Physiology Core

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Abstract The Physiology Core provides a series of graded in vitro technologies for studying the function and regulation of transport and other membrane resident proteins with progressive degrees of complexity from single molecules to model systems to native epithelia. To gain understanding of biological processes that mediate kidney function in health and disease states, in light of the phenotypic diversity and functional complexity of this organ, the use of strategies that allow specific segment/cell/protein to be studied in isolation under defined conditions is required. The overall goal of this core is to elucidate at a molecular and cellular level the function and regulation of key proteins involved in kidney health (including development) and disease. To accomplish this goal, the Core will provide investigators with: a) microdissected tubules for analysis of RNA expression and/or abundance, protein expression, immunolocalization, and enzyme/transporter microassays, b) functional fluorescence assays of channel/transporter function in individually identified cells in isolated tubules microperfused in vitro, c) measurements of transepithelial ion/solute fluxes across isolated tubules microperfused in vitro, d) electrophysiological assays for functional analysis of transport proteins in heterologous expression systems, either wild-type or mutated, alone or in combination with putative regulatory proteins, e) analyses of transcellular and paracellular transport in model and native epithelia in Ussing chambers, f) technologies for analysis of post-translational modification of proteins including phosphorylation, ubiquitination, palmitoylation and glycosylation, and g) technologies to study composition, dynamics, and regulatory mechanisms through the assessment of transcript abundance in isolated single cells. A central function of the Core is to provide instruction in all the techniques performed by the Core and the use of the Core equipment. The Physiology Core will interact synergistically with the other Cores to: a) extend the studies conducted in either heterologous expression systems or isolated tubules to animals models, b) identify potential therapeutic compounds that target transporters and associated regulatory pathways, and c) perform detailed and quantitative analysis using imaging technologies of the expression of transport and regulatory proteins in heterologous expression systems, model and native epithelia, and isolated tubules.