Huiran Zhang

Activation of the Cl- channel ANO1 by localized calcium signals in nociceptive sensory neurons requires coupling with the IP3 receptor.

Positioning of an excitatory channel where the endoplasmic reticulum and plasma membrane meet enables pain-sensing neurons to distinguish among calcium signals. Insulating Pain Signals Activation of damage-sensing neurons evokes pain. Jin et al. describe a mechanism by which the depolarizing chloride channel anoctamin 1 (ANO1) is coupled only to those calcium signals that arise from activation of proinflammatory G protein (heterotrimeric guanine nucleotide–binding protein)–coupled receptors (GPCRs), rather than by global calcium signals mediated by voltage-gated calcium channels (VGCCs). ANO1, which is present in the plasma membrane, was found in a lipid raft–based complex that contained the activating GPCRs and the endoplasmic reticulum (ER)–resident calcium channel, IP3R1 (inositol 1,4,5-trisphosphate receptor 1). The interaction of ANO1 with IP3R1 and the ability of the local calcium signals to stimulate ANO1 activity were lost when lipid microdomains were chemically disrupted. Instead, ANO1 became receptive to VGCC activation in the absence of lipid rafts. Thus, a plasma membrane–ER microdomain complex limits activation of nociceptive neurons to only the appropriate signals. We report that anoctamin 1 (ANO1; also known as TMEM16A) Ca2+-activated Cl− channels in small neurons from dorsal root ganglia are preferentially activated by particular pools of intracellular Ca2+. These ANO1 channels can be selectively activated by the G protein–coupled receptor (GPCR)–induced release of Ca2+ from intracellular stores but not by Ca2+ influx through voltage-gated Ca2+ channels. This ability to discriminate between Ca2+ pools was achieved by the tethering of ANO1-containing plasma membrane domains, which also contained GPCRs such as bradykinin receptor 2 and protease-activated receptor 2, to juxtamembrane regions of the endoplasmic reticulum. Interaction of the carboxyl terminus and the first intracellular loop of ANO1 with IP3R1 (inositol 1,4,5-trisphosphate receptor 1) contributed to the tethering. Disruption of membrane microdomains blocked the ANO1 and IP3R1 interaction and resulted in the loss of coupling between GPCR signaling and ANO1. The junctional signaling complex enabled ANO1-mediated excitation in response to specific Ca2+signals rather than to global changes in intracellular Ca2+.

Membrane microdomain determines the specificity of receptor-mediated modulation of Kv7/M potassium currents.

The Kv7/M current is one of the major mechanisms controlling neuronal excitability, which can be modulated by activation of the G protein-coupled receptor (GPCR) via distinct signaling pathways. Membrane microdomains known as lipid rafts have been implicated in the specificity of various cell signaling pathways. The aim of this study was to understand the role of lipid rafts in the specificity of Kv7/M current modulation by activation of GPCR. Methyl-β-cyclodextrin (MβCD), often used to disrupt the integrity of lipid rafts, significantly reduced the bradykinin receptor (B2R)-induced but not muscarinic receptor (M1R)-induced inhibition of the Kv7/M current. B2R and related signaling molecules but not M1R were found in caveolin-containing raft fractions of the rat superior cervical ganglia. Furthermore, activation of B2R resulted in translocation of additional B2R into the lipid rafts, which was not observed for the activation of M1R. The increase of B2R-induced intracellular Ca(2+) was also greatly reduced after MβCD treatment. Finally, B2R but not M1R was found to interact with the IP3 receptor. In conclusion, the present study implicates an important role for lipid rafts in mediating specificity for GPCR-mediated inhibition of the Kv7/M current.

Tannic acid modulates excitability of sensory neurons and nociceptive behavior and the Ionic mechanism.

