Quercetin Inhibits Pacemaker Potentials via Nitric Oxide/cGMP-Dependent Activation and TRPM7/ANO1 Channels in Cultured Interstitial Cells of Cajal from Mouse Small Intestine

Abstract

Background: Quercetin regulates gastrointestinal (GI) motor activity but the molecular mechanism involved has not been determined. The authors investigated the effects of quercetin, a flavonoid present in various foods, on the pacemaker activities of interstitial cells of Cajal (ICCs) in murine small intestine in vitro and on GI motility in vivo. Materials and Methods: Enzymatic digestion was used to dissociate ICCs from mouse small intestines. The whole-cell patch-clamp configuration was used to record pacemaker potentials in cultured ICCs in the absence or presence of quercetin and to record membrane currents of transient receptor potential melastatin (TRPM) 7 or transmembrane protein 16A (Tmem16A, anoctamin1 (ANO1)) overexpressed in human embryonic kidney (HEK) 293 cells. The in vivo effects of quercetin on GI motility were investigated by measuring the intestinal transit rates (ITRs) of Evans blue in normal mice. Results: Quercetin (100-200 μM) decreased the amplitudes and frequencies of pacemaker activity in a concentration-dependent manner in current clamp mode, but this action was blocked by naloxone (a pan-opioid receptor antagonist) and by GDPβS (a GTP-binding protein inhibitor). However, potassium channels were not involved in these inhibitory effects of quercetin. To study the quercetin signaling pathway, we examined the effects of 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ), an inhibitor of guanylate cyclase, and of RP-8-CPT-cGMPS, an inhibitor of protein kinase G (PKG). These inhibitors blocked the inhibitory effects of quercetin on pacemaker activities. Also, L-NAME (100 μM), a non-selective NO synthase (NOS) inhibitor, blocked the effects of quercetin on pacemaker activity and quercetin stimulated cGMP production. Furthermore, quercetin inhibited both Ca²⁺-activated Cl^- channels (TMEM16A, ANO1) and TRPM7 channels. In vivo, quercetin (10-100 mg/kg, p.o.) decreased ITRs in normal mice in a dose-dependent manner. Conclusions: Quercetin inhibited ICC pacemaker activities by inhibiting TRPM7 and ANO1 via opioid receptor signaling pathways in cultured murine ICCs. The study shows quercetin attenuates GI tract motility, and suggests quercetin be considered the basis for the development of novel spasmolytic agents for the prevention or alleviation of GI motility dysfunctions.