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Journal of anatomy2022; 241(5); 1211-1218; doi: 10.1111/joa.13719

Cultured dissociated primary dorsal root ganglion neurons from adult horses enable study of axonal transport.

Abstract: Neurological disorders are prevalent in horses, but their study is challenging due to anatomic constraints and the large body size; very few host-specific in vitro models have been established to study these types of diseases, particularly from adult donor tissue. Here we report the generation of primary neuronal dorsal root ganglia (DRG) cultures from adult horses: the mixed, dissociated cultures, containing neurons and glial cells, remained viable for at least 90 days. Similar to DRG neurons in vivo, cultured neurons varied in size, and they developed long neurites. The mitochondrial movement was detected in cultured cells and was significantly slower in glial cells compared to DRG-derived neurons. In addition, mitochondria were more elongated in glial cells than those in neurons. Our culture model will be a useful tool to study the contribution of axonal transport defects to specific neurodegenerative diseases in horses as well as comparative studies aimed at evaluating species-specific differences in axonal transport and survival.
Publication Date: 2022-06-21 PubMed ID: 35728923PubMed Central: PMC9558156DOI: 10.1111/joa.13719Google Scholar: Lookup
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  • Journal Article
  • Research Support
  • Non-U.S. Gov't

Summary

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The study focused on creating a laboratory model using neurons from the dorsal root ganglia (DRG) of adult horses to investigate neurodegenerative diseases, specifically focusing on axonal transport mechanisms.

Objective of the Study

The research aimed to create a viable in-vitro model using mixed, dissociated primary dorsal root ganglia (DRG) cultures from adult horses. It sought to observe neuron and glial cell development and compare mitochondrial movement and structure between the two.

This model is intended for studying the impact of axonal transport defects in horse-related neurodegenerative diseases. It can also be used in comparative studies to understand species-specific differences in axonal transport and survival.

Methods Employed

  • The researchers generated dissociated primary DRG cultures from adult horses. These cultures included neurons and glial cells.
  • The viability of such cells was maintained over a period of 90 days or more.
  • They observed neuron development, particularly the growth of long neurites, akin to those seen in vivo.
  • The study also monitored and compared the movement of mitochondria between neurons and glial cells. Taken together, these methodologies produced in-depth insights into the behavior of cells in culture.

Key Findings

  • In cultures, just as in the in vivo setting, DRG neurons showed variations in size.
  • Mitochondrial movement was noted in cultured cells. Interestingly, glial cells featured significantly slower mitochondrial motion than DRG-derived neurons.
  • Moreover, mitochondria in glial cells were also more elongated than those in neurons.

Future Implications

The findings from this research herald new avenues for similar future studies. The novel culture model established here is expected to greatly contribute to the research field of horse-specific neurodegenerative diseases, elucidating the role of axonal transport abnormalities in these conditions.

Furthermore, it opens the way for comparative studies to examine and understand species-related differences in axonal transport and cell survival. This will provide greater depth to our knowledge of disease processes and potential therapies in horses specifically, and potentially other species as well.

Cite This Article

APA
Adalbert R, Cahalan S, Hopkins EL, Almuhanna A, Loreto A, Pór E, Körmöczy L, Perkins J, Coleman MP, Piercy RJ. (2022). Cultured dissociated primary dorsal root ganglion neurons from adult horses enable study of axonal transport. J Anat, 241(5), 1211-1218. https://doi.org/10.1111/joa.13719

Publication

ISSN: 1469-7580
NlmUniqueID: 0137162
Country: England
Language: English
Volume: 241
Issue: 5
Pages: 1211-1218

Researcher Affiliations

Adalbert, Robert
  • Comparative Neuromuscular Diseases Laboratory, Department of Clinical Science and Services, Royal Veterinary College, London, UK.
  • Department of Anatomy, Histology, and Embryology, Faculty of Medicine, University of Szeged, Szeged, Hungary.
Cahalan, Stephen
  • Comparative Neuromuscular Diseases Laboratory, Department of Clinical Science and Services, Royal Veterinary College, London, UK.
Hopkins, Eleanor L
  • John van Geest Centre for Brain Repair, Department of Clinical Neurosciences, University of Cambridge, Cambridge, UK.
Almuhanna, Abdulaziz
  • Comparative Neuromuscular Diseases Laboratory, Department of Clinical Science and Services, Royal Veterinary College, London, UK.
Loreto, Andrea
  • John van Geest Centre for Brain Repair, Department of Clinical Neurosciences, University of Cambridge, Cambridge, UK.
  • Andrea Loreto, Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford, UK.
Pór, Erzsébet
  • Department of Anatomy, Histology, and Embryology, Faculty of Medicine, University of Szeged, Szeged, Hungary.
Körmöczy, Laura
  • Department of Anatomy, Histology, and Embryology, Faculty of Medicine, University of Szeged, Szeged, Hungary.
Perkins, Justin
  • Comparative Neuromuscular Diseases Laboratory, Department of Clinical Science and Services, Royal Veterinary College, London, UK.
Coleman, Michael P
  • John van Geest Centre for Brain Repair, Department of Clinical Neurosciences, University of Cambridge, Cambridge, UK.
Piercy, Richard J
  • Comparative Neuromuscular Diseases Laboratory, Department of Clinical Science and Services, Royal Veterinary College, London, UK.

