Repeated cocaine administration upregulates CB2 receptor expression in striatal medium-spiny neurons that express dopamine D1 receptors in mice
Acta pharmacologica Sinica
Zhang, HY;De Biase, L;Chandra, R;Shen, H;Liu, QR;Gardner, E;Lobo, MK;Xi, ZX;
PMID: 34316031 | DOI: 10.1038/s41401-021-00712-6
Cannabinoid CB2 receptors (CB2R) are importantly involved in drug reward and addiction. However, the cellular mechanisms underlying CB2R action remain unclear. We have previously reported that cocaine self-administration upregulates CB2R expression in midbrain dopamine (DA) neurons. In the present study, we investigated whether cocaine or heroin also alters CB2R expression in striatal medium-spiny neurons that express dopamine D1 or D2 receptors (D1-MSNs, D2-MSNs) and microglia. Due to the concern of CB2R antibody specificity, we developed three mouse CB2-specific probes to detect CB2R mRNA using quantitative RT-PCR and RNAscope in situ hybridization (ISH) assays. We found that a single injection of cocaine failed to alter, while repeated cocaine injections or self-administration dose-dependently upregulated CB2R gene expression in both brain (cortex and striatum) and periphery (spleen). In contrast, repeated administration of heroin produced a dose-dependent reduction in striatal CB2 mRNA expression. RNAscope ISH assays detected CB2R mRNA in striatal D1- and D2-MSNs, not in microglia. We then used transgenic CX3CR1eGFP/+ microglia reporter mice and D1- or D2-Cre-RiboTag mice to purify striatal microglia or ribosome-associated mRNAs from CX3CR1eGFP/+, D1-MSNs, or D2-MSNs, respectively. We found that CB2R upregulation occurred mainly in D1-MSNs, not in D2-MSNs or microglia, in the nucleus accumbens rather than the dorsal striatum. These findings indicate that repeated cocaine exposure may upregulate CB2R expression in both brain and spleen, with regional and cell type-specific profiles. In the striatum, CB2R upregulation occurs mainly in D1-MSNs in the nucleus accumbens. Given the important role of D1-MSNs in brain reward function, the present findings provide new insight into mechanisms by which brain CB2Rs modulate cocaine action.
Choi MH, Na JE, Yoon YR, Lee HJ, Yoon S, Rhyu IJ, Baik JH.
PMID: 28912499 | DOI: 10.1038/s41598-017-10173-9
Dopaminergic systems play a major role in reward-related behavior and dysregulation of dopamine (DA) systems can cause several mental disorders, including depression. We previously reported that dopamine D2 receptor knockout (D2R-/-) mice display increased anxiety and depression-like behaviors upon chronic stress. Here, we observed that chronic stress caused myelin loss in wild-type (WT) mice, while the myelin level in D2R-/- mice, which was already lower than that in WT mice, was not affected upon stress. Fewer mature oligodendrocytes (OLs) were observed in the corpus callosum of stressed WT mice, while in D2R-/- mice, both the control and stressed group displayed a decrease in the number of mature OLs. We observed a decrease in the number of active β-catenin (ABC)-expressing and TCF4-expressing cells among OL lineage cells in the corpus callosum of stressed WT mice, while such regulation was not found in D2R-/- mice. Administration of lithium normalized the behavioral impairments and myelin damage induced by chronic stress in WT mice, and restored the number of ABC-positive and TCF4-positive OLs, while such effect was not found in D2R-/- mice. Together, our findings indicate that chronic stress induces myelin loss through the Wnt/β-catenin signaling pathway in association with DA signaling through D2R.
