Hydroxy-α sanshool induces colonic motor activity in rat proximal colon: a possible involvement of KCNK9.
Bottom Line: Hydroxy-α sanshool (HAS), a readily absorbable active ingredient of TU-100 and a KCNK3/KCNK9/KCNK18 blocker as well as TRPV1/TRPA1 agonist, has been investigated for its effects on colonic motility.The effect of HAS was unaltered by TRPV1/TRPA1 antagonists and desensitization.HAS may evoke strong LDC possibly via blockage of the neural KCNK9 channel in the colonic myenteric plexus.
Affiliation: Tsumura Research Laboratories, Tsumura & Co., Ibaraki, Japan;Show MeSH
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Mentions: To investigate the possible involvement of these KCNK channels in HAS-induced motility, RT-PCR was performed with specific primers for rat KCNK3, KCNK9, and KCNK18. Our results show that KCNK3 and KCNK9 mRNAs are expressed in the muscle layer, whereas KCNK18 mRNA is barely detectable in the rat proximal colon (Fig. 4A). The localization of KCNK3 and KCNK9 was evaluated by ISH and immunohistochemistry. As shown in Fig. 4B, KCNK3 mRNA was localized in the longitudinal muscle (LM) layer and myenteric plexus (MP) and KCNK9 mRNA in MP (Fig. 4B). A neuronal marker PGP9.5 mRNA coexisted with KCNK9 mRNA (Fig. 4B). Immunohistochemistry of whole-mount preparation of colonic muscle layers confirmed the predominant localization of KCNK9 in MP and in the enteric nerves in the circular muscle (CM) layer (Fig. 5A), and that of KCNK3 in LM SMCs (Fig. 5B). Coimmunostaining analysis with anti-KCNK9 and anti-PGP9.5 antibodies (Fig. 5C) showed that KCNK9 is expressed in 35.8% of PGP9.5+ MP cells (n = 148). More than a quarter of KCNK9+ cells in MP are PGP9.5−. Furthermore, in coimmunostaining experiments with KCNK9 and an ICC marker c-Kit (Fig. 5D), 22.4% of c-Kit+ cells in MP were found to be KCNK9+ (n = 156) though the double-positive cells had relatively weaker c-Kit immunosignals (i.e., most of the cells with the strongest c-Kit immunosignals lacked KCNK9 immunosignal). Nearly half of KCNK9+ cells in MP were c-Kit− (often located adjacent to c-Kit+ cells). KCNK9 signals were not detected in PDGFR-α-positive fibroblast-like interstitial cells (data not shown), which are thought to participate in inhibitory neurotransmission of enteric nerves.
Affiliation: Tsumura Research Laboratories, Tsumura & Co., Ibaraki, Japan;