The dopamine receptor D1 inhibitor, SKF83566, suppresses GBM stemness and invasion through the DRD1-c-Myc-UHRF1 interactions.

BACKGROUND: Extensive local invasion of glioblastoma (GBM) cells within the central nervous system (CNS) is one factor that severely limits current treatments. The aim of this study was to uncover genes involved in the invasion process, which could also serve as therapeutic targets. For the isolation of invasive GBM cells from non-invasive cells, we used a three-dimensional organotypic co-culture system where glioma stem cell (GSC) spheres were confronted with brain organoids (BOs). Using ultra-low input RNA sequencing (ui-RNA Seq), an invasive gene signature was obtained that was exploited in a therapeutic context. METHODS: GFP-labeled tumor cells were sorted from invasive and non-invasive regions within co-cultures. Ui-RNA sequencing analysis was performed to find a gene cluster up-regulated in the invasive compartment. This gene cluster was further analyzed using the Connectivity MAP (CMap) database. This led to the identification of SKF83566, an antagonist of the D1 dopamine receptor (DRD1), as a candidate therapeutic molecule. Knockdown and overexpression experiments were performed to find molecular pathways responsible for the therapeutic effects of SKF83566. Finally, the effects of SKF83566 were validated in orthotopic xenograft models in vivo. RESULTS: Ui-RNA seq analysis of three GSC cell models (P3, BG5 and BG7) yielded a set of 27 differentially expressed genes between invasive and non-invasive cells. Using CMap analysis, SKF83566 was identified as a selective inhibitor targeting both DRD1 and DRD5. In vitro studies demonstrated that SKF83566 inhibited tumor cell proliferation, GSC sphere formation, and invasion. RNA sequencing analysis of SKF83566-treated P3, BG5, BG7, and control cell populations yielded a total of 32 differentially expressed genes, that were predicted to be regulated by c-Myc. Of these, the UHRF1 gene emerged as the most downregulated gene following treatment, and ChIP experiments revealed that c-Myc binds to its promoter region. Finally, SKF83566, or stable DRD1 knockdown, inhibited the growth of orthotopic GSC (BG5) derived xenografts in nude mice. CONCLUSIONS: DRD1 contributes to GBM invasion and progression by regulating c-Myc entry into the nucleus that affects the transcription of the UHRF1 gene. SKF83566 inhibits the transmembrane protein DRD1, and as such represents a candidate small therapeutic molecule for GBMs.

Structural and expression analysis of the dopamine receptors reveals their crucial roles in regulating the insulin signaling pathway in oysters.

Dopamine performs its critical role upon binding to receptors. Since dopamine receptors are numerous and versatile, understanding their protein structures and evolution status, and identifying the key receptors involved in the modulation of insulin signaling will provide essential clues to investigate the molecular mechanism of neuroendocrine regulating the growth in invertebrates. In this study, seven dopamine receptors were identified in the Pacific oysters (Crassostrea gigas) and were classified into four subtypes according to their protein secondary and tertiary structures, and ligand-binding activities. Of which, DR2 (dopamine receptor 2) and D(2)RA-like (D(2) dopamine receptor A-like) were considered the invertebrate-specific type 1 and type 2 dopamine receptors, respectively. Expression analysis indicated that the DR2 and D(2)RA-like were highly expressed in the fast-growing oyster "Haida No.1". After in vitro incubation of ganglia and adductor muscle with exogenous dopamine and dopamine receptor antagonists, the expression of these two dopamine receptors and ILPs (insulin-like peptides) was also significantly affected. Dual-fluorescence in situ hybridization results showed that D(2)RA-like and DR2 were co-localized with MIRP3 (molluscan insulin-related peptide 3) and MIRP3-like (molluscan insulin-related peptide 3-like) in the visceral ganglia, and were co-localized with ILP (insulin-like peptide) in the adductor muscle. Furthermore, the downstream components of dopamine signaling, including PKA, ERK, CREB, CaMKK1, AKT, and GSK3ß were also significantly affected by the exogenous dopamine and dopamine receptor antagonists. These findings confirmed that dopamine might affect the secretion of ILPs through the invertebrate-specific dopamine receptors D(2)RA-like and DR2, and thus played crucial roles in the growth regulation of the Pacific oysters. Our study establishes the potential regulatory relationship between the dopaminergic system and insulin-like signaling pathway in marine invertebrates.