Maynard, JP;Godwin, TN;Lu, J;Vidal, I;Lotan, TL;De Marzo, AM;Joshu, CE;Sfanos, KS;
PMID: 35971807 | DOI: 10.1002/pros.24424
Black men are two to three times more likely to die from prostate cancer (PCa) than White men. This disparity is due in part to discrepancies in socioeconomic status and access to quality care. Studies also suggest that differences in the prevalence of innate immune cells and heightened function in the tumor microenvironment of Black men may promote PCa aggressiveness.We evaluated the spatial localization of and quantified CD66ce+ neutrophils by immunohistochemistry and CD68+ (pan), CD80+ (M1), and CD163+ (M2) macrophages by RNA in situ hybridization on formalin-fixed paraffin-embedded tissues from organ donor "normal" prostate (n = 9) and radical prostatectomy (n = 38) tissues from Black and White men. Neutrophils were quantified in PCa and matched benign tissues in tissue microarray (TMA) sets comprised of 560 White and 371 Black men. Likewise, macrophages were quantified in TMA sets comprised of tissues from 60 White and 120 Black men. The phosphatase and tensin homolog (PTEN) and ETS transcription factor ERG (ERG) expression status of each TMA PCa case was assessed via immunohistochemistry. Finally, neutrophils and macrophage subsets were assessed in a TMA set comprised of distant metastatic PCa tissues collected at autopsy (n = 6) sampled across multiple sites.CD66ce+ neutrophils were minimal in normal prostates, but were increased in PCa compared to benign tissues, in low grade compared to higher grade PCa, in PCa tissues from White compared to Black men, and in PCa with PTEN loss or ERG positivity. CD163+ macrophages were the predominant macrophage subset in normal organ donor prostate tissues from both Black and White men and were significantly more abundant in organ donor compared to prostatectomy PCa tissues. CD68,+ CD80,+ and CD163+ macrophages were significantly increased in cancer compared to benign tissues and in cancers with ERG positivity. CD68+ and CD163+ macrophages were increased in higher grade cancers compared to low grade cancer and CD80 expression was significantly higher in benign prostatectomy tissues from Black compared to White men.Innate immune cell infiltration is increased in the prostate tumor microenvironment of both Black and White men, however the composition of innate immune cell infiltration may vary between races.
Neuropsychopharmacology : official publication of the American College of Neuropsychopharmacology
McNulty, CJ;Fallon, IP;Amat, J;Sanchez, RJ;Leslie, NR;Root, DH;Maier, SF;Baratta, MV;
PMID: 36076018 | DOI: 10.1038/s41386-022-01443-w
Stress-linked disorders are more prevalent in women than in men and differ in their clinical presentation. Thus, investigating sex differences in factors that promote susceptibility or resilience to stress outcomes, and the circuit elements that mediate their effects, is important. In male rats, instrumental control over stressors engages a corticostriatal system involving the prelimbic cortex (PL) and dorsomedial striatum (DMS) that prevent many of the sequelae of stress exposure. Interestingly, control does not buffer against stress outcomes in females, and here, we provide evidence that the instrumental controlling response in females is supported instead by the dorsolateral striatum (DLS). Additionally, we used in vivo microdialysis, fluorescent in situ hybridization, and receptor subtype pharmacology to examine the contribution of prefrontal dopamine (DA) to the differential impact of behavioral control. Although both sexes preferentially expressed D1 receptor mRNA in PL GABAergic neurons, there were robust sex differences in the dynamic properties of prefrontal DA during controllable stress. Behavioral control potently attenuated stress-induced DA efflux in males, but not females, who showed a sustained DA increase throughout the entire stress session. Importantly, PL D1 receptor blockade (SCH 23390) shifted the proportion of striatal activity from the DLS to the DMS in females and produced the protective effects of behavioral control. These findings suggest a sex-selective mechanism in which elevated DA in the PL biases instrumental responding towards prefrontal-independent striatal circuitry, thereby eliminating the protective impact of coping with stress.
American journal of physiology. Endocrinology and metabolism
Abdelmoez, AM;Dmytriyeva, O;Zurke, YX;Trauelsen, M;Marica, AA;Savikj, M;Smith, JAB;Monaco, C;Schwartz, TW;Krook, A;Pillon, NJ;
PMID: 36812387 | DOI: 10.1152/ajpendo.00009.2023
Succinate is released by skeletal muscle during exercise and activates SUCNR1/GPR91. Signaling of SUCNR1 is involved in metabolite-sensing paracrine communication in skeletal muscle during exercise. However, the specific cell types responding to succinate and the directionality of communication are unclear. We aim to characterize the expression of SUCNR1 in human skeletal muscle. De novo analysis of transcriptomic datasets demonstrated that SUCNR1 mRNA is expressed in immune, adipose, and liver tissues, but scarce in skeletal muscle. In human tissues, SUCNR1 mRNA was associated with macrophage markers. Single-cell RNA sequencing and fluorescent RNAscope demonstrated that in human skeletal muscle, SUCNR1 mRNA is not expressed in muscle fibers but coincided with macrophage populations. Human M2-polarized macrophages exhibit high levels of SUCNR1 mRNA and stimulation with selective agonists of SUCNR1 triggered Gq- and Gi-coupled signaling. Primary human skeletal muscle cells were unresponsive to SUCNR1 agonists. In conclusion, SUCNR1 is not expressed in muscle cells and its role in the adaptive response of skeletal muscle to exercise is most likely mediated via paracrine mechanisms involving M2-like macrophages within the muscle.
