Journal: Frontiers in Pharmacology

Article Title: Activation of Dopamine D1-D2 Receptor Complex Attenuates Cocaine Reward and Reinstatement of Cocaine-Seeking through Inhibition of DARPP-32, ERK, and ΔFosB

doi: 10.3389/fphar.2017.00924

Figure Lengend Snippet: The effects of D1-D2 heteromer stimulation and inactivation on basal conditioned place preference. (A) Vehicle-conditioned rats did not exhibit a preference toward a particular chamber. D1-D2 heteromer stimulation by SKF 83959 (1.5 mg/kg, s.c.) induced conditioned place aversion (CPA) as the animals spent significantly less time in the drug paired chamber. (B) SKF 83959-induced CPA was abolished by pre-treatment by the D1-D2 heteromer selective disrupting peptide, TAT-D1, but not the control TAT-Sc peptide. (C) Inactivation of D1-D2 heteromer by TAT-D1 resulted in conditioned place preference (CPP) as the rats spent significantly more time in the drug paired chamber, not observed with the control TAT-Sc. (D) Representative western blots (inset) and histogram showing the amount of D1R co-immunoprecipitated with D2R from the NAc of rats treated with saline or SKF 83959. Pretreatment with TAT-D1 led to decreased co-immunoprecipitated receptors. An aliquot of each sample was used as a control for WB (input control). (E,F) The CPA induced by D1-D2 heteromer stimulation was abolished by Cdk5 inhibitor roscovitine pre-treatment (200 nmol, i.c.v, E ) or intra-accumbal injection (30 nmol, F ). (G) Representative western blot and histogram showing the density of Thr75-DARPP-32 phosphorylation (pT75) relative to GAPDH (as loading control). Data represent means ± SEM of n = 8–10 rats/group. ( * p < 0.05, ** p < 0.01: compared to saline).

Article Snippet: Protein homogenates (300 μg /each condition) from rat NAc were incubated with an anti-D2R antibody (Rabbit, Alomone) at 4°C overnight under gentle rotation.

Techniques: Conditioned Place Preference, Western Blot, Immunoprecipitation, Injection

Journal: Frontiers in Pharmacology

Article Title: Activation of Dopamine D1-D2 Receptor Complex Attenuates Cocaine Reward and Reinstatement of Cocaine-Seeking through Inhibition of DARPP-32, ERK, and ΔFosB

doi: 10.3389/fphar.2017.00924

Figure Lengend Snippet: Activation of Thr75-DARRP-32 by D1-D2 heteromer in rat NAc. (A,B) Rats ( n = 8/group) were injected with saline or SKF 83959 (1.5 mg/kg, s.c.), sacrificed 15, 45, or 90 min later, and phospho-Thr34-DARPP-32 (pT34) or phospho-Thr75-DARPP-32 (pT75) analyzed by western blot with GAPDH as loading control. (A) Representative blots of pT34 and pT75. (B) Quantification of blots from all animals represented as % mean ± SEM of control (saline values), ( ** p < 0.05). (C–F) Rats were injected with saline or SKF 83959 (1.5 mg/kg), sacrificed 15 min later, and immunohistochemistry performed using anti-pT34 or anti-pT75 assessed in the three types of MSNs: D1R-only (red arrow), Enk-only (D2R, green arrow) or D1R and ENK (D1-D2 heteromer)-expressing neurons (yellow arrow). (C) Representative confocal images of pT75-DARPP-32. (D) Quantification of pT75-DARPP-32 fluorescence in MSNs ( n = numbers of neurons from at least N = 3 rats/condition). (E) Representative confocal images of pT34-DARPP-32. (F) Quantification of pT34 fluorescence in MSNs. Data represents means ± SEM after removal of non-specific background. ( ** p < 0.01).

Article Snippet: Protein homogenates (300 μg /each condition) from rat NAc were incubated with an anti-D2R antibody (Rabbit, Alomone) at 4°C overnight under gentle rotation.

Techniques: Activation Assay, Injection, Western Blot, Immunohistochemistry, Expressing, Fluorescence

