Dopamine (DA) neurons in the substantia nigra (SN) exhibit a slow intrinsic pacemaker activity that is modified by synaptic input resulting in regular irregular, and burst firing important for movement and motivated behaviors. Voltage-gated Ca2+ (CaV) channels contribute to pacing in SN DA neurons directly by conducting depolarizing currents and indirectly by activation of Ca2+-activated channels. The L-, P/Q- and T-type CaV channels have been found to significantly contribute to pacing, but only the L-type CaV current has been studied in depth. The goal of the proposed research is to evaluate the role of the P/Q- and T-type CaV channels in the slow pacemaker activity of SNc DA neurons and determine whether their contribution to pacing is mediated by the depolarizing current or the secondary effects of the Ca2+ carrying the current. This goal will be achieved by using a combination of pharmacology and voltage-, action potential-, current-, and dynamic- clamp electrophysiology in acutely dissociated SN DA neurons. Action potential-clamp is a modified form of the voltage-clamp technique where an action potential or group of action potentials are used as the voltage command allowing one to determine when and how much of a particular current is flowing during activity. Dynamic-clamp is a modified form of the current-clamp technique that dynamically adjusts the current injected in response to real-time changes in the membrane potential based on equations that model channel behavior allowing one to add virtual channels to a living cell. As a result, these virtual channels produce currents that mimic the native channe but without the ion selectivity. To achieve the goal of this application, these techniques will be used to address the following specific aims:
Aim 1. Characterize the voltage-dependent kinetics of P/Q- and T-type CaV currents in SNc DA neurons.
Aim 2. Quantify the amount and timing of P/Q- and T-type CaV currents flowing during the AP and pacing in SNc DA neurons.
Aim 3. Determine the relative importance of the depolarizing current and secondary effects of Ca2+ entry for the pacemaker activity of SNc DA neurons for the P/Q- and T-type CaV currents. The results of the proposed experiments will allow the scientific community to: 1. Understand the mechanism by which these channels contribute to normal SNc DA neuronal activity. 2. Understand how drugs used to block these channels, such as those used to treat migraines, neuropathic pain and epilepsy, might disrupt the normal activity of these neurons. 3. Develop novel drugs to treat movement and motivation disorders.
Abnormalities of substantia nigra dopamine neuron function have been implicated in a variety of neurodegenerative and psychiatric diseases including Parkinson's disease, Tourette's syndrome and schizophrenia. By understanding how changes in ion channel function modulate the excitability of these neurons, novel ion channel drug targets can be found, resulting in potent selective drugs for these diseases.