Imaging activity in nerves that mediate pain from gastrointestinal organs

Czech version

Authors: Lipták P. ¹; Pavelková N. ^2,3; Harsányiová J. ^2,3; Bánovčin P. jr. ¹; Kollárik M. ³
Authors‘ workplace: Interná klinika – gastroenterologická JLF UK a UN Martin, Slovenská republika ¹; Ústav patologickej fyziológie, JLF UK v Bratislave, Slovenská republika ²; Department of Molecular Pharmacology & Physiology, Morsani College of Medicine, University of South Florida, USA ³
Published in: Gastroent Hepatol 2019; 73(4): 313-318
Category: Clinical and Experimental Gastroenterology: Review Article

Overview

Background: Visceral afferent nerves project from internal organs and provide information to the central nervous system. The role of afferent nerves in the pathogenesis and symptoms of gastrointestinal diseases is often underappreciated. Nonetheless, pain, heartburn, fullness, nausea, and vomiting are all mediated by visceral afferent nerves. Furthermore, stimulation of visceral afferent nerves by inflammation in diseases disrupts the normal regulation of gastrointestinal secretion and motility, thereby contributing to symptoms ranging from dysphagia to diarrhea and constipation. Thus, therapeutic targeting of visceral afferent nerves may aid the treatment of many gastrointestinal diseases. However, the development of novel drugs targeting visceral afferent nerves is hindered by the limited knowledge of these nerves. For example, the identities of the acid receptors in esophageal nerves that cause heartburn and the ion channels responsible for mechanical activation in nerves that mediate colic pain have not been fully elucidated. The limited knowledge of visceral afferent nerves is largely attributable to difficulties associated with their study; only one nerve fiber can typically be studied at a time by classical electrophysiology.

Purpose: Here, we report the adaptation of a cutting-edge high-throughput method to study gastrointestinal afferent nerves that allows the activity in hundreds of neurons to be imaged simultaneously. This method is based on genetic expression of the fluorescent calcium indicator GCaMP6 and multiphoton microscopy in transgenic mice. In addition to providing fundamental information about visceral afferent nerves, future iterations of this method are expected to enable studies of nerve activity in biopsies from patients with gastrointestinal diseases.

Submitted: 11. 6. 2019

Accepted: 27. 7. 2019

Conflict of Interest: The authors declare that the article/manuscript complies with ethical standards, patient anonymity has been respected, and states that they have no financial, advisory or other commercial interests in relation to the subject matter.

Publication Ethics: The article/manuscript has not been published or is currently being submitted to another review.

The authors agree to publish their name and e-mail in the published article/manuscript.

Dedication: The article/manuscript is not supported by a grant nor has it been created with the support of any company.

The Editorial Board declares that the manuscript met the ICMJE „uniform requirements“ for biomedical papers.

Keywords:

Pain – innervation – Gastrointestinal tract – Microscopy

Sources

1. Ray K. Neurogastroenterology: a tale of two neurons – distinct functions of vagal afferents of the gut. Nat Rev Gastroenterol Hepatol 2016; 13 (8): 435. doi: 10.1038/nrgastro.2016.100.

2. Bonaz B, Sinniger V, Pellissier S. Vagal tone: effects on sensitivity, motility, and inflammation. Neurogastroenterol Motil 2016; 28 (4): 455–462. doi: 10.1111/nmo.12817.

3. Bonaz B, Bazin T, Pellissier S. The vagus nerve at the interface of the microbiota-gut-brain axis. Front Neurosci 2018; 12: 49. doi: 10.3389/fnins.2018.00049.

4. Vanner S, Greenwood-Van Meerveld B, Mawe G et al. Fundamentals of neurogastroenterology: basic science. Gastroenterology 2016; 150 (6): 1280–1291. doi: 10.1053/j.gastro.2016.02.018.

5. Brookes SJ, Spencer NJ, Costa M et al. Extrinsic primary afferent signalling in the gut. Nat Rev Gastroenterol Hepatol 2013; 10 (5): 286–296. doi: 10.1038/nrgastro.2013.29.

6. Rindi G, Leiter AB, Kopin AS et al. The „normal” endocrine cell of the gut: changing concepts and new evidences. Ann N Y Acad Sci 2004; 1014: 1–12. doi: 10.1196/annals.1294.001.

