Neural Calcium Channels-Regulation and Function

Brown University, Providence RI

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Abstract

[unreadable] DESCRIPTION (provided by applicant): In this renewal, we take full advantage of our expertise and tools developed to study splice forms of the neuronal N-type voltage-gated calcium channel. We transition from a general description of splice isoforms to determine the importance of alternative splicing in identified neuronal populations. At center stage is cell-specific alternative splicing and its importance in optimizing calcium channel activity for specific functions. In aim 1 we will assess the pattern of alternative splicing in two functionally distinct populations of hippocampal interneurons that use N-type and P-type channels respectively, to mediate fast GABAergic transmission, In aim 2, we extend our studies of a novel splice form of the N-type calcium channel expressed in nociceptive neurons of the dorsal root ganglia. This isoform generates larger N-type currents in nociceptive neurons. We will study how alternative splicing modifies current density. Our preliminary data suggests isoform-specific interactions with the ubiquitin proteosome degradation pathway, a mechanism that fits well with our functional data. Finally, dynamic changes in the pattern of alternative splicing of calcium channels will have profound effects on the responsiveness of the cell. We have evidence for temporal changes in the pattern of splicing of N-type calcium channels in sympathetic ganglia during development. We will study whether these changes depend on presynaptic input to sympathetic neurons. We will also determine whether temporal changes in the patterns of splicing occur in dorsal root ganglia following nerve injury, a process that results in neuropathic pain. The aims proposed in this application address the expression pattern, mechanism of action, and temporal changes in alternative splicing of the N-type calcium channel in identified cells. We have expertise in all techniques proposed in this application. Preliminary data are presented for methods developed recently, such as yeast two-hybrid screening and protein analyses. Alternative splicing is extensive in the mammalian nervous system and is essential for optimizing the function of proteins essential for neuronal signaling. Tissue-specific alternative fine-tunes the kinetics of channel activities and their association to downstream signaling pathways. Pharmacologically distinct splice isoforms preferentially expressed in specific tissues or cell types offer exciting new targets for the design of highly specific drugs [unreadable] [unreadable]