Figure 6.

Induction of autophagy, axonal degeneration, and cell death in neurons by GluD2^Lc and various mutants. A, Representative fluorescent images of EGFP-LC3 and mCherry in neurons. Cultured hippocampal neurons at 12 DIV were transfected with cDNAs encoding EGFP-LC3, mCherry, and one of GluD2^wt, GluD2^Lc, GluD2^Lc-Q618R, GluD2^Lc-V617R, GluD2^wt-ΔCT7, or GluD2^Lc-ΔCT7 and then analyzed after 15 h. Note that EGFP-LC3 clusters were observed in the cell bodies of neurons expressing GluD2^Lc, GluD2^Lc-Q618R, or GluD2^Lc-ΔCT7. B, Survival of neurons expressing GluD2^Lc and various mutants. At 15 h after transfection, the number of mCherry-positive neurons was counted and normalized by that in neurons expressing GluD2^wt. Error bars indicate the mean + SEM (from 5 independent experiments). C, Representative fluorescent images of axons of hippocampal neurons expressing GluD2^Lc, GluD2^Lc-Q618R, or GluD2^Lc-ΔCT7. Cultured hippocampal neurons were transfected with cDNAs encoding mCherry and GluD2^wt or GluD2 mutants as indicated, and then stained for the dendritic marker MAP2 (green) and the axonal marker tau-1 (blue). The regions indicated by the white boxes are enlarged in the right panels. Note that marked focal swellings were observed along the MAP2-negative and tau-1-positive axons of hippocampal neurons expressing GluD2^Lc, GluD2^Lc-Q618R, and GluD2^Lc-ΔCT7. White arrows and arrowheads indicate the axons. Scale bars: A, 10 μm; C, 100 μm. **p < 0.01.