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Modulation of N-methyl-D-aspartate receptor-mediated increases in cytosolic calcium in cultured rat cerebellar granule cells.
- Source :
-
Brain research [Brain Res] 1991 Jun 21; Vol. 552 (1), pp. 13-22. - Publication Year :
- 1991
-
Abstract
- The concentration of intracellular free Ca2+ ([Ca2+]i) was measured in rat cerebellar granule cells using the fluorescent indicator fura-2. Culturing the cells as monolayers on plastic squares which could be placed into cuvettes allowed measurements of [Ca2+]i to be performed on large and homogeneous populations of CNS neurons. Granule cells so cultured maintained low levels of [Ca2+]i (around 90 nM) which increased promptly upon the addition of various excitatory amino acids including N-methyl-D-aspartate (NMDA). Increases in [Ca2+]i elicited by NMDA were inhibited by Mg2+ (1 mM) and often potentiated by glycine (1 microM). The addition of TTX or strychnine (5 microM each) did not alter responses to NMDA or NMDA plus glycine. Cytosolic Ca2+ responses to NMDA/glycine were dependent on the presence of extracellular Ca2+ and were unaffected by concentrations of nifedipine or verapamil that blocked increases in [Ca2+]i elicited by K+ depolarization. Responses elicited by NMDA/glycine were inhibited competitively by 2-amino-5-phosphonovalerate or 3-((+-)-2-carboxypiperazin-4-yl)-propyl-1- phosphonic acid and non-competitively by MK-801 or Mg2+. HA-966 and 7-chlorokynurenate inhibited responses to NMDA alone and blocked competitively the potentiating effects of glycine. The results demonstrate NMDA-mediated increases in [Ca2+]i in cerebellar granule cells that arise solely from influx of extracellular Ca2+ through dihydropyridine-insensitive channels. The strict dependence of the NMDA-evoked response on extracellular Ca2+ provides little evidence for a coupling of NMDA receptors to inositol phosphate metabolism and mobilization of intracellular Ca2+. The effect of various agents on NMDA/glycine-induced increases in [Ca2+]i parallels their effects on ligand binding to or current flow through the NMDA receptor-channel complex. The measurement of cytosolic Ca2+ in this preparation of neuronal cells thus appears especially well suited for assessing, on a functional level, the regulation of NMDA receptors in the CNS.
- Subjects :
- Animals
Calcium Channel Blockers pharmacology
Cells, Cultured
Cerebellum cytology
Cerebellum drug effects
Cytosol metabolism
Egtazic Acid pharmacology
Fura-2
Glycine pharmacology
Kinetics
Potassium Chloride pharmacology
Rats
Rats, Inbred Strains
Receptors, N-Methyl-D-Aspartate drug effects
Spectrometry, Fluorescence
Calcium metabolism
Cerebellum metabolism
N-Methylaspartate pharmacology
Receptors, N-Methyl-D-Aspartate physiology
Subjects
Details
- Language :
- English
- ISSN :
- 0006-8993
- Volume :
- 552
- Issue :
- 1
- Database :
- MEDLINE
- Journal :
- Brain research
- Publication Type :
- Academic Journal
- Accession number :
- 1833031
- Full Text :
- https://doi.org/10.1016/0006-8993(91)90653-d