Fig. 3

C9orf72 immunohistochemistry reveals synaptic staining pattern with enrichment in hippocampal mossy fiber terminals. Immunohistochemistry with anti-C9orf72 mAb 1C1 (a-j); immunohistochemistry with anti-synaptoporin antibody (k). In the adult mouse brain (a-h), strong immunoreactivity for C9orf72 is seen in the hippocampal mossy fiber system (a) with labeling in the hilus (asterisk), stratum lucidum (arrow) and infrapyramidal mossy fiber bundles (arrowhead). (b) Higher magnification of punctate staining pattern of mossy fiber terminals in suprapyramidal (SPB) and infrapyramidal (IPB) mossy fiber bundles. Robust staining was also observed in the globus pallidus (GP) (c and d) while the caudate putamen (CPu) (c) and other gray matter regions showed weaker immunoreactivity of the neuropil as shown for frontal cortex (e and f) and cerebellum with predominant staining in the molecular and granular layer (g). No immunoreactivity is seen in the white matter and internal capsule (ic). In addition to punctate neuropil staining, neurons with large cytoplasm such as motor neurons in the spinal cord showed several cytoplasmic puncta (h). (i and j): Specificity of anti-C9orf72 immunohistochemistry was validated by the complete absence of immunoreactivity in brain sections from C9orf72 knock-out mice as shown for hippocampus (i) and cerebellum (j). Note the strikingly similar staining pattern of the mossy fiber terminals in the hippocampus for C9orf72 (a) and for the presynaptic marker protein synaptoporin (k). Scale bar: 533 μm (c); 400 μm (a, i, k); 267 μm (e); 80 μm (d, j, insert k); 40 μm (b, f, g); 20 μm (h); 6,5 μm (insert h)