Calcium-dependent and -independent release of glutamate from synaptosomes monitored by continuous fluorometry.
An enzyme-linked fluorometric assay is described for the continuous monitoring of the unidirectional efflux of glutamate from guinea-pig synaptosomes. Glutamate efflux from freshly suspended, polarized synaptosomes occurs at 0.35-0.39 nmol min-1 mg of protein-1 and is not significantly affected by external Ca2+. KCl depolarization (30 mMKCl) in the absence of Ca2+ doubles this rate, whereas in the presence of Ca2+, the initial kinetics of the assay are consistent with the release in the first 5 s of 0.6 nmol mg of protein-1. The final extent of Ca2+-dependent release amounts to 1.9 nmol mg of protein-1, or 8.5% of the total intrasynaptosomal glutamate content. Preincubation of synaptosomes at 30 degrees C for 2 h before depolarization leads to a decrease in Ca2+-independent release and an increase in Ca2+-dependent release, consistent with an intrasynaptosomal relocation of the amino acid.
D000818:Animals; D002118:Calcium / Q000502:physiology*; D002851:Chromatography, High Pressure Liquid; D005260:Female; D005470:Fluorometry; D005969:Glutamate Dehydrogenase; D005971:Glutamates / Q000378:metabolism*; D018698:Glutamic Acid; D006168:Guinea Pigs; D008297:Male; D013574:Synaptosomes / Q000378:metabolism*
D005971:Glutamates; D018698:Glutamic Acid; D005969:Glutamate Dehydrogenase; D002118:Calcium