Time-varying SUVr reflects the dynamics of dopamine increases during methylphenidate challenges in humans.

Dopamine facilitates cognition and is implicated in reward processing. Methylphenidate, a dopamine transporter blocker widely used to treat attention-deficit/hyperactivity disorder, can have rewarding and addictive effects if injected. Since methylphenidate's brain uptake is much faster after intravenous than oral intake, we hypothesize that the speed of dopamine increases in the striatum in addition to its amplitude underly drug reward. To test this we use simulations and PET data of [11C]raclopride's binding displacement with oral and intravenous methylphenidate challenges in 20 healthy controls. Simulations suggest that the time-varying difference in standardized uptake value ratios for [11C]raclopride between placebo and methylphenidate conditions is a proxy for the time-varying dopamine increases induced by methylphenidate. Here we show that the dopamine increase induced by intravenous methylphenidate (0.25 mg/kg) in the striatum is significantly faster than that by oral methylphenidate (60 mg), and its time-to-peak is strongly associated with the intensity of the self-report of "high". We show for the first time that the "high" is associated with the fast dopamine increases induced by methylphenidate.

Gut commensal E. coli outer membrane proteins activate the host food digestive system through neural-immune communication.

The gastrointestinal tract facilitates food digestion, with the gut microbiota playing pivotal roles in nutrient breakdown and absorption. However, the microbial molecules and downstream signaling pathways that activate food digestion remain unexplored. Here, by establishing a food digestion system in C. elegans, we discover that food breakdown is regulated by the interaction between bacterial outer membrane proteins (OMPs) and a neural-immune pathway. E. coli OmpF/A activate digestion by increasing the neuropeptide NLP-12 that acts on the receptor CCKR. NLP-12 is homologous to mammalian cholecystokinin, known to stimulate dopamine, and we found that loss of dopamine receptors or addition of a dopamine antagonist inhibited OMP-mediated digestion. Dopamine and NLP-12-CKR-1 converge to inhibit PMK-1/p38 innate immune signaling. Moreover, directly inhibiting PMK-1/p38 boosts food digestion. This study uncovers a role of bacterial OMPs in regulating animal nutrient uptake and supports a key role for innate immunity in digestion.

Effort-based decision making in response to high-dose androgens: role of dopamine receptors.

INTRODUCTION: Anabolic-androgenic steroids (AAS) are performance-enhancing drugs used by both world-class and rank-and-file athletes. AAS abuse has been linked with risky decision-making, ranging from drunk driving to abusing multiple drugs. Our lab uses operant behavior in rats to test the effects of AAS (testosterone) on decision making. In our previous study, testosterone caused rats to work harder for food reward during an effort discounting (ED) task. ED is sensitive to dopamine in the nucleus accumbens, and AAS alter accumbens dopamine receptor expression. Accordingly, we determined if testosterone increases response to dopamine receptor antagonists during ED. METHODS: Rats were treated chronically with high-dose testosterone (7.5 mg/kg; n = 9) or vehicle (n = 9). We measured baseline preference for the large reward in an ED task, where rats choose between a small easy reward (one lever press for one sugar pellet) and a large difficult reward (2, 5, 10, or 15 presses for three pellets). Preference for the large reward was measured after administration of D1-like (SCH23390, 0.01 mg/kg) or D2-like (eticlopride, 0.06 mg/kg) receptor antagonists. RESULTS: At baseline, testosterone- and vehicle-treated rats showed similar preference for the large reward lever (FR5, testosterone: 68.6 ± 9.7% and vehicle: 85.7 ± 2.5%). SCH23390 reduced large reward preference significantly in both groups (FR5, testosterone: 41.3 ± 9.2%; vehicle: 49.1 ± 8.2%; F(1,16) = 17.7; P < 0.05). Eticlopride decreased large reward preference in both groups, but more strongly in testosterone-treated rats (FR5: testosterone: 37.0 ± 9.7%; vehicle: 56.3 ± 7.8%; F(1,16) = 35.3; P < 0.05). CONCLUSION: Testosterone increases response to dopamine D2-like receptor blockade, and this contributes to previously observed changes in decision-making behaviors.

