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Isolation of metabolically distinct synaptosomes on Percoll gradients

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Abstract

Synaptosomes were prepared from whole rat brain by six different methods based on gradients of sucrose, Ficoll or Percoll. In these, the synthesis and calcium-specific release of amino acids were assessed by two different procedures. Preparations based on sucrose showed the least calcium-specific release, followed by Ficoll-derived synaptosomes. As previously described, Percoll gave two separate populations of synaptosomes, both very active in terms of release of aspartate, glutamate, and GABA. The data involving release and synthesis were not identical, but did agree in the following: in low-density synaptosomes, haloperidol blocked both the release and synthesis of glutamate, but was without effect in the heavier populatin. 2-chloroadenosine and 2-oxoglutarate affected both release and synthesis only in the high-density population. Dopamine blocked aspartate release and synthesis only in the high-density population. These results suggest that haloperidol interferes with glutamate release and synthesis via a mechanism which may not involve adenosine, serotonin, or dopamine.

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References

  1. Gray, E. G., and Whittaker, V. P. 1962. The isolation of nerve endings from brain. An electron microscopic study of cell fragments divided by homogenization and centrifugation. J. Anat. (Lond.) 96:79–88.

    Google Scholar 

  2. De Robertis, E., Pellegrino de Iraldi, A., Rodriguez de Lorez Arnaiz, G., and Salganicoff, L. 1962. Cholinergic and non-cholinergic nerve endings in rat brain. I Isolation and subcellular distribution of acetylcholine and acetylcholine esterase. J. Neurochem. 9:23–25.

    PubMed  Google Scholar 

  3. Whittaker, V. P. 1969. The synaptosome. Pages 327–364,in Lajtha A. (ed.), Handbook of Neurochemistry, Plenum Press, NY.

    Google Scholar 

  4. Kurokawa, M. T., Sakamoto, M. T., and Kato, M. 1965. Distribution of sodium-plus-potassium adenosine-triphosphatese activity in isolated nerve-ending particles. Biochem. J. 97:833–844.

    PubMed  Google Scholar 

  5. Verity, M. A. 1972. Cation modulation of synaptosomal respiration. J. Neurochem. 19:1305–1317.

    PubMed  Google Scholar 

  6. Booth, R. F. G., and Clark, J. B. 1978. A rapid method for preparation of relatively pure metabolically competent synaptosomes from rat brain. Biochem. J. 176:365–370.

    PubMed  Google Scholar 

  7. Cooper, A. J., and Perry, S. 1980. The applicability of freeze-Percoll gradients to whole-cell isopycnic fractionations. Preliminary results. J. Immunol. Meth. 37:353–362.

    Google Scholar 

  8. Pertoft, H., and Laurent, T. C. 1982. Sedimentation of cells in celloidal silica (Percoll)in Pretlow, T. G. and Pretlow, T. P. (eds.) Cell separation: methods and selected applications, Academic Press, New York.

    Google Scholar 

  9. Nagy, A., and Delgado-Escueta, A. V. 1984. Rapid preparation of synaptosomes from mammalian brain using nontoxic iso-osmotic gradient material (Percoll). J. Neurochem. 43:1114–1123.

    PubMed  Google Scholar 

  10. Dunkley, P. R., Jarvie, P. A., Heath, J. W., Kidd, G. J., and Rostas, J. A. 1986. A rapid method for isolation of synaptosomes on Percoll gradients. Brain Res. 372:115–129.

    PubMed  Google Scholar 

  11. Robinson, P. J., Lovenberg, W., and Dunkley, P. R. 1985. The release of dopamine and serotonin from two populations of synaptosomes isolated on Percoll gradients, Soc. Neurosci. Abstr. 11:303.

    Google Scholar 

  12. Sherman, A. D., and Mott, J. 1984. Direct effects of neuroleptics on glutamate release. Neuropharmacol. 23:1253–1259.

    Google Scholar 

  13. Sherman, A. D., and Mott, J. 1986. Effects of glutaminase inhibition on release of endogenous glutamic acid. Neuropharmacol. 25:1353–1357.

