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1.
Biomolecules ; 13(11)2023 11 17.
Article in English | MEDLINE | ID: mdl-38002340

ABSTRACT

L-DOPA is the mainstay of treatment for Parkinson's disease (PD). However, over time this drug can produce dyskinesia. A useful acute PD model for screening novel compounds for anti-parkinsonian and L-DOPA-induced dyskinesia (LID) are dopamine-depleted dopamine-transporter KO (DDD) mice. Treatment with α-methyl-para-tyrosine rapidly depletes their brain stores of DA and renders them akinetic. During sensitization in the open field (OF), their locomotion declines as vertical activities increase and upon encountering a wall they stand on one leg or tail and engage in climbing behavior termed "three-paw dyskinesia". We have hypothesized that L-DOPA induces a stereotypic activation of locomotion in DDD mice, where they are unable to alter the course of their locomotion, and upon encountering walls engage in "three-paw dyskinesia" as reflected in vertical counts or beam-breaks. The purpose of our studies was to identify a valid index of LID in DDD mice that met three criteria: (a) sensitization with repeated L-DOPA administration, (b) insensitivity to a change in the test context, and (c) stimulatory or inhibitory responses to dopamine D1 receptor agonists (5 mg/kg SKF81297; 5 and 10 mg/kg MLM55-38, a novel compound) and amantadine (45 mg/kg), respectively. Responses were compared between the OF and a circular maze (CM) that did not hinder locomotion. We found vertical counts and climbing were specific for testing in the OF, while oral stereotypies were sensitized to L-DOPA in both the OF and CM and responded to D1R agonists and amantadine. Hence, in DDD mice oral stereotypies should be used as an index of LID in screening compounds for PD.


Subject(s)
Dyskinesia, Drug-Induced , Parkinson Disease , Mice , Animals , Levodopa/pharmacology , Levodopa/therapeutic use , Dopamine Agonists/pharmacology , Dopamine Agonists/therapeutic use , Dopamine , Dopamine Plasma Membrane Transport Proteins/genetics , Dyskinesia, Drug-Induced/drug therapy , Mice, Knockout , Parkinson Disease/drug therapy , Parkinson Disease/genetics , Amantadine/pharmacology
2.
Neuropharmacology ; 234: 109544, 2023 08 15.
Article in English | MEDLINE | ID: mdl-37055008

ABSTRACT

Strong expression of the G protein-coupled receptor (GPCR) neurotensin receptor 1 (NTR1) in ventral tegmental area (VTA) dopamine (DA) neurons and terminals makes it an attractive target to modulate DA neuron activity and normalize DA-related pathologies. Recent studies have identified a novel class of NTR1 ligand that shows promising effects in preclinical models of addiction. A lead molecule, SBI-0654553 (SBI-553), can act as a positive allosteric modulator of NTR1 ß-arrestin recruitment while simultaneously antagonizing NTR1 Gq protein signaling. Using cell-attached recordings from mouse VTA DA neurons we discovered that, unlike neurotensin (NT), SBI-553 did not independently increase spontaneous firing. Instead, SBI-553 blocked the NT-mediated increase in firing. SBI-553 also antagonized the effects of NT on dopamine D2 auto-receptor signaling, potentially through its inhibitory effects on G-protein signaling. We also measured DA release directly, using fast-scan cyclic voltammetry in the nucleus accumbens and observed antagonist effects of SBI-553 on an NT-induced increase in DA release. Further, in vivo administration of SBI-553 did not notably change basal or cocaine-evoked DA release measured in NAc using fiber photometry. Overall, these results indicate that SBI-553 blunts NT's effects on spontaneous DA neuron firing, D2 auto-receptor function, and DA release, without independently affecting these measures. In the presence of NT, SBI-553 has an inhibitory effect on mesolimbic DA activity, which could contribute to its efficacy in animal models of psychostimulant use.


