Extended Data Figure 6. Metabolic inhibition in astrocyte causes neuronal cell death and retards neurite outgrowth in vitro.

a, Astrocytic aconitase was inhibited by fluorocitrate (FC) which disrupted astrocyte metabolism that was accompanied by SA-β-gal signal. b, Intracellular ATP was decreased in these metabolically-disrupted astrocytes (n=6). *P<0.05, **P<0.01 vs FC 0 mM. c, PI staining showed that fluorocitrate (0.5 mM) did not induce cell death in astrocytes. d, Metabolically-disrupted astrocytes significantly decreased mitochondrial membrane potential. Red: aggregated JC1, Green: monomer JC1. Scale: 20 μm. e, Rat cortical neurons were co-cultured with JC1-labeled astrocytes. After 24 hours co-culture, control astrocytes transferred mitochondria which had a high-membrane potential (aggregated JC1), but metabolically-disrupted astrocytes released and transferred dysfunctional mitochondria into neurons (n=3). f, Metabolically-disrupted astrocytes could not support neural viability under starvation in the co-culture (n=4). g, Co-culture between astrocytes and neurons was conducted for 48 hours to test neurite outgrowth. Immunocytochemistry showed that metabolically-disrupted astrocytes retarded neurite outgrowth and increased neuronal cell death (n=3). h, LDH assay indicated that fluorocitrate (0.5 mM) did not affect cell viability in either rat cortical astrocytes (n=4) or rat cortical neurons (n=4). All values are mean +/− SEM.