Figure 3. Standardization of the lactate assay in neurons and astrocytes in slices of inferior colliculus from adult rat brain.

(a) Impaling a single cell with a micropipette containing known concentrations of resorufin gives a stable fluorescence response with repeated sampling with 100 ms exposures at 30 s intervals (n=3/group). (b) The linear ranges of standard curves were identified by diffusing known concentrations of resorufin into single neurons and astrocytes (n = 5/data point); the equation of the linear regression line is y = 28.8x + 14.1. The inset shows the non-linear fluorescence response of higher resorufin concentrations. (c) Area labeled by co-diffusion of Lucifer yellow plus resorufin into single astrocytes (n = 5/group; paired t test). (d) Net increase in fluorescence (ΔF) due to lactate uptake from extracellular fluid into astrocytes and neurons located at the indicated distances from the tip of the micropipette containing L-lactate (10 mmol/L). P values for indicated comparisons were determined by ANOVA and Tukey’s test (n=5 for astrocytes and neurons at 2 µm, n=4 for astrocytes at 50 µm). Values are means and vertical bars represent ±1SD; if not visible, SDs are smaller than the symbol.