Dopamine D2 Receptor Expression Research Articles

Studies of the Effects of Monoamines on Orexinergic Neurons in the Hypothalamus of Rat Embryos

Hypothalamic orexinergic neurons are involved in controlling various body functions, particularly energy balance, feeding behavior, wakefulness, and awakening. The direct effects of brain dopamine and serotonin on orexinergic neurons in the hypothalamus are mediated via various subtypes of the corresponding receptors, whose expression has previously been demonstrated in the bodies of orexinergic neurons. Studies were performed on 18-day rat embryos to assess the possibility that these interactions form during the embryonic period of development. Western blotting was used to demonstrate expression of dopamine D1 and serotonin 2C receptor proteins in the developing hypothalamus. Studies using double fluorescence immunolabeling and confocal microscopy confirmed the expression of dopamine D1 and serotonin 2C receptors in the bodies of developing orexin-immunopositive cells in rat embryos. These results provide evidence that the development of morphofunctional interactions between the monoaminergic systems and the orexinergic system occur during the embryonic period of development.

Targeting dopamine receptor D2 as a novel therapeutic strategy in endometrial cancer

BackgroundONC201 is a dopamine receptor D2 (DRD2) antagonist that inhibits tumor growth in preclinical models through ClpP activation to induce integrated stress response pathway and mitochondrial events related to inhibition of cell growth, which is being explored in clinical trials for solid tumors and hematological malignancies. In this study, we investigated the anti-tumorigenic effect of ONC201 in endometrial cancer cell lines and a genetically engineered mouse model of endometrial cancer.MethodsCell proliferation was assessed by MTT and colony formation assays. Cell cycle and apoptosis were evaluated by Cellometer. Invasion capacity was tested using adhesion, transwell and wound healing assays. LKB1fl/flp53fl/fl mouse model of endometrial cancer were fed a control low fat diet versus a high fat diet to mimic diet-induced obesity. Following tumor onset, mice were treated with placebo or ONC201. Metabolomics and lipidomics were used to identify the obesity-dependent effects of ONC201 in the mouse endometrial tumors. DRD2 expression was analyzed by immunohistochemistry in human endometrioid and serous carcinoma specimens. DRD2 mRNA expression from the Cancer Genome Atlas (TCGA) database was compared between the four molecular subtypes of endometrial cancer.ResultsIncreasing DRD2 expression in endometrial cancer was significantly associated with grade, serous histology and stage, as well as worse progression free survival and overall survival. Higher expression of DRD2 mRNA was found for the Copy Number High (CNH) subtype when compared to the other subtypes. ONC201 inhibited cell proliferation, induced cell cycle G1 arrest, caused cellular stress and apoptosis and reduced invasion in endometrial cancer cells. Diet-induced obesity promoted endometrial tumor growth while ONC201 exhibited anti-tumorigenic efficacy in the obese and lean LKB1fl/fl/p53fl/fl mice. Metabolomic analysis demonstrated that ONC201 reversed the obesity-driven upregulation of lipid biosynthesis and reduced protein biosynthesis in obese and lean mice.ConclusionONC201 has anti-tumorigenic effects in endometrial cancer cells and a transgenic mouse model of endometrial cancer, and DRD2 expression was documented in both human serous and endometrioid endometrial cancer. These studies support DRD2 antagonism via ONC201 as a promising therapeutic strategy for endometrial cancer that has already demonstrated pharmacodynamic activity and clinical benefit in both serous and endometrioid endometrial cancer patients.

Czy intensywny wysiłek fizyczny może być lekiem w chorobie Parkinsona?

