Journal: International Journal of Molecular Sciences

Article Title: Traumatic Brain Injury Induces Nociceptin/Orphanin FQ and Nociceptin Opioid Peptide Receptor Expression within 24 Hours

doi: 10.3390/ijms25031658

Figure Lengend Snippet: NOP receptor protein expression was unchanged ( a ) but mRNA expression increased ( b ) 24 h after ModTBI compared to sham and mTBI groups. ( a ) NOP expression was determined as described in Methods. Representative blots are shown below the graph. Kruskal–Wallis test indicates a significant effect of injury ( a ; p = 0.0249), but no differences between groups were found. Values are presented as mean ± SD ( n = 6 per group; * p < 0.05). ( b ) NOP receptor gene expression from all groups was normalized to GAPDH and fold change in mRNA compared to sham was determined as described in Methods. Values are presented as mean ± SEM and compared using one-way ANOVA with Tukey’s multiple comparisons test.

Article Snippet: The protein concentration of the supernatants was determined as described above, prior to solubilization in 4X sample loading buffer (LI-COR Biosciences, Lincoln, NE, USA) heated to 65 °C for 20 min. Soluble protein samples (20 µg of total protein) were resolved by Novex™ WedgeWell™ 8–16% Tris-Glycine gradient gels (Thermo Fisher Scientific Inc., Waltham, MA, USA), transferred to nitrocellulose membranes, and probed with antibodies directed against the following proteins: NOP receptor (bs-0181R, 1:500; Bioss Inc., Woburn, MA, USA), GFAP (GPCA-GFAP, 1:4000; EnCor Biotechnology, Gainesville, FL, USA), UCH-L1 (sc-271639, 1:200), NF-L (sc-20012, 1:200), and cofilin (sc-376476, 1:200) from Santa Cruz Biotechnology, Inc., Dallas, TX, USA; ERK1/2 (4696S, 1:2000) and phospho-ERK1/2 (4370S, 1:500 and phospho-cofilin (Ser3) (#3313S; 1:250) from Cell Signaling Technology, Inc. (Danvers, MA, USA); and actin (A3853, 1:2000; Sigma-Aldrich, Inc, St. Louis, MO, USA).

Techniques: Expressing

Journal: International Journal of Molecular Sciences

Article Title: Traumatic Brain Injury Induces Nociceptin/Orphanin FQ and Nociceptin Opioid Peptide Receptor Expression within 24 Hours

doi: 10.3390/ijms25031658

Figure Lengend Snippet: Schematic representation of CCI TBI-induced changes in N/OFQ and NOP receptor protein and mRNA as a function of injury severity and time. The color of the arrows matches the color of the data points on graphs for mild (light blue) and moderate (dark blue) TBI; ↑ arrows represent increase, ↓ arrows represent decrease, and ↔ represents no significant change compared to sham. The width of the arrow represents the size of the change in NOP or N/OFQ levels. Dashed arrows represent mRNA, while solid arrows represent protein or peptide levels. * represents change from contralateral (undamaged) tissue. Data noted in the 3 h and 8-day time points are from recent published publications; the day 1 data are from the current study on WT rats. This figure was created in BioRender.com (accessed on 22 January 2024).

Article Snippet: The protein concentration of the supernatants was determined as described above, prior to solubilization in 4X sample loading buffer (LI-COR Biosciences, Lincoln, NE, USA) heated to 65 °C for 20 min. Soluble protein samples (20 µg of total protein) were resolved by Novex™ WedgeWell™ 8–16% Tris-Glycine gradient gels (Thermo Fisher Scientific Inc., Waltham, MA, USA), transferred to nitrocellulose membranes, and probed with antibodies directed against the following proteins: NOP receptor (bs-0181R, 1:500; Bioss Inc., Woburn, MA, USA), GFAP (GPCA-GFAP, 1:4000; EnCor Biotechnology, Gainesville, FL, USA), UCH-L1 (sc-271639, 1:200), NF-L (sc-20012, 1:200), and cofilin (sc-376476, 1:200) from Santa Cruz Biotechnology, Inc., Dallas, TX, USA; ERK1/2 (4696S, 1:2000) and phospho-ERK1/2 (4370S, 1:500 and phospho-cofilin (Ser3) (#3313S; 1:250) from Cell Signaling Technology, Inc. (Danvers, MA, USA); and actin (A3853, 1:2000; Sigma-Aldrich, Inc, St. Louis, MO, USA).

Techniques:

Journal: Neuropharmacology

Article Title: Pharmacological blockage of NOP receptors decreases ventral tegmental area dopamine neuronal activity through GABA B receptor-mediated mechanism

doi: 10.1016/j.neuropharm.2024.109866

Figure Lengend Snippet: (A-E) The graphs show representative and averaged traces of the spontaneous firing from putative VTA DA neuron before, during, and after LY2817412 (1 μM) slice treatment in (A) absence, and in presence of (B) picrotoxin (100 μM) + CGP55845 (1 μM), (C) picrotoxin (100 μM), (D) CGP55845 (1 μM), or (E) naloxone (10 μM). Scale bar: 1 s. (F) Bar graphs represent the averaged maximal effects of LY2817412 on the firing rate of VTA DA neurons in the same experimental conditions. *** p < 0.001, ### p < 0.001.

Article Snippet: The NOP receptor antagonist LY2817412 was synthesized and kindly provided by Eli Lilly (IN, USA; [ 66 ]).

Techniques:

Journal: Neuropharmacology

Article Title: Pharmacological blockage of NOP receptors decreases ventral tegmental area dopamine neuronal activity through GABA B receptor-mediated mechanism

doi: 10.1016/j.neuropharm.2024.109866

Figure Lengend Snippet: (A) Representative traces of spontaneous firing activity recorded in cell-attached configuration from a putative VTA DA neuron before (Baseline), during (LY2817412 1 μM), and after (Wash out) LY2817412 slice perfusion. Scale bar: 2 s. (B) The bar graph shows the temporal changes of firing rate (Hz) of the same representative putative VTA DA cell before, during and after LY2817412 slice perfusion. (C) The graph reports the concentration-dependent effect of LY2817412 on putative VTA DA neuronal firing rate. Note that, except for the 10−2 nM concentration, only data from LY2817412-responding cells are reported in this graph.

Article Snippet: The NOP receptor antagonist LY2817412 was synthesized and kindly provided by Eli Lilly (IN, USA; [ 66 ]).

Techniques: Activity Assay, Concentration Assay

Journal: Neuropharmacology

Article Title: Pharmacological blockage of NOP receptors decreases ventral tegmental area dopamine neuronal activity through GABA B receptor-mediated mechanism

doi: 10.1016/j.neuropharm.2024.109866

Figure Lengend Snippet: (A) Representative sIPSCs traces recorded from putative VTA DA neurons before (Baseline), during LY2817412 (1 μM), and during picrotoxin (100 μM) slice perfusion. Scale bars: 50 pA × 2 s. (B-C) Bar graphs showing the effects of LY2817412 (1 μM) on frequency (D) and amplitude (E) of sIPSCs. (D). Representative mIPSCs traces recorded from putative VTA DA neurons before (vehicle), during TTX (1 μM), TTX + LY2817412 (1 μM), and during TTX + LY2817412 + picrotoxin (100 μM) slice perfusion. Scale bars: 50 pA × 2 s. (E-F) Bar graphs showing the effects of LY2817412 (1 μM) on frequency (D) and amplitude (E) of mIPSCs.

Article Snippet: The NOP receptor antagonist LY2817412 was synthesized and kindly provided by Eli Lilly (IN, USA; [ 66 ]).

Techniques: