The Natural Compound Cinnamaldehyde is a Novel Activator of Calcium-Activated Chloride Channel.

Calcium-activated chloride channels (CaCCs) play important roles in a multitude of physiological processes, and in many cells types, TMEM16A was identified as the molecular basis of CaCC. Abnormal CaCC function has been implicated in variety of diseases, which reinforces the need for modulators of CaCCs/TMEM16A. However, there are few specific, clinical modulators of CaCCs. Here, we identified a potent novel activator of TMEM16A from a bank of traditional Chinese medicines (TCM) and explored its mechanism of activation by laser confocal scanning microscopy and patch clamping. Fluorescence data demonstrated that among the 36 tested TCM medicines, one compound, cinnamaldehyde (CA), can activate the TMEM16A channel in a dose-dependent manner. To determine the mechanism by which CA activates the TMEM16A channel, we performed an excised patch clamp experiment and measured the intracellular calcium concentration in fluorescence experiments. Our data show that CA activates TMEM16A channels by elevating the intracellular concentrations of calcium ions. The results of the whole-cell patch clamping showed that CA dose-dependently activates these channels, with an EC50 of 9.73 ± 5.64 µM at + 80 mV, and prolongs the deactivation of TMEM16A. Finally, we found that CA can strengthen contractions of the ileum in guinea pigs by activating TMEM16A. The results demonstrate that CA is a novel, natural activator of TMEM16A.

Hydrocinnamic Acid Inhibits the Currents of WT and SQT3 Syndrome-Related Mutants of Kir2.1 Channel.

Gain of function in mutations, D172N and E299V, of Kir2.1 will induce type III short QT syndrome. In our previous work, we had identified that a mixture of traditional Chinese medicine, styrax, is a blocker of Kir2.1. Here, we determined a monomer, hydrocinnamic acid (HA), as the effective component from 18 compounds of styrax. Our data show that HA can inhibit the currents of Kir2.1 channel in both excised inside-out and whole-cell patch with the IC50 of 5.21 ± 1.02 and 10.08 ± 0.46 mM, respectively. The time course of HA blockage and washout are 2.3 ± 0.6 and 10.5 ± 2.6 s in the excised inside-out patch. Moreover, HA can also abolish the currents of D172N and E299V with the IC50 of 6.66 ± 0.57 and 5.81 ± 1.10 mM for D172N and E299V, respectively. Molecular docking results determine that HA binds with Kir2.1 at K182, K185, and K188, which are phosphatidylinositol 4,5-bisphosphate (PIP2) binding residues. Our results indicate that HA competes with PIP2 to bind with Kir2.1 and inhibits the currents.

Styrax blocks inward and outward current of Kir2.1 channel.

Kir2.1 plays key roles in setting rest membrane potential and modulation of cell excitability. Mutations of Kir2.1, such as D172N or E299V, inducing gain-of-function, can cause type3 short QT syndrome (SQT3) due to the enlarged outward currents. So far, there is no clinical drug target to block the currents of Kir2.1. Here, we identified a novel blocker of Kir2.1, styrax, which is a kind of natural compound selected from traditional Chinese medicine. Our data show that styrax can abolish the inward and outward currents of Kir2.1. The IC50 of styrax for WT, D172N and E299V are 0.0113 ± 0.00075, 0.0204 ± 0.0048 and 0.0122 ± 0.0012 (in volume), respectively. The results indicate that styrax can serve as a novel blocker for Kir2.1.