Visualization of D1 dopamine receptors on living nucleus accumbens neurons and their colocalization with D2 receptors

To examine the substrate for dopamine (DA) synaptic action in the nucleus accumbens (nAcc), we visualized the cellular and subcellular distribution of DA receptors on postnatal nAcc neurons in culture using fluoroprobe derivatives of DA receptor ligands. Previously, we have shown that rhodamine-N-(p-aminophenethyl)-spiperone (NAPS) (10 nM), a derivative of the D2 antagonist spiperone, labels D2-like receptors on living nAcc neurons. We now show that rhodamine-Sch-23390 (30 nM), a derivative of the D1 antagonist, labels D1-like receptors. Putative specific membrane labeling reached a plateau after about 20 min. Labeling was stereospecific, as it was unaffected by competition with (-)-butaclamol, but blocked with (+)-butaclamol. We found that 52 +/- 7% of nAcc medium-sized neurons showed D1 labeling, which extended onto the dendrites. Labeling was also seen on presynaptic terminals, often abutting D1-positive and D1-negative cell bodies, consistent with a presynaptic modulatory role for D1 receptors. Larger neurons, which may be GABAergic or cholinergic interneurons, were also labeled. By sequential labeling first with rhodamine-Sch-23390 and then rhodamine-NAPS, we found that 38 +/- 6% of medium-sized neurons express both D1- and D2-like receptors, indicating that D1-D2 interactions may occur at the level of single postsynaptic neurons.     

Subcellular and subsynaptic distribution of the NR1 subunit of the NMDA receptor in the neostriatum and globus pallidus of the rat: co-localization at synapses with the GluR2/3 subunit of the AMPA receptor

Glutamatergic neurotransmission in the neostriatum and the globus pallidus is mediated through NMDA-type as well as other glutamate receptors and is critical in the expression of basal ganglia function. In order to characterize the cellular, subcellular and subsynaptic localization of NMDA receptors in the neostriatum and globus pallidus, multiple immunocytochemical techniques were applied using antibodies that recognize the NR1 subunit of the NMDA receptor. In order to determine the spatial relationship between NMDA receptors and AMPA receptors, double labelling was performed with the NR1 antibodies and an antibody that recognizes the GluR2 and 3 subunits of the AMPA receptor. In the neostriatum all neurons with characteristics of spiny projection neurons, some interneurons and many dendrites and spines were immunoreactive for NR1. In the globus pallidus most perikarya and many dendritic processes were immunopositive. Immunogold methods revealed that most NR1 labelling is associated with asymmetrical synapses and, like the labelling for GluR2/3, is evenly spread across the synapse. Double immunolabelling revealed that in neostriatum, over 80% of NR1-positive axospinous synapses are also positive for GluR2/3. In the globus pallidus most NR1-positive synapses are positive for GluR2/3. In both regions many synapses labelled only for GluR2/3 were also detected. These results, together with previous data, suggest that NMDA and AMPA receptor subunits are expressed by the same neurons in the neostriatum and globus pallidus and that NMDA and AMPA receptors are, at least in part, colocalized at individual asymmetrical synapses. The synaptic responses to glutamate in these regions are thus likely be mediated by both AMPA and NMDA receptors at the level of individual synapses.     

The effects of D1 or D2 dopamine receptor antagonism in the medial preoptic area, ventral pallidum, or nucleus accumbens on the maternal retrieval response and other aspects of maternal behavior in rats.

The medial preoptic area (MPOA), ventral pallidum (VP), and nucleus accumbens (NA) receive dopaminergic afferents and are involved in maternal behavior. Experiments investigated whether dopamine (DA) receptor antagonism in NA disrupts maternal behavior, determined the type of DA receptor involved, and investigated the involvement of drug spread to VP or MPOA. Injection of SCH 23390 (D1 DA receptor antagonist) into NA of postpartum rats disrupted retrieving at dosage levels that were ineffective when injected into MPOA or VP. Motor impairment was not the cause of the deficit. Injection of eticlopride (D2 DA receptor antagonist) into NA or VP was without effect. Results emphasize the importance of DA action on D1 receptors in NA for retrieval behavior. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Carbamazepine reduces dopamine-mediated behavior in chronic neuroleptic-treated and untreated rats: Implications for treatment of tardive dyskinesia and hyperdopaminergic states.

Chronic treatment with neuroleptic drugs such as haloperidol (HAL) can result in a syndrome of abnormal involuntary movements known as tardive dyskinesia (TD). The authors have obtained evidence that TD in humans is reduced in patients also taking anticonvulsant drugs, primarily carbamazepine (CBZ). To test for a causal role of CBZ in this effect, the authors quantified abnormal movements elicited by dopamine (DA) receptor stimulation in rats (Rattus norvegicus) withdrawn from chronic treatment with HAL or CBZ alone or in combination. The expected increased behavioral responsiveness to combined D1/D2 stimulation in rats treated with HAL for 8 weeks was significantly attenuated by chronic CBZ, which also attenuated behavioral responsiveness in otherwise untreated rats. Striatal D2 DA receptor density was elevated in rats treated chronically with HAL but unaffected by CBZ. Striatal D1 DA receptor density was elevated by chronic CBZ but unaffected by HAL. These findings suggest that by reducing DA supersensitivity, CBZ may be useful in treating TD and other hyperdopaminergic states. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Medial prefrontal cortical D2 and striatolimbic D4 dopamine receptors: common targets for typical and atypical antipsychotic drugs

1. In vitro receptor autoradiography was used to examine the long-term effects of a typical (fluphenazine), atypical (clozapine), or potential atypical antipsychotic (S[+]-N-n-propylnorapomorphine; [+]-NPA) on different dopamine (DA) receptor subtypes. 2. D1-Like and D3 receptor levels were not changed with any treatment in any brain region examined. 3. D2 Receptors in caudate-putamen (CPu), nucleus accumbens (NAc) and olfactory tubercle (OT) were significantly increased by long-term treatment with fluphenazine, but not with clozapine or S[+]-NPA. 4. D2 Receptor levels in medial prefrontal cortex (MPC), but not dorsolateral frontal cortex (DFC), were elevated after repeated daily administration of fluphenazine, clozapine, and S[+]-NPA. 5. D4-Like receptors, assayed under D4-selective conditions, were increased by fluphenazine, clozapine and S(+)-NPA in both NAc and CPu, but by none of these treatments in OT, DFC or MPC. 6. These results support a common role for medial prefrontal cortical D2 and striatolimbic D4 receptors in mediating the clinical actions of typical and atypical antipsychotic drugs.     

D2 dopamine receptor antisense oligodeoxynucleotide inhibits the synthesis of a functional pool of D2 dopamine receptors

In vivo administration of an antisense oligonucleotide targeted toward the D2 dopamine (DA) receptor mRNA (D2 AS) markedly inhibited D2 receptor-mediated behaviors but produced only a relatively small reduction in the levels of D2 DA receptors in mouse striatum. This apparent dissociation between DA receptor-mediated behaviors and the levels of D2 DA receptors was addressed by inhibiting the total number of D2 DA receptors by intraperitoneal administration of the selective, irreversibly acting D2 DA receptor antagonist fluphenazine-N-mustard (FNM) and then determining the effects of D2 AS, administered intracerebroventricularly, on the rate of synthesis of D2 DA receptors and on the recovery of D2 receptor-mediated behaviors. FNM inactivated approximately 90% of D2 DA receptors within 4 hr of treatment, after which the receptors returned to normal levels by approximately 8 days. D2 AS treatment significantly inhibited the rate of recovery of D2 DA receptors in striatum of FNM-treated mice. FNM treatment also produced a number of behavioral alterations, including catalepsy, and the inhibition of stereotypic behavior induced by the D2/D3 DA receptor agonist quinpirole. Both of these behaviors returned to normal within 8 days after FNM treatment. D2 AS treatment delayed the restoration of these FNM-induced behaviors. Thus, it reduced the rate of disappearance of the cataleptic behavior induced by FNM and significantly delayed the restoration of the stereotypic behavior induced by quinpirole. The changes induced by D2 AS on D2 receptor-mediated behaviors were reversed on cessation of D2 AS treatment. A random oligomer given in the same amount and for the same length of time as that of the D2 AS had no significant effects on either D2 DA receptor synthesis or DA receptor-mediated behaviors. These studies demonstrate that in vivo administration of D2 AS decreased the rate of recovery of D2 DA receptors and inhibited the recovery of D2 DA receptor-mediated behaviors after irreversible receptor inactivation and suggest that D2 AS treatment inhibits the synthesis of a functional pool of D2 DA receptors.     