Journal: Frontiers in Pharmacology

Article Title: Activation of Dopamine D1-D2 Receptor Complex Attenuates Cocaine Reward and Reinstatement of Cocaine-Seeking through Inhibition of DARPP-32, ERK, and ΔFosB

doi: 10.3389/fphar.2017.00924

Figure Lengend Snippet: Signaling pathways involved in D1-D2 heteromer modulation of cocaine-induced behaviors: DARPP-32, ERK and ΔFosB. (A) Representative immunoblots of pT34-DARPP-32 (top panel) or pT75-DARPP-32 (lower panel) obtained from NAc of rats conditioned with saline or cocaine (10 mg/kg, i.p.) and injected on the test day with saline or SKF 83959. Loading controls (GAPDH) are shown. Quantification of pT34- and pT75-DARPP-32 immunoblots is shown. (B) Representative immunoblot of pERK44/42 obtained from the NAc of rats conditioned as in (A) is shown. Quantification of pERK44/42 immunoblots obtained from all animals is shown. Results in (A,B) represent the mean ± SEM from 8 to 9 rats/condition. * p < 0.05 and ** p < 0.01 represent significant differences from control. (C) Representative confocal images of pERK assessed in the three types of MSNs: D1R-only (red arrow), Enk-only (D2R, green arrow) or D1R and ENK (D1-D2 heteromer)-expressing neurons (yellow arrow) in MSNs from NAc of saline- or SKF 83959-treated rats. (D) Quantification of pERK fluorescence in MSNs in the NAc. Results are the mean ± SEM of data after removing the non-specific background ( n = number of MSNs from N = at least 3 rats/condition). ( ** p < 0.001; *** p < 0.0001). (E) Representative immunohistochemistry images and their quantification obtained using an antibody against ΔFosB and a secondary antibody conjugated to Alexa-488. Nuclei are stained by DAPI. Rats were treated for 7 days with cocaine (10 mg/kg, i.p.) without or with co-injection of SKF 83959 (1 mg/kg, s.c.). Disrupting the heteromer by repeated injections of TAT-D1 had the same effect as repeated injections of cocaine. Results are means ± SD obtained by the analysis of n = 1,500–1,700 neurons from the NAc of N = 3 rats/condition.

Article Snippet: Protein homogenates (300 μg /each condition) from rat NAc were incubated with an anti-D2R antibody (Rabbit, Alomone) at 4°C overnight under gentle rotation.

Techniques: Western Blot, Injection, Expressing, Fluorescence, Immunohistochemistry, Staining

Journal: Frontiers in Pharmacology

Article Title: Activation of Dopamine D1-D2 Receptor Complex Attenuates Cocaine Reward and Reinstatement of Cocaine-Seeking through Inhibition of DARPP-32, ERK, and ΔFosB

doi: 10.3389/fphar.2017.00924

Figure Lengend Snippet: Evidence for the existence of dopamine D1-D2 receptor heteromer in rat NAc. (A) Proximity ligation assay (PLA) was used to visualize and detect D1R and D2R close proximity. (A1) A scheme depicts the PLA probes used in the present study. (A2–A4) Representative images of PLA signals (red dots) in neurons (nuclei stained by DAPI) in rat caudate putamen (CPu), nucleus accumbens core (NAc-core) and shell (NAc-shell) subregions. (A5) Graph representing the percent of neurons with a positive PLA signal. (A6) Representative image of PLA signals in neurons (nuclei stained by DAPI) in rat NAc-core using the second set of antibodies. (B) Representative images of immunohistochemistry using D1R antibody (D1R-Ab) or D2R antibody (D2R-Ab) directly conjugated to Alexa-488 or Alexa-568, respectively, in the NAc-shell. Direct confocal FRET analysis was performed, reflected by FRET efficiency (FRET E) and the distance between the dipoles, less than 10 nm (100 Å). (C) A representative close-up of a single MSN cell body from NAc showing D1R-D2R colocalization (left), D1-D2 heteromer FRET efficiency (center) and relative distance between receptors (right). (D,E) Histograms showing FRET E ratios (D) and distance (E) obtained from MSN cell bodies from NAc ( n = 24). Bars are 10 μm.

Article Snippet: Protein homogenates (300 μg /each condition) from rat NAc were incubated with an anti-D2R antibody (Rabbit, Alomone) at 4°C overnight under gentle rotation.

Techniques: Proximity Ligation Assay, Staining, Immunohistochemistry

Journal: International Journal of Molecular Sciences

Article Title: Expression Profiles of Dopamine-Related Genes and miRNAs Regulating Their Expression in Breast Cancer

doi: 10.3390/ijms25126546

Figure Lengend Snippet: Venn diagram. DRD2 , dopamine receptor 2; DRD3 , dopamine receptor 3; DRD5 , dopamine receptor 5; TGF-β-2 , transforming growth factor beta 2; CAV2 , caveolin 2; SLC22A1 , solute carrier family 22 (organic cation transporter), member 1; CXCL1 2, chemokine (C-X-C motif) ligand 12; DRD1 , dopamine receptor 1; NR4A2 , nuclear receptor subfamily 4, group A, member 2; HRH2 , histamine receptor H2; NSG1 , neuron-specific gene family member 1.