Subject(s)
Dopamine D2 Receptor Antagonists , Dopamine , Dopaminergic Neurons , Neurotensin , Nucleus Accumbens , Receptors, Neurotensin , Ventral Tegmental Area , Ventral Tegmental Area/metabolism , Ventral Tegmental Area/physiology , Dopaminergic Neurons/drug effects , Dopaminergic Neurons/metabolism , Dopaminergic Neurons/physiology , Nucleus Accumbens/metabolism , Dopamine/metabolism , Male , Female , Animals , Mice , Mice, Inbred C57BL , Presynaptic Terminals/metabolism , Presynaptic Terminals/physiology , Action Potentials/drug effects , Receptors, Neurotensin/antagonists & inhibitors , Receptors, Neurotensin/metabolism , Neurotensin/metabolism , Neurotensin/pharmacology , Ligands , Dopamine D2 Receptor Antagonists/metabolism , Dopamine D2 Receptor Antagonists/pharmacology
3.
Biochemistry ; 62(7): 1233-1248, 2023 04 04.
Article in English | MEDLINE | ID: mdl-36917754

ABSTRACT

The NTSR1 neurotensin receptor (NTSR1) is a G protein-coupled receptor (GPCR) found in the brain and peripheral tissues with neurotensin (NTS) being its endogenous peptide ligand. In the brain, NTS modulates dopamine neuronal activity, induces opioid-independent analgesia, and regulates food intake. Recent studies indicate that biasing NTSR1 toward ß-arrestin signaling can attenuate the actions of psychostimulants and other drugs of abuse. Here, we provide the cryoEM structures of NTSR1 ternary complexes with heterotrimeric Gq and GoA with and without the brain-penetrant small-molecule SBI-553. In functional studies, we discovered that SBI-553 displays complex allosteric actions exemplified by negative allosteric modulation for G proteins that are Gα subunit selective and positive allosteric modulation and agonism for ß-arrestin translocation at NTSR1. Detailed structural analysis of the allosteric binding site illuminated the structural determinants for biased allosteric modulation of SBI-553 on NTSR1.


Subject(s)
Neurotensin , Receptors, Neurotensin , Receptors, Neurotensin/chemistry , Receptors, Neurotensin/metabolism , Neurotensin/metabolism , Signal Transduction , Peptides/metabolism , beta-Arrestins/metabolism
4.
Trends Endocrinol Metab ; 34(2): 106-118, 2023 02.
Article in English | MEDLINE | ID: mdl-36567228

ABSTRACT

As allosteric complexes, G-protein-coupled receptors (GPCRs) respond to extracellular stimuli and pleiotropically couple to intracellular transducers to elicit signaling pathway-dependent effects in a process known as biased signaling or functional selectivity. One such GPCR, the ghrelin receptor (GHSR1a), has a crucial role in restoring and maintaining metabolic homeostasis during disrupted energy balance. Thus, pharmacological modulation of GHSR1a bias could offer a promising strategy to treat several metabolism-based disorders. Here, we summarize current evidence supporting GHSR1a functional selectivity in vivo and highlight recent structural data. We propose that precise determinations of GHSR1a molecular pharmacology and pathway-specific physiological effects will enable discovery of GHSR1a drugs with tailored signaling profiles, thereby providing safer and more effective treatments for metabolic diseases.


Subject(s)
Receptors, Ghrelin , Signal Transduction , Humans , Receptors, Ghrelin/genetics , Receptors, Ghrelin/metabolism , Signal Transduction/physiology , Ghrelin/metabolism
5.
NMR Biomed ; 36(2): e4842, 2023 02.
Article in English | MEDLINE | ID: mdl-36259728