MeSH Terms

  • Animals
  • Axonal Transport
  • Cells, Cultured
  • Ganglia, Spinal
  • Horses
  • Neurites / physiology
  • Neurons

Grant Funding

  • UK Horserace Betting Levy Board

References

This article includes 51 references
  1. Adalbert R, Kaieda A, Antoniou C, Loreto A, Yang X, Gilley J, Hoshino T, Uga K, Makhija MT, Coleman MP. Novel HDAC6 Inhibitors Increase Tubulin Acetylation and Rescue Axonal Transport of Mitochondria in a Model of Charcot-Marie-Tooth Type 2F.. ACS Chem Neurosci 2020 Feb 5;11(3):258-267.
    pubmed: 31845794doi: 10.1021/acschemneuro.9b00338google scholar: lookup
  2. Adalbert R, Milde S, Durrant C, Ando K, Stygelbout V, Yilmaz Z, Gould S, Brion JP, Coleman MP. Interaction between a MAPT variant causing frontotemporal dementia and mutant APP affects axonal transport.. Neurobiol Aging 2018 Aug;68:68-75.
  3. Aleman M, Williams DC, Brosnan RJ, Nieto JE, Pickles KJ, Berger J, Lecouteur RA, Holliday TA, Madigan JE. Sensory nerve conduction and somatosensory evoked potentials of the trigeminal nerve in horses with idiopathic headshaking.. J Vet Intern Med 2013 Nov-Dec;27(6):1571-80.
    pubmed: 24107198doi: 10.1111/jvim.12191google scholar: lookup
  4. Andrews S, Gilley J, Coleman MP. Difference Tracker: ImageJ plugins for fully automated analysis of multiple axonal transport parameters.. J Neurosci Methods 2010 Nov 30;193(2):281-7.
  5. Baccaglini PI, Hogan PG. Some rat sensory neurons in culture express characteristics of differentiated pain sensory cells.. Proc Natl Acad Sci U S A 1983 Jan;80(2):594-8.
    pmc: PMC393426pubmed: 6188155doi: 10.1073/pnas.80.2.594google scholar: lookup
  6. Bayat FK, Polat Budak B, Yiğit EN, Öztürk G, Gülçür HÖ, Güveniş A. Adult mouse dorsal root ganglia neurons form aberrant glutamatergic connections in dissociated cultures.. PLoS One 2021;16(3):e0246924.
  7. Bertoni-Freddari C, Fattoretti P, Paoloni R, Caselli U, Giorgetti B, Solazzi M. Inverse correlation between mitochondrial size and metabolic competence: a quantitative cytochemical study of cytochrome oxidase activity.. Naturwissenschaften 2003 Feb;90(2):68-71.
    pubmed: 12590300doi: 10.1007/s00114-002-0398-8google scholar: lookup
  8. Bilsland LG, Sahai E, Kelly G, Golding M, Greensmith L, Schiavo G. Deficits in axonal transport precede ALS symptoms in vivo.. Proc Natl Acad Sci U S A 2010 Nov 23;107(47):20523-8.
    pmc: PMC2996651pubmed: 21059924doi: 10.1073/pnas.1006869107google scholar: lookup
  9. Cottrell DF, McGorum BC, Pearson GT. The neurology and enterology of equine grass sickness: a review of basic mechanisms.. Neurogastroenterol Motil 1999 Apr;11(2):79-92.
  10. Crawford LK, Caterina MJ. Functional Anatomy of the Sensory Nervous System: Updates From the Neuroscience Bench.. Toxicol Pathol 2020 Jan;48(1):174-189.
    pubmed: 31554486doi: 10.1177/0192623319869011google scholar: lookup
  11. Darlington PJ, Goldman JS, Cui QL, Antel JP, Kennedy TE. Widespread immunoreactivity for neuronal nuclei in cultured human and rodent astrocytes.. J Neurochem 2008 Mar;104(5):1201-9.
  12. De Koninck P, Carbonetto S, Cooper E. NGF induces neonatal rat sensory neurons to extend dendrites in culture after removal of satellite cells.. J Neurosci 1993 Feb;13(2):577-85.
  13. Delree P, Leprince P, Schoenen J, Moonen G. Purification and culture of adult rat dorsal root ganglia neurons.. J Neurosci Res 1989 Jun;23(2):198-206.
