Distribution of intracellular pH in the rat brain cortex after global ischemia as measured by color film histophotometry of neutral red

Tissue acidosis is an important determinant of cell viability following cerebral ischemia. Because of the heterogeneity of tissue response to metabolic stress, a method for measuring intracellular pH (pH i) that preserves spatil information would be desirable. Histophotometry of the pH indicator dye Neutral red offers such a possibility. The purpose of our study was to determine the distribution of pH i following complete irreversible ischemia and show the correlation of mean pH i measured by Neutral red and [ 31P]NMR in the same brain. Three rats were studied in the anesthetized state. A pH i range was obtained by total cerebral ischemia at various pre-arrest plasma glucose concentrations. The data show that mean pH i calculated by Neutral red was strongly correlated to pH i determined from [ 31P]NMR (slope:0.99 ± 0.08; P < 0.001, r 2 = 0.96). Within each brain, 80–110 discrete samples were analyzed by histophotometry. The pH i distribution of those samples broadened in those rat brains with greater acidosis, suggesting a heterogeneity of response by the tissue to ischemia and the presence of multiple pH i pools. Our results demonstrate the need to use methods which maintain spatial resolution such as is available with histophotometry.