ABSTRACT

The United States is experiencing a dramatic increase in maternal opioid misuse and, consequently, the number of individuals exposed to opioids in utero. Prenatal opioid exposure has both acute and long-lasting effects on health and wellbeing. Effects on the brain, often identified at school age, manifest as cognitive impairment, attention deficit, and reduced scholastic achievement. The neurobiological basis for these effects is poorly understood. Here, we examine how in utero exposure to heroin affects brain development into early adolescence in a mouse model. Pregnant C57BL/6J mice received escalating doses of heroin twice daily on gestational days 4-18. The brains of offspring were assessed on postnatal day 28 using 9.4 T diffusion MRI of postmortem specimens at 36 µm resolution. Whole-brain volumes and the volumes of 166 bilateral regions were compared between heroin-exposed and control offspring. We identified a reduction in whole-brain volume in heroin-exposed offspring and heroin-associated volume changes in 29 regions after standardizing for whole-brain volume. Regions with bilaterally reduced standardized volumes in heroin-exposed offspring relative to controls include the ectorhinal and insular cortices. Regions with bilaterally increased standardized volumes in heroin-exposed offspring relative to controls include the periaqueductal gray, septal region, striatum, and hypothalamus. Leveraging microscopic resolution diffusion tensor imaging and precise regional parcellation, we generated whole-brain structural MRI diffusion connectomes. Using a dimension reduction approach with multivariate analysis of variance to assess group differences in the connectome, we found that in utero heroin exposure altered structure-based connectivity of the left septal region and the region that acts as a hub for limbic regulatory actions. Consistent with clinical evidence, our findings suggest that prenatal opioid exposure may have effects on brain morphology, connectivity, and, consequently, function that persist into adolescence. This work expands our understanding of the risks associated with opioid misuse during pregnancy and identifies biomarkers that may facilitate diagnosis and treatment.


Subject(s)
Opioid-Related Disorders , Prenatal Exposure Delayed Effects , Humans , Pregnancy , Female , Animals , Mice , Heroin/adverse effects , Diffusion Tensor Imaging/methods , Analgesics, Opioid/pharmacology , Mice, Inbred C57BL , Brain
6.
Sci Rep ; 12(1): 21422, 2022 Dec 11.
Article in English | MEDLINE | ID: mdl-36503898

ABSTRACT

Genetically tractable animal models provide needed strategies to resolve the biological basis of drug addiction. Intravenous self-administration (IVSA) is the gold standard for modeling psychostimulant and opioid addiction in animals, but technical limitations have precluded the widespread use of IVSA in mice. Here, we describe IVSA paradigms for mice that capture the multi-stage nature of the disorder and permit predictive modeling. In these paradigms, C57BL/6J mice with long-standing indwelling jugular catheters engaged in cocaine- or remifentanil-associated lever responding that was fixed ratio-dependent, dose-dependent, extinguished by withholding the drug, and reinstated by the presentation of drug-paired cues. The application of multivariate analysis suggested that drug taking in both paradigms was a function of two latent variables we termed incentive motivation and discriminative control. Machine learning revealed that vulnerability to drug seeking and relapse were predicted by a mouse's a priori response to novelty, sensitivity to drug-induced locomotion, and drug-taking behavior. The application of these behavioral and statistical-analysis approaches to genetically-engineered mice will facilitate the identification of neural circuits driving addiction susceptibility and relapse and focused therapeutic development.


Subject(s)
Drug-Seeking Behavior , Mice , Animals , Mice, Inbred C57BL , Administration, Intravenous , Self Administration , Models, Animal
7.
Sci Signal ; 15(726): eabg5203, 2022 03 22.
Article in English | MEDLINE | ID: mdl-35316095

ABSTRACT

G protein-coupled receptors (GPCRs) are the largest family of cell surface receptors and signal through the proximal effectors, G proteins and ß-arrestins, to influence nearly every biological process. The G protein and ß-arrestin signaling pathways have largely been considered separable; however, direct interactions between Gα proteins and ß-arrestins have been described that appear to be part of a distinct GPCR signaling pathway. Within these complexes, Gαi/o, but not other Gα protein subtypes, directly interacts with ß-arrestin, regardless of the canonical Gα protein that is coupled to the GPCR. Here, we report that the endogenous biased chemokine agonists of CXCR3 (CXCL9, CXCL10, and CXCL11), together with two small-molecule biased agonists, differentially formed Gαi:ß-arrestin complexes. Formation of the Gαi:ß-arrestin complexes did not correlate well with either G protein activation or ß-arrestin recruitment. ß-arrestin biosensors demonstrated that ligands that promoted Gαi:ß-arrestin complex formation generated similar ß-arrestin conformations. We also found that Gαi:ß-arrestin complexes did not couple to the mitogen-activated protein kinase ERK, as is observed with other receptors such as the V2 vasopressin receptor, but did couple with the clathrin adaptor protein AP-2, which suggests context-dependent signaling by these complexes. These findings reinforce the notion that Gαi:ß-arrestin complex formation is a distinct GPCR signaling pathway and enhance our understanding of the spectrum of biased agonism.