Badania naukowe wskazują, że aktywność fizyczna wpływa nie tylko na polepszenie ogólnego stanu zdrowia u ludzi, ale dowodzą również specyficznego wpływu ćwiczeń fizycznych na "zdrowie mózgu". Doniesienia ostatnich lat pokazują, że uprawianie aktywności fizycznej w młodym wieku obniża ryzyko zachorowania na choroby neurodegeneracyjne, np. na chorobę Parkinsona. Jednocześnie istnieje wiele badań naukowych, prowadzonych na zwierzętach i ludziach wskazujących, że ćwiczenia fizyczne wywołują korzystne zmiany neuroplastyczne w mózgu zarówno u ludzi zdrowych jak i u chorujących na choroby neurodegeneracyjne. Niniejszy artykuł przedstawia dowody naukowe na zmiany neuroprotekcyjne i neuronaprawcze w mózgu wywołane ćwiczeniami fizycznymi. Zagadnieniem które szerzej podjęto w niniejszym artykule jest wpływ ćwiczeń fizycznych o wysokiej intensywności na zmiany neuroplastyczne i mechanizmy neurofizjologiczne ośrodkowego układu nerwowego oraz na zachowania psychomotoryczne w chorobie Parkinsona. Przedstawione w niniejszym artykule dowody naukowe na pozytywne efekty wysiłku fizycznego na ośrodkowy układ nerwowy to: (i) wzrost wydzielania czynników neurotroficznych, (ii) wzrost ekspresji receptorów dopaminergicznych D2 w prążkowiu, (iii) spadek aktywności transportera dopaminergicznego, (iv) wydłużenie korowo-rdzeniowego okresu ciszy u pacjentów z chorobą Parkinsona zbadanego za pomocą metody przezczaszkowej stymulacji magnetycznej, (v) poprawa sprawności czynności ruchowych oburęcznych, (vi) poprawa funkcji kognitywnych oraz (viii) złagodzenie symptomów ruchowych i pozaruchowych choroby Parkinsona. Wiedza ta jest ważna dla fizjoterapeutów poszukujących skutecznych metod terapii ruchem opartych na dowodach naukowych, wskazujących na wpływ odpowiednio dozowanego wysiłku na patomechanizm choroby Parkinsona, jak również dla modyfikacji systemu opieki zdrowotnej. Niniejszy artykuł ma również uświadomić każdemu z nas, jak wielki wpływ mamy na nasze "zdrowie mózgu" gdy regularnie uprawiamy aktywność fizyczną na każdym etapie naszego życia.

Nucleus Accumbens Medium Spiny Neuron Subtypes Differentially Regulate Stress-Associated Alterations in Sleep Architecture

BackgroundStress is implicated in the pathophysiology of major depression and posttraumatic stress disorder. These conditions share core features, including motivational deficits, heighted anxiety, and sleep dysregulation. Chronic stress produces these same features in rodents, with some individuals being susceptible or resilient, as seen in humans. While stress-induced neuroadaptations within the nucleus accumbens are implicated in susceptibility-related dysregulation of motivational and emotional behaviors, their effects on sleep are unclear. MethodsWe used chemogenetics (DREADDs [designer receptors exclusively activated by designer drugs]) to examine the effects of selective alterations in activity of nucleus accumbens medium spiny neurons expressing dopamine D1 receptors (D1-MSNs) or dopamine D2 receptors (D2-MSNs) on sleep-related end points. Mice were implanted with wireless transmitters enabling continuous collection of data to quantify vigilance states over a 20-day test period. Parallel cohorts were examined in behavioral tests assessing stress susceptibility. ResultsD1- and D2-MSNs play dissociable roles in sleep regulation. Stimulation of inhibitory or excitatory DREADDs expressed in D1-MSNs exclusively affects rapid eye movement sleep, whereas targeting D2-MSNs affects slow wave sleep. The combined effects of D1-MSN inhibition and D2-MSN activation on sleep resemble those of chronic social defeat stress. Alterations in D1-MSN function also affect stress susceptibility in social behavior tests. Elevation of CREB (cAMP response element-binding protein) within D1-MSNs is sufficient to produce stress-like effects on rapid eye movement sleep. ConclusionsIn addition to regulation of motivational and emotional behaviors, the nucleus accumbens also influences sleep, an end point with high translational relevance. These findings provide a neural basis for comorbidity in key features of stress-related illness.

Quinpirole-Mediated Regulation of Dopamine D2 Receptors Inhibits Glial Cell-Induced Neuroinflammation in Cortex and Striatum after Brain Injury.

Brain injury is a significant risk factor for chronic gliosis and neurodegenerative diseases. Currently, no treatment is available for neuroinflammation caused by the action of glial cells following brain injury. In this study, we investigated the quinpirole-mediated activation of dopamine D2 receptors (D2R) in a mouse model of traumatic brain injury (TBI). We also investigated the neuroprotective effects of quinpirole (a D2R agonist) against glial cell-induced neuroinflammation secondary to TBI in adult mice. After the brain injury, we injected quinpirole into the TBI mice at a dose of 1 mg/kg daily intraperitoneally for 7 days. Our results showed suppression of D2R expression and deregulation of downstream signaling molecules in ipsilateral cortex and striatum after TBI on day 7. Quinpirole administration regulated D2R expression and significantly reduced glial cell-induced neuroinflammation via the D2R/Akt/glycogen synthase kinase 3 beta (GSK3-β) signaling pathway after TBI. Quinpirole treatment concomitantly attenuated increase in glial cells, neuronal apoptosis, synaptic dysfunction, and regulated proteins associated with the blood–brain barrier, together with the recovery of lesion volume in the TBI mouse model. Additionally, our in vitro results confirmed that quinpirole reversed the microglial condition media complex-mediated deleterious effects and regulated D2R levels in HT22 cells. This study showed that quinpirole administration after TBI reduced secondary brain injury-induced glial cell activation and neuroinflammation via regulation of the D2R/Akt/GSK3-β signaling pathways. Our study suggests that quinpirole may be a safe therapeutic agent against TBI-induced neurodegeneration.

Striatal bilateral control of skilled forelimb movement

Skilled motor behavior requires bihemispheric coordination, and participation of striatal outputs originating from two neuronal groups identified by distinctive expression of D1 or D2 dopamine receptors. We trained mice to reach for and grasp a single food pellet and determined how the output pathways differently affected forelimb trajectory and task efficiency. We found that inhibition and excitation of D1-expressing spiny projection neurons (D1SPNs) have a similar effect on kinematics results, as if excitation and inhibition disrupt the whole ensemble dynamics and not exclusively one kind of output. In contrast, D2SPNs participate in control of target accuracy. Further, exvivo electrophysiological comparison of naive mice and mice exposed to the task showed stronger striatal neuronal connectivity for ipsilateral D1 and contralateral D2 neurons in relation to the paw used. In summary, while the output pathways work together to smoothly execute skill movements, practice of the movement itself changes synaptic patterns.

Whole-Brain Mapping of Direct Inputs to Dopamine D1 and D2 Receptor-Expressing Medium Spiny Neurons in the Posterior Dorsomedial Striatum.

The posterior dorsomedial striatum (pDMS) is mainly composed of medium spiny neurons (MSNs) expressing either dopamine D1 receptors (D1Rs) or D2Rs. Activation of these two MSN types produces opposing effects on addictive behaviors. However, it remains unclear whether pDMS D1-MSNs or D2-MSNs receive afferent inputs from different brain regions or whether the extrastriatal afferents express distinct dopamine receptors. To assess whether these afferents also contained D1Rs or D2Rs, we generated double transgenic mice, in which D1R-expressing and D2R-expressing neurons were fluorescently labeled. We used rabies virus-mediated retrograde tracing in these mice to perform whole-brain mapping of direct inputs to D1-MSNs or D2-MSNs in the pDMS. We found that D1-MSNs preferentially received inputs from the secondary motor, secondary visual, and cingulate cortices, whereas D2-MSNs received inputs from the primary motor and primary sensory cortices, and the thalamus. We also discovered that the bed nucleus of the stria terminalis (BNST) and the central nucleus of the amygdala (CeA) contained abundant D2R-expressing, but few D1R-expressing, neurons in a triple transgenic mouse model. Remarkably, although limited D1R or D2R expression was observed in extrastriatal neurons that projected to D1-MSNs or D2-MSNs, we found that cortical structures preferentially contained D1R-expressing neurons that projected to D1-MSNs or D2-MSNs, while the thalamus, substantia nigra pars compacta (SNc), and BNST had more D2R-expressing cells that projected to D2-MSNs. Taken together, these findings provide a foundation for future understanding of the pDMS circuit and its role in action selection and reward-based behaviors.

Dopamine D1+D3 receptor density may correlate with parkinson disease clinical features.

ObjectiveDopamine D2‐like receptors – mainly dopamine D2 receptors (D2R) and dopamine D3 receptors (D3R) – are believed to be greatly involved in the pathology of Parkinson disease (PD) progression. However, these receptors have not been precisely examined in PD patients. Our aim was to quantitatively calculate the exact densities of dopamine D1 receptors (D1R), D2R, and D3R in control, Alzheimer disease (AD), and Lewy body disease (LBD) patients (including PD, Dementia with Lewy bodies, and Parkinson disease dementia); and analyze the relationship between dopamine receptors and clinical PD manifestations.MethodsWe analyzed the densities of D1R, D2R, and D3R in the striatum and substantia nigra (SN) using a novel quantitative autoradiography procedure previously developed by our group. We also examined the expression of D2R and D3R mRNA in the striatum by in situ hybridization.ResultsThe results showed that although no differences of striatal D1R were found among all groups; D2R was significantly decreased in the striatum of PD patients when compared with control and AD patients. Some clinical manifestations: age of onset, PD stage, dopamine responsiveness, and survival time after onset; showed a better correlation with striatal D1R + D3R densities combined compared to D1R or D3R alone.InterpretationThere is a possibility that we may infer the results in diagnosis, treatment, and prognosis of PD by detecting D1R + D3R as opposed to using dopamine D1 or D3 receptors alone. This is especially true for elderly patients with low D2R expression as is common in this disease.

Accumbal D2R-medium spiny neurons regulate aversive behaviors through PKA-Rap1 pathway

The nucleus accumbens (NAc) plays a crucial role in various mental activities, including positive and negative reinforcement. We previously hypothesized that a balance between dopamine (DA) and adenosine signals regulates the PKA-Rap1 pathway in medium spiny neurons expressing DA D1 receptors (D1R-MSNs) or D2 receptors (D2R-MSNs) and demonstrated that the PKA-Rap1 pathway in D1R-MSNs is responsible for positive reinforcement. Here, we show the role of the PKA-Rap1 pathway in accumbal D2R-MSNs in negative reinforcement. Mice were exposed to electric foot shock as an aversive stimulus. We monitored the phosphorylation level of Rap1gap S563, which leads to the activation of Rap1. Electric foot shocks increased the phosphorylation level of GluN1 S897 and Rap1gap S563 in the NAc. The aversive stimulus-evoked phosphorylation of Rap1gap S563 was detected in accumbal D2R-MSNs and inhibited by pretreatment with adenosine A2a receptor (A2aR) antagonist. A2aR antagonist-treated mice showed impaired aversive memory in passive avoidance tests. AAV-mediated inhibition of PKA, Rap1, or MEK1 in accumbal D2R-MSNs impaired aversive memory in passive avoidance tests, whereas activation of this pathway potentiated aversive memory. Optogenetic inactivation of mesolimbic DAergic neurons induced place aversion in real-time place aversion tests. Aversive response was attenuated by inhibition of PKA-Rap1 signaling in accumbal D2R-MSNs. These results suggested that accumbal D2R-MSNs regulate aversive behaviors through the A2aR-PKA-Rap1-MEK pathway. Our findings provide a novel molecular mechanism for regulating negative reinforcement.

Dopamine receptor D2 inhibition alleviates diabetic hepatic stellate cells fibrosis by regulating the TGF-β1/Smads and NFκB pathways.

Diabetic hepatic fibrosis (DHF) is a progressive liver disease and a chronic complication of diabetes mellitus. The main cause of DHF is the activation of quiescent hepatic stellate cells (HSCs) by high glucose stimulation. Dopamine receptor D2 (DRD2)-mediated dopamine signalling can be involved in the regulation of diabetic liver disease, but the exact role of DRD2 in DHF is still poorly understood. This study aimed to investigate the protective effect of DRD2 inhibition on diabetic liver fibrosis and the potential mechanism. We established both streptozotocin (STZ)-induced type 1 diabetes (T1D, fed for 20weeks) rat model and high glucose (HG, 40 mmol/L)-stimulated HSCs model. The results from both the rats with STZ and the HSCs treated with HG showed increased expression of DRD2, NOX-5, inflammation-related proteins (IL-6 and TNFα) and fibrosis-related proteins (TGF-β1, CO-Ⅰ/Ⅲ/ IV, MMP-2/9 and fibronectin). In vivo, the serum aspartate aminotransferase (AST), alanine aminotransferase (ALT) and total antioxidant capacity (T-AOC) levels were significantly increased, and hematoxylin-eosin (HE) staining, Masson staining, and electron microscopy revealed liver lesions and hepatocyte injury. In addition, HG-treated HSCs exhibited altered oxidative stress - related indexes, including superoxide dismutase (SOD), malondialdehyde (MDA) and reactive oxygen species (ROS), changed and abnormally proliferated in vitro. TGF-β1, the phosphorylated Smad2, nuclear NFκB-p65, phosphorylated NFκB-p65 and phosphorylated IκBα were also increased. Interestingly, haloperidol (DRD2 inhibitor) and n-acetyl-L-cysteine (NAC, an active oxygen scavenger) reduced the above-mentioned changes. In conclusion, DRD2 inhibition can reduce diabetic HSCs oxidative damage and fibrotic proliferation partly via the TGF-β1/Smads and NFκB pathways.

