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Fig. 1 | Molecular Brain

Fig. 1

From: Mutation of copper binding sites on cellular prion protein abolishes its inhibitory action on NMDA receptors in mouse hippocampal neurons

Fig. 1

a NMDAR-mediated currents from hippocampal neuron cultures of PrPc knock-in Tga20 mice, PrPc knock-out PrPc KO mice versus wild-type C57. Neurons were held at − 60 mV throughout and currents were evoked by application of 500 µM NMDA and 1 μM glycine. The dashed lines indicate baseline, steady state current, and peak current, and the arrows indicate the magnitude of the non-desentitizing (steady state) current. b Glycine dose response curve of the percentage of steady-state current (normalized to peak) in wild-type C57, Tga20 and PrPc KO muse neurons (n = 5). Asterisks denote statistical significance for C57 vs Tga20, and number symbols indicate statistical significance between C57 and PrPc KO at the 0.05, 0.1 and 0.001 levels for one, two and three symbols, respectively (one way ANOVA with Bonferroni post hoc test). c Structure of PrPc illustrating the location of copper binding motifs in the unstructured N-terminal region (taken from [11]). d Illustration of constructs of recombinant AAV-GFP-PrPc and the various copper site mutants. e Confocal images of hippocampal neurons from PrPc KO mice transduced with AAV9 expressing eGFP (left), mPrPc (middle), and mPrPc-6HA (right). Green signal reflects eGFP fluorescence and thus PrPc expression. The primary hippocampal neurons were cultured for 3 days before transduction and confocal images were collected 8 days later. The dose was 1 × 1011 GC/ml for all constructs. Scale bar = 50 µm. f Representative traces of NMDAR currents from hippocampal neurons of PrPc KO mice infected with AAV-GFP, AAV-GFP-PrPc, and AAV-GFP-PrPc-6HA. Neurons were held at − 60 mV throughout and currents were evoked by application of 500 µM NMDA and 1 μM glycine g. Glycine dose response curve of the percentage of steady-state current (normalized to peak) in neurons from PrPc KO mice transduced with AAV-GFP, AAV-GFP-PrPc, AAV-GFP-PrPc-6HA, AAV-GFP-PrPc-4HA and AAV-GFP-PrPc-2HA (n = 6 for AAV-GFP and AAV-GFP-PrPc, n = 5 for AAV-GFP-PrPc-6HA, AAV-GFP-PrPc-4HA and AAV-GFP-PrPc-2HA). Asterisks refer to statistical difference relative to AAV-GFP-PrPc (One way ANOVA with Tamhane-Dunnett's Test [13]), with colour of the asterisks corresponding to the colour denoting the various PrPc mutant constructs

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