Dopaminergic neurons project to the nucleus accumbens regulates anxiety-like behaviors through dopamine D1 signaling.

Dopamine (DA) transmission is important in the regulation of mood and anxiety behaviors. However, how specific dopaminergic signaling pathways respond to anxiogenic stimuli as well as regulate behaviors remains unknown. To understand how DA regulates the animal behaviors under anxiety we performed retrograde labeling and c-Fos staining of midbrain DA neurons. Our c-Fos labeling results showed that DA neurons projected to nucleus accumbens (NAc) are activated in animals treated with the elevated plus-maze (EPM). Real-time measurement of DA release using fast scanning cyclic voltammetry (FSCV) in NAc of freely behaving mice showed that increased DA release and more DA transients in the close arms than the open arms in the EPM. Meanwhile, we also observed a reduction of DA level from the close arms to the open arms. Local infusion of DA D1 receptor antagonist, SCH23390 in the core of NAc, leads to an anxiolytic-like effect in the open-field and EPM. These anxiolytic effects were not observed in animals received D2 receptor antagonist sulpiride infusion in the core of NAc. Taken together, our results reveal a novel function of the mesolimbic DA pathway through the D1 receptor in the regulation of anxiety-like behaviors.

Novel Dual-Target µ-Opioid Receptor and Dopamine D3 Receptor Ligands as Potential Nonaddictive Pharmacotherapeutics for Pain Management.

The need for safer pain-management therapies with decreased abuse liability inspired a novel drug design that retains µ-opioid receptor (MOR)-mediated analgesia, while minimizing addictive liability. We recently demonstrated that targeting the dopamine D3 receptor (D3R) with highly selective antagonists/partial agonists can reduce opioid self-administration and reinstatement to drug seeking in rodent models without diminishing antinociceptive effects. The identification of the D3R as a target for the treatment of opioid use disorders prompted the idea of generating a class of ligands presenting bitopic or bivalent structures, allowing the dual-target binding of the MOR and D3R. Structure-activity relationship studies using computationally aided drug design and in vitro binding assays led to the identification of potent dual-target leads (23, 28, and 40), based on different structural templates and scaffolds, with moderate (sub-micromolar) to high (low nanomolar/sub-nanomolar) binding affinities. Bioluminescence resonance energy transfer-based functional studies revealed MOR agonist-D3R antagonist/partial agonist efficacies that suggest potential for maintaining analgesia with reduced opioid-abuse liability.

Low convergent validity of [11C]raclopride binding in extrastriatal brain regions: A PET study of within-subject correlations with [11C]FLB 457.

Dopamine D2 receptors (D2-R) in extrastriatal brain regions are of high interest for research in a wide range of psychiatric and neurologic disorders. Pharmacological competition studies and test-retest experiments have shown high validity and reliability of the positron emission tomography (PET) radioligand [11C]FLB 457 for D2-R quantification in extrastriatal brain regions. However, this radioligand is not available at most research centers. Instead, the medium affinity radioligand [11C]raclopride, which has been extensively validated for quantification of D2-R in the high-density region striatum, has been applied also in studies on extrastriatal D2-R. Recently, the validity of this approach has been questioned by observations of low occupancy of [11C]raclopride in extrastriatal regions in a pharmacological competition study with quetiapine. Here, we utilise a data set of 16 healthy control subjects examined with both [11C]raclopride and [11C]FLB 457 to assess the correlation in binding potential (BPND) in extrastriatal brain regions. BPND was quantified using the simplified reference tissue model with cerebellum as reference region. The rank order of mean regional BPND values were similar for both radioligands, and corresponded to previously reported data, both post-mortem and using PET. Nevertheless, weak to moderate within-subject correlations were observed between [11C]raclopride and [11C]FLB 457 BPND extrastriatally (Pearson's R: 0.30-0.56), in contrast to very strong correlations between repeated [11C]FLB 457 measurements (Pearson's R: 0.82-0.98). In comparison, correlations between repeated [11C]raclopride measurements were low to moderate (Pearson's R: 0.28-0.75). These results are likely related to low signal to noise ratio of [11C]raclopride in extrastriatal brain regions, and further strengthen the recommendation that extrastriatal D2-R measures obtained with [11C]raclopride should be interpreted with caution.