Ultrastructural immunocytochemical localization of neurotensin and the dopamine D2 receptor in the rat nucleus accumbens

The neuroleptic-like effects of neurotensin (NT) are thought to be due to interactions with dopamine (DA) acting primarily at D2 receptors within the nucleus accumbens septi (Acb). Using electron microscopic dual labeling immunocytochemistry, we sought to demonstrate cellular substrates for functional interactions involving NT and DA D2 receptors in the adult rat Acb. Peroxidase reaction product representing D2 receptor-like immunoreactivity (D2-LI) was seen along membranes of Golgi lamellae and multivesicular bodies of perikarya containing immunogold labeling representing NT-LI. Dually labeled somata usually contained highly indented nuclei, a characteristic of aspiny neurons. Dendrites also occasionally colocalized the two immunomarkers. Other somata, dendrites, and all axon terminals were singly labeled with either NT-LI or D2-LI. In distinct sets of terminals, NT-LI was commonly associated with large, dense-cored vesicles, whereas D2-LI was found along the plasmalemma and over nearby small clear vesicles. Each type of terminal comprised approximately 20% of synaptic input to NT-immunoreactive dendrites. Similar proportions of terminals containing NT-LI or D2-LI contacted unlabeled (approximately 55%) or NT-labeled (approximately 35%) dendrites and, occasionally, were observed converging onto common dendrites. Terminals containing NT-LI or D2-LI also were often closely apposed. These findings provide the first ultrastructural evidence that: (1) NT and D2 receptors are colocalized in aspiny neurons and dendrites, (2) NT may produce a direct postsynaptic effect on neurons receiving input from terminals which are presynaptically modulated by DA via D2 receptors, and (3) NT and DA acting at D2 receptors may interact through presynaptic modulation of common axon terminals.     

Hexadecylphosphocholine and 2-modified 1,3-diacylglycerols as effectors of phospholipase D

The effects of mesulergine (100 and 200 microg/kg s.c.), SB 206553 (1 and 2.5 mg/kg i.p.), RP 62203 (2.5 and 4 mg/kg i.p.) and ritanserin (630 microg/kg i.p.) were studied on the extracellular concentration of dopamine (DA) and dihydroxyphenylacetic acid (DOPAC) in the nucleus accumbens of chloral hydrate-anesthetized rats, using intracerebral microdialysis. Mesulergine, a non selective serotonin2C/2B/2A (5-HT2C/2B/2A) receptor antagonist, significantly increased DA release, which reached a peak level (+ 20%) 60 min after drug injection and slowly returned back to baseline values. Mesulergine also caused a dose-dependent increase in DOPAC outflow. Pretreatment with mesulergine (200 microg/kg) did not change the inhibition of DA release induced by apomorphine (100 microg/kg), whereas it prevented the reduction of DOPAC outflow induced by apomorphine (100 microg/kg). Administration of SB 206553, a selective blocker of 5-HT2C/2B receptors, dose-dependently increased DA outflow. The dose of 2.5 mg/kg SB 206553 caused a linear increase of DA output which reached a peak (+75%) 40 min after injection, while 1 mg/kg induced a more gradual increase of DA release which peaked (+54%) 60 min after administration of the drug. Treatment with RP 62203, a selective 5-HT2A receptor antagonist, did not produce any significant effect on DA outflow. Administration of ritanserin, a mixed 5-HT2A/2C receptor antagonist, did not cause any significant change of DA and DOPAC outflow. Taken together, these data indicate that selective blockade of 5-HT2/2B receptor subtypes increases DA release in the rat nucleus accumbens.     