Social isolation in rats: Effects on animal welfare and molecular markers for neuroplasticity.

Early life stress compromises brain development and can contribute to the development of mental illnesses. A common animal model used to study different facets of psychiatric disorders is social isolation from early life on. In rats, this isolation can induce long-lasting alterations in molecular expression and in behavior. Since social isolation models severe psychiatric symptoms, it is to be expected that it affects the overall wellbeing of the animals. As also promoted by the 3Rs principle, though, it is pivotal to decrease the burden of laboratory animals by limiting the number of subjects (reduce, replace) and by improving the animals’ wellbeing (refine). The aim of this study was therefore to test possible refinement strategies such as resocialization and mere adult social isolation. We examined whether the alternatives still triggered the necessary phenotype while minimizing the stress load on the animals. Interestingly, we did not find reduced wellbeing-associated burrowing performance in isolated rats. The hyperactive phenotype seen in socially isolated animals was observed for rats undergoing the adult-only isolation, but resocializing ameliorated the locomotor abnormality. Isolation strongly affected markers of neuroplasticity in the prefrontal cortex independent of timing: mRNA levels of Arc, Bdnf and the pool of Bdnf transcripts with the 3’ long UTR were reduced in all groups. Bdnf splice variant IV expression was reduced in lifelong-isolated animals. Some of these deficits normalized after resocialization; likewise, exon VI Bdnf mRNA levels were reduced only in animals persistently isolated. Conversely, social deprivation did not affect the expression of Gad67 and Pvb, two GABAergic markers, whereas changes occurred in the expression of dopamine d1 and d2 receptors. As adult isolation was sufficient to trigger the hyperactive phenotype and impaired neuroplasticity in the prefrontal cortex, it could be a candidate for a refinement strategy for certain research questions. To fully grade the severity of post-weaning social isolation and the alternatives, adult isolation and resocialization, a more profound and multimodal assessment approach is necessary.

Influence of serum concentration in retinoic acid and phorbol ester induced differentiation of SH-SY5Y human neuroblastoma cell line.

Numerous protocols to establish dopaminergic phenotype in SH-SY5Y cells have been reported. In most of these protocols there are variations in concentration of serum used. In this paper, we compared the effects of high (10%), low (3%) and descending (2.5%/1%) serum concentration in differentiation medium containing different proportion of retinoic acid (RA) and 12-O-Tetradecanoylphorbol-13-acetate (TPA) or RA-only on the undifferentiated SH-SY5Y cells with regards to cell morphology, biochemical and gene expression alterations. Cells differentiated in culture medium containing low and descending serum concentrations showed increased number of neurite projections and reduced proliferation rates when compared to undifferentiated cells. The SH-SY5Y cells differentiated in culture medium containing 3% RA and low serum or descending (2.5%/1% RA/TPA) were found to be more susceptible to 6-hydroxydopamine (6-OHDA) induced cytotoxicity. Cells differentiated with RA/TPA or RA differentiated showed increased production of the α-synuclein (SNCA) neuroprotein and dopamine neurotransmitter compared to undifferentiated cells, regardless serum concentrations used. There was no significant difference in the expression of tyrosine hydroxylase (TH) gene between undifferentiated and differentiated SH-SY5Y cells. However, the expression of dopamine receptor D2 (DRD2) gene was markedly increased (p<0.05) in differentiated cells with 3% serum and RA only when compared to undifferentiated cells. In conclusion, to terminally differentiate SH-SY5Y cells to be used as a cell-based model to study Parkinson's disease (PD) to investigate molecular mechanisms and drug discovery, the optimal differentiation medium should contain 3% serum in RA-only.

Dopamine D1 receptor in the NAc shell is involved in delayed emergence from isoflurane anesthesia in aged mice.

