Simultaneous kinetic analyses of neuronal connectivities

DESCRIPTION provided by applicant Aberrant neuronal connectivity is associated with a number of neuropathological and neuropsychiatric diseases disorders Inefficient developmental connectivity misdirected connectivity or disrupted and degenerated connectivity can account for losses of normal neuronal communication and subsequent losses of mental or physical health Altered expression of axonal guidance proteins neuroinflammation neuronal toxicity and glial disturbances may be involved in inadequate neuronal connectivity The goal of the work described in this application is to develop an instrument platform that can both enumerate development or loss of axons and the kinetics of these changes in response to molecular and cellular modifiers The proposed grating coupled surface plasmon resonance GCSPR and grating coupled surface plasmon coupled emission GCSPCE instrumentation with a microflow biosensor chip will be developed and validated for quantification of developmental axonal outgrowths among punches of brain regional sections as well as loss of connections previously established Up to five mm punches can be placed on the cm biosensor gold chip precoated with extracellular matrix proteins and specific molecules guidance proteins or cells microglia spotted at specific locations Lack or loss of axonal connections will be kinetically monitored by SPR and presynaptic and postsynaptic antigens can be assayed by SPCE with fluorochrome conjugated antibodies to neuronal antigens The geometrical placement of specific brain regions will be identified so that preferential regional interactions can be quantified The planned device to assess neuronal circuitry is built upon extensive experience with SPR microarrays that can characterize presence of single cells and their released products The work described will produce instrumentation capable of providing a more complete and coherent picture of differential neuronal connectivities to provide a more comprehensive view of neuronal communications which will reveal new opportunities for therapeutic interventions PUBLIC HEALTH RELEVANCE The proposed surface plasmon resonance SPR assessment of axonal outgrowth between brain regions on a biosensor chip takes a step beyond what is currently possible with microscopic techniques With the instrumentation and software the differential kinetics of axonal projections toward certain brain regions is measured by SPR and coupled fluorescence Additionally proteins and cells spotted at precise areas on the chip can be evaluated for their influences and modifiers can be added or removed from the flowcell with microfluidic exchanges