Inhibition of STAT-mediated cytokine responses to chemically-induced colitis prevents inflammation-associated neurobehavioral impairments.

Depression can be associated with chronic systemic inflammation, and production of peripheral proinflammatory cytokines and upregulation of the kynurenine pathway have been implicated in pathogenesis of depression. However, the mechanistic bases for these comorbidities are not yet well understood. As tryptophan 2,3-dioxygenase (TDO) and indoleamine 2,3-dioxygenase (IDO), which convert tryptophan to kynurenine, are rate-limiting enzymes of the kynurenine pathway, we screened TDO or IDO inhibitors for effects on the production of proinflammatory cytokines in a mouse macrophage cell line. The TDO inhibitor 680C91 attenuated LPS-induced pro-inflammatory cytokines including IL-1ß and IL-6. Surprisingly, this effect was TDO-independent, as it occurred even in peritoneal macrophages from TDO knockout mice. Instead, the anti-inflammatory effects of 680C91 were mediated through the suppression of signal transducer and activator of transcription(STAT) signaling. Furthermore, 680C91 suppressed production of proinflammatory cytokines and STAT signaling in an animal model of inflammatory bowel disease. Specifically, 680C91 effectively attenuated acute phase colon cytokine responses in male mice subjected to dextran sulfate sodium (DSS)-induced colitis. Interestingly, this treatment also prevented the development of anxiodepressive-like neurobehaviors in DSS-treated mice during the recovery phase. The ability of 680C91 to prevent anxiodepressive-like behavior in response to chemically-induced colitis appeared to be due to rescue of attenuated dopamine responses in the nucleus accumbens. Thus, inhibition of STAT-mediated, but TDO-independent proinflammatory cytokines in macrophages can prevent inflammation-associated anxiety and depression. Identification of molecular mechanisms involved may facilitate the development of new treatments for gastrointestinal-neuropsychiatric comorbidity.

Subregion-Specific Regulation of Dopamine D1 Receptor Signaling in the Striatum: Implication for L-DOPA-Induced Dyskinesia.

The striatum is the main structure of the basal ganglia. The striatum receives inputs from various cortical areas, and its subregions play distinct roles in motor and emotional functions. Recently, striatal maps based on corticostriatal connectivity and striosome-matrix compartmentalization were developed, and we were able to subdivide the striatum into seven subregions. Dopaminergic modulation of the excitability of medium spiny neurons (MSNs) is critical for striatal function. In this study, we investigated the functional properties of dopamine signaling in seven subregions of the striatum from male mice. By monitoring the phosphorylation of PKA substrates including DARPP-32 in mouse striatal slices, we identified two subregions with low D1 receptor signaling: the dorsolateral portion of the intermediate/rostral part (DL-IR) and the intermediate/caudal part (IC). Low D1 receptor signaling in the two subregions was maintained by phosphodiesterase (PDE)10A and muscarinic M4 receptors. In an animal model of 6-hydroxydopamine (6-OHDA)-induced hemi-parkinsonism, D1 receptor signaling was upregulated in almost all subregions including the DL-IR, but not in the IC. When L-DOPA-induced dyskinesia (LID) was developed, D1 receptor signaling in the IC was upregulated and correlated with the severity of LID. Our results suggest that the function of the striatum is maintained through the subregion-specific regulation of dopamine D1 receptor signaling and that the aberrant activation of D1 receptor signaling in the IC is involved in LID. Future studies focusing on D1 receptor signaling in the IC of the striatum will facilitate the development of novel therapeutics for LID.SIGNIFICANCE STATEMENT Recent progress in striatal mapping based on corticostriatal connectivity and striosome-matrix compartmentalization allowed us to subdivide the striatum into seven subregions. Analyses of D1 receptor signaling in the seven subregions identified two unique subregions with low D1 receptor signaling: the dorsolateral portion of the intermediate/rostral part (DL-IR) and the intermediate/caudal part (IC). Aberrant activation of D1 receptor signaling in the IC is involved in L-DOPA-induced dyskinesia (LID). Previous studies of LID have mainly focused on the DL-IR, but not on the IC of the striatum. Future studies to clarify aberrant D1 receptor signaling in the IC are required to develop novel therapeutics for LID.

