Evaluation of hexose monophosphate shunt activity in isolated murine lens by monitoring the potential of the ferricyanide--ferrocyanide system.

Abstract

A method for evaluation of the activity of the hexose monophosphate shunt (HMS) in isolated lens is presented. The measurement of HMS activity is based on continuous monitoring of the potential of the ferricyanide--ferrocyanide system (where ferricyanide is an artificial electron acceptor) in the presence of a lens. The rate of reduction of ferricyanide increased after the addition of methylene blue (MB) or saponin. The ferricyanide reduction rate did not correlate with GSH content in the contralateral lenses of the same mouse in the absence of MB or saponin. Correlations between the ferricyanide reduction rate and GSH content in the lens were 0.67 (beta = 0.93) in the presence of MB and 0.82 (beta = 0.95) in the presence of saponin. We think that the measured curves of ferricyanide reduction are representative of: 1) normal level of HMS activity (in the absence of methylene blue and saponin); 2) maximal HMS activity (in the presence of methylene blue); 3) the intracellular concentration of reducing equivalents (in the presence of saponin).