    Google Scholar 

  14. Mott, J. and Sherman, A. D. 1987. Specificity of the effects of neuroleptics on the release of glutamate from the rat amygdala. Drug Dev. Res. 11:235–241.

    Google Scholar 

  15. Hamberger, A., Chiang, G. H., Nylen, E. S., Scherr, S. W., and Cotman, C. W. 1979. Glutamate as a CNS transmitter I: Evaiuation of glucose and glutamine as precursors for the synthesis of preferentially released glutamate. Brain Res. 168:513–530.

    PubMed  Google Scholar 

  16. Baughman, R. W., and Gilbert, C. D. 1980. Aspartate and glutamate as possible neurotransmitters of cells in layer 5 of the visual cortex. Nature 287:848–860.

    PubMed  Google Scholar 

  17. Mitchell, P. R., and Doggett, N. S. 1980. Modulation of striatal [3H]glutamic acid release by dopaminergic drugs. Life Sci. 26:2073–2081.

    PubMed  Google Scholar 

  18. Rowland, R. J., and Roberts, P. J. 1980. Activation of dopamine receptors inhibits calcium-dependent release from cortico-striaral terminals in vitro. Eur. J. Pharmacol. 62:241–242.

    PubMed  Google Scholar 

  19. Katz, I. A. 1983. Glutamine inhibits the accumulation and hydroxylation of tryptophan in rat striatal synaptosomes. Brain Res. 264:160–173.

    PubMed  Google Scholar 

  20. Dolphin, A. C., and Archer, E. R. 1983. An adenosine agonist inhibits and a cyclic AMP analog enhances the release of glutamate, but not GABA from slices of rat dentage gyrus. Neurosci. Lett. 43:49–54.

    PubMed  Google Scholar 

  21. Hajos, F. 1975. An improved method for preparation of synaptosomal fractions of high purity. Brain Res. 93:485–493.

    PubMed  Google Scholar 

  22. Cotman, C. W., and Matthews, D. A. 1971. Synaptic plasma membranes from rat brain synaptosomes: isolation and initial characterization. Biochem. Biophys. Acta. 249:380–392.

    PubMed  Google Scholar 

  23. Nichols, D. G., Sihra, T. S., and Sanchez-Prieto, J. 1987. Calcium-dependent and independent release of glutamate from synaptosomes monitored by continuous fluorimetry. J. Neurochem. 49:50–57.

    PubMed  Google Scholar 

  24. Sanchez-Prieto, J., Sihra, T. S., and Nichols, D. G. 1987. Characterization of the exocytotic release of glutamate from guinea pig cortical synaptosomes. J. Neurochem. 49:58–64.

    PubMed  Google Scholar 

  25. Lenda, K., and Svenneby, G. 1980. Rapid high-performance liquid chromatographic determination of amino acids in synaptosomal extracts. J. Chromatog. 198:516–519.

    Google Scholar 

  26. Kvamme, E., and Lenda, K. 1982. Regulation of glutaminase by exogenous glutamate, ammonia, and 2-oxoglutarate in synaptosomal enriched preparations of rat brain. Neurochem. Res. 7:667–678.

    PubMed  Google Scholar 

  27. Vyas, S., and Bradford, H. F. 1987. Co-release of acetylcholine, glutamate, and tautine from synaptosomes ofTorpedo electric organ. Neurosci. Lett. 82:58–64.

    PubMed  Google Scholar 

  28. Dochtery, M., Bradford, H. F., and Wu, J.-Y. 1987. Co-release of glutamate and aspartate from cholinergic and GABAergic synaptosomes. Nature 330:64–66.

    PubMed  Google Scholar 

  29. Nicholls, D. G., and Sihra, T. S. 1986. Synaptosomes possess an exocytotic pool of glutamate. Nature 321:772–774.

    PubMed  Google Scholar 

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Sherman, A.D. Isolation of metabolically distinct synaptosomes on Percoll gradients. Neurochem Res 14, 97–101 (1989). https://doi.org/10.1007/BF00969765

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