Cell and Tissue Research

, Volume 348, Issue 3, pp 397–405 | Cite as

Identification of neurons that express ghrelin receptors in autonomic pathways originating from the spinal cord

  • John B. Furness
  • Hyun-Jung Cho
  • Billie Hunne
  • Haruko Hirayama
  • Brid P. Callaghan
  • Alan E. Lomax
  • James A. Brock
Regular Article

Abstract

Functional studies have shown that subsets of autonomic preganglionic neurons respond to ghrelin and ghrelin mimetics and in situ hybridisation has revealed receptor gene expression in the cell bodies of some preganglionic neurons. Our present goal has been to determine which preganglionic neurons express ghrelin receptors by using mice expressing enhanced green fluorescent protein (EGFP) under the control of the promoter for the ghrelin receptor (also called growth hormone secretagogue receptor). The retrograde tracer Fast Blue was injected into target organs of reporter mice under anaesthesia to identify specific functional subsets of postganglionic sympathetic neurons. Cryo-sections were immunohistochemically stained by using anti-EGFP and antibodies to neuronal markers. EGFP was detected in nerve terminal varicosities in all sympathetic chain, prevertebral and pelvic ganglia and in the adrenal medulla. Non-varicose fibres associated with the ganglia were also immunoreactive. No postganglionic cell bodies contained EGFP. In sympathetic chain ganglia, most neurons were surrounded by EGFP-positive terminals. In the stellate ganglion, neurons with choline acetyltransferase immunoreactivity, some being sudomotor neurons, lacked surrounding ghrelin-receptor-expressing terminals, although these terminals were found around other neurons. In the superior cervical ganglion, the ghrelin receptor terminals innervated subgroups of neurons including neuropeptide Y (NPY)-immunoreactive neurons that projected to the anterior chamber of the eye. However, large NPY-negative neurons projecting to the acini of the submaxillary gland were not innervated by EGFP-positive varicosities. In the celiaco-superior mesenteric ganglion, almost all neurons were surrounded by positive terminals but the VIP-immunoreactive terminals of intestinofugal neurons were EGFP-negative. The pelvic ganglia contained groups of neurons without ghrelin receptor terminal innervation and other groups with positive terminals around them. Ghrelin receptors are therefore expressed by subgroups of preganglionic neurons, including those of vasoconstrictor pathways and of pathways controlling gut function, but are absent from some other neurons, including those innervating sweat glands and the secretomotor neurons that supply the submaxillary salivary glands.

Keywords

Autonomic ganglia Ghrelin Immunohistochemistry Preganglionic neurons Retrograde tracing Mouse 

Notes

Acknowledgements

We thank Dr. Colin Anderson and Mr. David Gonsalves for their advice and assistance with the Fast Blue injections and for their comments on the manuscript and Dr. Trung Nguyen for advice on the statistics.