Obligatory roles of dopamine D1 receptors in the dentate gyrus in antidepressant actions of a selective serotonin reuptake inhibitor, fluoxetine.

Depression is a leading cause of disability. Current pharmacological treatment of depression is insufficient, and development of improved treatments especially for treatment-resistant depression is desired. Understanding the neurobiology of antidepressant actions may lead to development of improved therapeutic approaches. Here, we demonstrate that dopamine D1 receptors in the dentate gyrus act as a pivotal mediator of antidepressant actions in mice. Chronic administration of a selective serotonin reuptake inhibitor (SSRI), fluoxetine, increases D1 receptor expression in mature granule cells in the dentate gyrus. The increased D1 receptor signaling, in turn, contributes to the actions of chronic fluoxetine treatment, such as suppression of acute stress-evoked serotonin release, stimulation of adult neurogenesis and behavioral improvement. Importantly, under severely stressed conditions, chronic administration of a D1 receptor agonist in conjunction with fluoxetine restores the efficacy of fluoxetine actions on D1 receptor expression and behavioral responses. Thus, our results suggest that stimulation of D1 receptors in the dentate gyrus is a potential adjunctive approach to improve therapeutic efficacy of SSRI antidepressants.

[Improvement of the learning effectiveness of pharmacology role-play by introducing P-drug reports].

Pharmacology role-play works well for students playing the role of the medical doctor or patient, but students without any roles behave just like observers, resulting in a relatively low learning effectiveness. To improve this issue, a personal drug (P-drug) report was introduced to the role-play program. To examine to what extent the P-drug report affected the learning effectiveness of role-play, we performed questionnaire surveys for players and audiences and subsequent nominal logistic regression analysis. The questionnaire topics were (1) understanding of medical treatment, (2) understanding patient's feelings, (3) improvement of awareness and motivation as a medical doctor, and (4) positive influence upon study attitude. In the topics (1) and (2), the statistical analyses in audiences showed significant relationship between the introduction of the report and observer's recognition of the learning effectiveness, indicating the improvement of learning effectiveness after the introduction of the P-drug report. In players, the percentage of high marks was higher than that in audiences, and no significant differences were found between before and after the introduction of the report. In addition, in the free description, many students realized the importance of selection of therapeutic drugs based on P-drug methods. These results suggest that the introduction of the P-drug report seems useful to make all students participate in the activity of role-play with understanding the selection process of therapeutic drugs, and improves the learning effectiveness of role-play especially in observers. It may be useful to combine P-drug with pharmacology role-play in practical pharmacotherapy education.

DARPP-32/protein phosphatase 1 regulates Rasgrp2 as a novel component of dopamine D1 receptor signaling in striatum.

Dopamine regulates psychomotor function by D1 receptor/PKA-dependent phosphorylation of DARPP-32. DARPP-32, phosphorylated at Thr34 by PKA, inhibits protein phosphatase 1 (PP1), and amplifies the phosphorylation of other PKA/PP1 substrates following D1 receptor activation. In addition to the D1 receptor/PKA/DARPP-32 signaling pathway, D1 receptor stimulation is known to activate Rap1/ERK signaling. Rap1 activation is mediated through the phosphorylation of Rasgrp2 (guanine nucleotide exchange factor; activation) and Rap1gap (GTPase-activating protein; inhibition) by PKA. In this study, we investigated the role of PP1 inhibition by phospho-Thr34 DARPP-32 in the D1 receptor-induced phosphorylation of Rasgrp2 and Rap1gap at PKA sites. The analyses in striatal and NAc slices from wild-type and DARPP-32 knockout mice revealed that the phosphorylation of Rasgrp2 at Ser116/Ser117 and Ser586, but not of Rasgrp2 at Ser554 or Rap1gap at Ser441 or Ser499 induced by a D1 receptor agonist, is under the control of the DARPP-32/PP1. The results were supported by pharmacological analyses using a selective PP1 inhibitor, tautomycetin. In addition, analyses using a PP1 and PP2A inhibitor, okadaic acid, revealed that all sites of Rasgrp2 and Rap1gap were regulated by PP2A. Thus, the interactive machinery of DARPP-32/PP1 may contribute to efficient D1 receptor signaling via Rasgrp2/Rap1 in the striatum.

