Skip to main content
SpringerLink
Log in

Distribution and morphology of cholinergic cells in the branchial epithelium of zebrafish (Danio rerio)

  • Regular Article
  • Published:
Cell and Tissue Research Aims and scope Submit manuscript

Abstract

Acetylcholine is an excitatory neurotransmitter important for oxygen sensing in mammals. A cholinergic mechanism in the fish gill has been implicated in the hyperventilatory response to acute hypoxia; however, the identity and distribution of acetylcholine-containing cells in the gills is poorly defined. We test the hypothesis that cholinergic cells are present in the gill filament epithelium in zebrafish (Danio rerio), a model vertebrate for which oxygen chemoreceptors are well characterized, and that these cells would receive nervous innervation. Using immunohistochemistry and confocal microscopy, we observed 10.2 ± 0.6 cells immunoreactive for the vesicular acetylcholine transporter (VAChT) on the efferent aspect of each gill filament, where a high density of serotonergic oxygen-sensitive neuroepithelial cells (NECs) were located. VAChT-positive cells of the efferent epithelium were positioned within 10 μm of NECs. On the afferent aspect of the gill filaments, VAChT-positive cells were greater in number (30.8 ± 3.1 per filament). On the efferent and afferent filament aspects, VAChT-positive cells did not contain serotonin, but did express choline acetyltransferase (ChAT), the enzyme that synthesizes ACh, and were often closely apposed to nerve fibers labeled with the neuronal marker, zn-12. We conclude that cholinergic cells in the zebrafish gills were present in the primary epithelium of gill filaments, and formed contacts with nerve fibers. These studies provide morphological evidence for the presence of a cholinergic system in the zebrafish gill. Such a pathway may contribute to the reflex hyperventilatory response during hypoxia.

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
Fig. 8

Similar content being viewed by others

References

  • Abdallah SJ, Jonz MG, Perry SF (2015) Extracellular H+ induces Ca2+ signals in respiratory chemoreceptors of zebrafish. Pflugers Arch 467:399–413

    Article  CAS  PubMed  Google Scholar 

  • Bailly Y, Dunel-Erb S (1986) The sphincter of the efferent filament artery in teleost gills: I. structure, and parasympathetic innervation. J Morphol 187:219–237

    Article  Google Scholar 

  • Bailly Y, Dunel-Erb S, Laurent P (1992) The neuroepithelial cells of the fish gill filament: indolamine-immunocytochemistry and innervation. Anat Rec 233:143–161

    Article  CAS  PubMed  Google Scholar 

  • Booth JH (1978) The distribution of blood flow in the gills of fish: application of a new technique to rainbow trout (Salmo gairdneri). J Exp Biol 73:119–129

    Google Scholar 

  • Booth JH (1979) The effects of oxygen supply, epinephrine, and acetylcholine on the distribution of blood flow in trout gills. J Exp Biol 83:31–39

    CAS  Google Scholar 

  • Buckler KJ (2007) TASK-like potassium channels and oxygen sensing in the carotid body. Respir Physiol Neurobiol 157:55–64

    Article  CAS  PubMed  Google Scholar 

  • Burleson ML, Milsom WK (1995a) Cardio-ventilatory control in rainbow trout: I. Pharmacology of branchial, oxygen-sensitive chemoreceptors. Respir Physiol 100:231–238

    Article  CAS  PubMed  Google Scholar 

  • Burleson ML, Milsom WK (1995b) Cardio-ventilatory control in rainbow trout: II. Reflex effects of exogenous neurochemicals. Respir Physiol 101:289–299

    Article  CAS  PubMed  Google Scholar 

  • Clemente D, Porteros A, Weruaga E, Alonso JR, Arenzana FJ, Aijón J, Arévalo R (2004) Cholinergic elements in the zebrafish central nervous system: histochemical and immunohistochemical analysis. J Comp Neurol 474:75–107

    Article  PubMed  Google Scholar 

  • Coolidge EH, Ciuhandu CS, Milsom WK (2008) A comparative analysis of putative oxygen-sensing cells in the fish gill. J Exp Biol 211:1231–1242

    Article  PubMed  Google Scholar 

  • Cutz E, Pan J, Yeger H, Domnik NJ, Fisher JT (2013) Recent advances and contraversies on the role of pulmonary neuroepithelial bodies as airway sensors. Semin Cell Dev Biol 24:40–50

    Article  PubMed  Google Scholar 

  • Dasso LL, Buckler KJ, Baughan-Jones RD (1997) Muscarinic and nicotinic receptors raise intracellular Ca2+ levels in rat carotid body type I cells. J Physiol 498:327–338