BackgroundDelayed emergence after general anesthesia tends to occur in the elderly population, but the mechanism remains unclear. Apart from age‐related pharmacokinetic changes, the aging‐induced structural and functional alterations in the arousal‐promoting neural substrates should be considered. The nucleus accumbens (NAc) is a crucial arousal‐related nucleus, in which activating medium spiny neurons (MSNs) expressing dopamine D1 receptor (D1R) could facilitate the arousal from natural sleep. Meanwhile, the dopaminergic systems decline with aging in multiple brain regions. However, whether the age‐related decline in D1R in the NAc shell attenuates its arousal‐promoting capacity from general anesthesia remains to be elucidated.MethodsWe first verified the delayed emergence from isoflurane anesthesia and examined the corresponding changes of electroencephalogram (EEG) power in aged mice. In turn, the arousal‐modulating capacity of D1R was characterized in the young and aged cohorts by microinjection of D1R agonist/antagonist into the NAc shell. Furthermore, to address the possible mechanism responsible for the attenuated arousal‐modulating capacity of the aged NAc, the expression of D1R in the NAc shell was measured and compared between young and aged mice.ResultsOur data indicated that compared with young mice, the emergence time in aged mice was notably longer, while EEG power in δ band (1‐4Hz) was significantly higher and power in β band (12‐25Hz) was lower. Activating or inhibiting D1R in the NAc shell by microinjection D1R agonist/antagonist promoted or delayed the emergence process in young mice. Nevertheless, this modulation capacity of D1R in the NAc shell declined in aged mice, respectively. Meanwhile, downregulation of D1R expression in the NAc shell was detected in the aged brain.ConclusionTogether, these results suggest that aging attenuates the arousal‐modulating capacity of D1R in the NAc shell probably through downregulation of D1R expression therein, which may provide a potential explanation and a therapeutic target for increased sensitivity to anesthetics in the elderly patients.

Effects of Dopamine on the Immature Neurons of the Adult Rat Piriform Cortex.

The layer II of the adult piriform cortex (PCX) contains a numerous population of immature neurons. Interestingly, in both mice and rats, most, if not all, these cells have an embryonic origin. Moreover, recent studies from our laboratory have shown that they progressively mature into typical excitatory neurons of the PCX layer II. Therefore, the adult PCX is considered a “non-canonical” neurogenic niche. These immature neurons express the polysialylated form of the neural cell adhesion molecule (PSA-NCAM), a molecule critical for different neurodevelopmental processes. Dopamine (DA) is a relevant neurotransmitter in the adult CNS, which also plays important roles in neural development and adult plasticity, including the regulation of PSA-NCAM expression. In order to evaluate the hypothetical effects of pharmacological modulation of dopaminergic neurotransmission on the differentiation of immature neurons of the adult PCX, we studied dopamine D2 receptor (D2r) expression in this region and the relationship between dopaminergic fibers and immature neurons (defined by PSA-NCAM expression). In addition, we analyzed the density of immature neurons after chronic treatments with an antagonist and an agonist of D2r: haloperidol and PPHT, respectively. Many dopaminergic fibers were observed in close apposition to PSA-NCAM-expressing neurons, which also coexpressed D2r. Chronic treatment with haloperidol significantly increased the number of PSA-NCAM immunoreactive cells, while PPHT treatment decreased it. These results indicate a prominent role of dopamine, through D2r and PSA-NCAM, on the regulation of the final steps of development of immature neurons in the adult PCX.

GC-TOF-MS-Based Metabolomic Analysis and Evaluation of the Effects of HX106, a Nutraceutical, on ADHD-Like Symptoms in Prenatal Alcohol Exposed Mice.

Attention deficit hyperactivity disorder (ADHD) is a neurodevelopmental disorder that occurs in children characterized by inattention and hyperactivity. Prenatal alcohol exposure (PAE) can disrupt fetal neuronal development and cause an ADHD-like hyperactive behavior in the offspring. In this study, we hypothesized that metabolic disturbance would involve in ADHD neuropathology and aimed to investigate the changes in metabolite profile in PAE-induced ADHD-like model and the effects of HX106, a nutraceutical, on ADHD-like pathophysiology and metabolite changes. To this end, we administered HX106 to the mouse offspring affected by PAE (OPAE) and assessed the hyperactivity using the open field test. We observed that HX106-treated OPAE showed less hyperactive behavior than vehicle-treated OPAE. The effects of HX106 were found to be related to the regulation of dopamine transporter and D2 dopamine receptor expression. Furthermore, using gas chromatography time-of-flight mass spectrometry-based metabolomics, we explored the metabolite changes among the experimental groups. The metabolite profile, particularly related with the amino acids, linoleic acid and amino sugar pathways, was altered by PAE and reversed by HX106 treatment partially similar to that observed in the control group. Overall, this study suggest that metabolite alteration would be involved in ADHD pathology and that HX106 can be an efficient supplement to overcome ADHD by regulating dopamine signaling-related protein expression and metabolite changes.

