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The release of the major excitatory and inhibitory neurotransmitters, glutamate and GABA from rat caudate-putamen (CP) tissue slices and GABA release from fetal rat primary striatal cultures were studied to ascertain key mechanisms underlying the regulation of their release. An improved, highly sensitive method of high performance liquid chromatography and electrochemical detection (LCED) was developed to enable the reliable measurement of these endogenous amino acids from tissue release extracts.The assay permits detection of endogenous amino acids from one milligram of CP tissue in vitro. Release of both amino acids from CP tissue was evoked by elevated levels of potassium while the potassium channel blocker, 4-aminopyridine (4-AP) selectively released GABA. Glu was selectively released by the excitatory amino acid (EAA) receptor agonist N-methyl-D-aspartate (NMDA) independently of external Ca2+ concentration. Ca2+-dependent release of Glu and GABA by potassium- and 4-AP evoked depolarization was shown to be solely dependent on external Ca2+ influx through the P-type Ca2+ channel (P