The goals of the proposed experiments are to obtain information about the basic mechanisms underlying long-term changes in the synaptic properties of mammalian CNS neurons. Such long-term alterations are thought to be responsible for the enduring enhancement in the efficacy of synaptic transmission underlying learning and memory formation. The present study will examine the role of intraneuronal Ca2+ homeostasis, in particular the contribution of Ca2+ sequestering organelles, to the control of synaptic excitability. Long-term potentiation (LTP), a use-dependent increase in synaptic efficacy critically linked to Ca2+, will serve as the model for plastic cellular alterations. The study will take advantage of the newly developed whole-cell patch clamp recording technique in conventional brain slice preparations. In addition to improving the quality and resolution of electrophysiological recordings, this technique is best suited for introducing chemicals and drugs into the neurons' internal environment. Whole-cell patch (current- and voltage-clamp) and extracellular recordings will be obtained in neurons of the rat hippocampal formation. Spontaneous or evoked excitatory or inhibitory postsynaptic currents will be recorded under various conditions of intraneuronal Ca2+ homeostasis. LTP will be induced by tetanic stimulation of afferent pathways. Considering the ubiquitous role of Ca2+ in regulating cellular function, the modulation of synaptic events by Ca2+ originating from Ca2+-sequencing organelles will be especially emphasised. This can be accomplished by studying how intraneuronally applied chemicals, which alter the release of Ca2+ from intracellular sequestering organelles, modify basal or long-term potentiated synaptic responses. Furthermore, the induction and maintenance phases of currents and fluctuations in steady excitatory current noise in potentiated neurons will permit the examination of long-term synaptic alterations at the single receptor/channel level. The study proposes to establish the nature of the relationship between intraneuronal Ca2+ homeostasis and the control of synaptic excitability intrinsic to the nervous system. The regulation of neuronal excitability by Ca2+ released from the neuronal endoplasmic reticulum or other Ca2+ sequestering organelles has not yet been investigated in detail. Only recently, by analogy to the function of the sarcoplasmic reticulum in muscle, studies have begun to stress the importance of Ca2+ buffering and release mechanisms in neurons. By extending these novel findings and examining previously unexplored aspects of intraneuronal Ca2+ homeostasis, the present experiments will provide a better understanding of the long-term regulation of neuronal function.
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