Selective loss of pallidal dopamine D2 receptor density in hepatic encephalopathy

The binding parameters of [3H]SCH 23390 and [3H]spiperone (radioligands for dopamine D1 and D2 receptors, respectively) were investigated in autopsied frontal cortex, caudate nucleus and globus pallidus/putamen of cirrhotic patients who died in hepatic coma as well as in age- and sex-matched controls. Specific [3H]SCH 23390 binding site densities were unchanged in all regions; in contrast, specific [3H]spiperone binding site density was decreased (by 44%, P < 0.001) in the globus pallidus/putamen of patients with HE. Decreased densities of pallidal D2 binding sites could relate to the motor dysfunctions commonly encountered in human HE.     

Neostriatal dopamine and sensory inattention.

Damage to the mesotelencephalic dopamine (DA)-containing projection results in a sensory inattention, characterized by impairments in orientation toward somatosensory, visual, and olfactory stimuli. Experiments with Sprague-Dawley rats investigated which branch of the dopaminergic system is responsible for these deficits. In the 1st approach, DA-innervated forebrain sites were damaged by 6-hydroxydopamine (6-OHDA) injection into, or by electrolytic lesions of, these regions. In the 2nd, Ss were given tegmental 6-OHDA injections that damaged the entire mesotelencephalic projection and subsequently received intracerebral injections of the DA agonist apomorphine into forebrain sites in an attempt to reinstate orientation. Results demonstrate that dopaminergic terminals in the neostriatum are critical for orientation. Unilateral neostriatal 6-OHDA injections or electrolytic lesions reduced orientation to contralateral touch, whereas similar damage to other DA-innervated forebrain structures did not. Results also suggest that dopaminergic terminals in the anterior neostriatum are important for orientation to the touch of the rostral body surface, while those in the posterior neostriatum are most critical for orientation to caudal touch. (32 ref) (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Beauty is in the soul of the beholder: psychological implications of beauty and African American women

An in vitro receptor binding and in vivo microdialysis study was performed to further investigate the modulation of dopamine (DA) D2 receptors by neurotensin (NT) peptides. Saturation experiments with the D2 agonist [3H]NPA (N-propylnorapomorphine) showed that 10 nM of NT, 10 nM of neuromedin N (NN) and 1 nM of the C-terminal NT-(8-13) fragment significantly increased the KD values by 125%, 181%, and 194%, respectively without significantly affecting the Bmax value of the [3H]NPA binding sites in coronal sections of rat ventral forebrain mainly containing the nucleus accumbens (Acb) and the olfactory tubercle. In line with the previous findings that NT can increase GABA release in the Acb and that NT receptors are not found on DA terminals in this brain region, the present in vivo microdialysis study demonstrated that local perfusion of NT (1 nM) counteracted the D2 agonist pergolide (2 mu M) induced inhibition of GABA, but not of DA release in the rat Acb. This result indicates that NT counteracts the D2 agonist induced inhibition of GABA release in the rat Acb, via an antagonistic postsynaptic NT/D2 receptor interaction as also suggested by the inhibitory regulation of D2 receptor affinity in the Acb by the NT peptides demonstrated in the present receptor binding experiments. Thus, the neuroleptic and potential antipsychotic profile of the NT peptides may involve an antagonistic NT/D2 receptor regulation in the ventral striatum.     

Neurogenesis in the mammalian neostriatum and nucleus accumbens: parvalbumin-immunoreactive GABAergic interneurons

