Display options
Cite
Dixon CL, Zhang Y, Lynch JW. Generation of Functional Inhibitory Synapses Incorporating Defined Combinations of GABA(A) or Glycine Receptor Subunits. Front Mol Neurosci. 2015;8:80doi: 10.3389/fnmol.2015.00080.
Dixon, C. L., Zhang, Y., & Lynch, J. W. (2015). Generation of Functional Inhibitory Synapses Incorporating Defined Combinations of GABA(A) or Glycine Receptor Subunits. Frontiers in molecular neuroscience, 880. https://doi.org/10.3389/fnmol.2015.00080
Dixon, Christine L, et al. "Generation of Functional Inhibitory Synapses Incorporating Defined Combinations of GABA(A) or Glycine Receptor Subunits." Frontiers in molecular neuroscience vol. 8 (2015): 80. doi: https://doi.org/10.3389/fnmol.2015.00080
Dixon CL, Zhang Y, Lynch JW. Generation of Functional Inhibitory Synapses Incorporating Defined Combinations of GABA(A) or Glycine Receptor Subunits. Front Mol Neurosci. 2015 Dec 23;8:80. doi: 10.3389/fnmol.2015.00080. eCollection 2015. PMID: 26778954; PMCID: PMC4688394.
Copy Download .nbib Format: NLM
Share it on

Front Mol Neurosci. 2015 Dec 23;8:80. doi: 10.3389/fnmol.2015.00080. eCollection 2015.

Generation of Functional Inhibitory Synapses Incorporating Defined Combinations of GABA(A) or Glycine Receptor Subunits.

Frontiers in molecular neuroscience

Christine L Dixon, Yan Zhang, Joseph W Lynch

Affiliations

  1. Queensland Brain Institute, University of Queensland Brisbane, QLD, Australia.
  2. Queensland Brain Institute, University of QueenslandBrisbane, QLD, Australia; School of Biomedical Sciences, University of QueenslandBrisbane, QLD, Australia.

PMID: 26778954 PMCID: PMC4688394 DOI: 10.3389/fnmol.2015.00080

Abstract

Fast inhibitory neurotransmission in the brain is mediated by wide range of GABAA receptor (GABAAR) and glycine receptor (GlyR) isoforms, each with different physiological and pharmacological properties. Because multiple isoforms are expressed simultaneously in most neurons, it is difficult to define the properties of individual isoforms under synaptic stimulation conditions in vivo. Although recombinant expression systems permit the expression of individual isoforms in isolation, they require exogenous agonist application which cannot mimic the dynamic neurotransmitter profile characteristic of native synapses. We describe a neuron-HEK293 cell co-culture technique for generating inhibitory synapses incorporating defined combinations of GABAAR or GlyR subunits. Primary neuronal cultures, prepared from embryonic rat cerebral cortex or spinal cord, are used to provide presynaptic GABAergic and glycinergic terminals, respectively. When the cultures are mature, HEK293 cells expressing the subunits of interest plus neuroligin 2A are plated onto the neurons, which rapidly form synapses onto HEK293 cells. Patch clamp electrophysiology is then used to analyze the physiological and pharmacological properties of the inhibitory postsynaptic currents mediated by the recombinant receptors. The method is suitable for investigating the kinetic properties or the effects of drugs on inhibitory postsynaptic currents mediated by defined GABAAR or GlyR isoforms of interest, the effects of hereditary disease mutations on the formation and function of both types of synapses, and synaptogenesis and synaptic clustering mechanisms. The entire cell preparation procedure takes 2-5 weeks.

Keywords: GABAergic; IPSC; electrophysiology; glycinergic; inhibitory postsynaptic current; neuropharmacology; synaptogenesis

References

  1. Eur J Neurosci. 2009 Sep;30(6):1077-91 - PubMed
  2. Curr Drug Targets. 2015;16(7):735-46 - PubMed
  3. J Neurophysiol. 1998 Nov;80(5):2608-20 - PubMed
  4. Nat Neurosci. 2006 Oct;9(10 ):1294-301 - PubMed
  5. J Biol Chem. 2005 Jun 10;280(23):22365-74 - PubMed
  6. Science. 2004 May 7;304(5672):884-7 - PubMed
  7. J Cell Mol Med. 2008 Dec;12(6B):2848-66 - PubMed
  8. J Neurosci Res. 1995 Dec;42(5):674-83 - PubMed
  9. Int J Parasitol. 2010 Nov;40(13):1477-81 - PubMed
  10. Front Mol Neurosci. 2009 Nov 09;2:23 - PubMed
  11. Mol Pharmacol. 1996 Nov;50(5):1253-61 - PubMed
  12. J Biol Chem. 2014 Feb 28;289(9):5399-411 - PubMed
  13. Br J Pharmacol. 2015 Jul;172(14):3522-36 - PubMed
  14. Mol Cell Neurosci. 2006 Jul;32(3):254-73 - PubMed
  15. Curr Top Med Chem. 2012;12(4):254-69 - PubMed
  16. Nat Protoc. 2007;2(3):670-6 - PubMed
  17. Curr Opin Investig Drugs. 2006 Jan;7(1):48-53 - PubMed
  18. J Biol Chem. 2007 Nov 9;282(45):33052-63 - PubMed
  19. J Neurosci Res. 1993 Aug 1;35(5):567-76 - PubMed
  20. J Biol Chem. 2012 Aug 10;287(33):27417-30 - PubMed
  21. Nat Neurosci. 2003 Jul;6(7):708-16 - PubMed
  22. Nat Neurosci. 2005 Jun;8(6):736-44 - PubMed
  23. Neuropharmacology. 2015 Feb;89:391-7 - PubMed
  24. Biochemistry. 2012 Jul 3;51(26):5229-31 - PubMed
  25. Mol Cell Neurosci. 2007 May;35(1):14-23 - PubMed
  26. Int J Biochem Cell Biol. 2014 Aug;53:218-23 - PubMed
  27. Eur J Neurosci. 2013 Oct;38(8):3146-58 - PubMed
  28. J Physiol. 2010 Jun 1;588(Pt 11):1861-9 - PubMed
  29. Science. 2002 Aug 30;297(5586):1525-31 - PubMed
  30. Neuron. 2006 Nov 22;52(4):679-90 - PubMed
  31. Neuropharmacology. 2009 Jan;56(1):141-8 - PubMed
  32. Cell. 2004 Dec 29;119(7):1013-26 - PubMed
  33. Cell. 2000 Jun 9;101(6):657-69 - PubMed
  34. J Pharmacol Toxicol Methods. 2005 May-Jun;51(3):187-200 - PubMed
  35. Hum Mol Genet. 2011 Aug 1;20(15):3042-51 - PubMed
  36. Pharmacol Ther. 2006 Nov;112(2):513-28 - PubMed
  37. J Neurosci. 2012 Sep 12;32(37):12915-20 - PubMed
  38. Neurosci Lett. 2011 Jan 13;488(1):31-5 - PubMed
  39. J Neurosci. 2005 Jan 5;25(1):260-70 - PubMed
  40. Cell Mol Life Sci. 2005 Sep;62(18):2027-35 - PubMed
  41. J Neurosci. 2006 May 3;26(18):4880-90 - PubMed
  42. Mol Brain. 2014 Jan 09;7:2 - PubMed
  43. Neuropharmacology. 2009 Jan;56(1):303-9 - PubMed
  44. J Vis Exp. 2014 Nov 14;(93):e52115 - PubMed
  45. Mol Psychiatry. 2016 Jul;21(7):936-45 - PubMed

Publication Types