Estrogen Deficiency and Colonic Function: Surgical Menopause and Sex Differences in Angiotensin and Dopamine Receptor Interaction.

The physiopathological mechanisms that regulate menopausal and sex differences in colonic transit, inflammatory processes, and efficacy of treatments have not been clarified. The dopaminergic system and renin-angiotensin system coexist in the gut and regulate different processes such as motility, absorption/secretion, and inflammation. We investigated the changes in expression of major angiotensin and dopamine receptors in the colon of male, female, and ovariectomized female mice. Possible interaction between both systems was investigated using male and female mice deficient (ko) for major angiotensin and dopamine receptors. In wild-type mice, colonic tissue from females showed lower angiotensin type 1/angiotensin type 2 ratio (an index of pro-inflammatory/anti-inflammatory renin-angiotensin system balance), lower dopamine D1 and D2 receptor expression, and lower levels of pro-inflammatory and pro-oxidative markers relative to males. Interestingly, ovariectomy increased the expression of pro-inflammatory angiotensin type 1 receptor expression and decreased anti-inflammatory angiotensin type 2 receptor expression, increased D1 and D2 receptor expression, and increased the levels of pro-inflammatory and pro-oxidative markers. Ovariectomy-induced changes were blocked by estrogen replacement. The present results suggest a mutual regulation between colonic angiotensin and dopamine receptors and sex differences in this mutual regulation. Estrogen regulates changes in both angiotensin and dopamine receptor expression, which may be involved in sex- and surgical menopause-related effects on gut motility, permeability, and vulnerability to inflammatory processes.

Lack of dopamine D4 receptor participation in mouse hyperdopaminergic locomotor response

Chronic methamphetamine (METH) treatment induces behavioral sensitization in rodents. During this process, hyperactivation of the mesolimbic dopamine system plays a central role, and dopamine D2-like receptor-based antipsychotics are known to alleviate the behavioral hyperactivity. The atypical antipsychotic, clozapine (Clz), acts partially as a dopamine D4 receptor (D4R) antagonist and mitigates hyperdopaminergic drug addiction and/or comorbid psychotic symptoms; however, it remains unclear whether D4R blockade contributes to the therapeutic effects of Clz. Here, we evaluated the potential role of D4R in regulating hyperdopaminergia-induced behavioral hyperactivity in METH behavioral sensitization and dopamine transporter (DAT) knockdown (KD) mice. Clz or a D4R-selective antagonist, L-745,870, were co-administered to mice with daily METH in a METH sensitization model, and Clz or L-745,870 were administered alone in a DAT KD hyperactivity model. Locomotor activity and accumbal D4R expression were analyzed. Clz suppressed both the initiation and expression of METH behavioral sensitization, as well as DAT KD hyperactivity. However, repetitive Clz treatment induced tolerance to the suppression effect on METH sensitization initiation. In contrast, D4R inhibition by L-745,870 had no effect on METH sensitization or DAT KD hyperactivity. Accumbal D4R expression was similar between METH-sensitized mice with and without Clz co-treatment. In sum, our results suggest the mesolimbic D4R does not participate in behavioral sensitization encoded by hyperdopaminergia, a finding which likely extends to the therapeutic effects of Clz. Therefore, molecular targets other than D4R should be prioritized in the development of future therapeutics for treatment of hyperdopaminergia-dependent neuropsychiatric disorders.

(±)VK4‐40, a novel dopamine D3 receptor partial agonist, attenuates cocaine reward and relapse in rodents