Neural bases for attenuation of morphine withdrawal by Heantos-4: role of l-tetrahydropalmatine.

Severe withdrawal symptoms triggered by cessation of long-term opioid use deter many individuals from seeking treatment. Opioid substitution and α2-adrenergic agonists are the current standard of pharmacotherapy for opioid use disorder in western medicine; however, each is associated with significant complications. Heantos-4 is a non-opioid botanical formulation used to facilitate opioid detoxification in Vietnam. While ongoing clinical use continues to validate its safety and effectiveness, a mechanism of action accounting for these promising effects remains to be specified. Here, we assess the effects of Heantos-4 in a rat model of morphine-dependence and present evidence that alleviation of naloxone-precipitated somatic withdrawal signs is related to an upregulation of mesolimbic dopamine activity and a consequent reversal of a hypodopaminergic state in the nucleus accumbens, a brain region implicated in opioid withdrawal. A central dopaminergic mechanism is further supported by the identification of l-tetrahydropalmatine as a key active ingredient in Heantos-4, which crosses the blood-brain barrier and shows a therapeutic efficacy comparable to its parent formulation in attenuating withdrawal signs. The anti-hypodopaminergic effects of l-tetrahydropalmatine may be related to antagonism of the dopamine autoreceptor, thus constituting a plausible mechanism contributing to the effectiveness of Heantos-4 in facilitating opioid detoxification.

Effects of Doxycycline in Swiss Mice Predictive Models of Schizophrenia.

Schizophrenia patients show very complex symptoms in several psychopathological domains. Some of these symptoms remain poorly treated. Therefore, continued effort is needed to find novel pharmacological strategies for improving schizophrenia symptoms. Recently, minocycline, a second-generation tetracycline, has been suggested as an adjunctive treatment for schizophrenia. The antipsychotic-like effect of doxycycline, a minocycline analog, was investigated here. We found that both minocycline and doxycycline prevented amphetamine-induced prepulse inhibition (PPI) disruption. However, neither of them blocked MK801-induced effects, albeit doxycycline had a modest impact against ketamine-induced effects. Neither c-Fos nor nNOS expression, which was evaluated in limbic regions, were modified after acute or sub-chronic treatment with doxycycline. Therefore, apomorphine inducing either PPI disruption and climbing behavior was not prevented by doxycycline. This result discards a direct blockade of D2-like receptors, also suggested by the lack of doxycycline cataleptic-induced effect. Contrasting, doxycycline prevented SKF 38393-induced effects, suggesting a preferential doxycycline action at D1-like rather than D2-like receptors. However, doxycycline did not bind to the orthosteric sites of D1, D2, D3, D4, 5-HT2A, 5-HT1A, and A2A receptors suggesting no direct modulation of these receptors. Our data corroborate the antipsychotic-like effect of doxycycline. However, these effects are probably not mediated by doxycycline direct interaction with classical receptors enrolled in the antipsychotic effect.

OATP1A2 and OATP2B1 Are Interacting with Dopamine-Receptor Agonists and Antagonists.