Subject(s)
GTP-Binding Proteins , Receptors, G-Protein-Coupled , GTP-Binding Proteins/metabolism , Receptors, G-Protein-Coupled/genetics , Receptors, G-Protein-Coupled/metabolism , Signal Transduction , beta-Arrestin 1/genetics , beta-Arrestin 1/metabolism , beta-Arrestins/metabolism
8.
eNeuro ; 9(5)2022.
Article in English | MEDLINE | ID: mdl-36635920

ABSTRACT

The protease caspase-3 is a key mediator of apoptotic programmed cell death. But weak or transient caspase activity can contribute to neuronal differentiation, axonal pathfinding, and synaptic long-term depression. Despite the importance of sublethal, or nonapoptotic, caspase activity in neurodevelopment and neural plasticity, there has been no simple method for mapping and quantifying nonapoptotic caspase activity (NACA) in rodent brains. We therefore generated a transgenic mouse expressing a highly sensitive and specific fluorescent reporter of caspase activity, with peak signal localized to the nucleus. As a proof of concept, we first obtained evidence that NACA influences neurophysiology in an amygdalar circuit. Then focusing on the amygdala, we were able to quantify a sex-specific persistent elevation in caspase activity in females after restraint stress. This simple in vivo caspase activity reporter will facilitate systems-level studies of apoptotic and nonapoptotic phenomena in behavioral and pathologic models.


Subject(s)
Apoptosis , Brain , Male , Female , Mice , Animals , Apoptosis/physiology , Mice, Transgenic , Neuronal Plasticity , Caspase 9
9.
Front Endocrinol (Lausanne) ; 12: 714561, 2021.
Article in English | MEDLINE | ID: mdl-34484125

ABSTRACT

ß-arrestins are partners of the G protein-coupled receptors (GPCRs), regulating their intracellular trafficking and signaling. Development of biased GPCR agonists, selectively targeting either G protein or ß-arrestin pathways, are in the focus of interest due to their therapeutic potential in different pathological conditions. The CB2 cannabinoid receptor (CB2R) is a GPCR involved in various functions in the periphery and the central nervous system. Two common occurring variants of CB2R, harboring Q63R or L133I missense mutations, have been implicated in the development of a diverse set of disorders. To evaluate the effect of these mutations, we characterized the binding profile of these mutant CB2 receptors to G proteins and ß-arrestin2. Although their ability to inhibit cAMP signaling was similar, the Q63R mutant had increased, whereas the L133I mutant receptor had decreased ß-arrestin2 binding. In line with these observations, the variants also had altered intracellular trafficking. Our results show that two common variants of the CB2 receptor have biased signaling properties, which may contribute to the pathogenesis of the associated disorders and may offer CB2R as a target for further development of biased receptor activation strategies.


Subject(s)
Mutation, Missense , Receptor, Cannabinoid, CB2/metabolism , beta-Arrestins/metabolism , HEK293 Cells , Humans , Protein Binding , Protein Transport , Receptor, Cannabinoid, CB2/chemistry , Receptor, Cannabinoid, CB2/genetics , beta-Arrestins/genetics
10.
Mol Cancer Res ; 19(10): 1699-1711, 2021 10.
Article in English | MEDLINE | ID: mdl-34131071

ABSTRACT

HER2-positive breast cancers are among the most heterogeneous breast cancer subtypes. The early amplification of HER2 and its known oncogenic isoforms provide a plausible mechanism in which distinct programs of tumor heterogeneity could be traced to the initial oncogenic event. Here a Cancer rainbow mouse simultaneously expressing fluorescently barcoded wildtype (WTHER2), exon-16 null (d16HER2), and N-terminally truncated (p95HER2) HER2 isoforms is used to trace tumorigenesis from initiation to invasion. Tumorigenesis was visualized using whole-gland fluorescent lineage tracing and single-cell molecular pathology. We demonstrate that within weeks of expression, morphologic aberrations were already present and unique to each HER2 isoform. Although WTHER2 cells were abundant throughout the mammary ducts, detectable lesions were exceptionally rare. In contrast, d16HER2 and p95HER2 induced rapid tumor development. d16HER2 incited homogenous and proliferative luminal-like lesions which infrequently progressed to invasive phenotypes whereas p95HER2 lesions were heterogenous and invasive at the smallest detectable stage. Distinct cancer trajectories were observed for d16HER2 and p95HER2 tumors as evidenced by oncogene-dependent changes in epithelial specification and the tumor microenvironment. These data provide direct experimental evidence that intratumor heterogeneity programs begin very early and well in advance of screen or clinically detectable breast cancer. IMPLICATIONS: Although all HER2 breast cancers are treated equally, we show a mechanism by which clinically undetected HER2 isoforms program heterogenous cancer phenotypes through biased epithelial specification and adaptations within the tumor microenvironment.


Subject(s)
Breast Neoplasms/genetics , Carcinogenesis/genetics , Protein Isoforms/genetics , Receptor, ErbB-2/genetics , Animals , Female , Gene Expression Regulation, Neoplastic/genetics , Mice , Mice, Knockout , Tumor Microenvironment/genetics
12.
Trends Pharmacol Sci ; 42(4): 283-299, 2021 04.
Article in English | MEDLINE | ID: mdl-33581873

ABSTRACT

G protein-coupled receptors (GPCRs) are the largest class of cell surface receptors in the genome and the most successful family of targets of FDA-approved drugs. New frontiers in GPCR drug discovery remain, however, as achieving receptor subtype selectivity and controlling off- and on-target side effects are not always possible with classic agonist and antagonist ligands. These challenges may be overcome by focusing development efforts on allosteric ligands that confer signaling bias. Biased allosteric modulators (BAMs) are an emerging class of GPCR ligands that engage less well-conserved regulatory motifs outside the orthosteric pocket and exert pathway-specific effects on receptor signaling. The unique ways that BAMs texturize receptor signaling present opportunities to fine-tune physiology and develop safer, more selective therapeutics. Here, we provide a conceptual framework for understanding the pharmacology of BAMs, explore their therapeutic potential, and discuss strategies for their discovery.


Subject(s)
Drug Discovery , Receptors, G-Protein-Coupled , Allosteric Regulation , Ligands , Signal Transduction
13.
Science ; 371(6534)2021 03 12.
Article in English | MEDLINE | ID: mdl-33479120

ABSTRACT

Heterotrimeric guanine nucleotide-binding protein (G protein)-coupled receptors (GPCRs) are common drug targets and canonically couple to specific Gα protein subtypes and ß-arrestin adaptor proteins. G protein-mediated signaling and ß-arrestin-mediated signaling have been considered separable. We show here that GPCRs promote a direct interaction between Gαi protein subtype family members and ß-arrestins regardless of their canonical Gα protein subtype coupling. Gαi:ß-arrestin complexes bound extracellular signal-regulated kinase (ERK), and their disruption impaired both ERK activation and cell migration, which is consistent with ß-arrestins requiring a functional interaction with Gαi for certain signaling events. These results introduce a GPCR signaling mechanism distinct from canonical G protein activation in which GPCRs cause the formation of Gαi:ß-arrestin signaling complexes.


Subject(s)
GTP-Binding Protein alpha Subunits, Gi-Go/metabolism , Receptors, G-Protein-Coupled/metabolism , beta-Arrestins/metabolism , Bioluminescence Resonance Energy Transfer Techniques , Cell Movement , Extracellular Signal-Regulated MAP Kinases/metabolism , HEK293 Cells , Humans , Signal Transduction
14.
J Clin Invest ; 130(12): 6616-6630, 2020 12 01.
Article in English | MEDLINE | ID: mdl-33164988