Article Snippet: To accurately quantify protein concentrations, we used specific ELISA kits in accordance with the manufacturer’s instructions: Human Dopamine Receptor D5 Kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS724527), Human Dopamine Receptor D2 Kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS723432), Human Dopamine Receptor D3 Kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS722010), Human Caveolin 2 kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS1607726) and Human TGF beta 2 Kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS824902).

Techniques:

Journal: International Journal of Molecular Sciences

Article Title: Expression Profiles of Dopamine-Related Genes and miRNAs Regulating Their Expression in Breast Cancer

doi: 10.3390/ijms25126546

Figure Lengend Snippet: Changes in the expression profile of genes differentiating tumor samples compared to control tissues (−3.0 < FC > 3.0; p < 0.05).

Article Snippet: To accurately quantify protein concentrations, we used specific ELISA kits in accordance with the manufacturer’s instructions: Human Dopamine Receptor D5 Kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS724527), Human Dopamine Receptor D2 Kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS723432), Human Dopamine Receptor D3 Kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS722010), Human Caveolin 2 kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS1607726) and Human TGF beta 2 Kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS824902).

Techniques: Expressing

Journal: International Journal of Molecular Sciences

Article Title: Expression Profiles of Dopamine-Related Genes and miRNAs Regulating Their Expression in Breast Cancer

doi: 10.3390/ijms25126546

Figure Lengend Snippet: Changes in the expression profile of mRNAs in different breast cancer subtype samples compared to control samples obtained via RT-PCR. DRD2 , dopamine receptor 2; DRD3 , dopamine receptor 3; DRD5 , dopamine receptor 5; TGF-β-2 , transforming growth factor beta 2; CAV2 , caveolin 2. Data are presented as mean ± standard deviation.

Article Snippet: To accurately quantify protein concentrations, we used specific ELISA kits in accordance with the manufacturer’s instructions: Human Dopamine Receptor D5 Kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS724527), Human Dopamine Receptor D2 Kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS723432), Human Dopamine Receptor D3 Kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS722010), Human Caveolin 2 kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS1607726) and Human TGF beta 2 Kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS824902).

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Standard Deviation

Journal: International Journal of Molecular Sciences

Article Title: Expression Profiles of Dopamine-Related Genes and miRNAs Regulating Their Expression in Breast Cancer

doi: 10.3390/ijms25126546

Figure Lengend Snippet: The expression profile of miRNAs potentially regulated by selected mRNAs in different breast cancer subtype tissues in comparison to control tissues.

Article Snippet: To accurately quantify protein concentrations, we used specific ELISA kits in accordance with the manufacturer’s instructions: Human Dopamine Receptor D5 Kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS724527), Human Dopamine Receptor D2 Kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS723432), Human Dopamine Receptor D3 Kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS722010), Human Caveolin 2 kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS1607726) and Human TGF beta 2 Kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS824902).

Techniques: Expressing, Comparison

Journal: International Journal of Molecular Sciences

Article Title: Expression Profiles of Dopamine-Related Genes and miRNAs Regulating Their Expression in Breast Cancer

doi: 10.3390/ijms25126546

Figure Lengend Snippet: The concentration of selected proteins related to the dopaminergic system in different subtypes of breast cancer and control tissue.

Article Snippet: To accurately quantify protein concentrations, we used specific ELISA kits in accordance with the manufacturer’s instructions: Human Dopamine Receptor D5 Kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS724527), Human Dopamine Receptor D2 Kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS723432), Human Dopamine Receptor D3 Kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS722010), Human Caveolin 2 kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS1607726) and Human TGF beta 2 Kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS824902).

Techniques: Concentration Assay

Journal: International Journal of Molecular Sciences

Article Title: Expression Profiles of Dopamine-Related Genes and miRNAs Regulating Their Expression in Breast Cancer

doi: 10.3390/ijms25126546

Figure Lengend Snippet: Overall survival analysis for DRD2 in every group.

Article Snippet: To accurately quantify protein concentrations, we used specific ELISA kits in accordance with the manufacturer’s instructions: Human Dopamine Receptor D5 Kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS724527), Human Dopamine Receptor D2 Kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS723432), Human Dopamine Receptor D3 Kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS722010), Human Caveolin 2 kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS1607726) and Human TGF beta 2 Kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS824902).

Techniques:

Journal: International Journal of Molecular Sciences

Article Title: Expression Profiles of Dopamine-Related Genes and miRNAs Regulating Their Expression in Breast Cancer

doi: 10.3390/ijms25126546

Figure Lengend Snippet: The sequence of primers used in qRT-PCR.

