Increased degradation of glutamate receptors in ALS2-/- neurons following AMPA treatment. A. The biotinylated internalized cell surface proteins were affinity purified and immunoblotted with antibodies against glutamate receptors: GluR1, GluR2, GluR2/3, and NR1 (labeled as "internalized"). The neurons were treated with mock (AMPA-) or AMPA (AMPA+) prior to the biotinylation. The expression of each glutamate receptor and β-tubulin in total cell lysate (labeled as "total") was also blotted to normalize the accumulation of internalized proteins. B. Similar to A, except that neurons were pre-incubated with lysosome inhibitor leupeptin. C. Histogram reveals a significant decrease of internalized glutamate receptors in KO neurons as compared with WT controls (n > 6). Error bars represent SEM. ** P < 0.01. D. Histogram shows that in the presence of leupeptin, no significant alteration of internalized glutamate receptors was observed between WT and KO neurons (n > 6). Error bars represent SEM.