Skip to main content

Advertisement

SpringerLink
Log in

Cystic fibrosis transmembrane conductance regulator modulates enteric cholinergic activities and is abnormally expressed in the enteric ganglia of patients with slow transit constipation

  • Original Article—Alimentary Tract
  • Published:
Journal of Gastroenterology Aims and scope Submit manuscript

Abstract

Background

Cystic fibrosis transmembrane conductance regulator (CFTR) was recently found in the enteric nervous system, where its role is unclear. We aimed to identify which enteric neuronal structures express CFTR, whether CFTR modulates enteric neurotransmission and if altered CFTR expression is associated with slow transit constipation (STC).

Methods

Immunofluorescence double labeling was performed to localize CFTR with various neuronal and glial cell markers in the human colon. The immunoreactivity (IR) of CFTR and choline acetyltransferase (ChAT) on myenteric plexus of control and STC colon was quantitatively analyzed. In control colonic muscle strips, electrical field stimulation (EFS) evoked contractile responses and the release of acetylcholine (ACh) was measured in the presence of the CFTR channel inhibitor, CFTR(inh)-172.

Results

CFTR-IR was densely localized to myenteric ganglia, where it was co-localized with neuronal markers HuC/D and β-tubulin, and glial marker S-100 but little with glial fibrillary acidic protein. Vesicular ACh transport was almost exclusively co-localized with CFTR, but neurons expressing nitric oxide synthase were CFTR negative. Significant reductions of CFTR-IR (P < 0.01) and ChAT-IR (P < 0.05) were observed on myenteric ganglia of STC compared to control. Pre-treatment of colonic muscle strips with CFTR(inh)-172 (10 µM) significantly inhibited EFS-evoked contractile responses (P < 0.01) and ACh release (P < 0.05).

Conclusions

Co-localization of CFTR-IR with cholinergic markers, inhibition of EFS-induced colonic muscle contractility and ACh release by CFTR(inh)-172 suggest that CFTR modulates enteric cholinergic neurotransmission. The downregulation of CFTR and ChAT in myenteric ganglia of STC correlated with the impaired contractile responses to EFS.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

Similar content being viewed by others

References

  1. Gustafsson JK, Ermund A, Ambort D, et al. Bicarbonate and functional CFTR channel are required for proper mucin secretion and link cystic fibrosis with its mucus phenotype. J Exp Med. 2012;209:1263–72.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  2. Char JE, Wolfe MH, Cho HJ, et al. A little CFTR goes a long way: CFTR-dependent sweat secretion from G551D and R117H–5T cystic fibrosis subjects taking ivacaftor. PLoS ONE. 2014;9:e88564.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  3. Oca F, Dreux S, Gerard B, et al. Amniotic Fluid Digestive Enzyme Analysis Is Useful for Identifying CFTR Gene Mutations of Unclear Significance. Clin Chem. 2009;55:2214–7.

    Article  CAS  PubMed  Google Scholar 

  4. Sorio C, Buffelli M, Angiari C, et al. Defective CFTR expression and function are detectable in blood monocytes: development of a new blood test for cystic fibrosis. PLoS ONE. 2011;6:e22212.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  5. Johansson J, Vezzalini M, Verze G, et al. Detection of CFTR protein in human leukocytes by flow cytometry. Cytometry A. 2014;85:611–20.

    Article  CAS  PubMed  Google Scholar 

  6. Verloo P, Kocken CH, Van der Wel A, et al. Plasmodium falciparum-activated chloride channels are defective in erythrocytes from cystic fibrosis patients. J Biol Chem. 2004;279:10316–22.

    Article  CAS  PubMed  Google Scholar 

  7. Su M, Guo Y, Zhao Y, et al. Expression of cystic fibrosis transmembrane conductance regulator in paracervical ganglia this paper is one of a selection of papers published in this special issue entitled “Second International Symposium on Recent Advances in Basic, Clinical, and Social Medicine” and has undergone the journal's usual peer review process. Biochem Cell Biol. 2010;88:747–55.

    Article  CAS  PubMed  Google Scholar 

  8. Riordan JR, Rommens JM, Kerem B, et al. Identification of the cystic fibrosis gene: cloning and characterization of complementary DNA. Science. 1989;245:1066–73.

