ANO1, CaV1.2, and IP3R form a localized unit of EC-coupling in mouse pulmonary arterial smooth muscle.

Pulmonary arterial (PA) smooth muscle cells (PASMC) generate vascular tone in response to agonists coupled to Gq-protein receptor signaling. Such agonists stimulate oscillating calcium waves, the frequency of which drives the strength of contraction. These Ca2+ events are modulated by a variety of ion channels including voltage-gated calcium channels (CaV1.2), the Tmem16a or Anoctamin-1 (ANO1)-encoded calcium-activated chloride (CaCC) channel, and Ca2+ release from the sarcoplasmic reticulum through inositol-trisphosphate receptors (IP3R). Although these calcium events have been characterized, it is unclear how these calcium oscillations underly a sustained contraction in these muscle cells. We used smooth muscle-specific ablation of ANO1 and pharmacological tools to establish the role of ANO1, CaV1.2, and IP3R in the contractile and intracellular Ca2+ signaling properties of mouse PA smooth muscle expressing the Ca2+ biosensor GCaMP3 or GCaMP6. Pharmacological block or genetic ablation of ANO1 or inhibition of CaV1.2 or IP3R, or Ca2+ store depletion equally inhibited 5-HT-induced tone and intracellular Ca2+ waves. Coimmunoprecipitation experiments showed that an anti-ANO1 antibody was able to pull down both CaV1.2 and IP3R. Confocal and superresolution nanomicroscopy showed that ANO1 coassembles with both CaV1.2 and IP3R at or near the plasma membrane of PASMC from wild-type mice. We conclude that the stable 5-HT-induced PA contraction results from the integration of stochastic and localized Ca2+ events supported by a microenvironment comprising ANO1, CaV1.2, and IP3R. In this model, ANO1 and CaV1.2 would indirectly support cyclical Ca2+ release events from IP3R and propagation of intracellular Ca2+ waves.

Translational potential of targeting Anoctamin-1-Encoded Calcium-Activated chloride channels in hypertension.

Calcium-activated chloride channels (CaCC) provide a depolarizing stimulus to a variety of tissues through chloride efflux in response to a rise in internal Ca2+ and voltage. One of these channels, Anoctamin-1 (ANO1 or TMEM16A) is now recognized to play a central role in promoting smooth muscle tone in various types of blood vessels. Its role in hypertension, and thus the therapeutic promise of targeting ANO1, is less straightforward. This review gives an overview of our current knowledge about the potential role ANO1 may play in hypertension within the systemic, portal, and pulmonary vascular systems and the importance of this information when pursuing potential treatment strategies. While the role of ANO1 is well-established in several forms of pulmonary hypertension, its contributions to both the generation of vascular tone and its role in hypertension within the systemic and portal systems are much less clear. This, combined with ANO1's various roles throughout a multitude of tissues throughout the body, command caution when targeting ANO1 as a therapeutic target and may require tissue-selective strategies.