Interaction with the dopaminergic system in the central nervous system is either therapeutically intended or it is a side effect. In both cases, dopamine-receptor agonists (DRA) like the ergoline derivative bromocriptine and dopamine-receptor antagonists (DRAn) like metoclopramide have to cross the blood-brain barrier (BBB). The organic anion transporting polypeptides (OATP) 1A2 and 2B1 are cellular uptake carriers for a variety of endogenous and xenobiotic compounds. As both transporters are expressed in endothelial cells of the BBB, the aim of the present study was to determine whether the DRA bromocriptine, cabergoline, and pergolide and the DRAn metoclopramide and domperidone are interacting with OATP1A2 and 2B1 and could therefore be candidate genes modifying wanted and unwanted effects of these drugs. Localization of both transporters in the brain was confirmed using LC-MS/MS and immunofluorescence stainings. For the functional studies, MDCKII cells stably expressing OATP1A2 or 2B1 were used. Initial interaction studies with the well-characterized transporter substrate estrone 3-sulfate revealed that all tested compounds except pergolide inhibit the transport function of both proteins with the most potent effect for bromocriptine (IC50 = 2.2 µM (OATP1A2) and IC50 = 2.5 µM (OATP2B1)). Further studies using the indirect competitive counterflow method identified bromocriptine, cabergoline, and domperidone as substrates of both transporters, whereas metoclopramide was only transported by OATP1A2. These findings were verified for domperidone by direct measurements using its tritium-labeled form as a tracer. Moreover, the transporter-mediated uptake of this compound was sensitive to the OATP1A2 and OATP2B1 inhibitor naringin. In conclusion, this study suggests that OATP1A2 and 2B1 may play a role in the uptake of DR agonists and antagonists into the brain.

Design and Synthesis of Bitopic 2-Phenylcyclopropylmethylamine (PCPMA) Derivatives as Selective Dopamine D3 Receptor Ligands.

2-Phenylcyclopropylmethylamine (PCPMA) analogues have been reported as selective serotonin 2C agonists. On the basis of the same scaffold, we designed and synthesized a series of bitopic derivatives as dopamine D3R ligands. A number of these new compounds show a high binding affinity for D3R with excellent selectivity. Compound (1R,2R)-22e and its enantiomer (1S,2S)-22e show a comparable binding affinity for the D3R, but the former is a potent D3R agonist, while the latter acts as an antagonist. Molecular docking studies revealed different binding poses of the PCPMA moiety within the orthosteric binding pocket of the D3R, which might explain the different functional profiles of the enantiomers. Compound (1R,2R)-30q shows a high binding affinity for the D3R (Ki = 2.2 nM) along with good selectivity, as well as good bioavailability and brain penetration properties in mice. These results reveal that the PCPMA scaffold may serve as a privileged scaffold for the design of aminergic GPCR ligands.

Deep Brain Stimulation of the Medial Forebrain Bundle in a Rodent Model of Depression: Exploring Dopaminergic Mechanisms with Raclopride and Micro-PET.

BACKGROUND: Deep brain stimulation (DBS) of the medial forebrain bundle (MFB) can reverse depressive-like symptoms clinically and in experimental models of depression, but the mechanisms of action are unknown. OBJECTIVES: This study investigated the role of dopaminergic mechanisms in MFB stimulation-mediated behavior changes, in conjunction with raclopride administration and micropositron emission tomography (micro-PET). METHODS: Flinders Sensitive Line (FSL) rats were allocated into 4 groups: FSL (no treatment), FSL+ (DBS), FSL.R (FSL with raclopride), and FSL.R+ (FSL with raclopride and DBS). Animals were implanted with bilateral electrodes targeting the MFB and given 11 days access to raclopride in the drinking water with or without concurrent continuous bilateral DBS over the last 10 days. Behavioral testing was conducted after stimulation. A PET scan using [18F]desmethoxyfallypride was performed to determine D2 receptor availability before and after raclopride treatment. Changes in gene expression in the nucleus accumbens and the hippocampus were assessed using quantitative polymerase chain reaction. RESULTS: Micro-PET imaging showed that raclopride administration blocked 36% of the D2 receptor in the striatum, but the relative level of blockade was reduced/modulated by stimulation. Raclopride treatment enhanced depressive-like symptoms in several tasks, and the MFB DBS partially reversed the depressive-like phenotype. The raclopride-treated MFB DBS animals had increased levels of mRNA coding for dopamine receptor D1 and D2 suggestive of a stimulation-mediated increase in dopamine receptors. CONCLUSION: Data suggest that chronic and continuous MFB DBS could act via the modulation of the midbrain dopaminergic transmission, including impacting on the postsynaptic dopamine receptor profile.