We study the neurogenesis of a distinct subclass of rat striatum gamma-aminobutyric acid (GABA)ergic interneurons marked by the calcium-binding protein parvalbumin (PV). Timed pregnant rats are given an intraperitoneal injection of bromodeoxyuridine (BrdU), a marker of cell proliferation, on designated days between embryonic day (E) 11 and E22. Birthdate of PV neurons is determined in the adult neostriatum and nucleus accumbens by using a BrdU-PV double-labeling immunohistochemical technique. PV-immunoreactive interneurons of the neostriatum show maximum birthrates (>10% double-labeling) between E14-E17, whereas PV-immunoreactive interneurons of the nucleus accumbens show maximum double-labeling between E16-E19. In the neostriatum, caudal PV-immunoreactive neurons are born before those at rostral levels, and lateral PV-immunoreactive neurons become postmitotic before medial neurons. In the postcommissural striatum, ventral PV-immunoreactive neurons become postmitotic before dorsal neurons. In the precommissural striatum, ventral neurons are born before dorsal neurons laterally, but a dorsoventral gradient is seen medially. At corresponding coronal levels, PV-immunoreactive neurons of the nucleus accumbens are born shortly after PV neurons of the neostriatum. Analysis of BrdU labeling intensity in the nucleus accumbens shows that medium spiny projection neurons of the shell become postmitotic before neurons of the core. Similarly, PV-immunoreactive interneurons of the nucleus accumbens shell are born before PV interneurons of the core. Compared with cholinergic interneurons of the neostriatum, PV-immunoreactive interneurons are born later, but neurogenetic gradients are similar. The period of striatum PV interneuron genesis encompasses the period for somatostatin interneurons, although the latter neurons do not show neurogenetic gradients, possibly due to heterogeneous subtypes. Consideration of basal telencephalon neurogenesis suggests that subpopulations of striatum interneurons may share common neurogenetic features with phenotypically similar populations in the basal forebrain, with final morphology and connectivity depending on local cues provided by the host environment.     

CI-1007, a dopamine partial agonist and potential antipsychotic agent. I. Neurochemical effects

The receptor binding and biochemical effects of the putative dopamine (DA) partial agonist CI-1007 ([R(+)-1,2,3,6-tetrahydro-4-phenyl- 1-[(3-phenyl-3-cyclohexen-1-yl)methyl]pyridine] maleate) and potential antipsychotic were evaluated with a variety of biochemical methods. In receptor binding studies, CI-1007 bound to rat striatal DA receptors exhibiting a Ki of 3 nM as assessed by inhibition of [3H]N-propylnorapomorphine binding. CI-1007 also exhibited high affinity for cloned human D2L (Ki = 25.5 nM) and D3 (Ki = 16.6 nM) receptors with less affinity for D4.2 receptors (Ki = 90.9 nM). The affinity for serotonin-1A (5-HT-1A), alpha-2 adrenergic and 5-HT-2 receptors was moderate (submicromolar range) and slight or negligible for alpha-1, DA D1 and various other receptors. Unlike dopamine, the inhibition of [3H]spiperone binding was monophasic for CI-1007 and only slightly affected by the addition of Gpp-(NH)p. In vitro CI-1007 antagonized the forskolin-induced increases in cyclic AMP levels in GH4C1 cells expressing the human D2L receptor, having an intrinsic activity of 53% of that seen with the full agonist quinpirole. In vivo CI-1007 antagonized the gamma-butyrolactone (GBL)-induced accumulation of L-3,4-dihydroxyphenylalanine in striatum and mesolimbic regions of rat brain, causing a maximal 64% reversal in striatum, consistent with a partial agonist profile. In microdialysis studies it decreased DA overflow in both striatum and nucleus accumbens, indicating decreased release of DA. CI-1007 also reduced brain DA synthesis (DOPA accumulation), metabolism (DOPAC and HVA) and utilization (after tyrosine hydroxylase inhibition with alpha-methyl-p-tyrosine). CI-1007 did not affect striatal acetylcholine levels indicating lack of potent postsynaptic DA actions. CI-1007 seemed to be selective for DA neurons as it did not alter rat brain norepinephrine (NE) synthesis in the NE-enriched brainstem or NE utilization in the mesolimbic region. In addition, it did not affect in general 5-HT synthesis and metabolism in striatum and mesolimbic regions. These neurochemical results demonstrate that CI-1007 is a selective potent brain dopamine partial agonist with limited agonist activity at postsynaptic DA receptors.     

