Over the last several decades, our understanding of the diversity of ion channels and their role in electrical signaling in neurons has increased exponentially. Such advances have helped understand and treat some brain disorders, but the mechanisms underlying many other common brain disorders remain to be discovered. In- deed, to the extent examined, many of these conditions are associated with synaptic or channelopathic abnormalities, including ones that emerge in adolescence, with aging, or as a result of experience (e.g., drug expo- sure, post-traumatic stress disorder). Deepening our understanding of the links among ion channels, synapses, dendrites, and brain disease is therefore a fundamental goal of both basic and clinical neuroscience research. One critical bottleneck to such an endeavor is that thin dendrites remain inaccessible to patch-clamp physiology, the technique responsible for much of our understanding with regard to dendritic and neuronal integration. Consequently, our understanding of neuronal and dendritic function is based primarily on a combination of inferences from large-diameter dendritic recordings and computational models. To circumvent such limitations and fill crucial knowledge gaps, this project will determine the expression patterns of several key ligand- and voltage-gated ion channels for entire, complete dendrites from hippocampal CA1 pyramidal neurons using at or near ultrastructural resolution techniques. To accomplish this, the proposed experiments will combine field emission scanning electron microscopy (FESEM) and array tomography (AT) with immunogold or immunoflourescence channel detection, respectively. Such an approach will fill limiting gaps in our knowledge of the single-dendrite proteome in both mice and humans. And, if successful, the proposed approach has the poten- tial to revolutionize our understanding of the role of ion channels in dendritic/neuronal function because trafficking networks and expression levels are likely to differ intradendritically in single neurons throughout the brain. Finally, to validate the potential clinical/translational relevance of such an approach, a channelopathy that emerges with age in two different mouse models of Alzheimer's disease will be acutely reversed pharmacologically, and then probed with FESEM and AT to determine whether expression is indeed rendered normal again, or whether such a treatment induces a functional, but not a restorative, reversal of the channelopathy.

Public Health Relevance

The thin dendrites of most neurons are the major sites of synapses and ion channel expression, but remain inaccessible to patch-clamp recordings. Such inaccessibility severely limits our understanding of the role of synaptic and dendritic ion channels in neuronal integration and the pathogenesis of brain diseases. The experiments in this proposal will overcome this critical limitation using large-scale microscopic reconstructions of dendrites and synapses from healthy neurons and other neurons affected by Alzheimer's disease-linked molecular processing, probed proteomically for several important ligand- and voltage-gated ion channels with immunogold particles and fluorescence markers.

Agency
National Institute of Health (NIH)
Institute
National Institute on Aging (NIA)
Type
Research Project (R01)
Project #
4R01AG047073-04
Application #
9104056
Study Section
Special Emphasis Panel (ZNS1)
Program Officer
Wise, Bradley C
Project Start
2013-09-30
Project End
2017-06-30
Budget Start
2016-07-15
Budget End
2017-06-30
Support Year
4
Fiscal Year
2016
Total Cost
Indirect Cost
Name
Rush University Medical Center
Department
Neurosurgery
Type
Schools of Medicine
DUNS #
068610245
City
Chicago
State
IL
Country
United States
Zip Code
60612
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