Pyridine nucleotides of rabbit cornea with histotoxic anoxia: Chemical analysis, non-invasive fluorometry and physiological correlates

• Published in: Experimental eye research Vol. 37; no. 1; pp. 1 - 9
• Main Authors: Masters, Barry R., Riley, Michael V., Fischbarg, Jorge, Chance, Britton
• Format: Journal Article
• Language: English
• Published: England Elsevier Ltd 01.07.1983
• Subjects: Animals; Azides - pharmacology; Biological Transport - drug effects; cornea; Cornea - drug effects; Cornea - metabolism; cycling assay; endothelium; Endothelium - metabolism; epithelium; Epithelium - metabolism; histotoxic anoxia; In Vitro Techniques; Male; Membrane Potentials - drug effects; NAD - metabolism; NADP - metabolism; Oxygen; Perfusion; pyridine nucleotides; Rabbits; redox fluorometry; Sodium Azide; Spectrometry, Fluorescence; cycling assay; endothelium; redox fluorometry; histotoxic anoxia; cornea; epithelium; pyridine nucleotides
• ISSN: 0014-4835; 1096-0007
• DOI: 10.1016/0014-4835(83)90144-6

The pyridine nucleotides from both the epithelium and the endothelium of rabbit cornea were measured by the cycling assay. Sodium azide (10 m m) applied for 1 hr to induce histotoxic anoxia decreased the endothelial NAD + NADH ratio from 4·62 to 1·49 and decreased the epithelial NAD + NADH ratio from 2·56 to 1·08. The larger NAD + NADH ratio for the endothelium as compared to the epithelium corresponds to a more oxidized state. The corresponding ratios for NADP + NADPH were 1·2 for the endothelium and 0·70 for the epithelium. Sodium azide had no effect on the NADP + NADPH ratio for the endothelium, but decreased the epithelial ratio to 0·62. Pyridine nucleotide fluorescence was measured with a difference corneal fluorometer on the perfused whole cornea preparation and the perfused everted corneal preparation. Sodium azide (10 m m) for 30 min resulted in a 19·4 ± 0·7% increase in the pyridine nucleotide fluorescence from the whole corneal preparation and a 4·5 ± 0·6% increase from the everted endothelial preparation. Corneal anoxia induced by stopping the perfusion on the endothelial side resulted in a 18·7 ± 0·6% increase in pyridine nucleotide fluorescence for the whole corneal preparation. Sodium azide (10 m m) resulted in a 35% decrease in the transendothelial potential difference and a 76% decrease in the rate of transendothelial fluid transport. A comparison is made between invasive chemical analysis and real time, non-invasive fluorometry to measure histotoxic corneal anoxia.