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Dunel-Erb S, Bailly Y, Laurent P (1982) Neuroepithelial cells in fish gill primary lamellae. J Appl Physiol Respir Environ Exerc Physiol 53:1342–1353

    CAS  PubMed  Google Scholar 

  • Fu XW, Nurse CA, Wong V, Cutz E (2002) Hypoxia-induced secretion of serotonin from intact pulmonary neuroepithelial bodies in neonatal rabbit. J Physiol 539:503–510

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Houser A, McNair C, Piccinini R, Luxhoj A, Bell WE, Turner JE (2011) Effects of estrogen on the neuromuscular system in the embryonic zebrafish (Danio rerio). Brain Res 1381:106–116

    Article  CAS  PubMed  Google Scholar 

  • Iturriaga R, Alcayaga J (2004) Neurotransmission in the carotid body: transmitters and modulators between glomus cells and petrosal ganglion nerve terminals. Brain Res Brain Res Rev 47:46–53

    Article  CAS  PubMed  Google Scholar 

  • Jonz MG (2014) Oxygen sensing. In: Evans DH, Claiborne JB, Currie S (eds) The physiology of fishes, 4th edn. CRC, Boca Raton, pp 149–175

    Google Scholar 

  • Jonz MG, Nurse CA (2003) Neuroepithelial cells and associated innervation of the zebrafish gill: a confocal immunofluorescence study. J Comp Neurol 461:1–17

    Article  PubMed  Google Scholar 

  • Jonz MG, Nurse CA (2005) Development of oxygen sensing in the gills of zebrafish. J Exp Biol 208:1537–1549

    Article  PubMed  Google Scholar 

  • Jonz MG, Zaccone G (2009) Nervous control of the gills. Acta Histochem 111:207–216

    Article  PubMed  Google Scholar 

  • Jonz MG, Fearon IM, Nurse CA (2004) Neuroepithelial oxygen chemoreceptors of the zebrafish gill. J Physiol 560:737–752

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Jonz MG, Zachar PC, Da Fonte DF, Mierzwa AS (2015) Peripheral chemoreceptors in fish: a brief history and a look ahead. Comp Biochem Physiol A 186:27–38

    Article  CAS  Google Scholar 

  • Kumar P, Prabhakar NR (2012) Peripheral chemoreceptors: function and plasticity of the carotid body. Compr Physiol 2:141–219

    PubMed  PubMed Central  Google Scholar 

  • Kusakabe T (2002) Carotid labyrinth of amphibians. Microsc Res Tech 59:207–226

    Article  PubMed  Google Scholar 

  • López-Barneo J, López-López JR, Ureña J, González C (1988) Chemotransduction in the carotid body: K+ current modulated by PO2 in type I chemoreceptor cells. Science 241:580–582

    Article  PubMed  Google Scholar 

  • Mauceri A, Fasulo S, Minniti F, Lo Cascio P, Maisano M, Zaccone G (2005) Neurochemical features of the innervation of respiratory organs in some air-breathing fishes. Ital J Zool 72:175–181

    Article  CAS  Google Scholar 

  • Milsom WK, Burleson ML (2007) Peripheral arterial chemoreceptors and the evolution of the carotid body. Respir Physiol Neurobiol 157:4–11

    Article  CAS  PubMed  Google Scholar 

  • Nilsson S (1984) Innervation and pharmacology of the gills. In: Hoar WS, Randall DJ (eds) Fish physiology, vol. XA. Academic, San Diego, pp 185–227

    Google Scholar 

  • Nurse CA (2010) Neurotransmitter and neuromodulatory mechanisms at peripheral arterial chemoreceptors. Exp Physiol 95:657–667

    Article  CAS  PubMed  Google Scholar 

  • Olsson C (2011) Calbindin immunoreactivity in the enteric nervous system of larval and adult zebrafish (Danio rerio). Cell Tissue Res 3441:31–40

    Article  Google Scholar 

  • Paulsen UJ, Tiedemann K, Mack AF (2010) Dendritic differentiation in the periphery of the growing zebrafish retina. Exp Eye Res 90:514–520

    Article  CAS  PubMed  Google Scholar 

  • Peers C, Wyatt CN, Evans AM (2010) Mechanisms for acute oxygen sensing in the carotid body. Respir Physiol Neurobiol 174:292–298

    Article  CAS  PubMed  Google Scholar 

  • Perry SF, Jonz MG, Gilmour KM (2009) Oxygen sensing and the hypoxic ventilatory response. In: Richards J, Farrell A, Brauner C (eds) Hypoxia. Academic, Waltham, pp 193–253

    Chapter  Google Scholar 

  • Porteus CS, Brink DL, Milsom WK (2012) Neurotransmitter profiles in fish gills: putative gill oxygen chemoreceptors. Respir Physiol Neurobiol 184:316–325

