Chloride channels are necessary for full platelet phosphatidylserine exposure and procoagulant activity

Platelets enhance thrombin generation at sites of vascular injury by exposing phosphatidylserine during necrosis-like cell death. Anoctamin 6 (Ano6) is required for Ca2+-dependent phosphatidylserine exposure and is defective in patients with Scott syndrome, a rare bleeding disorder. Ano6 may also form Cl− channels, though the role of Cl− fluxes in platelet procoagulant activity has not been explored. We found that Cl− channel blockers or removal of extracellular Cl− inhibited agonist-induced phosphatidylserine exposure. However, this was not due to direct inhibition of Ca2+-dependent scrambling since Ca2+ ionophore-induced phosphatidylserine exposure was normal. This implies that the role of Ano6 in Ca2+−dependent PS exposure is likely to differ from any putative function of Ano6 as a Cl− channel. Instead, Cl− channel blockade inhibited agonist-induced Ca2+ entry. Importantly, Cl− channel blockers also prevented agonist-induced membrane hyperpolarization, resulting in depolarization. We propose that Cl− entry through Cl− channels is required for this hyperpolarization, maintaining the driving force for Ca2+ entry and triggering full phosphatidylserine exposure. This demonstrates a novel role for Cl− channels in controlling platelet death and procoagulant activity.