ABSTRACT

Dysregulation of habit formation has been recently proposed as pivotal to eating disorders. Here, we report that a subset of patients suffering from restrictive anorexia nervosa have enhanced habit formation compared with healthy controls. Habit formation is modulated by striatal cholinergic interneurons. These interneurons express vesicular transporters for acetylcholine (VAChT) and glutamate (VGLUT3) and use acetylcholine/glutamate cotransmission to regulate striatal functions. Using mice with genetically silenced VAChT (VAChT conditional KO, VAChTcKO) or VGLUT3 (VGLUT3cKO), we investigated the roles that acetylcholine and glutamate released by cholinergic interneurons play in habit formation and maladaptive eating. Silencing glutamate favored goal-directed behaviors and had no impact on eating behavior. In contrast, VAChTcKO mice were more prone to habits and maladaptive eating. Specific deletion of VAChT in the dorsomedial striatum of adult mice was sufficient to phenocopy maladaptive eating behaviors of VAChTcKO mice. Interestingly, VAChTcKO mice had reduced dopamine release in the dorsomedial striatum but not in the dorsolateral striatum. The dysfunctional eating behavior of VAChTcKO mice was alleviated by donepezil and by l-DOPA, confirming an acetylcholine/dopamine deficit. Our study reveals that loss of acetylcholine leads to a dopamine imbalance in striatal compartments, thereby promoting habits and vulnerability to maladaptive eating in mice.


Subject(s)
Acetylcholine/metabolism , Corpus Striatum , Feeding and Eating Disorders/metabolism , Glutamic Acid/metabolism , Interneurons/metabolism , Adult , Animals , Corpus Striatum/metabolism , Corpus Striatum/physiopathology , Donepezil/pharmacology , Feeding Behavior/drug effects , Feeding and Eating Disorders/drug therapy , Feeding and Eating Disorders/genetics , Feeding and Eating Disorders/physiopathology , Female , Humans , Levodopa/pharmacology , Male , Mice , Mice, Knockout , Middle Aged , Vesicular Acetylcholine Transport Proteins/genetics , Vesicular Acetylcholine Transport Proteins/metabolism
15.
Cell ; 181(6): 1364-1379.e14, 2020 06 11.
Article in English | MEDLINE | ID: mdl-32470395

ABSTRACT

Small molecule neurotensin receptor 1 (NTSR1) agonists have been pursued for more than 40 years as potential therapeutics for psychiatric disorders, including drug addiction. Clinical development of NTSR1 agonists has, however, been precluded by their severe side effects. NTSR1, a G protein-coupled receptor (GPCR), signals through the canonical activation of G proteins and engages ß-arrestins to mediate distinct cellular signaling events. Here, we characterize the allosteric NTSR1 modulator SBI-553. This small molecule not only acts as a ß-arrestin-biased agonist but also extends profound ß-arrestin bias to the endogenous ligand by selectively antagonizing G protein signaling. SBI-553 shows efficacy in animal models of psychostimulant abuse, including cocaine self-administration, without the side effects characteristic of balanced NTSR1 agonism. These findings indicate that NTSR1 G protein and ß-arrestin activation produce discrete and separable physiological effects, thus providing a strategy to develop safer GPCR-targeting therapeutics with more directed pharmacological action.


Subject(s)
Behavior, Addictive/metabolism , Receptors, Neurotensin/metabolism , beta-Arrestins/metabolism , Allosteric Regulation/drug effects , Allosteric Regulation/physiology , Animals , Behavior, Addictive/drug therapy , Cell Line , Female , HEK293 Cells , Humans , Male , Mice , Mice, Inbred C57BL , Models, Animal , Receptors, G-Protein-Coupled/metabolism , Signal Transduction/drug effects , Signal Transduction/physiology , Small Molecule Libraries/pharmacology
16.
Biol Psychiatry ; 87(8): 745-755, 2020 04 15.
Article in English | MEDLINE | ID: mdl-31892408

