Abstract
Ca2+ uptake was measured in purified rat cerebral cortex synaptosomes (P3 pellets) using45Ca2+ as a tracer. Ca2+ influx increased in time, and with an increase in external K+ concentration and temperature. The net (external K+-induced, depolarization-dependent) uptake follows a two-component course. The exponential term, due to the opening of voltage-operated calcium channels (VOC), has a rate constant which increases with an increase in the depolarization level (1.04 versus 0.54 nmol/s/mg protein for 50 mM—versus 15 mM [K+]-dependent net influx). The linear term, due to the Na+/Ca2+ exchange system, has a similar rate constant at all depolarization levels (0.16+/−0.05 and 0.11+/−0.02 nmol/s/mg protein). Excitatory amino acids (glutamate, kainate and n-methyl-d-aspartate-NMDA-) were tested on this preparation at doses ranging between 5×10−5 M and 5×10−3M and at multiple incubation times, under resting conditions and under two depolarizing conditions (partial depolarization: 15 mM external K+ and maximal depolarization: 50 mM external K+). NMDA was also tested in the absence of Mg2+. No effect was detectable under any of these experimental conditions. Hypotheses to interpret these data are discussed. Further studies on other preparations are needed in order to directly investigate the presynaptic effects of excitatory amino acids.
Similar content being viewed by others
References
Fagg, G. E., Foster, A. C., and Ganong A. H. 1986. Excitatory amino acid synaptic mechanisms and neurological function. Trends Pharmacol. Sci. 7:357–363.
Dingledine, R. 1986. NMDA receptors: what do they do? Trends Neurosci. 9:47–49.
Collingridge, G. L., and Bliss, T. V. P. 1987. NMDA receptorstheir role in long-term potentiation. Trends Neurosci. 10:278–293.
Rothman, S. M., and Olney, J. W. 1987. Excitotoxicity and the NMDA receptor. Trends Neurosci. 10:299–302.
Cull-Candy, S. G., and Usowicz, M. M. 1987. Multiple-conductance channels activated by excitatory amino acids in cerebellar neurons. Nature 325:525–527.
Jahr, C. E., and Stevens, C. F. Glutamate activates multiple single channel conductances in hippocampal neurons. Nature 325:522–527.
Cull-Candy, S. G., and Usowicz, M. M. 1987. Patch clamp recording from single glutamate-receptor channels. Trends Pharmacol. Sci. 8:218–224.
Sladeczek, F., Recasens, M. and Bockaert, J. 1988. A new mechanism for glutamate receptor action: phosphoinositide hydrolysis. Trends Neurosci. 12:545–549.
Foster, A. C., Mena, E. E., Fagg, G. E., and Cotman, C. W. 1981. Glutamate and aspartate binding sites are enriched in synaptic junctions isolated from rat brain. J. Neurosci. 1:620–625.
Migani, P., Virgili, M., Contestabile, A., Poli, A., Villani, L., and Bernabei, O. 1985. [3H]Kainic acid binding sites in the synaptosomal-mitochondrial (P2) fraction from goldfish brain. Brain Res. 361:36–45.
Miwa, A., and Kawai, N. 1986. Presynaptic glutamate receptorpossible involvement of a K+ channel. Brain Res. 385:161–164.
Miller, R. J. 1987. Multiple calcium channels and neuronal function. Science 235:46–52.
Augustine, G. J., Charlton, M. P., and Smith, S. J. 1987. Calcium action in synaptic transmitter release. Ann. Rev. Neurosci. 10:633–693.
Tsien, R. W., Lipscombe, D., Madison, D. V., Bley, K. R., and Fox, A. P. 1988. Multiple types of neuronal calcium channels and their selective modulation. Trends Neurosci. 11:431–438.
Ashley, R. H., Brammer, M. J., and Marchbanks, R. 1984. Measurement of intrasynaptosomal free calcium by using the fluorescent indicator quin-2. Biochem. J. 219:149–158.
Blaustein, M. P. 1975. Effects of potassium, veratridine and scorpion venom on calcium accumulation and transmitter release by nerve terminals in vitro. J. Physiol. 247:617–655.
Nachshen, D. A., and Blaustein, M. P. 1980. Some properties of potassium-stimulated calcium influx in presynaptic nerve endings. J. Gen. Physiol. 76:709–727.
Nachshen, D. A., and Blaustein, M. P. 1980. Influx of calcium, strontium, and barium in presynaptic nerve endings. J. Gen. Physiol. 79:1065–1087.
Turner, T. J., and Goldin, S. M. 1985. Calcium channels in rat brain synaptosomes: identification and pharmacological characterization. J. Neurosci. 5:841–849.
Suszkiw, J. B., O'Leary, M. E., Murawsky, M. M., and Wang, T. 1986. Presynaptic calcium channels in rat cortical synaptosomes: fast-kinetics of phasic calcium influx, channel inactivation, and relationship to nitrendipine receptors. J. Neurosci. 6:1349–1357.
Harris, R. A. 1985. Effects of excitatory amino acids on calcium transport by brain membranes. Brain Res. 337:167–170.
Lazarewicz, J. W., Lehmann, A., Hagberg, H., and Hamberger, A. 1986. Effects of kainic acid on brain calcium fluxes studied in vivo and in vitro. J. Neurochem. 46:494–498.
Booth, R. F. G., and Clark, J. B. 1978. A rapid method for the preparation of relatively pure, metabolically competent synaptosomes from rat brain. Biochem. J. 176:365–370.
Koenig, M. L., and Jope, R. S. 1987. Aluminum inhibits the fast phase of voltage-dependent calcium influx into synaptosomes. J. Neurochem. 49:316–320.
Lowry, O. H., Rosebrough, N. J., Farr, A. L., and Randall, A. J. 1951. Protein measurement with the folin phenol reagent. J. Biol. Chem. 193:259–265.
Cotman, C. W., Monaghan, D. T., Ottersen, O. P., and Storm-Mathisen, J. 1987. Anatomical organization of excitatory amino acid receptors and their pathways. Trend Neutosci. 10:263–270.
O'Shaughnessy, C. T., and Lodge, D. 1988. N-methyl-D-aspartate receptor-mediated increase in intracellular calcium is reduced by ketamine and phencyclidine. Eur. J. Pharmacol. 153:201–209.
Nicoletti, F. 1988. Receptor-operated calcium channels activated by excitatory amino acids. Neurosci. Lett. Suppl. 33:5136.
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Simonato, M., Jope, R.S., Bianchi, C. et al. Lack of excitatory amino acid-induced effects on calcium fluxes measured with45Ca2+ in rat cerebral cortex synaptosomes. Neurochem Res 14, 677–682 (1989). https://doi.org/10.1007/BF00964878
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00964878