Thromboxane receptor activation enhances striatal dopamine release, leading to suppression of GABAergic transmission and enhanced sugar intake.

The extracellular dopamine level is regulated not only by synaptic inputs to dopamine neurons but also by local mechanisms surrounding dopaminergic terminals. However, much remains to be investigated for the latter mechanism. Thromboxane A(2) is one of the cyclooxygenase products derived from arachidonic acid, and acts on its cognate G protein-coupled receptor [thromboxane receptor (TP)]. We show here that TP in the striatum locally facilitates dopamine overflow. Intrastriatal injection of a TP agonist increased extracellular dopamine levels in the striatum as measured by in vivo microdialysis. TP stimulation also augmented electrically evoked dopamine overflow from striatal slices. Conversely, TP deficiency reduced dopamine overflow evoked by N-methyl-d-aspartic acid (NMDA) and acetylcholine in striatal slices. TP immunostaining showed that TP is enriched in vascular endothelial cells. Pharmacological blockade of nitric oxide (NO) synthesis and genetic deletion of endothelial NO synthase (eNOS) suppressed NMDA/acetylcholine-induced dopamine overflow. This involvement of NO was abolished in TP-deficient slices, suggesting a role for eNOS-derived NO synthesis in TP-mediated dopamine overflow. As a functional consequence of TP-mediated dopamine increase, a TP agonist suppressed GABAergic inhibitory postsynaptic currents in medium spiny neurons through a D2-like receptor-dependent mechanism. Finally, TP is involved in sucrose intake, a dopamine-dependent motivational behavior. These data suggest that TP stimulation in the striatum locally facilitates dopamine overflow evoked by synaptic inputs via NO synthesis in endothelial cells.

Sex Differences in Dendritic Spine Formation in the Hippocampus and Animal Behaviors in a Mouse Model of Hyperthyroidism.

Thyroid hormones are critical for the regulation of development and differentiation of neurons and glial cells in the central nervous system (CNS). We have previously reported the sex-dependent changes of glial morphology in the brain under the state of hyperthyroidism. Here, we examined sex-dependent changes in spine structure of granule neurons in the dentate gyrus of hippocampus in male and female mice with hyperthyroidism. Using FIB/SEM (focused ion beam/scanning electron microscopy), three-dimensional reconstructed structures of dendritic spines in dentate granule cells were analyzed. Dendritic spine density in granule cells increased significantly in both male and female mice with hyperthyroidism. The decrease in spine volume was observed only in female mice. These findings suggest that hyperthyroidism induces the formation of spines with normal size in male mice but the formation of spines with small size in female mice. To evaluate an outcome of neuronal and previously observed glial changes, behavioral tests were performed. Male mice with hyperthyroidism showed increased locomotor activity in the open field test, while female mice showed elevated immobility time in the tail suspension test, reflecting depression-like behavior. Although direct link between changes in spine and behavioral modifications requires further analysis, our results may help to understand gender-dependent neurological and psychological symptoms observed in patients with hyperthyroidism.

Adolescent psychosocial stress enhances sensitization to cocaine exposure in genetically vulnerable mice.

Development of drug addictive behaviors is modulated by both genetic and environmental risk factors. However, the molecular mechanisms remain unknown. To address the role of adolescent stress in the development of drug addiction, we combined a transgenic mouse model in which a putative dominant-negative form of DISC1 under expressional control of the prion protein promoter is used as a genetic risk factor and adolescent social isolation stress as a gene-environmental interaction (GXE). Repeated cocaine exposure induced greater locomotion in the GXE group than in the other groups. In a conditioned place preference (CPP) test, GXE mice exhibited a significant place preference to the cocaine-conditioned area compared with the other groups. In the nucleus accumbens (NAc) of GXE mice, we found increased enzyme activity of phosphodiesterase-4 (PDE4), predominantly located in NAc D2-receptor-expressing neurons, and enhanced effects of the PDE4 inhibitor rolipram, but not the D1 agonist SKF81297, on the phosphorylation of DARPP-32 and GluA1 at PKA sites. Rolipram injection before cocaine exposure completely inhibited cocaine-induced hyperlocomotion and CPP in the GXE group. These results indicate that GXE enhances sensitivity to repeated cocaine exposure via an increase in PDE4 activity in NAc D2-recptor-expressing neurons, leading to the development of cocaine addictive behaviors.

