Fig. 9
Depletion of glycogen within glial end-feet surrounding microvessels contributes to microvessel constrictions. a-f When microvessel constrictions emerged 1 hour after ischemia (n = 3 retinae), glycogen levels (detected by PAS staining) in microvascular glial end-feet (microvessels were detected by lectin staining, a, d; arrows in b, c, e) were significantly decreased (d-e) compared to the non-ischemic eye (n = 3 retinae) (a-c). PAS staining was performed after treatment with dimedone to block aldehyde groups on non-glycogen substances. The number of microvessel constrictions was highly correlated with low levels of end-feet glycogen during ischemia (Pearson product-moment correlation, R2 = 0.992; P < 0.001) (f). g and h Inhibition of glycogen utilization by DAB exacerbated the ischemia-induced constrictions so that they appeared as early as 30 min after ischemia (arrows, g). h The number of microvascular constrictions significantly increased 30 min after ischemia in DAB treated-retinae compared to DAB treatment or 30-min ischemia alone (DAB: n = 3 retinae; 30 min-ischemia: n = 3 retinae; 30-min-ischemia and DAB: n = 3 retinae; P < 0.001, ANOVA and Tukey’s test). Scale bars in a-e, g = 10 μm