    Article  CAS  PubMed  Google Scholar 

  • Porteus CS, Brink DL, Coolidge EH, Fong AY, Milsom WK (2013) Distribution of acetylcholine and catecholamines in fish gills and their potential roles in the hypoxic ventilatory response. Acta Histochem 115:158–169

    Article  CAS  PubMed  Google Scholar 

  • Prado MA, Reis RA, Prado VF, de Mello MC, Gomez MV, de Mello FG (2002) Regulation of acetylcholine synthesis and storage. Neurochem Int 41:291–299

    Article  CAS  PubMed  Google Scholar 

  • Qin Z, Lewis JE, Perry SF (2010) Zebrafish (Danio rerio) gill neuroepithelial cells are sensitive chemoreceptors for environmental CO2. J Physiol 588:861–872

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Regan KS, Jonz MG, Wright PA (2011) Neuroepithelial cells and the hypoxia emersion response in the amphibious fish Kryptolebias marmoratus. J Exp Biol 214:2560–2568

    Article  PubMed  Google Scholar 

  • Roghani A, Feldman J, Kohan SA, Shirzadi A, Gundersen CB, Brecha N, Edwards RH (1994) Molecular cloning of a putative vesicular transporter for acetylcholine. Proc Natl Acad Sci U S A 91:10620–10624

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Saltys HA, Jonz MG, Nurse CA (2006) Comparative study of gill neuroepithelial cells and their innervation in teleosts and Xenopus tadpoles. Cell Tissue Res 323:1–10

    Article  PubMed  Google Scholar 

  • Shakarchi K, Zachar PC, Jonz MG (2013) Serotonergic and cholinergic elements elements of the hypoxic ventilatory response in devloping zebrafish. J Exp Biol 216:869–880

    Article  CAS  PubMed  Google Scholar 

  • Shirahata M, Balbir A, Otsubo T, Fitzgerald RS (2007) Role of acetylcholine in neurotransmission of the carotid body. Respir Physiol Neurobiol 157:93–105

    Article  CAS  PubMed  Google Scholar 

  • Stoyek MR, Croll RP, Smith FM (2015) Intrinsic and extrinsic innervation of the heart in zebrafish (Danio rerio). J Comp Neurol 523:1683–1700

    Article  CAS  PubMed  Google Scholar 

  • Sundin L, Nilsson S (1997) Neurochemical mechanisms behind gill microcirculatory responses to hypoxia in trout: in vivo microscopy study. Am J Physiol 41:R576–R585

    Google Scholar 

  • Sundin L, Nilsson S (2002) Branchial innervation. J Exp Zool 293:232–248

    Article  PubMed  Google Scholar 

  • Trevarrow B, Marks DL, Kimmel CB (1990) Organization of hindbrain segments in the zebrafish embryo. Neuron 4:669–679

    Article  CAS  PubMed  Google Scholar 

  • Wyatt CN, Peers C (1993) Nicotinic acetylcholine receptors in isolated type I cells of the neonatal rat carotid body. Neuroscience 54:275–281

    Article  CAS  PubMed  Google Scholar 

  • Zaccone G, Ainis L, Mauceri A, Lo Cascio P, Lo Giudice F, Fasulo S (2003) NANC nerves in the respiratory air sac and branchial vasculature of the Indian catfish, Heteropneustes fossilis. Acta Histochem 105:151–163

    Article  PubMed  Google Scholar 

  • Zachar PC, Jonz MG (2012) Confocal imaging of Merkel-like basal cells in the taste buds of zebrafish. Acta Histochem 114:101–115

    Article  PubMed  Google Scholar 

  • Zhang L, Nurse CA, Jonz MG, Wood CM (2011) Ammonia sensing by neuroepithelial cells and ventilatory responses to ammonia in rainbow trout. J Exp Biol 214:2678–2689

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

This research was supported by the Natural Sciences and Engineering Research Council of Canada (grant number 342303), the Canadian Foundation for Innovation, and the Ontario Research Fund (grant number 16589).

Author information

Authors and Affiliations

  1. Department of Biology, University of Ottawa, 30 Marie Curie Pvt., Ottawa, ON, Canada, K1N 6N5

    Peter C. Zachar, Wen Pan & Michael G. Jonz

Authors
  1. Peter C. Zachar
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Wen Pan
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Michael G. Jonz
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Michael G. Jonz.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Zachar, P.C., Pan, W. & Jonz, M.G. Distribution and morphology of cholinergic cells in the branchial epithelium of zebrafish (Danio rerio). Cell Tissue Res 367, 169–179 (2017). https://doi.org/10.1007/s00441-016-2531-6

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00441-016-2531-6

Keywords

Search

Navigation