Gene-targeted, CREB-mediated induction of ΔFosB controls distinct downstream transcriptional patterns within D1 and D2 medium spiny neurons
Lardner, C;van der Zee, Y;Estill, M;Kronman, H;Salery, M;Cunningham, A;Godino, A;Parise, E;Kim, J;Neve, R;Shen, L;Hamilton, P;Nestler, E;
| DOI: 10.1016/j.biopsych.2021.06.017
Background The onset and persistence of addiction phenotypes are, in part, mediated by transcriptional mechanisms in the brain that affect gene expression and subsequently neural circuitry. ΔFosB is a transcription factor that accumulates in the nucleus accumbens (NAc) – a brain region responsible for coordinating reward and motivation – after exposure to virtually every known rewarding substance, including cocaine and opioids. ΔFosB has also been shown to directly control gene transcription and behavior downstream of both cocaine and opioid exposure, but with potentially different roles in D1 and D2 medium spiny neurons (MSNs) in NAc. Methods To clarify MSN subtype-specific roles for ΔFosB, and investigate how these coordinate the actions of distinct classes of addictive drugs in NAc, we developed a CRISPR/Cas9-based epigenome editing tool to induce endogenous ΔFosB expression in vivo in the absence of drug exposure. After inducing ΔFosB in D1 or D2 MSNs, or both, we performed RNA-sequencing on bulk male and female NAc tissue (N = 6-8/group). Results We find that ΔFosB induction elicits distinct transcriptional profiles in NAc by MSN subtype and by sex, establishing for the first time that ΔFosB mediates different transcriptional effects in males vs females. We also demonstrate that changes in D1 MSNs, but not in D2 MSNs or both, significantly recapitulate changes in gene expression induced by cocaine self-administration. Conclusions Together, these findings demonstrate the efficacy of a novel molecular tool for studying cell-type-specific transcriptional mechanisms, and shed new light on the activity of ΔFosB, a critical transcriptional regulator of drug addiction.
Zhou, K;Xu, H;Lu, S;Jiang, S;Hou, G;Deng, X;He, M;Zhu, Y;
PMID: 36271048 | DOI: 10.1038/s41467-022-33843-3
The nucleus accumbens (NAc) is critical in mediating reward seeking and is also involved in negative emotion processing, but the cellular and circuitry mechanisms underlying such opposing behaviors remain elusive. Here, using the recently developed AAV1-mediated anterograde transsynaptic tagging technique in mice, we show that NAc neurons receiving basolateral amygdala inputs (NAcBLA) promote positive reinforcement via disinhibiting dopamine neurons in the ventral tegmental area (VTA). In contrast, NAc neurons receiving paraventricular thalamic inputs (NAcPVT) innervate GABAergic neurons in the lateral hypothalamus (LH) and mediate aversion. Silencing the synaptic output of NAcBLA neurons impairs reward seeking behavior, while silencing of NAcPVT or NAcPVT→LH pathway abolishes aversive symptoms of opiate withdrawal. Our results elucidate the afferent-specific circuit architecture of the NAc in controlling reward and aversion.
Frontiers in cellular neuroscience
Giua, G;Lassalle, O;Makrini-Maleville, L;Valjent, E;Chavis, P;Manzoni, OJJ;
PMID: 37323585 | DOI: 10.3389/fncel.2023.1146647
Fragile X syndrome (FXS), resulting from a mutation in the Fmr1 gene, is the most common monogenic cause of autism and inherited intellectual disability. Fmr1 encodes the Fragile X Messenger Ribonucleoprotein (FMRP), and its absence leads to cognitive, emotional, and social deficits compatible with the nucleus accumbens (NAc) dysfunction. This structure is pivotal in social behavior control, consisting mainly of spiny projection neurons (SPNs), distinguished by dopamine D1 or D2 receptor expression, connectivity, and associated behavioral functions. This study aims to examine how FMRP absence differentially affects SPN cellular properties, which is crucial for categorizing FXS cellular endophenotypes.We utilized a novel Fmr1-/y::Drd1a-tdTomato mouse model, which allows in-situ identification of SPN subtypes in FXS mice. Using RNA-sequencing, RNAScope and ex-vivo patch-clamp in adult male mice NAc, we comprehensively compared the intrinsic passive and active properties of SPN subtypes.Fmr1 transcripts and their gene product, FMRP, were found in both SPNs subtypes, indicating potential cell-specific functions for Fmr1. The study found that the distinguishing membrane properties and action potential kinetics typically separating D1- from D2-SPNs in wild-type mice were either reversed or abolished in Fmr1-/y::Drd1a-tdTomato mice. Interestingly, multivariate analysis highlighted the compound effects of Fmr1 ablation by disclosing how the phenotypic traits distinguishing each cell type in wild-type mice were altered in FXS.Our results suggest that the absence of FMRP disrupts the standard dichotomy characterizing NAc D1- and D2-SPNs, resulting in a homogenous phenotype. This shift in cellular properties could potentially underpin select aspects of the pathology observed in FXS. Therefore, understanding the nuanced effects of FMRP absence on SPN subtypes can offer valuable insights into the pathophysiology of FXS, opening avenues for potential therapeutic strategies.
