TMEM16A/ANO1 Channels Contribute to the Myogenic Response in Cerebral Arteries

Abstract

Rationale Pressure-induced arterial depolarization and constriction (the myogenic response), is a smooth muscle cell (myocyte)-specific mechanism that controls regional organ blood flow and systemic blood pressure. Several different non-selective cation channels contribute to pressure-induced depolarization, but signaling mechanisms involved are unclear. Similarly uncertain is the contribution of anion channels to the myogenic response and physiological functions and mechanisms of regulation of recently discovered transmembrane 16A (TMEM16A) chloride (Cl−) channels in arterial myocytes. Objective Investigate the hypothesis that myocyte TMEM16A channels control membrane potential and contractility and contribute to the myogenic response in cerebral arteries. Methods and Results Cell swelling induced by hyposmotic bath solution stimulated Cl− currents in arterial myocytes that were blocked by TMEM16A channel inhibitory antibodies, RNAi-mediated selective TMEM16A channel knockdown, removal of extracellular calcium (Ca2+), replacement of intracellular EGTA with BAPTA, a fast Ca2+ chelator, and Gd3+ and SKF-96365, non-selective cation channel blockers. In contrast, nimodipine, a voltage-dependent Ca2+ channel inhibitor, or thapsigargin, which depletes intracellular Ca2+ stores, did not alter swelling-activated TMEM16A currents. Pressure (−40 mmHg)-induced membrane stretch activated ion channels in arterial myocyte cell-attached patches that were inhibited by TMEM16A antibodies and were of similar amplitude to recombinant TMEM16A channels. TMEM16A knockdown reduced intravascular pressure-induced depolarization and vasoconstriction, but did not alter depolarization (60 mmol/L K+)-induced vasoconstriction. Conclusions Membrane stretch activates arterial myocyte TMEM16A channels, leading to membrane depolarization and vasoconstriction. Data also provide a mechanism by which a local Ca2+ signal generated by non-selective cation channels stimulates TMEM16A channels to induce myogenic constriction.

Activation of Ca2+‐activated Cl− channel ANO1 by localized Ca2+ signals

“…2012) or other non‐selective Ca 2+ channels (Bulley et al . 2012) have been reported (reviewed in Leblanc et al . 2005; Bulley & Jaggar, 2014).…”

“…However, physiological activation of ANO1‐mediated CaCCs in smooth muscle is likely to depend on co‐localized Ca 2+ signals since Ca 2+ sensitivity of ANO1 at physiologically relevant voltages is lower than the physiological range of global [Ca 2+ ] i in smooth muscle cells (Knot & Nelson, 1998; Bulley et al . 2012). Thus, in cerebral artery smooth muscle cells ANO1 has been found to specifically couple to a Ca 2+ influx via non‐selective cation channels activated by cell swelling and pressure‐induced membrane stretch but not to VGCCs.…”

“…The ANO1 activation was blocked by BAPTA but not EGTA, suggesting close association between ANO1 and Ca 2+ influx channels (Bulley et al . 2012). …”

Functions of ANO1/TMEM16A, Ca2+-activated Cl- channels in Regulation of Blood Pressure and Vascular Remodeling

“…Participation of ANO1 in regulation of VSMC contractility is not only mediated by humoral factors but also by mechanosensitive mechanism which controls the intravascular pressure of arterioles and the blood flow volume through organs [19] . The mechanosensitive mechanism has been well-known to be contributed by cation channels, e.g., Ca 2+ , K + and TRP channels [20,21] .…”

“…However, the study on the involvement of anion channels in the process had been retarded due to unknown identity of CaCC channels which was regarded as an important factor in the process. Jaggar group recently made a major contribution to the elucidation of the role ANO1 played in the myogenic response of the cerebral arteries after ANO1 was identified as the endogenous CaCC channel [19] . With an ANO1 inhibitory antibody and siRNA, the group skillfully demonstrated that hyposmolarity or cell swelling-induced whole-cell Cl -currents and the Cl -currents activated by the stretch on the cell-attached membrane induced by negative pressure through recording pipettes, were produced by ANO1 expressed in cerebral arterial SMCs.…”