Distinct inactive conformations of the dopamine D2 and D3 receptors correspond to different extents of inverse agonism.

By analyzing and simulating inactive conformations of the highly homologous dopamine D2 and D3 receptors (D2R and D3R), we find that eticlopride binds D2R in a pose very similar to that in the D3R/eticlopride structure but incompatible with the D2R/risperidone structure. In addition, risperidone occupies a sub-pocket near the Na+ binding site, whereas eticlopride does not. Based on these findings and our experimental results, we propose that the divergent receptor conformations stabilized by Na+-sensitive eticlopride and Na+-insensitive risperidone correspond to different degrees of inverse agonism. Moreover, our simulations reveal that the extracellular loops are highly dynamic, with spontaneous transitions of extracellular loop 2 from the helical conformation in the D2R/risperidone structure to an extended conformation similar to that in the D3R/eticlopride structure. Our results reveal previously unappreciated diversity and dynamics in the inactive conformations of D2R. These findings are critical for rational drug discovery, as limiting a virtual screen to a single conformation will miss relevant ligands.

Almost a third of prescribed drugs work by acting on a group of proteins known as GPCRs (short for G-protein coupled receptors), which help to transmit messages across the cell's outer barrier. The neurotransmitter dopamine, for instance, can act in the brain and body by attaching to dopamine receptors, a sub-family of GPCRs. The binding process changes the three-dimensional structure (or conformation) of the receptor from an inactive to active state, triggering a series of molecular events in the cell. However, GPCRs do not have a single 'on' or 'off' state; they can adopt different active shapes depending on the activating molecule they bind to, and this influences the type of molecular cascade that will take place in the cell. Some evidence also shows that classes of GPCRs can have different inactive structures; whether this is also the case for the dopamine D₂ and D₃ receptors remained unclear. Mapping out inactive conformations of receptors is important for drug discovery, as compounds called antagonists can bind to inactive receptors and interfere with their activation. Lane et al. proposed that different types of antagonists could prefer specific types of inactive conformations of the dopamine D₂ and D₃ receptors. Based on the structures of these two receptors, the conformations of D₂ bound with the drugs risperidone and eticlopride (two dopamine antagonists) were simulated and compared. The results show that the inactive conformations of D₂ were very different when it was bound to eticlopride as opposed to risperidone. In addition D₂ and D₃ showed a very similar conformation when attached to eticlopride. The two drugs also bound to the inactive receptors in overlapping but different locations. These computational findings, together with experimental validations, suggest that D₂ and D₃ exist in several inactive states that only allow the binding of specific drugs; these states could also reflect different degrees of inactivation. Overall, the work by Lane et al. contributes to a more refined understanding of the complex conformations of GPCRs, which could be helpful to screen and develop better drugs.

Huntington's Disease: A Review of the Known PET Imaging Biomarkers and Targeting Radiotracers.

Huntington's disease (HD) is a fatal neurodegenerative disease caused by a CAG expansion mutation in the huntingtin gene. As a result, intranuclear inclusions of mutant huntingtin protein are formed, which damage striatal medium spiny neurons (MSNs). A review of Positron Emission Tomography (PET) studies relating to HD was performed, including clinical and preclinical data. PET is a powerful tool for visualisation of the HD pathology by non-invasive imaging of specific radiopharmaceuticals, which provide a detailed molecular snapshot of complex mechanistic pathways within the brain. Nowadays, radiochemists are equipped with an impressive arsenal of radioligands to accurately recognise particular receptors of interest. These include key biomarkers of HD: adenosine, cannabinoid, dopaminergic and glutamateric receptors, microglial activation, phosphodiesterase 10 A and synaptic vesicle proteins. This review aims to provide a radiochemical picture of the recent developments in the field of HD PET, with significant attention devoted to radiosynthetic routes towards the tracers relevant to this disease.

Multi-target dopamine D3 receptor modulators: Actionable knowledge for drug design from molecular dynamics and machine learning.

