Development of intracellular calcium responses to depolarization and to kainate and N-methyl-D-aspartate in cultured mouse hippocampal neurons.
We have investigated the initial appearance of voltage-gated Ca channels and kainate- and NMDA-type glutamate receptors in cultured embryonic mouse hippocampal neurons. The Ca-dependent fluorescence change of the dye fura-2 was used as a sensitive assay for the presence of functional channels and receptors. Expression of functional NMDA receptors was observed on some hippocampal neurons as early as E14. By the equivalent of E15-16, 40-50% of cells responded to Ko-depolarization (50 mM), indicating the presence of functional voltage-gated Ca channels, approximately 20% of cells responded to kainate (50 microM), and just under 20% responded to NMDA (50 microM; in the presence of glycine and strychnine). By the equivalent of the end of the embryonic period 70-80% of cells responded to all 3 stimuli. As approximately 20% of cells in these cultures are glia, these data indicate that by the time of birth close to 100% of neurons express functioning kainate and NMDA receptors, and voltage-gated Ca channels. Increases in [Ca2+]i in embryonic neurons after application of NMDA were sensitive to APV and to external Mg, as are responses in mature neurons. The IC50 for block by external Mg of the [Ca2+]i increase induced by NMDA was 130 microM, and there was a slight positive correlation between the amplitude of the response to NMDA and sensitivity to external Mg.
Barish
M E
ME
Department of Physiology and Biophysics, College of Medicine, University of California, Irvine 92717.
Mansdorf
N B
NB
D000375:Aging / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D006624:Hippocampus / Q000187:drug effects / Q000196:embryology / Q000378:metabolism*; D007091:Image Processing, Computer-Assisted; D007608:Kainic Acid / Q000494:pharmacology*; D008274:Magnesium / Q000494:pharmacology; D008564:Membrane Potentials / Q000502:physiology; D051379:Mice; D008856:Microscopy, Fluorescence; D016202:N-Methylaspartate / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D009474:Neurons / Q000187:drug effects / Q000378:metabolism*; D014741:Video Recording
D016202:N-Methylaspartate; D008274:Magnesium; D007608:Kainic Acid; D002118:Calcium