Despite widespread abuse of cocaine, there are no approved treatments for cocaine use disorder. Chronic cocaine use is associated with up-regulated dopamine D3 receptor expression in the brain. Therefore, most D3 -based medication development has focused on D3 antagonists. However, D3 antagonists do not attenuate cocaine intake under "easy" self-administration conditions, when response requirements are low. We evaluated a novel, highly selective and metabolically stable D3 partial agonist, (±)VK4-40, for its efficacy in reducing cocaine intake and relapse to drug seeking. The impact of (±)VK4-40 on cocaine intake and relapse was evaluated using intravenous self-administration procedures under a fixed-ratio 2 reinforcement schedule and cocaine-primed reinstatement conditions in rats. Optogenetic brain-stimulation reward procedures were used to evaluate the interaction of (±)VK4-40 and cocaine in the mesolimbic dopamine system in mice. Sucrose self-administration in rats and a conditioned place preference paradigm in mice were used to evaluate the abuse potential of (±)VK4-40 alone and other unwanted effects. (±)VK4-40 dose-dependently reduced cocaine self-administration and cocaine-primed reinstatement of drug-seeking behaviour. (±)VK4-40 also inhibited cocaine-enhanced brain-stimulation reward caused by optogenetic stimulation of dopamine neurons in the ventral tegmental area. (±)VK4-40 alone decreased brain-stimulation reward but produced neither conditioned place preference nor place aversion. This new D3 partial agonist also failed to alter oral sucrose self-administration. The novel D3 partial agonist, (±)VK4-40 attenuates cocaine reward and relapse in rodents, without significant unwanted effects. These findings support further investigation of D3 partial agonists as putative treatments for cocaine use disorder.

The Effect of PAOPA, a Novel Allosteric Modulator of Dopamine D2 Receptors, on Signaling Proteins Following Sub-Chronic Administration in Rats.

Allosteric modulators of G-protein coupled receptors regulate receptor activity by binding to sites other than the active site and have emerged as a new and highly desirable class of drugs. PAOPA (3(R)-[(2(S)-pyrrolidinylcarbonyl)amino]-2-oxo-1-pyrrolidineacetamide), a peptidomimetic analog of L-prolyl-L-leucyl-glycinamide, is a potent dopamine D2 receptor allosteric modulator. PAOPA has shown therapeutic effects in pre-clinical models of schizophrenia and extrapyramidal dysfunction. In this study, we sought to examine the biomolecular underpinnings of PAOPA's therapeutic outcomes in pre-clinical models of schizophrenia. Following sub-chronic (daily for 7 days) administration of PAOPA, we assessed levels of dopamine D2 receptors, receptor kinases (GRK2 (G protein-coupled receptor kinase 2) and Arrestin- 3), and phosphorylated mitogen-activated protein kinase (MAPKs), namely, extracellular signal- regulated kinases (ERK1/2) in the hippocampus, medial pre-frontal cortex, nucleus accumbens, pre-frontal cortex, and dorsal striatum via protein quantification. Following 7 days of daily PAOPA treatment, we observed decreased GRK2 and increased dopamine D2 receptor expression in the dorsal striatum. These findings potentially underscore the therapeutic mechanism of action of PAOPA for the positive-like symptoms of schizophrenia in pre-clinical animal models. Additionally, we observed a decline in GRK2 in the hippocampus and an increase in phosphorylated ERK1 in the pre-frontal cortex, suggesting a role of PAOPA in treating cognitive and/or affective dysfunction in pre-clinical models. While further studies are required to elucidate the mechanism of action of PAOPA, this study discusses prior investigations and develops an early framework to describe the therapeutic mechanism of action of PAOPA.

Characterization of dopamine D2 receptor binding, expression and signaling in different brain regions of control and schizophrenia-model Wisket rats

In previous studies we have shown that a three-hit animal model of schizophrenia (Wisket rat) has several behavioral impairments related to the disorder along with altered mu-opioid (MOP) and cannabinoid (CB1) receptor signaling. As the dopamine hypothesis of schizophrenia is central to research in the field, the goal of the present study was to investigate dopaminergic D2 receptor (D2R) functions (binding capacity, G-protein activation and expression) in several brain regions (hippocampus, prefrontal cortex, striatum, olfactory bulb, cerebellum, brainstem, cortex and diencephalon) of control (Wistar) and Wisket rats.It was found that the D2R mediated maximal activation of G-proteins was substantially higher in hippocampus, striatum and olfactory bulb membranes prepared from the Wisket than in control animals, which was accompanied with lower potency of the D2R-mediated G-protein activation. In contrast, enhanced potency was detected in the prefrontal cortex without changes in the maximal activation. In saturation binding assays the maximal binding capacity of D2Rs was higher in the model animals in cerebral cortex, striatum and lower in the brainstem, while no changes in the dissociation constant values were detected. The D2R mRNA expression showed a trend for greater level in the investigated areas, while the D2R protein expression was significantly higher of Wisket rats compared to Wistar animals in the hippocampus and in the prefrontal cortex but not in the cerebellum.This study proved that the Wisket animals show altered D2 receptor expression and function which might be related to the schizophrenia-like symptoms.