    Article  CAS  PubMed  Google Scholar 

  9. Dalli J, Rosignoli G, Hayhoe RP, et al. CFTR inhibition provokes an inflammatory response associated with an imbalance of the annexin A1 pathway. Am J Pathol. 2010;177:176–86.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  10. Malats N, Casals T, Porta M, et al. Cystic fibrosis transmembrane regulator (CFTR) ΔF508 mutation and 5T allele in patients with chronic pancreatitis and exocrine pancreatic cancer. Gut. 2001;48:70–4.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  11. McLatchie LM, Young JS, Fry CH. Regulation of ACH release from guinea pig bladder urothelial cells: potential role in bladder filling sensations. Br J Pharmacol. 2014;171:3394–403.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  12. Ochodnicky P, Michel MB, Butter JJ, et al. Bradykinin modulates spontaneous nerve growth factor production and stretch-induced ATP release in human urothelium. Pharmacol Res. 2013;70:147–54.

    Article  CAS  PubMed  Google Scholar 

  13. Lubowski DZ, Chen FC, Kennedy ML, et al. Results of colectomy for severe slow transit constipation. Dis Colon Rectum. 1996;39:23–9.

    Article  CAS  PubMed  Google Scholar 

  14. Wong SW, Lubowski DZ. Slow-transit constipation: evaluation and treatment. ANZ J Surg. 2007;77:320–8.

    Article  PubMed  Google Scholar 

  15. Preston DM, Lennardjones JE. Severe chronic constipation of young-women - idiopathic slow transit constipation. Gut. 1986;27:41–8.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  16. Bassotti G, Villanacci V, Maurer CA, et al. The role of glial cells and apoptosis of enteric neurones in the neuropathology of intractable slow transit constipation. Gut. 2006;55:41–6.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  17. Tomita R, Tanjoh K, Fujisaki S, et al. Regulation of the enteric nervous system in the colon of patients with slow transit constipation. Hepatogastroenterology. 2002;49:1540–4.

    PubMed  Google Scholar 

  18. Liu L, Shang F, Morgan MJ, et al. Cyclooxygenase-dependent alterations in substance P-mediated contractility and tachykinin NK1 receptor expression in the colonic circular muscle of patients with slow transit constipation. J Pharmacol Exp Ther. 2009;329:282–9.

    Article  CAS  PubMed  Google Scholar 

  19. Jung HK, Kim DY, Moon IH. Effects of gender and menstrual cycle on colonic transit time in healthy subjects. Korean J Intern Med. 2003;18:181.

    Article  PubMed Central  PubMed  Google Scholar 

  20. Björnsson ES, Chey WD, Hooper F, et al. Impaired gastrocolonic response and peristaltic reflex in slow-transit constipation: role of 5-HT3 pathways. Am J Physiol Gastrointest Liver Physiol. 2002;283:G400–G407407.

    Article  PubMed  Google Scholar 

  21. Frattini JC, Nogueras JJ. Slow transit constipation: a review of a colonic functional disorder. Clin Colon Rectal Surg. 2008;21:146–52.

    Article  PubMed Central  PubMed  Google Scholar 

  22. Wong SW, Lubowski DZJ. Slow‐transit constipation: evaluation and treatment. ANZ J Surg. 2007;77:320–8.

    Article  PubMed  Google Scholar 

  23. Strong TV, Boehm K, Collins FS. Localization of cystic fibrosis transmembrane conductance regulator mRNA in the human gastrointestinal tract by in situ hybridization. J Clin Invest. 1994;93:347–54.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  24. Beyder A, Farrugia G. Ion channelopathies in functional GI disorders. Am J Physiol Gastrointest Liver Physiol. 2016;311:G581–G586586.

    Article  PubMed Central  PubMed  Google Scholar 

  25. Cil O, Phuan PW, Lee S, et al. CFTR activator increases intestinal fluid secretion and normalizes stool output in a mouse model of constipation. Cell Mol Gastroenterol Hepatol. 2016;2:317–27.

    Article  PubMed Central  PubMed  Google Scholar 

  26. Sharma S, Sharma T, Dhingra R, et al. Linaclotide-a novel secretagogue in the treatment of irritable bowel syndrome with constipation and chronic idiopathic constipation. Mini Rev Med Chem. 2013;13:1685–90.