7. Raybould HE. Gut chemosensing: interactions between gut endocrine cells and visceral afferents. Auton Neurosci 2010; 153 (1–2): 41–46. doi: 10.1016/j.autneu.2009.07.007.

8. Berthoud HR, Blackshaw LA, Brookes SJ. Neuroanatomy of extrinsic afferents supplying the gastrointestinal tract. Neurogastroenterol Motil 2004; 16 (Suppl 1): S28–S33. doi: 10.1111/j.1743-3150.2004.00471.x.

9. Brierley SM, Hibberd TJ, Spencer NJ. Spinal afferent innervation of the colon and rectum. Front Cell Neurosci 2018; 12: 467. doi: 10.3389/fncel.2018.00467.

10. Yu S, Undem BJ, Kollarik M. Vagal afferent nerves with nociceptive properties in guinea-pig oesophagus. J Physiol 2005; 563: 831–842. doi: 10.1113/jphysiol.2004.079574.

11. Emery EC, Luiz AP, Sikandar S et al. In vivo characterization of distinct modality-specific subsets of somatosensory neurons using GCaMP. Sci Adv 2016; 2 (11): e1600990–e1600990. doi: 10.1126/sciadv.1600990.

12. Denk W, Strickler JH, Webb WW. Two-photon laser scanning fluorescence microscopy. Science 1990; 248 (4951): 73–76. doi: 10.1126/science.2321027.

13. Anderson M, Zheng Q, Dong X. Investigation of pain mechanisms by calcium imaging approaches. Neuroscience Bull 2018; 34 (1): 194–199. doi: 10.1007/s12264-017-01 39-9.

14. Benninger RK, Piston DW. Two-photon excitation microscopy for the study of living cells and tissues. Curr Protoc Cell Biol 2013; 4 (4): 1–24. doi: 10.1002/0471143030.cb0411s59.

15. Helmchen F, Denk W. Deep tissue two-photon microscopy. Nat Methods 2005; 2 (12): 932–940. doi: 10.1038/nmeth818.

16. Mittmann W, Wallace DJ, Czubayko U et al. Two-photon calcium imaging of evoked activity from L5 somatosensory neurons in vivo. Nat Neurosci 2011; 14 (8): 1089–1093. doi: 10.1038/nn.2879.

17. Huber D, Gutnisky DA, Peron S et al. Multiple dynamic representations in the motor cortex during sensorimotor learning. Nature 2012; 484 (7395): 473–478. doi: 10.1038/nature11 039.

18. Miller RE, Kim YS, Tran PB et al. Visualization of peripheral neuron sensitization in a surgical mouse model of osteoarthritis by in vivo calcium imaging. Arthritis Rheumatol 2018; 70 (1): 88–97. doi: 10.1002/art.40342.

19. Tian L, Hires SA, Looger LL. Imaging neuronal activity with genetically encoded calcium indicators. Cold Spring Harb Protoc 2012; 6: 647–656.

20. Kollarik M, Carr MJ, Ru F et al. Transgene expression and effective gene silencing in vagal afferent neurons in vivo using recombinant adeno-associated virus vectors. J Physiol 2010; 588: 4303–4315. doi: 10.1113/jphysiol. 2010.192971.

21. Kim YS, Anderson M, Park K et al. Coupled activation of primary sensory neurons contributes to chronic pain. Neuron 2016; 91 (5): 1085–1096. doi: 10.1016/j.neuron.2016.07.044.

22. Hoess RH, Abremski K. The Cre-lox recombination system. Nucleic Acids Mol Biol 1990; 4: 99–109. doi: 10.1007/978-3-642-84150-7_6.

23. Kim YS, Chu Y, Han L et al. Central terminal sensitization of TRPV1 by descending serotonergic facilitation modulates chronic pain. Neuron 2014; 81 (4): 873–887. doi: 10.1016/j.neuron.2013.12.011.

24. Kim AY, Tang Z, Liu Q et al. Pirt, a phospho-inositide-binding protein, functions as a regulatory subunit of TRPV1. Cell 2008; 133 (3): 475–485. doi: 10.1016/j.cell.2008.02.053.

25. Chen TW, Wardill TJ, Sun Y et al. Ultrasensitive fluorescent proteins for imaging neuronal activity. Nature 2013; 499 (7458): 295–300. doi: 10.1038/nature12354.