Parkinson's disease (PD) afflicts roughly 1 in every 1000 adults, rising exponentially in incidence after the age of fifty. Human and animal studies have made it clear that PD results from the degeneration of nigrostriatal dopaminergic neurons. This loss leads to molecular, cellular and systems level adaptations in the principal target of these neurons - the neostriatum. Our long-term goal is to characterize the molecular and cellular consequences of this denervation in the neostriatum. In this proposal, we will focus on adaptations in the cellular mechanisms mediating the actions of dopamine and acetylcholine on neostriatal neurons following the near total loss of their dopaminergic innervation. We propose to achieve our immediate aims by combining two experimental approaches in a rat model of advanced PD created by 6-OHDA lesioning or dopamine depletion. First, functionally significant adaptations in the ability of post-synaptic dopaminergic and cholinergic signaling pathways to modulate neuronal excitability will be studied using a patch-clamp analysis of acutely-isolated and cultured neostriatal neurons identified by retrograde labeling from the substantia nigra and globus pallidus. Second, in biophysically characterized neostriatal neurons, alterations in gene expression will determined using single cell mRNA amplification. Cellular profiles or fingerprints will be constructed by screening for specific mRNAs, including those coding for dopamine and acetylcholine receptors, signaling enzymes, and transmitters. These tools will be used to answer three basic questions. First, do near total 6-OHDA lesions of substantia nigra pars compacta or dopamine depletion induce alterations in the intrinsic excitability of neostriatal neurons? Second, do these lesions alter the neuromodulatory effects of dopamine and, if so, how are specific receptor signaling pathways altered? Lastly, do these lesions alter the neuromodulatory effects of acetylcholine and its interactions with dopamine in shaping excitability in neostriatal neurons? By combining anatomical, physiological and molecular analyses at the single cell level, we should be able to provide an unprecedented breadth of information about the molecular and cellular adaptations that take place in functionally relevant subsets of neostriatal neurons following dopaminergic denervation. The insights gained from this work should provide a cellular framework in which changes in the behavior of large collections of neurons and the neostriatum itself can be understood. In doing so, this work will be of value in the design of rational pharmacological and genetic therapies for Parkinson's disease and other disorders of dopaminergic signaling, such as schizophrenia.
