Skip to main content
Log in

K+-stimulated amino acid release from cultured cerebellar neurons: Comparison of static and dynamic stimulation paradigms

  • Original Articles
  • Published:
Neurochemical Research Aims and scope Submit manuscript

Abstract

The release of several endogenous amino acids and adenosine from rat cerebellar neuronal cultures following elevated K+ exposure in the presence and absence of added Ca2+ was studied. The amino acids aspartate (ASP), glutamate (GLU) and GABA were released from the cultures in a dose- and Ca2+-dependent manner. Taurine (TAU) and the nucleoside adenosine (ADN) efflux rates were dose-dependent but Ca2+-independent, and basal levels increased in the absence of Ca2+. The K+ depolarization induced release of serine (SER), alanine (ALA) and proline (PRO), was not dose-dependent and in the absence of extracellular Ca2+ (with added Mg2+) higher basal release of SER and ALA, but not PRO, was noted. These findings demonstrate that in addition to known cerebellar neurotransmitters, other neuroactive and neutral amino acids are released from cultured cerebellar neurons in response to K+ depolarization. Their observed efflux suggests they may have as yet unidentified roles in neuronal function with different classes of efflux corresponding to: neurotransmitter-type release (ASP, GLU, GABA), and osmoregulatory, possibly neuromodulatory-type release (TAU), a Ca2+-insensitive, possibly neuromodulatory-type release (ADN), and a depolarization-sensitive release (SER, ALA, PRO) of which SER and ALA are partially Ca2+-sensitive.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Similar content being viewed by others

References

  1. Nadi, N. S., McBride, W. J., and Aprison, M. H. 1977. Distribution of several amino acids in regions of the cerebellum of the rat. J. Neurochem. 28:453–455.

    Google Scholar 

  2. Kingsbury, A., Gallo, V., and Balazs, R. 1988. Stimulus-coupled release of amino acids from cerebellar granule cells in culture. Brain Res. 448:46–52.

    Google Scholar 

  3. Van Vliet, B. J., Sebben, M., Dumuis, A., Gabrion, J., Bockaert, J. and Pin, J. P. 1989. Endogenous amino acid release from cultured cerebellar neuronal cells: effect of tetanus toxin on glutamate release. J. Neurochem. 52:1229–1239.

    Google Scholar 

  4. Dutton, G. R. 1990. Isolation, Culture and Use of Viable Central Nervous System Perikarya. Pages 87–102,in Conn, P. M. (ed.), Methods in Neuroscience Vol. 2: Tissue Culture, Academic Press, San Diego.

    Google Scholar 

  5. Pearce, B. R., Currie, D. N., Dutton, G. R., Hussey, R. E. G., Beale, R., Pigott, R. 1981. A simple perfusion chamber for studying neurotransmitter release from cells maintained in monolayer culture. J. Neurosci. Meth. 3:255–259.

    Google Scholar 

  6. Rogers, K. L., Philibert, R. A., Allen, A. J., Molitor, J., Wilson, E. J., and Dutton, G. R. 1987. HPLC analysis of putative amino acid neurotransmitters from primary cerebellar cultures. J. Neurosci. Meth. 22:173–179.

    Google Scholar 

  7. Rogers, K. L., Philibert, R. A., and Dutton, G. R. 1990. Glutamate receptor agonists cause efflux of endogenous neuroactive amino acids from cerebellar neurons in culture. Eur. J. Pharm. 177:195–199.

    Google Scholar 

  8. Dutton, G. R., Currie, D. N., and Tear, K. 1981. An improved method for the bulk isolation of viable perikarya from postnatal cerebellum. J. Neurosci. Meth. 3:421–427.

    Google Scholar 

  9. Pearce, B. R., Currie, D. N., Beale, R., and Dutton, G. R. 1981. Potassium-stimulated, calcium-dependent release of [3H]GABA from neuron- and glial-enriched cultures of cells dissociated from rat cerebellum. Brain Res. 206:485–489.

    Google Scholar 

  10. Smith, P. K., Krohn, R. I., Hermanson, G. T., Mallia, A. K., Gatner, F. H., Provenzano, M. D., Fujimoto, E. K., Goeke, N. M., Olson, B. J., and Klenk, D. C. 1985. Measurement of protein using bicinchoninic acid. Anal. Biochem. 150:76–85, 1985.

    Google Scholar 

  11. Tennant, J. R. 1964. Evaluation of trypan blue technique for determination of cell viability. Transplantation 2:685–688.

    Google Scholar 

  12. Currie, D. N., and Dutton, G. R. 1980. [3H]GABA uptake as a marker for cell type in primary cultures of cerebellum and olfactory bulb. Brain Res. 199:473–481.

    Google Scholar 

  13. Philibert, R. A., and Dutton, G. R. 1989. Dihydorpyridines modulate K+-evoked amino acid and adenosine release from cerebellar neuronal cultures. Neurosci. Lett. 102:97–102.

