The corneal endothelium maintains the transparency of the cornea by transporting ions and fluid out of the corneal stroma to maintain the low level of hydration required for transparency. We propose to study the mechanisms of this transport to better understand the normal physiology of transport and thus how it is altered in disease. Transendothelial electrical potential difference, short circuit current, radioactive fluxes, and intracellular microelectrodes all measure ion flows through multiple pathways in the cell membrane simultaneously. Thus, the interpretation of those experimental results is often very complex and sometimes ambiguous. However, the recently invented patch clamp technique can measure the flows of ions through a single molecule, one pore in the cell membrane, and thus avoid these uncertainties. Unfortunately the patch clamp technique is now limited because 1) it takes a very long time to analyze the experimental data, 2) only patches with one pore can be used to measure pore kinetics, and 3) it cannot detect carriers (symports and antiports) or ion pumps (such as the soidum-potassium ATPase). We will overcome these limitations by designing and testing new mathematical techniques that will be able to 1) measure the kinetics of pores in patch clamp data much faster and more simply, 2) determine complex pore kinetics, 3) detect and measure net rates of cyclic pores, 4) extract kinetic data from patches with more than one pore, 5) identify pore-pore interactions, 6) study non-stationary kinetics, and 7) provide ways that ion pumps carriers and pumps could be measured using the patch clamp technique.
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