Prolonged Time Course of Glutamate Release from Nerve Terminals: Relationship Between Stimulus Duration and the Secretory Event

Abstract: The kinetics of synaptosomal ³H]glutamate release were measured on a subsecond time scale to study the relationship between the length of depolarization and the duration of the secretory event. The time course of release evoked by elevated K⁺ was complex, proceeding for several seconds after a 200-ms depolarization. We developed a protocol for depolarizing excitable membranes on a millisecond time scale to deliver brief depolarizations, termed the synthetic action potential, by using batrachotoxin to activate Na⁺ channels. Depolarization is achieved by superfusing with solutions containing elevated concentrations of Na⁺, and the duration of the depolarization is limited by including tetrodotoxin (TTX) in the superfusion solution to block Na⁺ entry. Direct measurements of the time courses of Na⁺ current and membrane depolarizations were made in batrachotoxin-treated sensory neurons using patch clamp recording methods. Rapid increases in Na⁺ and TTX concentrations produced transient increases in inward Na⁺ current that decayed with a time course proportional to TTX concentration. Current clamp measurements indicated that, with 10 µ M TTX, depolarizations last ～30 ms. Nonetheless, synaptosomal release of ³H]glutamate triggered by the synthetic action potential remained prolonged. Brief neuronal action potentials at some synapses may trigger transmitter release that persists for several seconds.