A dopamine/D1 receptor/protein kinase A/dopamine- and cAMP-regulated phosphoprotein (Mr 32 kDa)/protein phosphatase-1 pathway regulates dephosphorylation of the NMDA receptor

We have investigated the mechanism by which activation of dopamine (DA) receptors regulates the glutamate sensitivity of medium spiny neurons of the nucleus accumbens. Our results demonstrate that DA regulates the phosphorylation state of the NR1 subunit of NMDA-type glutamate receptors. The effect of DA was mimicked by SKF82526, a D1-type DA receptor agonist, and by forskolin, an activator of cAMP-dependent protein kinase (PKA), and was blocked by H-89, a PKA inhibitor. These data indicate that DA increases NR1 phosphorylation through a PKA-dependent pathway. DA-induced phosphorylation of NR1 was blocked in mice bearing a targeted deletion of the gene for dopamine- and cAMP-regulated phosphoprotein of Mr 32 kDa (DARPP-32), a phosphoprotein that is a potent and selective inhibitor of protein phosphatase-1, indicating that the effect of PKA is mediated, in part, by regulation of the DARPP-32/protein phosphatase-1 cascade. In support of this interpretation, NR1 phosphorylation was increased by calyculin A, a protein phosphatase-1/2A inhibitor. A model is proposed in which the ability of DA to regulate NMDA receptor sensitivity is attributable to a synergistic action involving increased phosphorylation and decreased dephosphorylation of the NR1 subunit of the NMDA receptor.     

Regional decreases in alpha-[3H]amino-3-hydroxy-5-methylisoxazole-4-propionic acid ([3H]AMPA) and 6-[3H]cyano-7-nitroquinoxaline-2,3-dione ([3H]CNQX) binding in response to chronic low-level lead exposure: reversal versus potentiation by chronic dopamine agonist treatment

This study evaluated the hypotheses that in vivo lead (Pb) exposure would alter alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptor binding and, based on known glutamate-dopamine interactions and Pb-induced changes in dopamine (DA) systems, that AMPA binding might be differentially influenced by DA agonist treatment under conditions of Pb exposure. Alterations in high-affinity ([3H]AMPA) versus total AMPA [6-[3H]cyano-7-nitroquinoxaline-2,3-dione ([3H]CNQX)] receptor binding were determined in medial frontal cortex, dorsal striatum, and nucleus accumbens of rats exposed to 0, 50, or 150 ppm of Pb acetate for 2 weeks or 8 months. Additional 8-month groups received chronic intermittent treatment with saline, the D1 agonist SKF82958, or the general DA agonist apomorphine. Two-week exposures increased AMPA receptor densities, whereas robust decreases occurred after 8 months of Pb; at the latter time point changes were more pronounced for high-affinity than total AMPA receptor binding, with high-affinity effects expressed preferentially in dorsal striatum and nucleus accumbens. DA agonist treatments almost fully reversed Pb-related declines in [3H]AMPA binding but either had no effect (apomorphine) or even further potentiated (SKF82958) the decreases in [3H]CNQX binding. One possible basis for the long-term (8-month) decrease in AMPA binding is a postsynaptic glutamatergic stimulation of non-NMDA receptors.     

D1 and D2 dopamine receptor mediation of amphetamine-induced acetylcholine release in nucleus accumbens

To assess the interaction of dopamine and acetylcholine systems in the rat nucleus accumbens in response to direct D-amphetamine administration, in vivo microdialysis measures of acetylcholine were used during reverse dialysis of amphetamine alone and in combination with D1 and D2 receptor antagonists SCH 23390 and sulpiride, respectively. During a 15-min exposure to amphetamine (50 microM) in the nucleus accumbens, acetylcholine increased to 33% above pre-infusion levels, became maximal at 15 min post-infusion (+41%) and gradually returned to baseline levels by 60 min post-amphetamine. Conversely, amphetamine (1 mM) administration caused a biphasic change in acetylcholine release with a trend toward a decrease (-14%) during exposure followed by a significant increase (+36%) at 30 min post-amphetamine that returned to baseline levels by 60 min after infusion. The increases observed during amphetamine (50 microM) exposure and during recovery from amphetamine (1 mM) were both blocked by co-administration with the D1 antagonist, SCH 23390 (10 microM), but not with the D2 antagonist, sulpiride (10 microM). Co-infusion of sulpiride eliminated the trend toward reduced acetylcholine release observed during 1 mM amphetamine whereas co-administration of SCH 23390 potentiated this decrease. A possible tonic D1 facilitation of nucleus accumbens acetylcholine release was indicated by the consistent reductions in acetylcholine release observed during infusion of SCH 23390. These results suggest that amphetamine administration in the nucleus accumbens induces a bidirectional change in acetylcholine release that is dependent on dose and opposing effects of nucleus accumbens D1 and D2 activation. In general, relatively low doses of amphetamine administered into the nucleus accumbens caused an increase in acetylcholine release that was dependent on dopamine D1 receptors whereas higher doses of amphetamine resulted in a D2-mediated decrease.     

