Dopamine D5 receptor agonist high affinity and constitutive activity profile conferred by carboxyl-terminal tail sequence.
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The mammalian dopamine D1-like receptor gene family is comprised of two members, termed D1/D1A and D5/D1B. In an attempt to define the role of the carboxyl terminal (CT) tail in the expression of D5 subtype-specific pharmacological and constitutive activity profiles, we examined a series of D5 receptor chimeras in which only the CT tail was swapped with corresponding sequences encoding human/vertebrate D1-like receptors. D5/D1(CT) or D5/D1D(CT) tail substitution mutants displayed a rank order of potency and agonist affinities virtually mimicking wild-type (wt) D1 receptors, as indexed by both ligand binding and dopamine-stimulated cAMP accumulation assays, and, similar to wt D1 receptors, did not exhibit receptor constitutive activity or responsiveness to inverse agonists. D1/D5(CT) or D1/D1D(CT) tail receptor mutants displayed agonist pharmacological and functional characteristics not significantly different from parental D1 or mutant D5/D1(CT) and D5/D1D(CT) receptors. The affinities for numerous antagonists remained essentially unchanged for all receptor chimeras relative to parental wt receptors. A series of stepwise D5-CT-tail truncation/deletion mutants identified the region encoded by amino acids 438-448 and particularly Gln(439), as necessary and sufficient for the full expression of high affinity agonist and functional D5 receptor characteristics. Site-directed mutagenesis of the highly conserved D5/D1B receptor residue Gln(439)-(Ala/Ile), converts the full-length D5 receptor to one displaying "super" D5 characteristics with expressed affinities for discriminating agonists approximately 4- to 5-fold higher than wt D5 but without any concomitant increases of agonist-independent basal cAMP accumulation or intrinsic activity. Taken together, these data suggest that, in addition to other well characterized receptor domains, the agonist pharmacological and functional signature of the D5/D1B receptor is modulated by sequence-specific motifs within the CT tail and that one conserved amino acid in this region can further regulate D5 agonist high affinity binding interactions independent of receptor constitutive activity. (+info)
Dopamine as a novel antioxidative agent for rat vascular smooth muscle cells through dopamine D(1)-like receptors.
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BACKGROUND: To elucidate the roles of vascular D(1)-like receptors in atherosclerosis, the effects of the specific D(1)-like agonists on platelet-derived growth factor (PDGF)-BB-mediated oxidative stress in vascular smooth muscle cells (VSMCs) were studied. METHODS AND RESULTS: Immunohistochemical studies demonstrated the coexistence of D(1A) and D(1B) dopamine receptors in VSMCs. Western blotting revealed a band of approximately 70 kDa for D(1A) and D(1B) dopamine receptors. VSMCs stimulated by PDGF-BB exhibited increased oxidative stress directly measured by flow cytometry. These effects were prevented by dopamine, SKF 38393, or YM 435, and this prevention was reversed by Sch 23390. These effects were blocked by a specific protein kinase A (PKA) inhibitor, N-(2-[p-bromocinnamylamino]ethyl)-5-isoquinolinesulfonamide (H 89). The PDGF-BB-mediated increase in oxidative stress of VSMCs was significantly suppressed by the indirect phospholipase D (PLD) inhibitor suramin or the specific protein kinase C (PKC) inhibitor calphostin C. Both antisense but neither sense nor scrambled oligonucleotides to D(1A) and D(1B) receptors inhibited dopamine-induced suppression of increase in oxidative stress of VSMCs induced by PDGF-BB. CONCLUSIONS: These findings suggest that vascular D(1)-like receptors (D(1A) and D(1B) receptors) inhibit any increase in oxidative stress of VSMCs, possibly through activation of PKA and suppression of PLD and PKC. (+info)
Severe deficiencies in dopamine signaling in presymptomatic Huntington's disease mice.
