Ca2+ channels mediate voltage-dependent Ca2+ influx in subcellular compartments of neurons, triggering such diverse processes such as neurotransmitter release, dendntic action potentials and excitation-transcnption coupling. One of the Ca2+ channels responsible for fast synaptic transmission is the P/Q-type Ca2+ channel. A fundamental question that remains unsolved is how Ca2+ channels and their associated modulatory proteins are targeted to the appropriate cellular compartments like presynaptic terminals to fulfil their designated function. In order to understand this question we will first determine which P/Q-type Ca2+ channel subunits and their intracellular domains are responsible for axonal/dendritic targeting of the Ca2+ channel complexes in hippocampal neurons. We will then correlate the axonal/dendritic targeting of P/Q-type Ca2+ channel complexes with their specific role in synaptic transmission using hippocampal neurons in culture from VQ-type channel knock out mice. These experiments will descnbe how changes in the localization and biophysical properties of wild type and mutated Ca2+ channels responsible for spinocerebellar ataxia 6 (SCA6) phenotypes effect synaptic transmission. Following these experiments we will analyze the specificity of the nodulation of these axonal/dendritic targeted Ca2+ channel complexes by G proteins and relate the specificity n modulation to the structure of the interacting proteins Protein interactions will be analyzed using a new developed two hybrid system and co-expression of Ca2+ channel complexes with G protein constructs in heterologous expression systems. The results will verify whether P/Q-type Ca2+ channels including their mutations have different specificity for G protein subunits and will identify the protein domains of the G protein subunits responsible for modulation of this presynaptic Ca2+ channel type. ? ? Elucidating the mechanisms that regulate Ca2+ channel targeting is critical to understanding both the basic physiology of neurons as well as several important neurological diseases. SCA6 appears to be caused by mutations in P/Q-type voltage-gated Ca2+ channels responsible for transmitter release. The identified mutations alter the biophysical properties of Ca2+ channel and change their potency to interact with intracellular modulating proteins like G proteins and Ca2+ channel ancillary subunits. Therefore a better understanding of the molecular epitopes underlying targeting, assembly and regulation of Ca2+ channels in subcellular compartments of neurons will help to design new strategies for treating ataxia and may identify new diseases related to ionic channel targeting.
