The overall aim of this proposal is to examine the role of internal calcium (Ca2+) stores in nerve terminals. It is well known that the release of neurotransmitters and hormones is tightly coupled to rises in the free intracellular Ca2+ concentration. While the influx of Ca2+ through voltage-gated Ca2+ channels is undoubtedly a very important source of Ca2+ affecting release, Ca2+ release from internal stores may provide another important source of Ca2+ for this process. Until recently these intracellular stores have attracted relatively little attention, and their very existence in nerve terminals was controversial. Recent work has shown that highly localized Ca2+ release events, like Ca2+ sparks of the muscle, can be seen in neuronal preparations. In neurohypophysial terminals, these Ca2+ release events appear to emanate from a ryanodine-sensitive intracellular Ca2+ pool, and depolarizing stimuli induce an increase in their frequency. In spite of all this information, the source of the released Ca2+, and a physiological role for this phenomenon, is unknown. Preliminary evidence suggests that these Ca2+ release events could represent mobilization of Ca2+ from vesicular stores. If so, localized Ca2+ release in the precise location of exocytosis should modulate release. It is our goal to determine the source of these ryanodine sensitive Ca2+ release events in neurohypophysial terminals, and to elucidate the physiological role of this mobilization of Ca2+ on neuropeptide secretion. To accomplish this we have been able to develop, for the first time, a technique capable of detecting quantal release events from individual nerve terminals. This amperometric technique allows us to simultaneously perform Ca2+ imaging and monitor transmitter release from a defined area of a single isolated nerve terminal. This project aims to add to the current body of knowledge of the physiology of the neurohypophysial terminals, and, more generally, will provide a more complete understanding of the role played by intracellular calcium and of the mechanism by which large dense core granular fusion occurs in the Central Nervous System. This knowledge could thus prove to be important for the understanding and treatment of synaptic pathologies.
Communication both within the brain and with its targets is via release of transmitters at nerve terminals, and such release is known to be dependent on the entry of extracellular calcium and the subsequent elevation of intraterminal calcium. We have recently shown that nerve terminals have intracellular calcium stores and in this proposal we utilize novel techniques to determine what are these intraterminal stores and whether they can regulate the release of transmitters from nerve terminals. This knowledge could prove to be important for the understanding and treatment of synaptic diseases such Eaton-Lambert Syndrome and Amyotrophic Lateral Sclerosis, as well as Alzheimer's disease and neuronal aging.