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In Huntington's disease (HD), mutation of huntingtin causes selective neurodegeneration of dopaminoceptive striatal medium spiny neurons. Transgenic HD model mice that express a portion of the disease-causing form of human huntingtin develop a behavioral phenotype that suggests dysfunction of dopaminergic neurotransmission. Here we show that presymtomatic mice have severe deficiencies in dopamine signaling in the striatum. These include selective reductions in total levels of dopamine- and cAMP-regulated phosphoprotein, M(r) 32 kDA (DARPP-32) and other dopamine-regulated phosphoprotein markers of medium spiny neurons. HD mice also show defects in dopamine-regulated ion channels and in the D(1) dopamine/DARPP-32 signaling cascade. These presymptomatic defects may contribute to HD pathology. (+info)
Regulation of phosphorylation of the GluR1 AMPA receptor in the neostriatum by dopamine and psychostimulants in vivo.
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The activation of cAMP-dependent protein kinase regulates the physiological activity of AMPA-type glutamate receptors. In this study, phosphorylation of the AMPA receptor subunit GluR1 at Ser(845) was increased in neostriatal slices by activation of D1-type dopamine receptors and by inhibitors of protein phosphatase 1/protein phosphatase 2A. In contrast, Ser(831), a residue which, when phosphorylated by protein kinase C or calcium/calmodulin-dependent kinase II, increases AMPA receptor channel conductance, was unaffected by either D1 or D2 receptor agonists in neostriatal slices. The phosphorylation of Ser(845), but not Ser(831), was strongly increased in neostriatum in vivo in response to the psychostimulants cocaine and methamphetamine. The effects of dopamine and psychostimulants on the phosphorylation of GluR1 were attenuated in dopamine and cAMP-regulated phosphoprotein M(r) 32 kDa (DARPP-32) knock-out mice. These results identify DARPP-32 and AMPA-type glutamate receptors as likely essential cellular effectors for psychostimulant actions. (+info)
In utero cocaine-induced dysfunction of dopamine D1 receptor signaling and abnormal differentiation of cerebral cortical neurons.
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Monoamines modulate neuronal differentiation, and alteration of monoamine neurotransmission during development produces specific changes in neuronal structure, function, and pattern formation. We have previously observed that prenatal exposure to cocaine in a clinically relevant animal model produces increased length of pyramidal neuron dendrites in the anterior cingulate cortex (ACC) postnatally. We now report that cocaine administered intravenously to pregnant rabbits at gestational stages preceding and during cortical histogenesis results in the early onset of hypertrophic dendritic outgrowth in the embryonic ACC. Confocal microscopy of DiI-labeled neurons revealed that the atypical, tortuous dendritic profiles seen postnatally in ACC-cocaine neurons already are apparent in utero. No defects in neuronal growth were observed in visual cortex (VC), a region lacking prominent dopamine innervation. In striking correlation with our in vivo results, in vitro experiments revealed a significant enhancement of spontaneous process outgrowth of ACC neurons isolated from cocaine-exposed fetuses but no changes in neurons derived from visual cortex. The onset of modified growth in vivo is paralleled by reduced D(1A) receptor coupling to its G-protein. These data suggest that the dynamic growth of neurons can be regulated by early neurotransmitter signaling in a selective fashion. Prenatal onset of defects in dopamine receptor signaling contributes to abnormal circuit formation and may underlie specific cognitive and behavioral dysfunction. (+info)
Dissociation of cocaine-antagonist properties and motoric effects of the D1 receptor partial agonists SKF 83959 and SKF 77434.
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Previous studies suggest that D1 receptor partial agonists may be viable candidates for development as pharmacotherapies for cocaine addiction. This study investigated the ability of the D1 receptor partial agonists SKF 83959 and SKF 77434 to modulate the behavioral effects of cocaine and compared these effects with those of the reference D1 receptor antagonist SCH 39166 and D1 receptor agonists SKF 81297 and 6-Br-APB. Squirrel monkeys were trained either to respond under a fixed-interval schedule of stimulus-shock termination or to discriminate cocaine from vehicle (procedures useful for evaluating the behavioral stimulant and subjective effects of cocaine, respectively). Additional monkeys were studied with quantitative observational techniques to evaluate the effects of the drugs on various forms of motor behavior. Like SCH 39166, but unlike SKF 81297 and 6-Br-APB, the D1 receptor partial agonists attenuated the behavioral stimulant and discriminative stimulus effects of cocaine in a dose-dependent manner, although maximum antagonism produced by SKF 77434 was not always as great as that produced by SKF 83959 or SCH 39166. In observational studies, SKF 83959 and SKF 77434 produced less severe disruptions in motor behavior than did SCH 39166 and, for SKF 83959, showed a greater separation between the dose required to antagonize the behavioral effects of cocaine and the dose that induced catalepsy (>/=33-fold). These results suggest that D1 receptor partial agonists can act as functional cocaine antagonists with less severe behavioral effects than D1 receptor antagonists. The prominent cocaine-antagonist properties and the low incidence of motoric side effects of SKF 83959 may reflect its unique binding profile at D1 as well as nondopaminergic receptors. (+info)
Regulation of human D(1), d(2(long)), d(2(short)), D(3) and D(4) dopamine receptors by amiloride and amiloride analogues.