Distinct roles of PDE4 and PDE10A in the regulation of cAMP/PKA signaling in the striatum.

Phosphodiesterase (PDE) is a critical regulator of cAMP/protein kinase A (PKA) signaling in cells. Multiple PDEs with different substrate specificities and subcellular localization are expressed in neurons. Dopamine plays a central role in the regulation of motor and cognitive functions. The effect of dopamine is largely mediated through the cAMP/PKA signaling cascade, and therefore controlled by PDE activity. We used in vitro and in vivo biochemical techniques to dissect the roles of PDE4 and PDE10A in dopaminergic neurotransmission in mouse striatum by monitoring the ability of selective PDE inhibitors to regulate phosphorylation of presynaptic [e.g., tyrosine hydroxylase (TH)] and postsynaptic [e.g., dopamine- and cAMP-regulated phosphoprotein of M(r) 32 kDa (DARPP-32)] PKA substrates. The PDE4 inhibitor, rolipram, induced a large increase in TH Ser40 phosphorylation at dopaminergic terminals that was associated with a commensurate increase in dopamine synthesis and turnover in striatum in vivo. Rolipram induced a small increase in DARPP-32 Thr34 phosphorylation preferentially in striatopallidal neurons by activating adenosine A(2A) receptor signaling in striatum. In contrast, the PDE10A inhibitor, papaverine, had no effect on TH phosphorylation or dopamine turnover, but instead robustly increased DARPP-32 Thr34 and GluR1 Ser845 phosphorylation in striatal neurons. Inhibition of PDE10A by papaverine activated cAMP/PKA signaling in both striatonigral and striatopallidal neurons, resulting in potentiation of dopamine D(1) receptor signaling and inhibition of dopamine D(2) receptor signaling. These biochemical results are supported by immunohistochemical data demonstrating differential localization of PDE10A and PDE4 in striatum. These data underscore the importance of individual brain-enriched cyclic-nucleotide PDE isoforms as therapeutic targets for neuropsychiatric and neurodegenerative disorders affecting dopamine neurotransmission.

Prostaglandin E2 acts on EP1 receptor and amplifies both dopamine D1 and D2 receptor signaling in the striatum.

Dopamine is involved in multiple neural functions including motor control, reward and motivational processing, learning and reinforcement, and cognitive attention. Dopamine binds to two distinct classes of receptors, namely D1 and D2, to exert these functions. Various endogenous substances regulate dopamine signaling, although their physiological functions are not fully understood. Here, we examined the role of prostaglandin E2 (PGE2) and one of its receptors, EP1, in dopaminergic function in the striatum. EP1 was expressed in both preprodynorphin-containing D1 and preproenkephalin-containing D2 neurons, and PGE2 was produced in striatal slices in response to both D1 and D2 dopamine receptor stimulation. EP1-deficient mice exhibited significant suppression of hyperlocomotion induced by cocaine or SKF81297 (6-chloro-2,3,4,5-tetrahydro-1-phenyl-1H-3-benzazepine hydrobromide), a D1 agonist, and significant attenuation of catalepsy induced by raclopride, a D2 antagonist. Despite these behavioral defects, the extracellular concentration of dopamine was not suppressed in the striatum of EP1-deficient mice, and the densities of D1 and D2 receptors in the striatum were not different between the two genotypes. Stimulation of the D1 receptor induced phosphorylation of dopamine and cAMP-regulated phosphoprotein of 32 kDa (DARPP-32) at Thr34 in striatal slices, and the addition of indomethacin, a PG synthesis inhibitor, attenuated the D1 agonist-induced increase in DARPP-32-Thr34 phosphorylation. The further addition of an EP1 agonist restored the indomethacin-attenuated phosphorylation. Furthermore, both D1- and D2-mediated changes in the DARPP-32-Thr34 phosphorylation were attenuated in EP1-/- slices. These results suggest that PGE2 is formed in response to dopamine receptor stimulation in the striatum and amplifies both D1 and D2 receptor signaling via EP1.

Regulation of DARPP-32 phosphorylation by three distinct dopamine D1-like receptor signaling pathways in the neostriatum.

