nmbr  (Alomone Labs)

Journal: Biochemical pharmacology

Article Title: Functional roles of neuromedin B and gastrin-releasing peptide in regulating itch and pain in the spinal cord of non-human primates

doi: 10.1016/j.bcp.2022.114972

Figure Lengend Snippet: A comparison of mRNA expression of GRP versus NMB, and GRPR versus NMBR, in the dorsal root ganglia and spinal dorsal horn of monkeys. mRNA levels were quantified using RT-qPCR and analyzed by the comparative CT method. (A) PCR amplification efficiency for each primer (NMB, GRP, NMBR, GRPR, and ACTB) was determined by the slope of standard curve generated from tenfold serial dilutions (0.02–20 ng) of the cDNA mixture from subjects. (B-E) Relative mRNA expression levels in the dorsal root ganglia (B, C) and spinal dorsal horn (D, E). Data represent the mean ± SD (n = 5), with dots representing individual NHP. Data were analyzed by the paired t-test. *p < 0.05, significantly different between GRP and NMB (B, D) or GRPR and NMBR (C, E).

Article Snippet: The tissue sections were washed in PBS containing 0.3% Triton X-100 (Sigma-Aldrich) and treated with TrueBlack Lipofusin Autofluorescence Quencher (Biotium, Fremont, CA, USA) for 2 min. After washing with PBS, we blocked the tissues with PBS containing 3% normal donkey serum (Jackson ImmunoResearch Laboratories, West Grove, PA, USA) at room temperature for 1 h. The sections were incubated with a primary antibody against GRPR (rabbit polyclonal, 1:500 dilution; MC-831, MBL International, Woburn, MA, USA), NMBR (rabbit polyclonal, 1:500 dilution; ABR-004, Alomone Labs, Jerusalem, Israel), and NeuN (mouse monoclonal, 1:500 dilution; MAB377, MilliporeSigma, Burlington, MA, USA; markers for neuronal nuclei) at 4 °C overnight.

Techniques: Comparison, Expressing, Quantitative RT-PCR, Amplification, Generated

Journal: Biochemical pharmacology

Article Title: Functional roles of neuromedin B and gastrin-releasing peptide in regulating itch and pain in the spinal cord of non-human primates

doi: 10.1016/j.bcp.2022.114972

Figure Lengend Snippet: Effects of intrathecal NMB and GRP on itch scratching responses in monkeys. (A, B) Time courses of itch scratching responses elicited by varying doses of NMB and GRP. (C) Total number of scratches summed from the four time points displayed in A and B. (D, E) Time courses of itch scratching responses elicited by NMB (100 nmol) or GRP (10 nmol), in combination with the GRPR antagonist RC-3095 (100 nmol) or NMBR antagonist PD168368 (300 nmol). Data represent the mean ± SD (n = 6), with dots representing individual NHP. Data were analyzed by the two- (A, B, D, and E) or one-way (C) repeated measures ANOVA, followed by Dunnett’s multiple comparison test. *p < 0.05, significantly different from the vehicle condition.

Article Snippet: The tissue sections were washed in PBS containing 0.3% Triton X-100 (Sigma-Aldrich) and treated with TrueBlack Lipofusin Autofluorescence Quencher (Biotium, Fremont, CA, USA) for 2 min. After washing with PBS, we blocked the tissues with PBS containing 3% normal donkey serum (Jackson ImmunoResearch Laboratories, West Grove, PA, USA) at room temperature for 1 h. The sections were incubated with a primary antibody against GRPR (rabbit polyclonal, 1:500 dilution; MC-831, MBL International, Woburn, MA, USA), NMBR (rabbit polyclonal, 1:500 dilution; ABR-004, Alomone Labs, Jerusalem, Israel), and NeuN (mouse monoclonal, 1:500 dilution; MAB377, MilliporeSigma, Burlington, MA, USA; markers for neuronal nuclei) at 4 °C overnight.

Techniques: Comparison

Journal: Biochemical pharmacology

Article Title: Functional roles of neuromedin B and gastrin-releasing peptide in regulating itch and pain in the spinal cord of non-human primates

doi: 10.1016/j.bcp.2022.114972

Figure Lengend Snippet: Effects of intrathecal NMBR and GRPR antagonists on peripherally elicited pain, compared with DAMGO ([D-Ala2,N-MePhe4,Gly-ol5]-enkephalin) in monkeys. Topical capsaicin-induced allodynia was evaluated by tail-withdrawal assays in 46 °C water following the intrathecal administration of μ-opioid receptor agonist DAMGO (1 and 3 nmol), GRPR antagonist RC-3095 (100 and 300 nmol), or NMBR antagonist PD168368 (300 and 1000 nmol). Data represent the mean ± SD (n = 5), and were analyzed via a one-way repeated measures ANOVA, followed by Dunnett’s multiple comparison test. *p < 0.05, significantly different from the vehicle condition.

Article Snippet: The tissue sections were washed in PBS containing 0.3% Triton X-100 (Sigma-Aldrich) and treated with TrueBlack Lipofusin Autofluorescence Quencher (Biotium, Fremont, CA, USA) for 2 min. After washing with PBS, we blocked the tissues with PBS containing 3% normal donkey serum (Jackson ImmunoResearch Laboratories, West Grove, PA, USA) at room temperature for 1 h. The sections were incubated with a primary antibody against GRPR (rabbit polyclonal, 1:500 dilution; MC-831, MBL International, Woburn, MA, USA), NMBR (rabbit polyclonal, 1:500 dilution; ABR-004, Alomone Labs, Jerusalem, Israel), and NeuN (mouse monoclonal, 1:500 dilution; MAB377, MilliporeSigma, Burlington, MA, USA; markers for neuronal nuclei) at 4 °C overnight.