ABSTRACT

BACKGROUND: Cortical dopaminergic systems are critically involved in prefrontal cortex (PFC) functions, especially in working memory and neurodevelopmental disorders such as schizophrenia. GSK-3ß (glycogen synthase kinase-3ß) is highly associated with cAMP (cyclic adenosine monophosphate)-independent dopamine D2 receptor (D2R)-mediated signaling to affect dopamine-dependent behaviors. However, the mechanisms underlying the GSK-3ß modulation of cognitive function via D2Rs remains unclear. METHODS: This study explored how conditional cell-type-specific ablation of GSK-3ß in D2R+ neurons (D2R-GSK-3ß-/-) in the brain affects synaptic function in the medial PFC (mPFC). Both male and female (postnatal days 60-90) mice, including 140 D2R, 24 D1R, and 38 DISC1 mice, were used. RESULTS: This study found that NMDA receptor (NMDAR) function was significantly increased in layer V pyramidal neurons in mPFC of D2R-GSK-3ß-/- mice, along with increased dopamine modulation of NMDAR-mediated current. Consistently, NR2A and NR2B protein levels were elevated in mPFC of D2R-GSK-3ß-/- mice. This change was accompanied by a significant increase in enrichment of activator histone mark H3K27ac at the promoters of both Grin2a and Grin2b genes. In addition, altered short- and long-term synaptic plasticity, along with an increased spine density in layer V pyramidal neurons, were detected in D2R-GSK-3ß-/- mice. Indeed, D2R-GSK-3ß-/- mice also exhibited a resistance of working memory impairment induced by injection of NMDAR antagonist MK-801. Notably, either inhibiting GSK-3ß or disrupting the D2R-DISC1 complex was able to reverse the mutant DISC1-induced decrease of NMDAR-mediated currents in the mPFC. CONCLUSIONS: This study demonstrates that GSK-3ß modulates cognition via D2R-DISC1 interaction and epigenetic regulation of NMDAR expression and function.


Subject(s)
Cognitive Dysfunction , Receptors, N-Methyl-D-Aspartate , Animals , Epigenesis, Genetic , Female , Glycogen Synthase Kinase 3 beta/genetics , Male , Mice , Nerve Tissue Proteins , Neuronal Plasticity , Prefrontal Cortex/metabolism , Receptors, Dopamine D2/genetics , Receptors, Dopamine D2/metabolism , Receptors, N-Methyl-D-Aspartate/genetics , Receptors, N-Methyl-D-Aspartate/metabolism
17.
Addict Biol ; 25(6): e12823, 2020 11.
Article in English | MEDLINE | ID: mdl-31441201

ABSTRACT

Psychostimulants and opioids increase dopamine (DA) neurotransmission, activating D1 and D2 G protein-coupled receptors. ß-arrestin2 (ßarr2) desensitizes and internalizes these receptors and initiates G protein-independent signaling. Previous work revealed that mice with a global or cell-specific knockout of ßarr2 have altered responses to certain drugs; however, the effects of ßarr2 on the excitability of medium spiny neurons (MSNs), and its role in mediating the rewarding effects of drugs of abuse are unknown. D1-Cre and D2-Cre transgenic mice were crossed with floxed ßarr2 mice to eliminate ßarr2 specifically in cells containing either D1 (D1ßarr2-KO ) or D2 (D2ßarr2-KO ) receptors. We used slice electrophysiology to characterize the role of ßarr2 in modulating D1 and D2 nucleus accumbens MSN intrinsic excitability in response to DA and tested the locomotor-activating and rewarding effects of cocaine and morphine in these mice. Eliminating ßarr2 attenuated the ability of DA to inhibit D2-MSNs and altered the DA-induced maximum firing rate in D1-MSNs. While D1ßarr2-KO mice had mostly normal drug responses, D2ßarr2-KO mice showed dose-dependent reductions in acute locomotor responses to cocaine and morphine, attenuated locomotor sensitization to cocaine, and blunted cocaine reward measured with conditioned place preference. Both D2ßarr2-KO and D1ßarr2-KO mice displayed an enhanced conditioned place preference for the highest dose of morphine. These results indicate that D1- and D2-derived ßarr2 functionally contribute to DA-induced changes in MSN intrinsic excitability and behavioral responses to psychostimulants and opioids dose-dependently.


Subject(s)
Analgesics, Opioid/pharmacology , Central Nervous System Stimulants/pharmacology , Nucleus Accumbens/drug effects , Receptors, Dopamine D2/metabolism , Reward , beta-Arrestin 2/metabolism , Analgesics, Opioid/administration & dosage , Animals , Central Nervous System Stimulants/administration & dosage , Cocaine/administration & dosage , Cocaine/pharmacology , Female , Locomotion/drug effects , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Morphine/administration & dosage , Morphine/pharmacology , Nucleus Accumbens/physiopathology , Receptors, Dopamine D1/genetics , Receptors, Dopamine D1/metabolism , Receptors, Dopamine D2/genetics
18.
Nat Commun ; 10(1): 5490, 2019 12 02.
Article in English | MEDLINE | ID: mdl-31792216