Article Snippet: To accurately quantify protein concentrations, we used specific ELISA kits in accordance with the manufacturer’s instructions: Human Dopamine Receptor D5 Kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS724527), Human Dopamine Receptor D2 Kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS723432), Human Dopamine Receptor D3 Kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS722010), Human Caveolin 2 kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS1607726) and Human TGF beta 2 Kit (MyBioSource, Inc., San Diego, CA, USA, Cat No. MBS824902).

Techniques: Sequencing

Journal: Journal for Immunotherapy of Cancer

Article Title: Pharmacologic targeting of the dopamine D2 receptor impacts the efficacy of immune checkpoint blockade in melanoma

doi: 10.1136/jitc-2025-014080

Figure Lengend Snippet: Graphical abstract. Figure created in BioRender. BRC, bromocriptine; D2R, D2 receptor; HAL, haloperidol; ICI, immune checkpoint inhibitor; IL, interleukin; MCP, metoclopramide; MHC, major histocompatibility complex; PRL, prolactin.

Article Snippet: Samples were loaded in 10% precast polyacrylamide gels (Bio-Rad, cat. #4561036), run at 100 V in Tris/Glycine/sodium dodecyl sulfate (SDS) buffer (Bio-Rad 1610732), transferred to polyvinylidene fluoride (PVDF) membrane (Bio-Rad, cat. #1704272), blocked for 1 hour with 5% dry milk in tris buffered saline-Tween (TBS-T), and probed for PRL (R&D, cat. #AF1445), D2R (Proteintech, cat. #55084-1-AP), and β-Actin-HRP (BioLegend, cat. #664803) in blocking buffer.

Techniques: Immunopeptidomics

Journal: Journal for Immunotherapy of Cancer

Article Title: Pharmacologic targeting of the dopamine D2 receptor impacts the efficacy of immune checkpoint blockade in melanoma

doi: 10.1136/jitc-2025-014080

Figure Lengend Snippet: Tumor immune microenvironment in prolactin-locus ICI non-responder and responder models. ( A ) Experimental scheme. B16F0-bearing (CC51xB6)F1 and B6 mice received 100 µg αCTLA-4 and 200 µg αPD-1 on days 3, 6, and 10 after tumor inoculation. On day 13 after tumor inoculation, tumors were enriched for CD45+ TILs then processed for scRNA-seq. ( B ) UMAP and ( C ) composition analysis showing the breakdown of B6U, B6T, CC51U, CC51T annotated cell types overlaying the UMAP ( B ) and proportions of each cell type cluster. For each experimental condition, cell proportions (Y axis) were calculated by dividing the number of cells of a given type by the total number of cells. B6U, B6 untreated; B6T, B6 ICI-treated; CC51U, (CC51xB6)F1 untreated; CC51T, (CC51xB6)F1 ICI-treated. Cell types annotated manually using canonical markers. ( D ) Expression levels of M1 and M2 markers in the MΦ cluster. ( E ) Volcano plot highlighting DEGs in the CD20− CD8+ T cluster between B6T and CC51T. Significant DEG (padj≤0.05, |log2FC | ≥ 1) were determined using FindMarkers and are colored (red=upregulated in B6T; purple=upregulated in CC51T). ( F ) Expression levels of Prlr, Drd2, Htr3a and Htr4 in all clusters. αCTLA-4, anti-cytotoxic T-lymphocyte-associated protein 4; αPD-1, anti-programmed cell death protein-1; B, B cells; B16, B16 F0 cells; CD4+ T, CD4+ T cells; CD20- CD8+ T, CD8+ T cells; CD20+ CD8+ T, CD20+ CD8 T cells; cDC1, conventional dendritic cells, type I; DEGs, differentially expressed genes; Drd2, dopamine D2 receptor; Htr3a, serotonin 5HT3 receptor; Htr4, 5HT4 receptors; ICI, immune checkpoint inhibitor; MΦ, macrophages; MDSC, myeloid-derived suppressor cells; Mono, monocytes; NK T, natural killer T cells; Prlr, prolactin receptor; scRNA-seq, single-cell RNA sequencing; TILs, tumor-infiltrating leukocytes.

Article Snippet: Samples were loaded in 10% precast polyacrylamide gels (Bio-Rad, cat. #4561036), run at 100 V in Tris/Glycine/sodium dodecyl sulfate (SDS) buffer (Bio-Rad 1610732), transferred to polyvinylidene fluoride (PVDF) membrane (Bio-Rad, cat. #1704272), blocked for 1 hour with 5% dry milk in tris buffered saline-Tween (TBS-T), and probed for PRL (R&D, cat. #AF1445), D2R (Proteintech, cat. #55084-1-AP), and β-Actin-HRP (BioLegend, cat. #664803) in blocking buffer.

Techniques: Expressing, Derivative Assay, Single Cell, RNA Sequencing