M/Kv7 K(+) channels, Ca(2+)-activated Cl(-) channels (CaCCs) and voltage gated Na(+) channels expressed in dorsal root ganglia (DRG) play an important role in nociception. Tannic acid has been proposed to be involved in multiple beneficial health effects; tannic acid has also been described to be analgesic. However the underlying mechanism is unknown. In this study, we investigated the effects of tannic acid on M/Kv7 K(+), Na(+) currents and CaCCs, and the effects on bradykinin-induced nociceptive behavior. A perforated patch technique was used. The bradykinin-induced rat pain model was used to assess the analgesic effect of tannic acid. We demonstrated that tannic acid enhanced M/Kv7 K(+) currents but inhibited bradykinin-induced activation of CaCC/TMEM16A currents in rat small DRG neurons. Tannic acid potentiated Kv7.2/7.3 and Kv7.2 currents expressed in HEK293B cells, with an EC50 of 7.38 and 5.40 µM, respectively. Tannic acid inhibited TTX-sensitive and TTX-insensitive currents of small DRG neurons with IC50 of 5.25 and 8.43 µM, respectively. Tannic acid also potently suppressed the excitability of small DRG neurons. Furthermore, tannic acid greatly reduced bradykinin-induced pain behavior of rats. This study thus demonstrates that tannic acid is an activator of M/Kv7 K(+) and an inhibitor of voltage-gated Na(+) channels and CaCC/TMEM16A, which may underlie its inhibitory effects on excitability of DRG neurons and its analgesic effect. Tannic acid could be a useful agent in treatment of inflammatory pain conditions such as osteoarthritis, rheumatic arthritis and burn pain.

Characterization and structure-activity relationship of natural flavonoids as hERG K+ channel modulators

Objectives: Flavonoids are present in varying concentrations in plant foods and have been reported to have numerous pharmacological activities, such as anti‐cancer, antioxidant, anti‐inflammatory, hepatoprotective, and vasodilator effects. We found that quercetin, fisetin, and some related flavonoid derivatives could inhibit human ether‐à‐go‐go‐related gene (hERG) K+ channels. Key findings: In this study, we tested the effects of a series of flavonoids on the hERG K+ channel expressed in HEK293 cells. For the first time, we demonstrate that quercetin and fisetin (Fise) are potent hERG current blockers. The 50% inhibiting concentration (IC50) and maximum efficacy (Emax) of quercetin were 11.8 ± 0.9 &mgr;M and 82 ± 2%, while those of fisetin were 38.4 ± 6 &mgr;M and 100 ± 6%, respectively. Luteolin (Lute) was a less potent inhibitor of hERG current (48 ± 1% at 100 &mgr;M). Galangin, kaempferol, and isorhamnetin (100 &mgr;M) showed weaker activity on the hERG currents. Conclusion: These results suggest that quercetin, fisetin, and luteolin are potent hERG K+ channel inhibitors and reveal the structure‐activity relationship of natural flavonoids. HighlightsFlavonoid derivatives quercetin, fisetin, and luteolin are potent hERG K+ channel inhibitors.The present work reveal the structure‐activity relationship of natural flavonoids.The results also provide new mechanistic evidence for understanding the effects of these natural products.

Inhibition of transmembrane member 16A calcium-activated chloride channels by natural flavonoids contributes to flavonoid anticancer effects.

Natural flavonoids are ubiquitous in dietary plants and vegetables and have been proposed to have antiviral, antioxidant, cardiovascular protective and anticancer effects. Transmembrane member 16A (TMEM16A)‐encoded Ca2+‐activated Cl− channels play a variety of physiological roles in many organs and tissues. Overexpression of TMEM16A is also believed to be associated with cancer progression. Therefore, inhibition of TMEM16A current may be a potential target for cancer therapy. In this study, we screened a broad spectrum of flavonoids for their inhibitory activities on TMEM16A currents.

Inhibition of TMEM16A calcium‐activated chloride channels by natural flavonoids contributes to flavonoid anticancer effects

Natural flavonoids are ubiquitous in dietary plants and vegetables and have been proposed to have antiviral, antioxidant, cardiovascular protective and anticancer effects. Transmembrane member 16A (TMEM16A)‐encoded Ca2+‐activated Cl− channels play a variety of physiological roles in many organs and tissues. Overexpression of TMEM16A is also believed to be associated with cancer progression. Therefore, inhibition of TMEM16A current may be a potential target for cancer therapy. In this study, we screened a broad spectrum of flavonoids for their inhibitory activities on TMEM16A currents.