ABSTRACT

Field cancerization is a premalignant process marked by clones of oncogenic mutations spreading through the epithelium. The timescales of intestinal field cancerization can be variable and the mechanisms driving the rapid spread of oncogenic clones are unknown. Here we use a Cancer rainbow (Crainbow) modelling system for fluorescently barcoding somatic mutations and directly visualizing the clonal expansion and spread of oncogenes. Crainbow shows that mutations of ß-catenin (Ctnnb1) within the intestinal stem cell results in widespread expansion of oncogenes during perinatal development but not in adults. In contrast, mutations that extrinsically disrupt the stem cell microenvironment can spread in adult intestine without delay. We observe the rapid spread of premalignant clones in Crainbow mice expressing oncogenic Rspondin-3 (RSPO3), which occurs by increasing crypt fission and inhibiting crypt fixation. Crainbow modelling provides insight into how somatic mutations rapidly spread and a plausible mechanism for predetermining the intratumor heterogeneity found in colon cancers.


Subject(s)
Colonic Neoplasms/genetics , Disease Models, Animal , Neoplastic Stem Cells/cytology , Animals , Carcinogenesis , Cell Proliferation , Colonic Neoplasms/metabolism , Colonic Neoplasms/physiopathology , Humans , Mice , Mutation , Neoplastic Stem Cells/metabolism , Oncogenes , Thrombospondins/genetics , Thrombospondins/metabolism
19.
J Med Chem ; 62(17): 8357-8363, 2019 09 12.
Article in English | MEDLINE | ID: mdl-31390201

ABSTRACT

Neurotensin receptor 1 (NTR1) is a G protein coupled receptor that is widely expressed throughout the central nervous system where it acts as a neuromodulator. Neurotensin receptors have been implicated in a wide variety of CNS disorders, but despite extensive efforts to develop small molecule ligands there are few reports of such compounds. Herein we describe the optimization of a quinazoline based lead to give 18 (SBI-553), a potent and brain penetrant NTR1 allosteric modulator.


Subject(s)
Central Nervous System Diseases/drug therapy , Drug Discovery , Quinazolines/pharmacology , Receptors, Neurotensin/antagonists & inhibitors , beta-Arrestins/pharmacology , Administration, Oral , Allosteric Regulation/drug effects , Animals , Biological Availability , Central Nervous System Diseases/metabolism , Dopamine Plasma Membrane Transport Proteins/deficiency , Dopamine Plasma Membrane Transport Proteins/metabolism , Dose-Response Relationship, Drug , Female , Locomotion/drug effects , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Molecular Structure , Quinazolines/administration & dosage , Quinazolines/chemistry , Rats , Receptors, Neurotensin/metabolism , Structure-Activity Relationship , beta-Arrestins/administration & dosage , beta-Arrestins/chemistry
20.
ACS Chem Neurosci ; 10(9): 4160-4182, 2019 09 18.
Article in English | MEDLINE | ID: mdl-31387346

ABSTRACT

Dopamine receptors are important G protein-coupled receptors (GPCRs) with therapeutic opportunities for treating Parkinson's Disease (PD) motor and cognitive deficits. Biased D1 dopamine ligands that differentially activate G protein over ß-arrestin recruitment pathways are valuable chemical tools for dissecting positive versus negative effects in drugs for PD. Here, we reveal an iterative approach toward modification of a D1-selective noncatechol scaffold critical for G protein-biased agonism. This approach provided enhanced understanding of the structural components critical for activity and signaling bias and led to the discovery of several novel compounds with useful pharmacological properties, including three highly GS-biased partial agonists. Administration of a potent, balanced, and brain-penetrant lead compound from this series results in robust antiparkinsonian effects in a rodent model of PD. This study suggests that the noncatechol ligands developed through this approach are valuable tools for probing D1 receptor signaling biology and biased agonism in models of neurologic disease.


Subject(s)
Dopamine Agonists/pharmacology , Dopamine/metabolism , Receptors, Dopamine D1/agonists , Receptors, Dopamine D1/drug effects , Animals , Antiparkinson Agents/pharmacology , Arrestins/metabolism , Cyclic AMP/metabolism , Receptors, Dopamine D2/metabolism , Receptors, G-Protein-Coupled/metabolism
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