    Article  CAS  PubMed  Google Scholar 

  27. Xue R, Gu H, Qiu Y, et al. Expression of cystic fibrosis transmembrane conductance regulator in ganglia of human gastrointestinal tract. Sci Rep. 2016;6:30926.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  28. Varghese F, Bukhari AB, Malhotra R, et al. IHC Profiler: an open source plugin for the quantitative evaluation and automated scoring of immunohistochemistry images of human tissue samples. PLoS ONE. 2014;9:e96801.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  29. Dhooghe B, Noel S, Bouzin C, et al. Correction of chloride transport and mislocalization of CFTR protein by vardenafil in the gastrointestinal tract of cystic fibrosis mice. PLoS ONE. 2013;8:e77314.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  30. Doucet L, Mendes F, Montier T, et al. Applicability of different antibodies for the immunohistochemical localization of CFTR in respiratory and intestinal tissues of human and murine origin. J Histochem Cytochem. 2003;51:1191–9.

    Article  CAS  PubMed  Google Scholar 

  31. Valdivieso AG, Marin MC, Clauzure M, et al. Measurement of cystic fibrosis transmembrane conductance regulator activity using fluorescence spectrophotometry. Anal Biochem. 2011;418:231–7.

    Article  CAS  PubMed  Google Scholar 

  32. Walsh DE, Harvey BJ, Urbach V. CFTR regulation of intracellular calcium in normal and cystic fibrosis human airway epithelia. J Membr Biol. 2000;177:209–19.

    Article  CAS  PubMed  Google Scholar 

  33. Billet A, Luo Y, Balghi H, et al. Role of tyrosine phosphorylation in the muscarinic activation of the cystic fibrosis transmembrane conductance regulator (CFTR). J Biol Chem. 2013;288:21815–233.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  34. Billet A, Hanrahan JW. The secret life of CFTR as a calcium-activated chloride channel. J Physiol. 2013;591:5273–8.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  35. Thiagarajah JR, Song Y, Haggie PM, et al. A small molecule CFTR inhibitor produces cystic fibrosis-like submucosal gland fluid secretions in normal airways. FASEB J. 2004;18:875–7.

    Article  CAS  PubMed  Google Scholar 

  36. Leveque M, Penna A, Le Trionnaire S, et al. Phagocytosis depends on TRPV2-mediated calcium influx and requires TRPV2 in lipids rafts: alteration in macrophages from patients with cystic fibrosis. Sci Rep-Uk. 2018;8:4310.

    Article  CAS  Google Scholar 

  37. Zhao YM, Migita K, Sun J, et al. MRP transporters as membrane machinery in the bradykinin-inducible export of ATP. N-S Arch Pharmacol. 2010;381:315–20.

    Article  CAS  Google Scholar 

  38. Braunstein GM, Roman RM, Clancy JP, et al. Cystic fibrosis transmembrane conductance regulator facilitates ATP release by stimulating a separate ATP release channel for autocrine control of cell volume regulation. J Biol Chem. 2001;276:6621–30.

    Article  CAS  PubMed  Google Scholar 

  39. Wedel T, Roblick UJ, Ott V, et al. Oligoneuronal hypoganglionosis in patients with idiopathic slow-transit constipation. Dis Colon Rectum. 2002;45:54–62.

    Article  CAS  PubMed  Google Scholar 

  40. Park HJ, Kamm MA, Abbasi AM, et al. Immunohistochemical study of the colonic muscle and innervation in idiopathic chronic constipation. Dis Colon Rectum. 1995;38:509–13.

    Article  CAS  PubMed  Google Scholar 

  41. Broadhead MJ, Bayguinov PO, Okamoto T, et al. Ca2+ transients in myenteric glial cells during the colonic migrating motor complex in the isolated murine large intestine. J Physiol. 2012;590:335–50.

    Article  CAS  PubMed  Google Scholar 

  42. Boesmans W, Lasrado R, Vanden Berghe P, et al. Heterogeneity and phenotypic plasticity of glial cells in the mammalian enteric nervous system. GLIA. 2015;63:229–41.

    Article  PubMed  Google Scholar 

  43. MacEachern SJ, Patel BA, McKay DM, et al. Nitric oxide regulation of colonic epithelial ion transport: a novel role for enteric glia in the myenteric plexus. J Physiol. 2011;589:3333–48.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  44. De Giorgio R, Giancola F, Boschetti E, et al. Enteric glia and neuroprotection: basic and clinical aspects. Am J Physiol Gastrointest Liver Physiol. 2012;303:G887–G89393.

