The relative ratio of anaerobic and aerobic corneal carbohydrate metabolism, as indicated by the lactate: pyruvate (L/P) ratio, increases during corneal epithelial hypoxia and contact lens wear, and this is acompanied by ATP depletion. During and following mid-term eyebank storage, corneal energy depletion and lactate accumulation have also been reported to accompany the corneal degradation of endothelial and epithelial function. The proposed experiments are designed to determine whether or not these type of edema are due to a) an osmotic effect of lactate accumulation due to an altered L/P ratio, b) a lactic acidosis, or c) a lactic acidosis accompanied by cellular """"""""rundown"""""""" of epithelial and/or stromal K+ gradients. If L, L/P, or stromal pH or K+ activity are altered by simulated contact lens wear, epthelial hypoxia, or eyebank-induced storage in isolated rabbit corneas, these timecourse and reversibility changes will be studied and correlated with corneal thickness changes (swelling or deswelling rates). Furthermore, """"""""stromal"""""""" loading in deepithelialized normoxic corneas will be used to determine whether the measured L, pH or K+ activity, placed at a normal stroma, can inhibit endothelium-mediated deturgescenece to account for the edema. If so, then their effects on endothelial hydraulic conductivity will be examined to determine whether the action is on """"""""pump"""""""" or """"""""leak"""""""". In other experiments two lactate dehydrogenase (LDH) inhibitors (3.2mM sodium oxamate and 3.2mM disodium oxalate) and two pyruvate dehydrogenase (PDH) stimulates (3.2mM sodium dichloracetate and 3.2mM sodium 2-chloropropionate) will be assessed for their ability to prevent or reverse these forms of corneal edema (and the associated ionic effects, if detected above). L and P will be assayed spectrophotometrically in homogenized corneal extracts and expressed both as umoles/g wet weight and as umoles/mg corneal protein. Stormal pH and K+ activity will be expressed in terms of pH and meq/liter, as determined from double barrelled glass liquid ion exchange microelectrodes. Specular microscopy will be used to determine corneal swelling and deswelling rates. Together, these studies will provide insight regarding the etiologies and prevention of contact lens- and eyebank storage-induced corneal edema.