Dopamine D(1)-like receptors play a key role in dopaminergic signaling. In addition to G(s/olf)/adenylyl cyclase (AC)-coupled D(1) receptors, the presence of D(1)-like receptors coupled to G(q)/phospholipase C (PLC) has been proposed. Benzazepine D(1) receptor agonists are known to differentially activate G(s/olf)/AC and G(q)/PLC signaling. By utilizing SKF83959 and SKF83822, we investigated the D(1)-like receptor signaling cascades, which regulate DARPP-32 phosphorylation at Thr34 (the PKA-site) in mouse neostriatal slices. Treatment with SKF83959 or SKF83822 increased DARPP-32 phosphorylation. The SKF83959- and SKF83822-induced increase in DARPP-32 phosphorylation was largely, but partially, antagonized by a D(1) receptor antagonist, SCH23390, and the residual SCH23390-insensitive increase was abolished by an adenosine A(2A) receptor antagonist. In addition, the SKF83959-induced, SCH23390-sensitive increase in DARPP-32 phosphorylation was enhanced by a PLC inhibitor. Analysis in slices from D(1)R/D(2)R-DARPP-32 mice revealed that both D(1) receptor agonists regulate DARPP-32 phosphorylation in striatonigral, but not in striatopallidal, neurons. Thus, dopamine D(1)-like receptors are coupled to three signaling cascades in striatonigral neurons: (i) SCH23390-sensitive G(s/olf)/AC/PKA, (ii) adenosine A(2A) receptor-dependent G(s/olf)/AC/PKA, and (iii) G(q)/PLC signaling. Interestingly, G(q)/PLC signaling interacts with SCH23390-sensitive G(s/olf)/AC/PKA signaling, resulting in its inhibition. Three signaling cascades activated by D(1)-like receptors likely play a distinct role in dopaminergic regulation of psychomotor functions.

Repeated administration of a dopamine D1 receptor agonist reverses the increased proportions of striatal dopamine D1High and D2High receptors in methamphetamine-sensitized rats.

Repeated administration of psychostimulants produces a behavioural sensitization. Amphetamine-sensitized animals are known to have a higher proportion of high-affinity states of dopamine D2 receptors (D2(High) receptors) in the striatum. We recently reported that repeated administration of a dopamine D1 receptor agonist, R-(+)-SKF38393, reverses the established behavioural sensitization to methamphetamine (MAP). To investigate the mechanisms for reversal of behavioural sensitization, we examined the effect of repeated administration of the dopamine D1 receptor agonist on the proportions of D2(High) receptors and the high-affinity states of dopamine D1 receptors (D1(High) receptors) in the striatum. In the striatum from the MAP-sensitized rats, the proportions of D1(High) and D2(High) receptors (28.5 +/- 1.96 and 57.5 +/- 3.58%) were higher than those in the saline-control rats (12.0 +/- 1.01 and 21.9 +/- 1.60%, respectively). Repeated administration of R-(+)-SKF38393 to the MAP-sensitized rats reduced the increased proportions of D1(High) and D2(High) receptors to 12.4 +/- 1.57 and 31.0 +/- 2.14%, respectively, which were similar to the proportions in the saline-control rats. The total densities of dopamine D1 and D2 receptors were not altered in each treatment condition. The results demonstrate that the proportions of D1(High) and D2(High) receptors in the striatum are elevated in MAP-sensitized rats, and that repeated administration of the dopamine D1 receptor agonist to the MAP-sensitized rats reverses the increased proportions of D1(High) and D2(High) receptors. The findings reveal postsynaptic mechanisms for the development of behavioural sensitization to MAP and the reversal of established sensitization by repeated administration of the dopamine D1 receptor agonist.

Potential for targeting dopamine/DARPP-32 signaling in neuropsychiatric and neurodegenerative disorders.

