title: Non-neuronal acetylcholine as an endogenous regulator of proliferation and differentiation of Lgr5-positive stem cells in mice.
abstract: Non-neuronal acetylcholine (ACh) is predicted to function as a local cell signaling molecule. However, the physiological significance of the synthesis of non-neuronal ACh in the intestine remains unclear. Here, experiments using crypt-villus organoids that lack nerve and immune cells in culture led us to suggest that endogenous ACh is synthesized in the intestinal epithelium to evoke growth and differentiation of the organoids through activation of muscarinic ACh receptors (mAChRs). The extracts of the cultured organoids showed a noticeable capacity for ACh synthesis that was sensitive to a potent inhibitor of choline acetyltransferase. Imaging MS revealed endogenous ACh localized in the epithelial layer in mouse small intestinal epithelium in vivo, suggesting that there are non-neuronal resources of ACh. Treatment of organoids with carbachol downregulated the growth of organoids and the expression of marker genes for epithelial cells. On the other hand, antagonists for mAChRs enhanced the growth and differentiation of organoids, indicating the involvement of mAChRs in regulating the proliferation and differentiation of Lgr5-positive stem cells. Collectively, our data provide evidence that endogenous ACh released from intestinal epithelium maintains homeostasis of intestinal epithelial cell growth and differentiation via mAChRs in mice.
authors.affiliation: Suntory Foundation for Life Sciences, Bioorganic Research Institute, Osaka, Japan.
authors.forename: Toshio
authors.initials: T
authors.lastname: Takahashi
authors.forename: Hiroe
authors.initials: H
authors.lastname: Ohnishi
authors.forename: Yuki
authors.initials: Y
authors.lastname: Sugiura
authors.forename: Kurara
authors.initials: K
authors.lastname: Honda
authors.forename: Makoto
authors.initials: M
authors.lastname: Suematsu
authors.forename: Takashi
authors.initials: T
authors.lastname: Kawasaki
authors.forename: Tomonori
authors.initials: T
authors.lastname: Deguchi
authors.forename: Takeshi
authors.initials: T
authors.lastname: Fujii
authors.forename: Kaoru
authors.initials: K
authors.lastname: Orihashi
authors.forename: Yoshitaka
authors.initials: Y
authors.lastname: Hippo
authors.forename: Takehiro
authors.initials: T
authors.lastname: Watanabe
authors.forename: Tohru
authors.initials: T
authors.lastname: Yamagaki
authors.forename: Shunsuke
authors.initials: S
authors.lastname: Yuba
mesh_terms: D000109:Acetylcholine / Q000494:pharmacology*; D000818:Animals; D015153:Blotting, Western; D002454:Cell Differentiation / Q000187:drug effects*; D049109:Cell Proliferation / Q000187:drug effects*; D002478:Cells, Cultured; D002795:Choline O-Acetyltransferase / Q000378:metabolism; D018679:Cholinergic Agonists / Q000494:pharmacology; D005434:Flow Cytometry; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D007124:Immunoenzyme Techniques; D019426:Integrases / Q000378:metabolism; D007413:Intestinal Mucosa / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D009940:Organoids / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D043562:Receptors, G-Protein-Coupled / Q000502:physiology*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013234:Stem Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism
chemical_list: D018679:Cholinergic Agonists; C493597:Lgr5 protein, mouse; D012333:RNA, Messenger; D043562:Receptors, G-Protein-Coupled; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D002795:Choline O-Acetyltransferase; C045073:Cre recombinase; D019426:Integrases; D000109:Acetylcholine
keywords: Lgr5; cholinergic system; gut; non-neuronal acetylcholine; organoid