    pubmed: 2754765doi: 10.1002/jnr.490230210google scholar: lookup
  14. Dráberová E, Del Valle L, Gordon J, Marková V, Smejkalová B, Bertrand L, de Chadarévian JP, Agamanolis DP, Legido A, Khalili K, Dráber P, Katsetos CD. Class III beta-tubulin is constitutively coexpressed with glial fibrillary acidic protein and nestin in midgestational human fetal astrocytes: implications for phenotypic identity.. J Neuropathol Exp Neurol 2008 Apr;67(4):341-54.
    pubmed: 18379434doi: 10.1097/NEN.0b013e31816a686dgoogle scholar: lookup
  15. Draper ACE, Piercy RJ. Pathological classification of equine recurrent laryngeal neuropathy.. J Vet Intern Med 2018 Jul;32(4):1397-1409.
    pmc: PMC6060325pubmed: 29691904doi: 10.1111/jvim.15142google scholar: lookup
  16. Dupraz S, Grassi D, Karnas D, Nieto Guil AF, Hicks D, Quiroga S. The insulin-like growth factor 1 receptor is essential for axonal regeneration in adult central nervous system neurons.. PLoS One 2013;8(1):e54462.
  17. Eva R, Crisp S, Marland JR, Norman JC, Kanamarlapudi V, ffrench-Constant C, Fawcett JW. ARF6 directs axon transport and traffic of integrins and regulates axon growth in adult DRG neurons.. J Neurosci 2012 Jul 25;32(30):10352-64.
  18. Fadda A, Bärtschi M, Hemphill A, Widmer HR, Zurbriggen A, Perona P, Vidondo B, Oevermann A. Primary Postnatal Dorsal Root Ganglion Culture from Conventionally Slaughtered Calves.. PLoS One 2016;11(12):e0168228.
  19. Frey E, Valakh V, Karney-Grobe S, Shi Y, Milbrandt J, DiAntonio A. An in vitro assay to study induction of the regenerative state in sensory neurons.. Exp Neurol 2015 Jan;263:350-63.
  20. George D, Ahrens P, Lambert S. Satellite glial cells represent a population of developmentally arrested Schwann cells.. Glia 2018 Jul;66(7):1496-1506.
    pubmed: 29520852doi: 10.1002/glia.23320google scholar: lookup
  21. Gerhauser I, Hahn K, Baumgärtner W, Wewetzer K. Culturing adult canine sensory neurons to optimise neural repair.. Vet Rec 2012 Jan;170(4):102.
    pubmed: 22068333doi: 10.1136/vr.100255google scholar: lookup
  22. Gilley J, Coleman MP. Endogenous Nmnat2 is an essential survival factor for maintenance of healthy axons.. PLoS Biol 2010 Jan 26;8(1):e1000300.
  23. Gilley J, Seereeram A, Ando K, Mosely S, Andrews S, Kerschensteiner M, Misgeld T, Brion JP, Anderton B, Hanger DP, Coleman MP. Age-dependent axonal transport and locomotor changes and tau hypophosphorylation in a "P301L" tau knockin mouse.. Neurobiol Aging 2012 Mar;33(3):621.e1-621.e15.
  24. Gusel'nikova VV, Korzhevskiy DE. NeuN As a Neuronal Nuclear Antigen and Neuron Differentiation Marker.. Acta Naturae 2015 Apr-Jun;7(2):42-7.
    pmc: PMC4463411pubmed: 26085943
  25. Hanani M. Satellite glial cells in sensory ganglia: from form to function.. Brain Res Brain Res Rev 2005 Jun;48(3):457-76.
  26. Herculano-Houzel S, Manger PR, Kaas JH. Brain scaling in mammalian evolution as a consequence of concerted and mosaic changes in numbers of neurons and average neuronal cell size.. Front Neuroanat 2014;8:77.
    pmc: PMC4127475pubmed: 25157220doi: 10.3389/fnana.2014.00077google scholar: lookup
  27. Ho KC, Gwozdz JT, Hause LL, Antuono PG. Correlation of neuronal cell body size in motor cortex and hippocampus with body height, body weight, and axonal length.. Int J Neurosci 1992 Jul-Aug;65(1-4):147-53.
    pubmed: 1341676doi: 10.3109/00207459209003286google scholar: lookup