INTRODUCTION: Alterations in dopamine neurotransmission has been implicated in pathophysiology of neuropsychiatric and neurodegenerative disorders, and DARPP-32 plays a pivotal role in dopamine neurotransmission. DARPP-32 likely influences dopamine-mediated behaviors in animal models of neuropsychiatric and neurodegenerative disorders and therapeutic effects of pharmacological treatment. Areas covered: We will review animal studies on the biochemical and behavioral roles of DARPP-32 in drug addiction, schizophrenia and Parkinson's disease. In general, under physiological and pathophysiological conditions, DARPP-32 in D1 receptor expressing (D1R) -medium spiny neurons (MSNs) promotes dopamine/D1 receptor/PKA signaling, whereas DARPP-32 in D2 receptor expressing (D2R)-MSNs counteracts dopamine/D2 receptor signaling. However, the function of DARPP-32 is differentially regulated in acute and chronic phases of drug addiction; DARPP-32 enhances D1 receptor/PKA signaling in the acute phase, whereas DARPP-32 suppresses D1 receptor/PKA signaling in the chronic phase through homeostatic mechanisms. Therefore, DARPP-32 plays a bidirectional role in dopamine neurotransmission, depending on the cell type and experimental conditions, and is involved in dopamine-related behavioral abnormalities. Expert opinion: DARPP-32 differentially regulates dopamine signaling in D1R- and D2R-MSNs, and a shift of balance between D1R- and D2R-MSN function is associated with behavioral abnormalities. An adjustment of this imbalance is achieved by therapeutic approaches targeting DARPP-32-related signaling molecules.

Reversal of methamphetamine-induced behavioral sensitization by repeated administration of a dopamine D1 receptor agonist.

Repeated intermittent administration of methamphetamine (MAP) produces an enduring hypersensitivity to the motor stimulant effect of MAP, termed behavioral sensitization. Dopamine plays a critical role in the development and expression of behavioral sensitization. Here, we investigated whether a dopamine D1 receptor agonist could reverse behavioral sensitization to MAP. Administration of MAP (1.0 mg/kg, i.p.) to rats once every 3 days for a total of 5 times (days 1-13) induced the enhancement of locomotor activity after MAP challenge (0.5 mg/kg, i.p.) on day 20, verifying the development of behavioral sensitization. The MAP-sensitized rats then received a dopamine D1 agonist, R-(+)-SKF38393 (3.0 mg/kg, i.p.), once a day for 7 consecutive days (days 21-27). Behavioral analysis on days 30 and 41 revealed that the enhanced locomotor activity was reversed by repeated R-(+)-SKF38393 administration. Moreover, repeated R-(+)-SKF38393 administration reversed the increased dopamine release in the striatum after MAP challenge on day 41. Thus, repeated administration of the dopamine D1 receptor agonist induces the reversal of established behavioral sensitization to MAP and of increased dopamine release in the striatum, lasting for at least 2 weeks. Dopamine D1 receptor agonists may be useful therapeutic agents for the treatment of psychostimulant addiction.

Chronic Fluoxetine Induces the Enlargement of Perforant Path-Granule Cell Synapses in the Mouse Dentate Gyrus.

A selective serotonin reuptake inhibitor is the most commonly prescribed antidepressant for the treatment of major depression. However, the mechanisms underlying the actions of selective serotonin reuptake inhibitors are not fully understood. In the dentate gyrus, chronic fluoxetine treatment induces increased excitability of mature granule cells (GCs) as well as neurogenesis. The major input to the dentate gyrus is the perforant path axons (boutons) from the entorhinal cortex (layer II). Through voltage-sensitive dye imaging, we found that the excitatory neurotransmission of the perforant path synapse onto the GCs in the middle molecular layer of the mouse dentate gyrus (perforant path-GC synapse) is enhanced after chronic fluoxetine treatment (15 mg/kg/day, 14 days). Therefore, we further examined whether chronic fluoxetine treatment affects the morphology of the perforant path-GC synapse, using FIB/SEM (focused ion beam/scanning electron microscopy). A three-dimensional reconstruction of dendritic spines revealed the appearance of extremely large-sized spines after chronic fluoxetine treatment. The large-sized spines had a postsynaptic density with a large volume. However, chronic fluoxetine treatment did not affect spine density. The presynaptic boutons that were in contact with the large-sized spines were large in volume, and the volumes of the mitochondria and synaptic vesicles inside the boutons were correlated with the size of the boutons. Thus, the large-sized perforant path-GC synapse induced by chronic fluoxetine treatment contains synaptic components that correlate with the synapse size and that may be involved in enhanced glutamatergic neurotransmission.

Acute effects of resveratrol to enhance cocaine-induced dopamine neurotransmission in the striatum.

