fragments [4,6,7]. The wide expression pattern of apelin/APJ receptors throughout the cardiovascular system strongly suggests an important role for the apelin/APJ system in cardiovascular homeostasis

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