Previous studies have demonstrated that cholinergic projection neurons from the brainstem pedunculopontine nucleus (PPN) modulate the excitability and firing rate of substantia nigra pars compacta (SNc) dopamine (DA) neurons. However, the underlying mechanisms by which acetylcholine (ACh) exerts its influence on DA cells remain unclear. We are interested in the role of cholinergic afferents in regulating DA cell electrical activity. Specifically, our goal is to understand the functional consequences of changes in afferent firing activity associated with muscarinic receptor activation during phasic (rapid) and sustained (tonic) stimulation. It is hypothesized that changes in membrane potential and calcium dynamics associated with DA neuron activation are differentially modulated by rapid (phasic) and sustained (tonic) muscarinic receptor stimulation. Using whole-cell voltage clamp recordings in acutely isolated DA neurons, the effects of phasic and tonic muscarine application (to mimic PPN output activity) on membrane potential and intracellular calcium dynamics will be assessed. The involvement of the interplay between calcium entry and release from intracellular stores will be investigated on the basis of the following specific aims: 1) To examine the effect of phasic muscarine receptor activation on whole-cell membrane currents and calcium dynamics; 2) to determine if the magnitude of the calcium rise in response to muscarinic receptor activation of DA neurons is dependent on basal intracellular calcium levels; and, 3) to determine the second messenger mechanisms associated with rapid muscarinic receptor activation of DA neurons. SNc DA activity is known to be highly regulated by afferent inputs and the delicate balance between the hyperpolarizing and depolarizing reponses of DA cells to ACh may be critical in the regulation of SNc activity and subsequent DA delivery to striatal targets. Findings from this study could have significant implications in understanding and developing treatments for movement and behavioral dysfunctions.
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