References

  1. Andrews ZB (2011) The extra-hypothalamic actions of ghrelin on neuronal function. Trends Neurosci 34:31–40PubMedCrossRefGoogle Scholar
  2. Asmus SE, Parsons S, Landis SC (2000) Developmental changes in the transmitter properties of sympathetic neurons that innervate the periosteum. J Neurosci 20:1495–1504PubMedGoogle Scholar
  3. Ferens DM, Yin L, Bron R, Hunne B, Ohashi-Doi K, Sanger GJ, Witherington J, Shimizu Y, Furness JB (2010a) Functional and in situ hybridisation evidence that preganglionic sympathetic vasoconstrictor neurons express ghrelin receptors. Neuroscience 166:671–679PubMedCrossRefGoogle Scholar
  4. Ferens DM, Yin L, Ohashi-Doi K, Habgood M, Bron R, Brock JA, Gale JD, Furness JB (2010b) Evidence for functional ghrelin receptors on parasympathetic preganglionic neurons of micturition control pathways in the rat. Clin Exp Pharmacol Physiol 37:926–932PubMedCrossRefGoogle Scholar
  5. Fujimiya M, Asakawa A, Ataka K, Chen C-Y, Kato I, Inui A (2010) Ghrelin, des-acyl ghrelin, and obestatin: regulatory roles on the gastrointestinal motility. Int J Pept 2010:1–8CrossRefGoogle Scholar
  6. Furness JB, Costa M (1979) Projections of intestinal neurons showing immunoreactivity for vasoactive intestinal polypeptide are consistent with these neurons being the enteric inhibitory neurons. Neurosci Lett 15:199–204PubMedCrossRefGoogle Scholar
  7. Furness JB, Hunne B, Matsuda N, Yin L, Russo D, Kato I, Fujimiya M, Patterson M, McLeod J, Andrews ZB, Bron R (2011) Investigation of the presence of ghrelin in the central nervous system of the rat and mouse. Neuroscience 193:1–9PubMedCrossRefGoogle Scholar
  8. Gibbins IL (1991) Vasomotor, pilomotor and secretomotor neurons distinguished by size and neuropeptide content in superior cervical ganglia of mice. J Auton Nerv Syst 34:171–184PubMedCrossRefGoogle Scholar
  9. Grkovic I, Edwards SL, Murphy SM, Anderson CR (1999) Chemically distinct preganglionic inputs to iris-projecting postganglionic neurons in the rat: a light and electron microscopic study. J Comp Neurol 412:606–616PubMedCrossRefGoogle Scholar
  10. Hirayama H, Shiina T, Shima T, Kuramoto H, Takewaki T, Furness JB, Shimizu Y (2010) Contrasting effects of ghrelin and des-acyl ghrelin on the lumbo-sacral defecation center and regulation of colorectal motility in rats. Neurogastroenterol Motil 22:1124–1131PubMedCrossRefGoogle Scholar
  11. Howard AD, Feighner SD, Cully DF, Arena JP, Liberator PA, Rosenblum CI, Hamelin M, Hreniuk DL, Palyha OC, Anderson J, Paress PS, Diaz C, Chou M, Liu KK, McKee KK, Pong SS, Chaung LY, Elbrecht A, Dashkevicz M, Heavens R, Rigby M, Sirinathsinghji DJ, Dean DC, Melillo DG, Patchett AA, Nargund R, Griffin PR, DeMartino JA, Gupta SK, Schaeffer JM, Smith RG, Van der Ploeg LH (1996) A receptor in pituitary and hypothalamus that functions in growth hormone release. Science 273:974–977PubMedCrossRefGoogle Scholar
  12. Keast JR (1999) Unusual autonomic ganglia: connections, chemistry, and plasticity of pelvic ganglia. Int Rev Cytol 193:1–69PubMedCrossRefGoogle Scholar
  13. Kojima M, Hosoda H, Date Y, Nakazato M, Matsuo H, Kangawa K (1999) Ghrelin is a growth-hormone-releasing acylated peptide from stomach. Nature 402:656–660PubMedCrossRefGoogle Scholar
  14. Langley JN, Anderson HK (1895) The innervation of the pelvic and adjoining viscera. IV. The internal generative organs. J Physiol (Lond) 19:122–130Google Scholar
  15. Lindh B, Hökfelt T, Elfvin LG (1988) Distribution and origin of peptide-containing nerve fibers in the celiac superior mesenteric ganglion of the guinea-pig. Neuroscience 26:1037–1071PubMedCrossRefGoogle Scholar
  16. Lomax AE, Sharkey KA, Furness JB (2010) The participation of the sympathetic innervation of the gastrointestinal tract in disease states. Neurogastroenterol Motil 22:7–18PubMedGoogle Scholar
  17. Lundberg JM, Hökfelt T, Anggard A, Terenius L, Elde R, Markey K, Goldstein M, Kimmel J (1982) Organization principles in the peripheral sympathetic nervous system: subdivision by coexisting peptides somatostatin-, avian pancreatic polypeptide-, and vasoactive intestinal polypeptide-like immunoreactive materials. Proc Natl Acad Sci USA 79:1303–1307PubMedCrossRefGoogle Scholar
  18. Maccarrone C, Jarrott B (1985) Differences in regional brain concentrations of neuropeptide Y in spontaneously hypertensive SH and Wistar Kyoto WKY rats. Brain Res 345:165–169PubMedCrossRefGoogle Scholar
  19. Macrae LM, Furness JB, Costa M (1986) Distribution of subgroups of noradrenaline neurons in the coeliac ganglion of the guinea-pig. Cell Tissue Res 244:173–180PubMedCrossRefGoogle Scholar
  20. Rocha-Sousa A, Saraiva J, Henriques-Coelho T, Falcão-Reis F, Correia-Pinto J, Leite-Moreira AF (2006) Ghrelin as a novel locally produced relaxing peptide of the iris sphincter and dilator muscles. Exp Eye Res 83:1179–1187PubMedCrossRefGoogle Scholar
  21. Schäfer MKH, Schutz B, Weihe E, Eiden LE (1997) Target-independent cholinergic differentiation in the rat sympathetic nervous system. Proc Natl Acad Sci USA 94:4149–4154PubMedCrossRefGoogle Scholar
  22. Schäfer MKH, Eiden LE, Weihe E (1998) Cholinergic neurons and terminal fields revealed by immunohistochemistry for the vesicular acetylcholine transporter. II. The peripheral nervous system. Neuroscience 84:361–376PubMedCrossRefGoogle Scholar
  23. Shimizu Y, Chang EC, Shafton AD, Ferens DM, Sanger GJ, Witherington J, Furness JB (2006) Evidence that stimulation of ghrelin receptors in the spinal cord initiates propulsive activity in the colon of the rat. J Physiol (Lond) 576:329–338CrossRefGoogle Scholar
  24. Szurszewski JH, Ermilov LG, Miller SM (2002) Prevertebral ganglia and intestinofugal afferent neurones. Gut 51:i6–i10PubMedCrossRefGoogle Scholar
  25. Tack J, Depoortere I, Bisschops R, Delporte C, Coulie B, Meulemans A, Janssens J, Peeters T (2006) Influence of ghrelin on interdigestive gastrointestinal motility in humans. Gut 55:327–333PubMedCrossRefGoogle Scholar
  26. Venables G, Hunne B, Bron R, Cho H-J, Brock JA, Furness JB (2011) Ghrelin receptors are expressed by distal tubules of the mouse kidney. Cell Tissue Res 346:135–139PubMedCrossRefGoogle Scholar
  27. Vidovic M, Hill CE, Hendry IA (1987) Developmental time course of the sympathetic postganglionic innervation of the rat eye. Dev Brain Res 32:133–138CrossRefGoogle Scholar
  28. Zhao T-J, Sakata I, Li RL, Liang G, Richardson JA, Brown MS, Goldstein JL, Zigman JM (2010) Ghrelin secretion stimulated by β1-adrenergic receptors in cultured ghrelinoma cells and in fasted mice. Proc Natl Acad Sci USA 107:15868–15873PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2012

Authors and Affiliations

  • John B. Furness
    • 1
  • Hyun-Jung Cho
    • 1
  • Billie Hunne
    • 1
  • Haruko Hirayama
    • 1
  • Brid P. Callaghan
    • 1
  • Alan E. Lomax
    • 2
  • James A. Brock
    • 1
  1. 1.Department of Anatomy & NeuroscienceUniversity of MelbourneParkvilleAustralia
  2. 2.Gastrointestinal Diseases Research UnitQueen’s UniversityKingstonCanada

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