Resveratrol is known as an activator of SIRT1, which leads to the deacetylation of histone and non-histone protein substrates, but also has other pharmacological profiles such as the inhibition of monoamine oxidase (MAO)-A and MAO-B. Resveratrol was previously demonstrated to potentiate the rewarding effects of chronic cocaine via activation of SIRT1. However, the role of resveratrol in cocaine responses in the acute phase remains unexplored. Therefore, we investigated the acute effects of resveratrol on cocaine-stimulated dopamine neurotransmission by analyzing protein phosphorylation in neostriatal slices. Treatment with resveratrol (50µM for 30min) enhanced cocaine-induced increases in the phosphorylation of DARPP-32 at Thr34 and GluA1 at Ser845, postsynaptic substrates for dopamine/D1 receptor/PKA signaling, and a cocaine-induced decrease in the phosphorylation of tyrosine hydroxylase at Ser40, a presynaptic substrate for dopamine/D2 receptor signaling. The inhibition of both MAO-A and MAO-B by clorgyline and pargyline, respectively, enhanced the effects of cocaine on DARPP-32 phosphorylation. The acute effect of resveratrol on cocaine-induced DARPP-32 phosphorylation was occluded with inhibition of MAO-A and MAO-B. In behavioral studies, resveratrol (40mg/kg, s.c.) enhanced the increase in locomotor activity induced by acute cocaine administration (10mg/kg, i.p.). Thus, this study provides pharmacological evidence that acute resveratrol enhances cocaine-induced dopamine neurotransmission and behavioral responses, presumably via mechanisms involving the inhibition of dopamine catabolism by MAO-A and MAO-B. Resveratrol may be useful to treat dysregulated dopamine neurotransmission, but it may enhance the risk of developing drug addiction.

Muscarinic receptors acting at pre- and post-synaptic sites differentially regulate dopamine/DARPP-32 signaling in striatonigral and striatopallidal neurons.

Muscarinic receptors, activated by acetylcholine, play critical roles in the functional regulation of medium spiny neurons in the striatum. However, the muscarinic receptor signaling pathways are not fully elucidated due to their complexity. In this study, we investigated the function of muscarinic receptors in the striatum by monitoring DARPP-32 (dopamine- and cAMP-regulated phosphoprotein of M(r) 32 kDa) phosphorylation at Thr34 (the PKA-site) using mouse striatal slices. Treatment of slices with a non-selective muscarinic receptor agonist, oxotremorine (10 µM), rapidly and transiently increased DARPP-32 phosphorylation. The increase in DARPP-32 phosphorylation was completely abolished either by a dopamine D(1) receptor antagonist (SCH23390), tetrodotoxin, genetic deletion of M5 receptors, muscarinic toxins for M1 and M4 receptors, or 6-hydroxydopamine lesioning of dopaminergic neurons, whereas it was enhanced by nicotine. Analysis in D(1)-DARPP-32-Flag/D(2)-DARPP-32-Myc transgenic mice revealed that oxotremorine increases DARPP-32 phosphorylation selectively in D(1)-type/striatonigral, but not in D(2)-type/striatopallidal, neurons. When D(1) and D(2) receptors were blocked by selective antagonists to exclude the effects of released dopamine, oxotremorine increased DARPP-32 Thr34 phosphorylation only in D(2)-type/striatopallidal neurons. This increase required activation of M1 receptors and was dependent upon adenosine A(2A) receptor activity. The results demonstrate that muscarinic receptors, especially M5 receptors, act at presynaptic dopaminergic terminals, regulate the release of dopamine in cooperation with nicotinic receptors, and activate D(1) receptor/DARPP-32 signaling in the striatonigral neurons. Muscarinic M1 receptors expressed in striatopallidal neurons interact with adenosine A(2A) receptors and activate DARPP-32 signaling.

Phosphodiesterase 4 inhibition enhances the dopamine D1 receptor/PKA/DARPP-32 signaling cascade in frontal cortex.

