Age-dependent neurodegeneration in forebrain-specific Atg7 -deficient mice. (a-b) Impaired macroautophagy in CamK-Atg7 cKO mouse forebrain tissues including hippocampus and cortex. a, ATG7 protein was significantly reduced in CamK-Atg7 cKO brains. Consistent with ATG7 change, mammalian Atg8 homologues (LC3, GABARAP and GABARAPL1) and macroautophagy substrate p62 were accumulated in CamK-Atg7 cKO brains. b, Poly-ubiquitinated proteins were accumulated in both 0.5% TritonX-100-soluble and insoluble fractions of CamK-Atg7 cKO forebrain. These results indicate that macroautophagy is impaired in the forebrains of CamK-Atg7 cKO mice. (c-d) Slow progressive loss of hippocampal CA1 pyramidal neurons in CamK-Atg7 cKO mice. c, Quantification of CA1 pyramidal neuron number. White bars, CamK-Atg7 cWT. Black bars, CamK-Atg7 cKO. n = 3 - 4 for each group. **, P < 0.01. d, Pyramidal neurons in the CA1 region of 8-month-old CamK-Atg7 cKO mice were cleaved caspase-3-positive. Bar, 40 μm. (e) Cytoplasmic inclusions in 6-month-old CamK-Atg7 cKO mice. Ubiquitin-positive (green) inclusions were present in MAP2-positive (red) hippocampal CA1 neurons of CamK-Atg7 cKO mice, but were never seen in control CamK-Atg7 cWT mice. Ubiquitin-positive inclusions were also positive for p62 (Additional file 1), as previously. Bar, 10 μm. (f) Electron microscopic analyses of cytoplasmic inclusions. The dashed squares outline two inclusions in the cytoplasm on neurons of Atg7 cKO mice. At higher magnification, these inclusions display fibrillar and vesicular components and lack an outer membrane. Inclusions were never observed in control cWT mice. Bar, 500 nm.