  28. Jackson JG, Robinson MB. Regulation of mitochondrial dynamics in astrocytes: Mechanisms, consequences, and unknowns.. Glia 2018 Jun;66(6):1213-1234.
    pmc: PMC5904024pubmed: 29098734doi: 10.1002/glia.23252google scholar: lookup
  29. Lin MT, Beal MF. Mitochondrial dysfunction and oxidative stress in neurodegenerative diseases.. Nature 2006 Oct 19;443(7113):787-95.
    pubmed: 17051205doi: 10.1038/nature05292google scholar: lookup
  30. Loreto A, Di Stefano M, Gering M, Conforti L. Wallerian Degeneration Is Executed by an NMN-SARM1-Dependent Late Ca(2+) Influx but Only Modestly Influenced by Mitochondria.. Cell Rep 2015 Dec 22;13(11):2539-2552.
    pubmed: 26686637doi: 10.1016/j.celrep.2015.11.032google scholar: lookup
  31. Malin SA, Davis BM, Molliver DC. Production of dissociated sensory neuron cultures and considerations for their use in studying neuronal function and plasticity.. Nat Protoc 2007;2(1):152-60.
    pubmed: 17401349doi: 10.1038/nprot.2006.461google scholar: lookup
  32. Mar FM, Simões AR, Leite S, Morgado MM, Santos TE, Rodrigo IS, Teixeira CA, Misgeld T, Sousa MM. CNS axons globally increase axonal transport after peripheral conditioning.. J Neurosci 2014 Apr 23;34(17):5965-70.
  33. Marinkovic P, Reuter MS, Brill MS, Godinho L, Kerschensteiner M, Misgeld T. Axonal transport deficits and degeneration can evolve independently in mouse models of amyotrophic lateral sclerosis.. Proc Natl Acad Sci U S A 2012 Mar 13;109(11):4296-301.
    pmc: PMC3306689pubmed: 22371592doi: 10.1073/pnas.1200658109google scholar: lookup
  34. Mellone M, Kestoras D, Andrews MR, Dassie E, Crowther RA, Stokin GB, Tinsley J, Horne G, Goedert M, Tolkovsky AM, Spillantini MG. Tau pathology is present in vivo and develops in vitro in sensory neurons from human P301S tau transgenic mice: a system for screening drugs against tauopathies.. J Neurosci 2013 Nov 13;33(46):18175-89.
  35. Milde S, Adalbert R, Elaman MH, Coleman MP. Axonal transport declines with age in two distinct phases separated by a period of relative stability.. Neurobiol Aging 2015 Feb;36(2):971-81.
  36. Misgeld T, Kerschensteiner M, Bareyre FM, Burgess RW, Lichtman JW. Imaging axonal transport of mitochondria in vivo.. Nat Methods 2007 Jul;4(7):559-61.
    pubmed: 17558414doi: 10.1038/nmeth1055google scholar: lookup
  37. Morris RL, Hollenbeck PJ. Axonal transport of mitochondria along microtubules and F-actin in living vertebrate neurons.. J Cell Biol 1995 Dec;131(5):1315-26.
    pmc: PMC2120647pubmed: 8522592doi: 10.1083/jcb.131.5.1315google scholar: lookup
  38. Muratori L, Ronchi G, Raimondo S, Geuna S, Giacobini-Robecchi MG, Fornaro M. Generation of new neurons in dorsal root Ganglia in adult rats after peripheral nerve crush injury.. Neural Plast 2015;2015:860546.
    pmc: PMC4333329pubmed: 25722894doi: 10.1155/2015/860546google scholar: lookup
  39. Muzio MR, Cascella M. Histology, Axon.. 2023 Jan;.
    pubmed: 32119275
  40. Nowicki M, Kosacka J, Spanel-Borowski K, Borlak J. Deferoxamine-induced neurite outgrowth and synapse formation in postnatal rat dorsal root ganglion (DRG) cell cultures.. Eur J Cell Biol 2009 Oct;88(10):551-62.
    pubmed: 19581022doi: 10.1016/j.ejcb.2009.05.003google scholar: lookup
  41. Osterloh JM, Yang J, Rooney TM, Fox AN, Adalbert R, Powell EH, Sheehan AE, Avery MA, Hackett R, Logan MA, MacDonald JM, Ziegenfuss JS, Milde S, Hou YJ, Nathan C, Ding A, Brown RH Jr, Conforti L, Coleman M, Tessier-Lavigne M, Züchner S, Freeman MR. dSarm/Sarm1 is required for activation of an injury-induced axon death pathway.. Science 2012 Jul 27;337(6093):481-4.