Frontiers in Neuroscience
Wang L, Lee G, Kuei C, Yao X, Harrington A, Bonaventure P, Lovenberg TW and Liu C
| DOI: 10.3389/fnins.2019.00281
GPR139, a Gq-coupled receptor that is activated by the essential amino acids L-tryptophan and L-phenylalanine, is predominantly expressed in the brain and pituitary. The physiological function of GPR139 remains elusive despite the availability of pharmacological tool agonist compounds and knock-out mice. Whole tissue RNA sequencing data from human, mouse and rat tissues revealed that GPR139 and the dopamine D2 receptor (DRD2) exhibited some similarities in their distribution patterns in the brain and pituitary gland. To determine if there was true co-expression of these two receptors, we applied double in situ hybridization in mouse tissues using the RNAscope™ technique. GPR139 and DRD2 mRNA co-localized in a majority of cells within part of the dopaminergic mesolimbic pathways (ventral tegmental area and olfactory tubercle), the nigrostriatal pathway (compact part of substantia nigra and caudate putamen) and also the tuberoinfundibular pathway (arcuate hypothalamic nucleus and anterior lobe of pituitary). Both receptors mRNA also co-localized in brain regions involved in responses to negative stimulus and stress, such as lateral habenula, lateral septum, interpeduncular nucleus and medial raphe nuclei. GPR139 mRNA expression was detected in the dentate gyrus and the pyramidal cell layer of the hippocampus as well as the paraventricular hypothalamic nucleus. The functional interaction between GPR139 and DRD2 was studied in vitro using a calcium mobilization assay in cells co-transfected with both receptors from several species (human, rat and mouse). The dopamine DRD2 agonist did not stimulate calcium response in cells expressing DRD2 alone consistent with the Gi signaling transduction pathway of this receptor. In cells co-transfected with DRD2 and GPR139 the DRD2 agonist was able to stimulate calcium response and its effect was blocked by either a DRD2 or a GPR139 antagonist supporting an in vitro interaction between GPR139 and DRD2. Taken together, these data showed that GPR139 and DRD2 are in position to functionally interact in native tissue.
J Ovarian Res. 2015 May 14;8(1):29
Abstract BACKGROUND: Folate receptor alpha (FOLR1/FRA) is expressed in a number of epithelial cancers and in particular epithelial ovarian cancer (EOC), especially of the serous histotype. Recent studies have shown that EOC originates from the fallopian tube fimbriae rather than from epithelial cells lining the ovary. We have previously shown by immunohistochemistry a strong correlation between FRA expression in EOC and normal and fallopian adenocarcinoma. Folate receptor beta (FOLR2/FRB) has been described to be expressed by macrophages both in inflammatory disorders and certain epithelial cancers. Given the high sequence identity of these two folate receptor family members we sought to investigate the architectural and cell-specific expression of these two receptors in gynecologic tissues. METHODS: RNA scope, a novel chromogenic in situ hybridization assay tool, was used to examine expression of the alpha (FOLR1) and beta (FOLR2) isoforms of folate receptor relative to each other as well as to the macrophage markers CD11b and CD68, in samples of normal fallopian tube and fallopian adenocarcinoma as well as normal ovary and EOC. RESULTS: We demonstrated expression of both FOLR1 and FOLR2 in EOC, normal fallopian tube and fallopian adenocarcinoma tissue while very little expression of either marker was observed in normal ovary. Furthermore, FOLR2 was shown to be expressed almost exclusively in macrophages, of both the M1 and M2 lineages, as determined by co-expression of CD11b and/or CD68, with little or no expression in epithelial cells. CONCLUSIONS: These findings further substantiate the hypothesis that the cell of origin of EOC is tubal epithelium and that the beta isoform of folate receptor is primarily restricted to macrophages. Further, macrophages expressing FOLR2 may represent tumor associated or infiltrating macrophages (TAMs) in epithelial cancers.
