Effects of Ca2+-Activated Cl- Channel ANO1inhibitors on Pacemaker Activity in Interstitial Cells of Cajal

Abstract

Background/Aims: Anoctamin1 (Ca²⁺-activated Cl^- channel, ANO1) is a specific marker of the interstitial cells of Cajal (ICC) in the gastrointestinal tract, and are candidate proteins that can function as pacemaker channels. Recently, novel selective ANO1 inhibitors were discovered and used to study Ca²⁺-activated Cl^- channels. Therefore, to investigate whether ANO1 channels function as pacemaker channels, selective ANO1 inhibitors were tested with respect to the pacemaker potentials in ICC. Methods: Whole-cell patch-clamp recording, RT-PCR, and intracellular Ca²⁺ ([Ca²⁺]_i) imaging were performed in cultured ICC obtained from mice. Results: Though CaCCinh-A01 (5 µM), T16Ainh-A01 (5 µM), and MONNA (5 µM) (selective ANO1 inhibitors) blocked the generation of pacemaker potentials in colonic ICC, they did not do so in small intestinal ICC. Though nifulmic acid (10 µM) and DIDS (10 µM) (classical Ca²⁺-activated Cl^- channel inhibitors) also had no effect in small intestinal ICC, they suppressed the generation of pacemaker potentials in colonic ICC. In addition, knockdown of ANO1 reduced the pacemaker potential frequency in colonic ICC alone. Though ANO1 inhibitors suppressed [Ca²⁺]_i oscillations in colonic ICC, they did not do so in small intestinal ICC. T-type Ca²⁺ channels were expressed in the both the small intestinal and colonic ICC, but mibefradil (5 µM) and NiCl₂ (30 µM) (T-type Ca²⁺ channel inhibitors) inhibited the generation of pacemaker potentials in colonic ICC alone. Conclusion: These results indicate that though ANO1 and T-type Ca²⁺ channels participate in generating pacemaker potentials in colonic ICC, they do not do so in small intestinal ICC. Therefore, the mechanisms underlying pacemaking in ICC might be different in the small intestine and the colon.

Differential sensitivity of gastric and small intestinal muscles to inducible knockdown of anoctamin 1 and the effects on gastrointestinal motility

“…A recent study of cultured mouse small intestinal ICC also reported a lack of sensitivity of slow waves to Ano1 antagonists (Choi et al . ). Thus, it appears that development of a cation conductance may occur as a function of age in small intestinal pacemaker ICC, and this process may be accelerated or possibly replace Ano1 in cell culture conditions.…”

Cyclic Stretch Promotes Proliferation and Contraction of Human Bladder Smooth Muscle Cells by Cajal-Mediated c-kit Expression in Interstitial Cells

“…Interstitial cells of Cajal (ICCs) are a group of special mesenchymal cells first discovered in the gastrointestinal tract. They are the pacemaker of the slow wave of the gastrointestinal tract, transmitting electrical signals and mediating the signal regulation between the intestinal nerves and smooth muscle [6,7]. ICCs were also in found in the urinary system of guinea pigs [8–10], and they exist in the autonomic nerve endings and the bladder smooth muscle cells.…”

TMEM16A in smooth muscle cells acts as a pacemaker channel in the internal anal sphincter

“…Two additional TMEM16A inhibitors, CaCCinh-A01 and Ani9, have region-specific effects in the GI 32 , 33 . Both, at 10 μM, block Cl Ca channels 34 , 35 .…”