Inhibition of TMEM16A by Docosahexaenoic Acid Plays a Crucial Role in Blood Vessel Relaxation

Abstract

Docosahexaenoic acid (DHA) is an omega-3 polyunsaturated fatty acid with beneficial cardiovascular effects, such as vasodilation. The mechanism underlying this vasodilation, however, is not fully defined. In arterial smooth muscle, TMEM16A chloride channels promote membrane depolarisation and vessel contraction. Cloned TMEM16A channels are inhibited by DHA. Here, we examine whether inhibition of native TMEM16A by DHA plays a role in aortic relaxation. Patch-clamp electrophysiology and isometric tension recordings were utilised to study the effect of DHA on cloned TMEM16A channels and aortic contractility. DHA inhibited TMEM16A currents in a dose dependent manner, IC50=15.4±1.5 μM (n=10). To study the regulation of TMEM16A by DHA in response to agonist-induced increase in [Ca2+]i, TMEM16A was co-expressed with the α1A-adrenoreceptor and currents recorded in response to 1 μM phenylephrine (PE). In the absence and presence of DHA (10 μM) the TMEM16A current increased by 7.6±1.8 fold (n=12) and 2.8±0.5 fold (n=6), respectively (p<0.05). In aortic rings, inhibition of TMEM16A with either 70 μM DHA or 1 μM 2-(4-Chloro-2-methylphenoxy)-N′-(2-methoxybenzylidene)acetohydrazide (Ani9) reduced PE-induced aortic contraction by 1.9±0.4 fold (n=9) and 2.0±0.4 fold (n=7), respectively. Inhibition of the NKCC1 transporter with 40 μM bumetanide, to lower intracellular Cl- concentration ([Cl-]), reduced aortic response to 1 μM PE by 3.3±0.4 fold (n=10). Conversely, lowering extracellular [Cl-] increased the aortic response to 1 μM PE by 9.3±3.4 fold (n=5); this effect was reduced to 2.8±1.0 fold (n=5) in the presence of 70 μM DHA (p<0.05), implicating TMEM16A channels in the response. DHA-mediated inhibition of TMEM16A channels reduces aortic contractility. This study highlights TMEM16A as a target for modulating vascular tone. Identification of the TMEM16A DHA binding site may enable development of new small molecules aiming to reduce blood pressure.