The Journal of Neuroscience, 8 April 2015, 35(14): 5625-5639
Rubio FJ, Liu QR, Li X, Cruz FC, Leão RM, Warren BL, Kambhampati S, Babin KR, McPherson KB, Cimbro R, Bossert JM, Shaham Y, Hope BT.
PMID: 25855177 | DOI: 10.1523/JNEUROSCI.4997-14.2015
Context-induced reinstatement of drug seeking is a well established animal model for assessing the neural mechanisms underlying context-induced drug relapse, a major factor in human drug addiction. Neural activity in striatum has previously been shown to contribute to context-induced reinstatement of heroin, cocaine, and alcohol seeking, but not yet for methamphetamine seeking. In this study, we found that context-induced reinstatement of methamphetamine seeking increased expression of the neural activity marker Fos in dorsal but not ventral striatum. Reversible inactivation of neural activity in dorsolateral but not dorsomedial striatum using the GABA agonists muscimol and baclofen decreased context-induced reinstatement. Based on our previous findings that Fos-expressing neurons play a critical role in conditioned drug effects, we assessed whether context-induced reinstatement was associated with molecular alterations selectively induced within context-activated Fos-expressing neurons. We used fluorescence-activated cell sorting to isolate reinstatement-activated Fos-positive neurons from Fos-negative neurons in dorsal striatum and used quantitative PCR to assess gene expression within these two populations of neurons. Context-induced reinstatement was associated with increased expression of the immediate early genes Fos and FosB and the NMDA receptor subunit gene Grin2a in only Fos-positive neurons. RNAscope in situ hybridization confirmed that Grin2a, as well as Grin2b, expression were increased in only Fos-positive neurons from dorsolateral, but not dorsomedial, striatum. Our results demonstrate an important role of dorsolateral striatum in context-induced reinstatement of methamphetamine seeking and that this reinstatement is associated with unique gene alterations in Fos-expressing neurons.
Egerod KL, Petersen N ,Timshel PN, Rekling JC, Wang Y, Liu Q, Schwartz TW, Gautron L.
PMID: - | DOI: 10.1016/j.molmet.2018.03.016
Abstract
Objectives
G protein-coupled receptors (GPCRs) act as transmembrane molecular sensors of neurotransmitters, hormones, nutrients, and metabolites. Because unmyelinated vagalafferents richly innervate the gastrointestinal mucosa, gut-derived molecules may directly modulate the activity of vagal afferents through GPCRs. However, the types of GPCRs expressed in vagal afferents are largely unknown. Here, we determined the expression profile of all GPCRs expressed in vagal afferents of the mouse, with a special emphasis on those innervating the gastrointestinal tract.
Methods
Using a combination of high-throughput quantitative PCR, RNA sequencing, and in situhybridization, we systematically quantified GPCRs expressed in vagal unmyelinated Nav1.8-expressing afferents.
Results
GPCRs for gut hormones that were the most enriched in Nav1.8-expressing vagal unmyelinated afferents included NTSR1, NPY2R, CCK1R, and to a lesser extent, GLP1R, but not GHSR and GIPR. Interestingly, both GLP1R and NPY2R were coexpressed with CCK1R. In contrast, NTSR1 was coexpressed with GPR65, a marker preferentially enriched in intestinal mucosal afferents. Only few microbiome-derived metabolite sensors such as GPR35 and, to a lesser extent, GPR119 and CaSR were identified in the Nav1.8-expressing vagal afferents. GPCRs involved in lipid sensing and inflammation (e.g. CB1R, CYSLTR2, PTGER4), and neurotransmitters signaling (CHRM4, DRD2, CRHR2) were also highly enriched in Nav1.8-expressing neurons. Finally, we identified 21 orphan GPCRs with unknown functions in vagal afferents.
Conclusion
Overall, this study provides a comprehensive description of GPCR-dependent sensing mechanisms in vagal afferents, including novel coexpression patterns, and conceivably coaction of key receptors for gut-derived molecules involved in gut-brain communication.