Techniques: Comparison

Journal: Biochemical pharmacology

Article Title: Functional roles of neuromedin B and gastrin-releasing peptide in regulating itch and pain in the spinal cord of non-human primates

doi: 10.1016/j.bcp.2022.114972

Figure Lengend Snippet: Expression of NMBR and GRPR protein in the spinal dorsal horn of monkeys. (A, B) Representative images of immunohistochemistry depict the expression of GRPR (left column) and NMBR (right column) in NeuN+ cells. (C) Higher magnification of the boxed areas in (B). Scale bar = 200 μm (A, B), and 50 μm (C).

Article Snippet: The tissue sections were washed in PBS containing 0.3% Triton X-100 (Sigma-Aldrich) and treated with TrueBlack Lipofusin Autofluorescence Quencher (Biotium, Fremont, CA, USA) for 2 min. After washing with PBS, we blocked the tissues with PBS containing 3% normal donkey serum (Jackson ImmunoResearch Laboratories, West Grove, PA, USA) at room temperature for 1 h. The sections were incubated with a primary antibody against GRPR (rabbit polyclonal, 1:500 dilution; MC-831, MBL International, Woburn, MA, USA), NMBR (rabbit polyclonal, 1:500 dilution; ABR-004, Alomone Labs, Jerusalem, Israel), and NeuN (mouse monoclonal, 1:500 dilution; MAB377, MilliporeSigma, Burlington, MA, USA; markers for neuronal nuclei) at 4 °C overnight.

Techniques: Expressing, Immunohistochemistry

Journal: Biochemical pharmacology

Article Title: Functional roles of neuromedin B and gastrin-releasing peptide in regulating itch and pain in the spinal cord of non-human primates

doi: 10.1016/j.bcp.2022.114972

Figure Lengend Snippet: Expression of NMBR and GRPR mRNA in the spinal dorsal horn of monkeys (A-C). Representative confocal images of in situ hybridization (RNAscope) depict the mRNA expression of NMBR in GRPR+ (A) or PKCG+ neurons (B) and the mRNA expression of GRPR in PKCG+ neurons (C). (D) Quantification of the distribution of NMBR+ and GRPR+ neurons in laminae I-III and IV-V of the spinal dorsal horn. (E, F) Bar graphs depict the percentage of co-expression. Data represent the mean ± SD (n = 3), with dots representing individual NHP. Scale bar = 20 μm.

Article Snippet: The tissue sections were washed in PBS containing 0.3% Triton X-100 (Sigma-Aldrich) and treated with TrueBlack Lipofusin Autofluorescence Quencher (Biotium, Fremont, CA, USA) for 2 min. After washing with PBS, we blocked the tissues with PBS containing 3% normal donkey serum (Jackson ImmunoResearch Laboratories, West Grove, PA, USA) at room temperature for 1 h. The sections were incubated with a primary antibody against GRPR (rabbit polyclonal, 1:500 dilution; MC-831, MBL International, Woburn, MA, USA), NMBR (rabbit polyclonal, 1:500 dilution; ABR-004, Alomone Labs, Jerusalem, Israel), and NeuN (mouse monoclonal, 1:500 dilution; MAB377, MilliporeSigma, Burlington, MA, USA; markers for neuronal nuclei) at 4 °C overnight.

Techniques: Expressing, In Situ Hybridization, RNAscope

Journal: Biochemical pharmacology

Article Title: Functional roles of neuromedin B and gastrin-releasing peptide in regulating itch and pain in the spinal cord of non-human primates

doi: 10.1016/j.bcp.2022.114972

Figure Lengend Snippet: Expression of VGLUT2 and VGAT mRNA in NMBR+ and GRPR+ neurons in the spinal dorsal horn of monkeys. (A-D) Representative confocal images of in situ hybridization (RNAscope) show the mRNA expression of VGLUT2 in NMBR+ (A) or GRPR+ (B) neurons and the mRNA expression of VGAT in NMBR+ (C) or GRPR+ (D) neurons. (E, F) Bar graphs show the percentage of co-expression. Data represent the mean ± SD (n = 3) with dots representing individual NHP. Scale bar = 20 μm.

Article Snippet: The tissue sections were washed in PBS containing 0.3% Triton X-100 (Sigma-Aldrich) and treated with TrueBlack Lipofusin Autofluorescence Quencher (Biotium, Fremont, CA, USA) for 2 min. After washing with PBS, we blocked the tissues with PBS containing 3% normal donkey serum (Jackson ImmunoResearch Laboratories, West Grove, PA, USA) at room temperature for 1 h. The sections were incubated with a primary antibody against GRPR (rabbit polyclonal, 1:500 dilution; MC-831, MBL International, Woburn, MA, USA), NMBR (rabbit polyclonal, 1:500 dilution; ABR-004, Alomone Labs, Jerusalem, Israel), and NeuN (mouse monoclonal, 1:500 dilution; MAB377, MilliporeSigma, Burlington, MA, USA; markers for neuronal nuclei) at 4 °C overnight.

Techniques: Expressing, In Situ Hybridization, RNAscope