Multiple components of synaptosomal [3H]-gamma-aminobutyric acid release resolved by a rapid superfusion system.
Release of [3H]-gamma-aminobutyric acid ([3H]GABA) from rat brain synaptosomes was studied with 60-ms time resolution, using a novel rapid superfusion method. Synaptosomes were prelabeled with [3H]GABA via an associated GABA uptake system. KCl depolarization stimulated at least three distinct components of GABA release: (1) a phasic Ca-dependent component, which develops rapidly and decays with a time constant of at most 60 ms; (2) a tonic Ca-dependent component that persists after KCl depolarization is ended; (3) a Ca-independent component. The three components of GABA release are pharmacologically distinct. The phasic component was selectively blocked by 50 microM Cd2+, while the tonic component was selectively blocked by 100 microM Ni2+. The Ca-independent component was selectively blocked by nipecotic acid (IC50 = 21 microM), a known inhibitor of Na+-dependent GABA uptake. The time course and amplitude of Ca-dependent GABA release evoked by the Ca2+ ionophore A23187 were nearly identical with Ca-dependent release evoked by depolarization. This result indicates that Ca-dependent GABA release depends primarily on Ca2+ entry into the nerve terminal, and not depolarization, per se. The properties of the phasic component suggest that it is normally initiated by a voltage-sensitive Ca2+ channel that is functionally and pharmacologically distinct from those previously described. The Ca-independent component of GABA release is probably mediated by reversal of the Na-dependent, electrogenic GABA uptake system. The ability to identify multiple components of GABA release on a physiologically relevant time scale may afford a more precise definition of the mechanism of action of drugs thought to affect neurotransmission in the brain.
Turner
T J
TJ
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts 02115.
Goldin
S M
SM
D000818:Animals; D001921:Brain / Q000378:metabolism*; D002104:Cadmium / Q000494:pharmacology; D002118:Calcium / Q000494:pharmacology; D007700:Kinetics; D008297:Male; D009532:Nickel / Q000494:pharmacology; D009557:Nipecotic Acids / Q000494:pharmacology; D011392:Proline / Q000031:analogs & derivatives*; D011865:Radioisotope Dilution Technique; D051381:Rats; D011919:Rats, Inbred Strains; D012964:Sodium / Q000494:pharmacology; D013574:Synaptosomes / Q000187:drug effects / Q000378:metabolism*; D014316:Tritium; D005680:gamma-Aminobutyric Acid / Q000378:metabolism*
D009557:Nipecotic Acids; D002104:Cadmium; D014316:Tritium; C030278:nipecotic acid; D005680:gamma-Aminobutyric Acid; D009532:Nickel; D011392:Proline; D012964:Sodium; C010983:homoproline; D002118:Calcium