Local changes in the structure of G-protein coupled receptors (GPCR) binders largely affect their pharmacological profile. While the sought efficacy can be empirically obtained by introducing local modifications, the underlining structural explanation can remain elusive. Here, molecular dynamics (MD) simulations of the eticlopride-bound inactive state of the Dopamine D3 Receptor (D3DR) have been clustered using a machine learning-based approach in the attempt to rationalize the efficacy change in four congeneric modulators. Accumulating extended MD trajectories of receptor-ligand complexes, we observed how the increase in ligand flexibility progressively destabilized the crystal structure of the inactivated receptor. To prospectively validate this model, a partial agonist was rationally designed based on structural insights and computational modeling, and eventually synthesized and tested. Results turned out to be in line with the predictions. This case study suggests that the investigation of ligand flexibility in the framework of extended MD simulations can assist and inform drug design strategies, highlighting its potential role as a powerful in silico counterpart to functional assays.

Binding of the D3-preferring antipsychotic candidate F17464 to dopamine D3 and D2 receptors: a PET study in healthy subjects with [11C]-(+)-PHNO.

RATIONALE: F17464, a dopamine D3 receptor antagonist with relatively high D3 selectivity (70 fold vs D2 in vitro), exhibits an antipsychotic profile in preclinical studies, and therapeutic efficacy was demonstrated in a randomized placebo-controlled clinical trial in patients with schizophrenia (Bitter et al. Neuropsychopharmacology 44(11):1917-1924, 2019). OBJECTIVE: This open-label study in healthy male subjects aimed at characterizing F17464 binding to D3/D2 receptors and the time course of receptor occupancy using positron emission tomography (PET) imaging with a D3-preferring tracer, [11C]-(+)-PHNO. METHODS: PET scans were performed at baseline and following a single 30 mg or 15 mg dose of F17464 (3 subjects/dose), and blood samples were collected for pharmacokinetic analysis. Receptor occupancy was calculated based upon reduction in binding potential of the tracer following F17464 administration. The relationship between plasma F17464 concentration and D3/D2 receptor occupancy was modeled and the plasma concentration corresponding to 50% receptor occupancy (EC50) calculated. RESULTS: Both doses of F17464 robustly blocked [11C]-(+)-PHNO D3 receptor binding, with substantial occupancy from 1 h post-administration, which increased at 6-9 h (89-98% and 79-87% for the 30 mg and 15 mg groups, respectively) and remained detectable at 22 h. In contrast, D2 binding was only modestly blocked at all time points (< 18%). F17464 exhibited a combination of rapid peripheral kinetics and hysteresis (persistence of binding 22 h post-dose despite low plasma concentration). The best estimate of the EC50 was 19 ng ml-1 (~ 40 nM). CONCLUSION: Overall, F17464 was strongly D3-selective in healthy volunteers, a unique profile for an antipsychotic candidate drug.

Striatal dopamine D2 binding correlates with locus of control: Preliminary evidence from [11C]raclopride Positron Emission Tomography.

The ability to exert control has been widely investigated as a hallmark of adaptive behaviour. Dopamine is recognized as the key neuromodulator mediating various control-related processes. The neural mechanisms underlying the subjective perception of being in control, or Locus of Control (LOC) are however less clear. LOC indicates the subjective tendency to attribute environmental outcomes to one's actions (internal LOC) or instead to external incontrollable factors (external LOC). Here we hypothesized that dopamine levels also relate to LOC. Previous work shows that dopamine signaling mediates learning of action-outcome relationships, outcome predictability, and opportunity cost. Prominent theories propose dopamine dysregulation as the key pathogenetic mechanism in schizophrenia and depression. Critically, external LOC is a risk factor for schizophrenia and depression, and predicts increased vulnerability to stress. However, a direct link between LOC and dopamine levels in healthy control had not been demonstrated. The purpose of our study was to investigate this link. Using [11C]raclopride Positron Emission Tomography we tested the relationship between D2 receptor binding in the striatum and LOC (measured with the Rotter Locus of Control scale) in 15 healthy volunteers. Our results show a large and positive correlation: increased striatal D2 binding was associated with External LOC. This finding opens promising avenues for the study of several psychological impairments that have been associated with both dopamine and LOC, such as addiction, schizophrenia, and depression.