    Google Scholar 

  14. Burry, R., and Lasher, R. S. 1978. A quantitative electron microscopic study of synapse formation in dispersed cell cultures of rat cerebellum stained either by Os-UL or by E-PTA. Brain Res. 147:1–15.

    Google Scholar 

  15. Beale, R., Dutton, G. R., and Currie, D. N. 1980. An ion flux assay of action potential sodium channels in neuron- and glial-enriched cultures of cells dissociated from rat cerebellum. Brain Res. 183:241–246.

    Google Scholar 

  16. Pearce, B. R., and Dutton, G. R. 1981. K+-stimulated release of endogenous glutamate, GABA and other amino acids from neuron-and glial-enriched cultures of the rat cerebellum. FEBS Lett. 135:215–218.

    Google Scholar 

  17. Gallo, V., Ciotti, M. T., Coletti, A., Aloisi, F., and Levi, G. 1982. Selective release of glutamate from cerebellar granule cells differentiating in culture. Proc. Natl. Acad. Sci. U.S.A. 79:7919–7923.

    Google Scholar 

  18. Toggenburger, G., Willund, L., Henke, H., and Cuenod, M. 1983. Release of endogenous and accumulated exogenous amino acids from slices of normal and climbing fibre-deprived rat cerebellar slices. J. Neurochem. 41:1606–1613.

    Google Scholar 

  19. Docherty, M., Bradford, H. F., Wu, J. Y. 1987. Co-release of glutamate and asparate from cholinergic and GABAergic synaptosomes. Nature 316:148–150.

    Google Scholar 

  20. Philibert, R. A., Rogers, K. L., Allen, A. J., and Dutton, G. R. 1988. Dose-dependent, K+-stimulated efflux of endogenous taurine from primary astrocyte cultures is Ca2+-dependent. J. Neurochem. 51:1122–1126.

    Google Scholar 

  21. Holopainen, I., Kontro, P., and Oja, S. S. 1989. Release of taurine from cultured cerebellar granule cells and astrocytes: Co-release with glutamate. Neurosci. 29:425–432.

    Google Scholar 

  22. Schousboe, A., Frandsen, A., and Drejer, J. 1989. Evidence for evoked release of adenosine and glutamate from cultured cerebellar granule cells. Neurochem. Res. 9:871–875.

    Google Scholar 

  23. Drejer, J., and Schousboe, A. 1989. Selection of a pure cerebellar granule cell culture by kainate treatment. Neurochem. Res. 14:751–754.

    Google Scholar 

  24. Schousboe, A., and Pasantes-Morales, H. 1989. Potassium-stimulated release of [3H]-taurine from cultured GABAergic and glutamatergic neurons. J. Neurochem. 53:1309–1315.

    Google Scholar 

  25. Simmons, M. L., and Dutton, G. R. 1990. Chronic kainate treatment decreases K+-stimulated release of endogenous amino acids from cultured cerebellar neurons. Soc. Neurosci. Abstr. 16:1183.

    Google Scholar 

  26. Clark, M., and Dar, M. S. 1989. Effect of acute ethanol on release of endogenous adenosine from rat cerebellar synaptosomes. J. Neurochem. 52:1857–1865.

    Google Scholar 

  27. Huxtable, R. J. 1989. Taurine in the central nervous system and the mammalian actions of taurine. Prog. Neurobiol. 32:471–533.

    Google Scholar 

  28. Enna, S. J. 1979. Amino acid neurotransmitter candidates. Pages 41–51,in Hess, H. J. (ed.), Ann. Rep. Med. Chem., Vol. 14 Academic Press, New York.

    Google Scholar 

  29. Nadler, J. V., Wang, A., and Hakin, A. 1988. Toxicity of L-proline toward rat hippocampal neurons. Brain Res. 456:168–172.

    Google Scholar 

  30. Pin, J-P., Weiss, S., Sebben, M., Kemp, D. E., and Bockaert, J. 1986. Release of endogenous amino acids from striatal cultures in primary culture. J. Neurochem. 47:594–603.

    Google Scholar 

  31. Kontro, P., Oja, S. S. 1987. Taurine and GABA release from mouse cerebral cortex slices: Effects of structural analogues and drugs. Neurochem. Res. 12:475–482.

    Google Scholar 

  32. Jacob, T. J. C., and Duncan, G. 1981. Calcium controls both sodium and potassium permeability of lens membranes. Exp. Eye. Res. 33:85–93.

    Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Rights and permissions

Reprints and permissions

About this article

Cite this article

Rogers, K.L., Philibert, R.A. & Dutton, G.R. K+-stimulated amino acid release from cultured cerebellar neurons: Comparison of static and dynamic stimulation paradigms. Neurochem Res 16, 899–904 (1991). https://doi.org/10.1007/BF00965539

Download citation

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF00965539

Key Words

Navigation