Ulcerative colitis is an inflammatory bowel disease (IBD) characterized by recurrent episodes of colonic inflammation and tissue degeneration in human or animal models. The contractile force generated by the smooth muscle is significantly attenuated, resulting in altered motility leading to diarrhea or constipation in IBD. The aim of this study is to clarify the altered contractility of circular and longitudinal smooth muscle layers in proximal colon of trinitrobenzen sulfonic acid (TNBS)-induced colitis mouse. Colitis was induced by direct injection of TNBS (120 mg/kg, 50% ethanol) in proximal colon of ICR mouse using a 30 G needle anesthetized with ketamin (50 mg/kg), whereas animals in the control group were injected of 50% ethanol alone. In TNBS-induced colitis, the wall of the proximal colon is diffusely thickened with loss of haustration, and showed mucosal and mucular edema with inflammatory infiltration. The colonic inflammation is significantly induced the reduction of colonic contractile activity including spontaneous contractile activity, depolarization-induced contractility, and muscarinic acetylcholine receptor-mediated contractile response in circular muscle layer compared to the longitudinal muscle layer. The inward rectification of currents, especially, important to Ca2+ and Na+ influx-induced depolarization and contraction, was markedly reduced in the TNBS-induced colitis compared to the control. The muscarinic acetylcholine-mediated contractile responses were significantly attenuated in the circular and longitudinal smooth muscle strips induced by the reduction of membrane expression of canonical transient receptor potential (TRPC) channel isoforms from the proximal colon of the TNBS-induced colitis mouse than the control.
Transient receptor potential canonical type 3 (TRPC3) channels are non-selective cation channels and regulate intracellular Ca2+ concentration. We examined the role of TRPC3 channels in agonist-, membrane depolarization (high K+)-, and mechanical (pressure)-induced vasoconstriction and vasorelaxation in mouse mesenteric arteries. Vasoconstriction and vasorelaxation of endothelial cells intact mesenteric arteries were measured in TRPC3 wild-type (WT) and knockout (KO) mice. Calcium concentration ([Ca2+]) was measured in isolated arteries from TRPC3 WT and KO mice as well as in the mouse endothelial cell line bEnd.3. Nitric oxide (NO) production and nitrate/nitrite concentrations were also measured in TRPC3 WT and KO mice. Phenylephrine-induced vasoconstriction was reduced in TRPC3 KO mice when compared to that of WT mice, but neither high K+- nor pressure-induced vasoconstriction was altered in TRPC3 KO mice. Acetylcholine-induced vasorelaxation was inhibited in TRPC3 KO mice and by the selective TRPC3 blocker pyrazole-3. Acetylcholine blocked the phenylephrine-induced increase in Ca2+ ratio and then relaxation in TRPC3 WT mice but had little effect on those outcomes in KO mice. Acetylcholine evoked a Ca2+ increase in endothelial cells, which was inhibited by pyrazole-3. Acetylcholine induced increased NO release in TRPC3 WT mice, but not in KO mice. Acetylcholine also increased the nitrate/nitrite concentration in TRPC3 WT mice, but not in KO mice. The present study directly demonstrated that the TRPC3 channel is involved in agonist-induced vasoconstriction and plays important role in NO-mediated vasorelaxation of intact mesenteric arteries.
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