    Article  CAS  PubMed  Google Scholar 

  45. Gulbransen BD, Sharkey KA. Novel functional roles for enteric glia in the gastrointestinal tract. Nat Rev Gastroenterol Hepatol. 2012;9:625–32.

    Article  CAS  PubMed  Google Scholar 

  46. Neunlist M, Van Landeghem L, Mahé MM, et al. The digestive neuronal–glial–epithelial unit: a new actor in gut health and disease. Nat Rev Gastroenterol Hepatol. 2013;10:90–100.

    Article  CAS  PubMed  Google Scholar 

  47. Bassotti G, Chiarioni G, Imbimbo BP, et al. Impaired colonic motor response to cholinergic stimulation in patients with severe chronic idiopathic (slow transit type) constipation. Dig Dis Sci. 1993;38:1040–5.

    Article  CAS  PubMed  Google Scholar 

  48. Wedel T, Spiegler J, Soellner S, et al. Enteric nerves and interstitial cells of Cajal are altered in patients with slow-transit constipation and megacolon. Gastroenterology. 2002;123:1459–67.

    Article  PubMed  Google Scholar 

  49. De Lisle RC. Altered transit and bacterial overgrowth in the cystic fibrosis mouse small intestine. Am J Physiol Gastrointest Liver Physiol. 2007;293:G104–G111111.

    Article  CAS  PubMed  Google Scholar 

  50. Lynch SV, Goldfarb KC, Wild YK, et al. Cystic fibrosis transmembrane conductance regulator knockout mice exhibit aberrant gastrointestinal microbiota. Gut Microbes. 2013;4:41–7.

    Article  PubMed Central  PubMed  Google Scholar 

  51. De Lisle RC, Meldi L, Flynn M, et al. Altered eicosanoid metabolism in the cystic fibrosis mouse small intestine. J Pediatr Gastroenterol Nutr. 2008;47:406–16.

    Article  PubMed  Google Scholar 

  52. De Lisle RC, Sewell R, Meldi L. Enteric circular muscle dysfunction in the cystic fibrosis mouse small intestine. Neurogastroenterol Motil. 2010;22:341–e87.

    Article  CAS  PubMed  Google Scholar 

  53. De Lisle RC, Borowitz D. The cystic fibrosis intestine. Cold Spring Harb Perspect Med. 2013;3:a009753.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  54. Cil O, Phuan PW, Son JH, et al. Phenylquinoxalinone CFTR activator as potential prosecretory therapy for constipation. Transl Res. 2017;182(14–26):e4.

    Google Scholar 

Download references

Acknowledgments

This study was supported by a project Grant from the National Health and Medical Research Council of Australia (APP1048885), and by the UNSW and Shanghai Jiao Tong University Collaborative Seed Grant. We would like to offer our special thanks to Stelina Drimousis and Erica Diezmos in specimen collection and preparation.

Author information

Authors and Affiliations

  1. Department of Pharmacology, Faculty of Medicine, School of Medical Sciences, University of New South Wales, Sydney, NSW, 2052, Australia

    Ka Ming Yeh, Huy Le, Karan Rao, Irit Markus & Lu Liu

  2. Faculty of Medicine and Health Sciences, Linköping University, Linköping, Sweden

    Olle Johansson

  3. Sydney Colorectal Associates, Sydney, Australia

    Dayashan Shevy Perera, David Zachary Lubowski & Denis Warwick King

  4. School of Biotechnology and Biomolecular Sciences, University of New South Wales, Sydney, Australia

    Li Zhang

  5. School of Medicine, Shanghai Jiao Tong University, Shanghai, China

    Hongzhuan Chen

Authors
  1. Ka Ming Yeh
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Olle Johansson
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Huy Le
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Karan Rao
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Irit Markus
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Dayashan Shevy Perera
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. David Zachary Lubowski
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Denis Warwick King
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Li Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Hongzhuan Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Lu Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Lu Liu.

Ethics declarations

Conflict of interest

There are no conflicts of interest.

Additional information

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary file1 (DOCX 4784 kb)

Supplementary file2 (DOCX 157 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Yeh, K.M., Johansson, O., Le, H. et al. Cystic fibrosis transmembrane conductance regulator modulates enteric cholinergic activities and is abnormally expressed in the enteric ganglia of patients with slow transit constipation. J Gastroenterol 54, 994–1006 (2019). https://doi.org/10.1007/s00535-019-01610-9

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00535-019-01610-9

Keywords

Search

Navigation