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1. The modulatory effects of the allosteric effectors methylisobutylamiloride (MIA), benzamil and amiloride have been examined at human D(1), D(2), D(3) and D(4) dopamine receptors. The subtype selectivity and the mechanism of action of this allosteric regulation was examined. 2. In radioligand dissociation experiments each modulator accelerated dissociation from all four receptor subtypes indicating allosteric regulation. MIA displayed selectivity for the D(3) subtype for acceleration of radioligand dissociation. 3. In equilibrium binding (pseudo-competition) experiments the three compounds inhibited radioligand binding at the four receptor subtypes. Inhibition curves for D(1), D(2(short)), D(2(long)) and D(3) receptors were described by Hill coefficients exceeding unity and data were fitted best by a model that assumes binding of modulator to both the primary and allosteric binding sites of the receptor (the allosteric/competitive model). 4. At the D(4) subtype, Hill coefficients of unity described the binding data for amiloride and benzamil, consistent with competitive inhibition. The Hill coefficient for MIA at the D(4) subtype was less than unity and data could be fitted well by the allosteric/competitive model, but it was not possible to define unambiguously the modulatory mechanism. For this effect a better definition of the mechanism could be obtained by simultaneous analysis of data obtained in the presence of a range of concentrations of a purely competitive ligand. 5. MIA reduced the potency with which dopamine stimulated [(35)S]-GTPgammaS binding at the D(2) receptor. The effects of MIA could be described by the allosteric/competitive model with effects of MIA to inhibit the binding of dopamine but not its ability to induce a response. (+info)
GABA in the deep layers of the superior Colliculus/Mesencephalic reticular formation mediates the enhancement of startle by the dopamine D1 receptor agonist SKF 82958 in rats.
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GABA transmission in the deep layers of the superior colliculus/deep mesencephalic reticular formation (deep SC/Me) mediates several motor responses, including those expressed after systemic administration of dopamine agonists. In the present study we examined the role of the deep SC/Me in the modulation of the acoustic startle reflex and its enhancement by the dopamine D(1) agonist SKF 82958. Rats were implanted with bilateral cannulas into the deep SC/Me or superficial layers of the SC (super SC) and 1 week later were infused with various compounds. The GABA(A) antagonist bicuculline (0, 5, and 10 ng) produced a dose- and time-dependent enhancement of startle after infusion into the deep SC/Me, but not the super SC. Infusion of the GABA(A) agonist muscimol (0.1 microg) into the deep SC/Me, but not the super SC, blocked the enhancement of startle by systemic SKF 82958 (1 mg/kg) but had no effect on baseline startle by itself. This effect was not produced by infusion of the D(1) antagonist SCH 23390(1 microg) or the glutamate antagonist NBQX (0.1 microg). Deposits of FluoroGold into the deep SC/Me, combined with immunohistochemistry for glutamic acid decarboxylase (GAD), confirmed a direct GABAergic input from the substantia nigra pars reticulata (SNr) to the deep SC/Me. These results suggest that GABA tone in the deep SC/Me modulates the expression of startle as well as the enhancement of startle by dopamine D(1) agonists. On the basis of these data and previous work, we have proposed a striatonigral-tectal-reticular neural pathway mediating the effects of dopamine D(1) agonists on startle. (+info)