Glutamate release from isolated synaptoneurosomes from the mouse cortex

doi:10.3969/j.issn.0529-1356.2014.06.023

Abstract

Abstract:

Objective To detect the release of glutamate from the isolated synaptoneurosomes from the mouse cortex. Methods The mouse cortex was dissected, homogenized and filtered through three layers of 147 μm nylon meshes. The synaptoneurosomes were resuspended after centrifuged. An enzyme-linked fluorescence detection method was used to detect the glutamate release, which utilized glutamate dehydrogenase, and the reduction of nicotinamide adenine dinucleotide phosphate(NADP) to NADPH. The glutamate standard curve was set up and then the glutamate release from isolated synaptoneurosomes was tested and analyzed in three independent experiments. Results A good linear relationship was found between the glutamate concentrations and fluorescence values. The release of glutamate from synaptoneurosomes significantly increased after depolarization agent KCl added and gradually reached a plateau. Glutamate release was increased more when Triton X-100 was added. The inactive synaptoneurosomes after one freeze-thaw cycle had a different curve of glutamate release. Conclusion The release of excitatory neurotransmitter glutamate from synaptoneurosomes of the mouse cortex isolated using the improved method is well studied.

Key words: Synaptoneurosome, Glutamate, Release, Enzymatic reaction, Mouse

JIN Guo-rong DING Cai-yun WANG Yi-min LIU Jian-rong WEI Hong-en*. Glutamate release from isolated synaptoneurosomes from the mouse cortex[J]. AAS, 2014, 45(6): 861-864.

References

［1］van Spronsen M, Hoogenraad CC. Synapse pathology in psychiatric and neurologic disease ［J］. Curr Neurol Neurosci Rep, 2010, 10(3): 207-214.
［2］Pereda AE. Electrical synapses and their functional interactions with chemical synapses ［J］. Nat Rev Neurosci, 2014, 15(4): 250-263.
［3］Spooren W, Lindemann L, Ghosh A, et al. Synapse dysfunction in autism: a molecular medicine approach to drug discovery in neurodevelopmental disorders ［J］. Trends Pharmacol Sci, 2012, 33(12): 669-684.
［4］Selkoe DJ. Alzheimer’s disease is a synaptic failure ［J］. Science, 2002, 298(5594): 789-791.
［5］Sudhof TC. Neurotransmitter release: the last millisecond in the life of a synaptic vesicle ［J］. Neuron, 2013, 80(3): 675-690.
［6］Meldrum BS. Glutamate as a neurotransmitter in the brain: review of physiology and pathology ［J］. J Nutr, 2000, 130(4S Suppl): 1007S-1015S.
［7］Chen X, Zhou SL, Gu XS, et al. Protective effects of salidroside on glutamate induced neurotoxicity in cultured hippocampal neurons ［J］. Acta Anatomica Sinica, 2008, 39(3): 355-359. (in Chinese)
陈霞, 周松林, 顾晓松, 等. 红景天苷对谷氨酸损伤海马神经元的保护作用［J］. 解剖学报, 2008, 39(3): 355-359.
［8］Westmark PR, Westmark CJ, Jeevananthan A, et al. Preparation of synaptoneurosomes from mouse cortex using a discontinuous percoll-sucrose density gradient ［J］. J Vis Exp, 2011, (55): e3196.
［9］Schwartz-Bloom RD, Engblom AC, Akerman KE, et al. Measurement of chloride movement in neuronal preparations［J］. Curr Protoc Neurosci, 2001, Chapter 7: Unit 7.10.
［10］Nicholls DG, Sihra TS, Sanchez-Prieto J. Calcium-dependent and -independent release of glutamate from synaptosomes monitored by continuous fluorometry ［J］. J Neurochem, 1987, 49(1): 50-57.
［11］Sim AT, Herd L, Proctor DT, et al. High throughput analysis of endogenous glutamate release using a fluorescence plate reader ［J］. J Neurosci Methods, 2006, 153(1): 43-47.
［12］Nicholls DG, Sihra TS. Synaptosomes possess an exocytotic pool of glutamate ［J］. Nature, 1986, 321(6072): 772-773.
［13］Cline H. Synaptogenesis: a balancing act between excitation and inhibition ［J］. Curr Biol, 2005, 15(6): R203-205.
［14］Hollingsworth EB, McNeal ET, Burton JL, et al. Biochemical characterization of a filtered synaptoneurosome preparation from guinea pig cerebral cortex: cyclic adenosine 3’:5’-monophosphate-generating systems, receptors, and enzymes ［J］. J Neurosci, 1985, 5(8): 2240-2253.
［15］Schwartz RD, Jackson JA, Weigert D, et al. Characterization of barbiturate-stimulated chloride efflux from rat brain synaptoneuro-somes ［J］. J Neurosci, 1985, 5(11): 2963-2970.
［16］Dunkley PR, Jarvie PE, Heath JW, et al. A rapid method for isolation of synaptosomes on Percoll gradients ［J］. Brain Res, 1986, 372(1): 115-129.
［17］Manchester KL. Glutamate dehydrogenase: a reappraisal ［J］. Biochem Educ, 1985, 13(3): 131-133.

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