Journal: eLife

Article Title: Cortico-striatal action control inherent of opponent cognitive-motivational styles

doi: 10.7554/eLife.100988

Figure Lengend Snippet: ( a ) Following initial handling and screening to identify the rat’s phenotype (17 goal trackers [GTs], 8 females; 12 sign-trackers [STs], 6 females; ), rats were infused with either a Cre-dependent inhibitory DREADD or the empty control vector (both expressing mCherry; b ) into the lower layers of the prelimbic cortex, and a retrogradely transported, Cre-expressing plasmid into the dorsomedial striatum (DMS) (expressing eGFP). These rats continued to undergo CTTT performance training, followed either by ( c ) a test of the effects of vehicle or CNO on CTTT performance (the blue syringes symbolizing CNO administration) or by ( d ) implantation of an microelectrode array for the measurement of glutamate concentrations in the DMS and following vehicle or CNO administration. Following recovery from that surgery, the effects of CNO and vehicle on performance and performance-associated glutamate concentrations were assessed. In GTs, administration of CNO significantly reduced the relative number of cued turns ( e ), but not cued stops ( f ; e and f show individual data, mean and 95% CI; post hoc multiple comparisons: ***: p < 0.001, Tukey’s Honest Significant Difference test). CNO had no effects on cued turns or stops in rats expressing an empty control vector (see Results). In GTs, CNO administration attenuated cue-locked maximum peak glutamate concentrations during (residual) cued turns ( g ) and decreased the number of traces with just one glutamate peak during the cue period while increasing the frequency of two peaks during this period ( h ). Following the administration of CNO, residual turns took significantly more time to be initiated in GTs when compared with vehicle-treated GTs and CNO-treated STs ( i ). When followed by failures to turn, or misses, turn cue-locked maximum peak concentrations were not affected in GTs but reduced in STs (significant interaction between the effects of phenotype and CNO; j ). Maximum peak glutamate concentrations locked to reward delivery again were higher in STs than in GTs, and CNO reduced these concentrations in both phenotypes (main effect of CNO, no interaction k ); (multiple comparisons: **, ***: p < 0.01, 0.001).

Article Snippet: Tukey’s Honest Significant Difference test was used to compare, post hoc, the effects of the first and second administration of CNO with the effects of the first and second administration of vehicle (GraphPad Prism).

Techniques: Control, Plasmid Preparation, Expressing, Microelectrode Array

Journal: Molecular Neurodegeneration

Article Title: Quantitative proteomic analysis of Parkin substrates in Drosophila neurons

doi: 10.1186/s13024-017-0170-3

Figure Lengend Snippet: Human Parkin and VPS35 interaction: VPS35 is ubiquitinated by wild type Parkin in human cells. a VPS35 is ubiquitinated by untagged WT Parkin in SH-SY5Y cells. YFP-VPS35 showed a significant increase in its ubiquitinated fraction when it is over-expressed with wild type untagged hParkin (WT), as shown with anti-FLAG (to FLAG-Ub) antibody Western blot ( red ) compared to control (C) or the single point (C431S) mutated hParkin (LD). The non-modified form of VPS35 was detected by GFP antibody ( green ). The bottom panel shows levels of Parkin protein in the whole cell extract before the isolation of the GFP-tagged proteins. The endogenous Parkin protein is barely detected. b Quantification of the ubiquitination status of VPS35 relative to the non-modified form was performed calculating the ratio FLAG:GFP with Image-J. The plot shows relative levels of VPS35 ubiquitination normalized to the GFP levels. Statistical significance in Western blotting semi-quantification was evaluated using an analysis of variance (ANOVA) complemented by Tukey’s honest significance difference test (Tukey’s HSD) performed in GraphPad PRISM software. Statistical significance differences (**, p < 0.01 (mean ± SEM, n = 4)) were observed for the WT Parkin sample relative to both control and LD Parkin samples. For LD Parkin sample ns indicates not significant differences relative to the control sample. c VPS35 knockdown does not affect Parkin - dependent mitophagy. hTERT-RPE1 cells stably over-expressing YFP-Parkin were subjected to control and VPS35 siRNA for 72 h or 165 h and treated with 10 μM CCCP for 2, 8 and 24 h. Samples were immunoblotted as indicated. High exp: high exposure

Article Snippet: Statistical significance in Western blotting quantification was evaluated using an analysis of variance (ANOVA) complemented by Tukey’s honest significance difference test (Tukey’s HSD) performed in GraphPad PRISM software.

Techniques: Western Blot, Control, Modification, Isolation, Ubiquitin Proteomics, Software, Knockdown, Stable Transfection, Expressing