Dopaminergic Mechanisms Underlying Normal Variation in Trait Anxiety.

Trait anxiety has been associated with altered activity within corticolimbic pathways connecting the amygdala and rostral anterior cingulate cortex (rACC), which receive rich dopaminergic input. Though the popular culture uses the term "chemical imbalance" to describe the pathophysiology of psychiatric conditions such as anxiety disorders, we know little about how individual differences in human dopamine neurochemistry are related to variation in anxiety and activity within corticolimbic circuits. We addressed this issue by examining interindividual variability in dopamine release at rest using [11C]raclopride positron emission tomography (PET), functional connectivity between amygdala and rACC using resting-state functional magnetic resonance imaging (fMRI), and trait anxiety measures in healthy adult male and female humans. To measure endogenous dopamine release, we collected two [11C]raclopride PET scans per participant. We contrasted baseline [11C]raclopride D2/3 receptor binding and D2/3 receptor binding following oral methylphenidate administration. Methylphenidate blocks the dopamine transporter, which increases extracellular dopamine and leads to reduced [11C]raclopride D2/3 receptor binding via competitive displacement. We found that individuals with higher dopamine release in the amygdala and rACC self-reported lower trait anxiety. Lower trait anxiety was also associated with reduced rACC-amygdala functional connectivity at baseline. Further, functional connectivity showed a modest negative relationship with dopamine release such that reduced rACC-amygdala functional connectivity was accompanied by higher levels of dopamine release in these regions. Together, these findings contribute to hypodopaminergic models of anxiety and support the utility of combining fMRI and PET measures of neurochemical function to advance our understanding of basic affective processes in humans.SIGNIFICANCE STATEMENT It is common wisdom that individuals vary in their baseline levels of anxiety. We all have a friend or colleague we know to be more "tightly wound" than others, or, perhaps, we are the ones marveling at others' ability to "just go with the flow." Although such observations about individual differences within nonclinical populations are commonplace, the neural mechanisms underlying normal variation in trait anxiety have not been established. Using multimodal brain imaging in humans, this study takes initial steps in linking intrinsic measures of neuromodulator release and functional connectivity within regions implicated in anxiety disorders. Our findings suggest that in healthy adults, higher levels of trait anxiety may arise, at least in part, from reduced dopamine neurotransmission.

Dopamine and serotonin antagonists fail to alter the discriminative stimulus properties of ±methylenedioxymethamphetamine.

Most studies on discriminative stimulus effects of 3,4-methylenedioxymethamphetamine (MDMA) have been conducted using a relatively low dose (1.5 mg/kg), and those studies have invariably implicated serotonergic mechanisms. In contrast, dopaminergic mechanisms mediate the discriminative stimulus effects of amphetamine (AMPH). Some studies have suggested that the discriminative stimulus effects of a higher (3.0 mg/kg) dose of MDMA might rely on both serotonergic and dopaminergic mechanisms. This study aimed to determine effects of selective dopamine (DA) and serotonin (5HT) antagonists on the discriminative stimulus properties of AMPH (0.5 mg/kg) and MDMA (3.0 mg/kg). Separate groups of rats were trained to discriminate AMPH (0.5 mg/kg) or MDMA (3.0 mg/kg) from saline using a food-reinforced drug-discrimination procedure. Effects of DA (SCH 23390: 0.003-0.03 mg/kg and eticlopride: 0.03-0.3 mg/kg) or 5HT (ritanserin: 1.0-10.0 mg/kg, WAY-100635: 0.3-1.0 mg/kg and GR129375: 1.0-3.0 mg/kg) antagonists on the discriminative stimulus effects of both drugs were determined. Both DA antagonists dose-dependently decreased the AMPH but not the MDMA discrimination. None of the 5HT antagonists altered the discriminative stimulus effects of either drug. The MDMA (3.0 mg/kg) stimulus comprises both a DAergic and 5HTergic response, and the results suggest that either one is sufficient, but not required, to maintain the stimulus effects.