    pmc: PMC5225956pubmed: 22678360doi: 10.1126/science.1223899google scholar: lookup
  42. Rinholm JE, Vervaeke K, Tadross MR, Tkachuk AN, Kopek BG, Brown TA, Bergersen LH, Clayton DA. Movement and structure of mitochondria in oligodendrocytes and their myelin sheaths.. Glia 2016 May;64(5):810-25.
    pubmed: 26775288doi: 10.1002/glia.22965google scholar: lookup
  43. Russo D, Bombardi C, Castellani G, Chiocchetti R. Characterization of spinal ganglion neurons in horse (Equus caballus). A morphometric, neurochemical and tracing study.. Neuroscience 2011 Mar 10;176:53-71.
  44. Saijilafu, Hur EM, Liu CM, Jiao Z, Xu WL, Zhou FQ. PI3K-GSK3 signalling regulates mammalian axon regeneration by inducing the expression of Smad1.. Nat Commun 2013;4:2690.
    pmc: PMC3836055pubmed: 24162165doi: 10.1038/ncomms3690google scholar: lookup
  45. Seo BJ, Yoon SH, Do JT. Mitochondrial Dynamics in Stem Cells and Differentiation.. Int J Mol Sci 2018 Dec 5;19(12).
    pmc: PMC6321186pubmed: 30563106doi: 10.3390/ijms19123893google scholar: lookup
  46. Takahashi K, Ninomiya T. Morphological changes of dorsal root ganglion cells in the process-forming period.. Prog Neurobiol 1987;29(4):393-410.
    pubmed: 3615909doi: 10.1016/0301-0082(87)90020-7google scholar: lookup
  47. Tongtako W, Lehmbecker A, Wang Y, Hahn K, Baumgärtner W, Gerhauser I. Canine dorsal root ganglia satellite glial cells represent an exceptional cell population with astrocytic and oligodendrocytic properties.. Sci Rep 2017 Oct 24;7(1):13915.
    pmc: PMC5654978pubmed: 29066783doi: 10.1038/s41598-017-14246-7google scholar: lookup
  48. Valtcheva MV, Copits BA, Davidson S, Sheahan TD, Pullen MY, McCall JG, Dikranian K, Gereau RW 4th. Surgical extraction of human dorsal root ganglia from organ donors and preparation of primary sensory neuron cultures.. Nat Protoc 2016 Oct;11(10):1877-88.
    pmc: PMC5082842pubmed: 27606776doi: 10.1038/nprot.2016.111google scholar: lookup
  49. Wong AW, K P Yeung J, Payne SC, Keast JR, Osborne PB. Neurite outgrowth in normal and injured primary sensory neurons reveals different regulation by nerve growth factor (NGF) and artemin.. Mol Cell Neurosci 2015 Mar;65:125-34.
    pubmed: 25752731doi: 10.1016/j.mcn.2015.03.004google scholar: lookup
  50. Wright DE, Snider WD. Neurotrophin receptor mRNA expression defines distinct populations of neurons in rat dorsal root ganglia.. J Comp Neurol 1995 Jan 16;351(3):329-38.
    pubmed: 7706545doi: 10.1002/cne.903510302google scholar: lookup
  51. Wu Y, Chen M, Jiang J. Mitochondrial dysfunction in neurodegenerative diseases and drug targets via apoptotic signaling.. Mitochondrion 2019 Nov;49:35-45.
    pubmed: 31288090doi: 10.1016/j.mito.2019.07.003google scholar: lookup

Citations

This article has been cited 2 times.
  1. Lang Q, Schiavo G, Sleigh JN. In vivo imaging of axonal transport in peripheral nerves of rodent forelimbs.. Neuronal Signal 2023 Mar;7(1):NS20220098.
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  2. Cahalan SD, Boehm I, Jones RA, Piercy RJ. Recognising the potential of large animals for modelling neuromuscular junction physiology and disease.. J Anat 2022 Nov;241(5):1120-1132.
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