RATIONALE: Alteration of dopamine neurotransmission in the prefrontal cortex, especially hypofunction of dopamine D1 receptors, contributes to psychotic symptoms and cognitive deficit in schizophrenia. D1 receptors signal through the cAMP/PKA second messenger cascade, which is modulated by phosphodiesterase (PDE) enzymes that hydrolyze and inactivate cyclic nucleotides. Though several PDEs are expressed in cortical neurons, the PDE4 enzyme family (PDE4A-D) has been implicated in the control of cognitive function. The best studied isoform, PDE4B, interacts with a schizophrenia susceptibility factor, disrupted in schizophrenia 1 (DISC1). OBJECTIVES: We explore the control of mouse frontal cortex dopamine D1 receptor signaling and associated behavior by PDE4. RESULTS: Inhibition of PDE4 by rolipram induced activation of cAMP/PKA signaling in cortical slices and in vivo, leading to the phosphorylation of DARPP-32 and other postsynaptic and presynaptic PKA-substrates. Rolipram also enhanced DARPP-32 phosphorylation invoked by D1 receptor activation. Immunohistochemical studies demonstrated PDE4A, PDE4B, and PDE4D expression in DARPP-32-positive neurons in layer VI of frontal cortex, most likely in D1 receptor-positive, glutamatergic corticothalamic pyramidal neurons. Furthermore, the ability of rolipram treatment to improve the performance of mice in a sensorimotor gating test was DARPP-32-dependent. CONCLUSIONS: PDE4, which is co-expressed with DARPP-32 in D1 receptor-positive cortical pyramidal neurons in layer VI, modulates the level of D1 receptor signaling and DARPP-32 phosphorylation in the frontal cortex, likely influencing cognitive function. These biochemical and behavioral actions of PDE4 inhibitors may contribute to the hypothesized antipsychotic actions of this class of compounds.

Treatment of the psychostimulant-sensitized animal model of schizophrenia.

Behavioral sensitization to psychostimulants in rodents is associated with the alteration of dopaminergic neurotransmission, and has been proposed as a useful model of schizophrenia due to its progressively intensifying, easily relapsing, and long-lasting features. Pharmacological treatments that reverse the established sensitization may have potential therapeutic values for schizophrenia. The present aim is to review pharmacological treatments that induce the reversal of established sensitization to psychostimulants. In addition, we discuss possible mechanisms for the reversal of sensitization. Reversal of sensitization is induced by chronic dopamine D1 receptor agonism, D2 or D1/D2 receptor agonism combined with mild N-methyl-D-aspartate (NMDA) receptor antagonism or serotonin (5-HT(2A) or 5-HT(3) ) receptor antagonism, 5-HT(1A) receptor agonism, and 5-HT(2A) or 5-HT(3) receptor antagonism. Chronic treatments with these drugs likely adjust altered dopaminergic neurotransmission in sensitized animals. Especially, chronic dopamine D1 receptor agonism, which may adjust mesolimbic hyperdopaminergic and mesocortical hypodopaminergic functions in sensitized animals, is an attractive therapeutic approach for schizophrenia.

Mechanisms for the modulation of dopamine d(1) receptor signaling in striatal neurons.

In the striatum, dopamine D(1) receptors are preferentially expressed in striatonigral neurons, and increase the neuronal excitability, leading to the increase in GABAergic inhibitory output to substantia nigra pars reticulata. Such roles of D(1) receptors are important for the control of motor functions. In addition, the roles of D(1) receptors are implicated in reward, cognition, and drug addiction. Therefore, elucidation of mechanisms for the regulation of dopamine D(1) receptor signaling is required to identify therapeutic targets for Parkinson's disease and drug addiction. D(1) receptors are coupled to G(s/olf)/adenylyl cyclase/PKA signaling, leading to the phosphorylation of PKA substrates including DARPP-32. Phosphorylated form of DARPP-32 at Thr34 has been shown to inhibit protein phosphatase-1, and thereby controls the phosphorylation states and activity of many downstream physiological effectors. Roles of DARPP-32 and its phosphorylation at Thr34 and other sites in D(1) receptor signaling are extensively studied. In addition, functional roles of the non-canonical D(1) receptor signaling cascades that coupled to G(q)/phospholipase C or Src family kinase become evident. We have recently shown that phosphodiesterases (PDEs), especially PDE10A, play a pivotal role in regulating the tone of D(1) receptor signaling relatively to that of D(2) receptor signaling. We review the current understanding of molecular mechanisms for the modulation of D(1) receptor signaling in the striatum.