Effects of the dopamine D3 antagonist PD 58491 and its interaction with the dopamine D3 agonist PD 128907 on brain dopamine synthesis in rat

The dopamine (DA) D3 receptor antagonist PD 58491 [3-[4-[1-[4-[2-[4-(3-diethylaminopropoxy)phenyl]benzoimidazol++ +-1-yl-butyl]-1H-benzoimidazol-2-yl]phenoxy]propyl]diethylamine] bound with high affinity and selectivity to recombinant human DA D3 versus D2L and D4.2 receptors transfected into Chinese hamster ovary cells: Ki values of 19.5 nM versus 2,362 and >3,000 nM, respectively. In contrast, the putative DA D3 receptor antagonist (+)-AJ76 displayed low affinity and selectivity for D3 versus D2L and D4.2 receptors (91 nM vs. 253 and 193 nM, respectively). In vitro, PD 58491 (1 nM-1 microM) exhibited D3 receptor antagonist activity, reversing the quinpirole (10 nM)-induced stimulation of [3H]thymidine uptake in D3 CHOpro-5 cells, but did not have any significant intrinsic activity by itself in this assay. PD 58491 did not decrease the gamma-butyrolactone-induced increase in DA synthesis (L-3,4-dihydroxyphenylalanine accumulation) in rat striatum, indicating that the compound possessed no in vivo DA D2/D3 receptor agonist action at DA autoreceptors. PD 58491 (3-30 mg/kg, i.p.) generally did not alter DA or serotonin synthesis in either the striatum or mesolimbic region of rat brain. The D3-preferring agonist PD 128907 decreased DA synthesis in striatum and mesolimbic regions, and this effect was attenuated by pretreatment with PD 58491. These findings support the hypothesis that DA D3 autoreceptors may in part modulate the synthesis and release of DA in striatum and mesolimbic regions.     

Comparative molecular field analysis-based prediction of drug affinities at recombinant D1A dopamine receptors

Determination of quantitative structure-activity relationship (QSAR) for affinity at particular dopamine (DA) receptors has become an even greater priority with the cloning of five DA receptor subtypes. The use of agonist affinity at recombinant receptors selectively expressed in clonal cells as the dependent variable in QSAR presents a unique opportunity for accuracy and precision in measurement of biological values. Bound conformations of 11 agonists (for which both affinity data at the recombinant D1A DA receptor and stereochemical configurations were available) were determined by alignment with a template compound, SKF38393, which shows high affinity and selectivity for D1A receptors and is fairly rigid in structure. These aligned structures suggested a 3-point pharmacophore map (one cationic nitrogen and two electronegative centers) of the D1A DA receptor. This map shows both similarities and differences when compared with a previously reported D2 DA receptor pharmacophore map based on biological data from rat brain and with a recently published map of the native D1 DA receptor using several semirigid compounds. Log(1/K(d)) values at recombinant D1A DA receptors were used as the target property for a CoMFA (comparative molecular field analysis) of the 11 aligned structures. The resulting CoMFA model yielded a cross-validated r(2)(q(2)) value of 0.829 and a simple r(2) = 0.96. In contrast, when a CoMFA model was developed for 10 of these compounds using striatal D1 K(d) values, the q(2) value was reduced to 0.178. These results are consistent with the idea that drug affinity data obtained from clonal cells expressing recombinant receptors may be superior to that obtained using heterogeneous mixtures of native receptors prepared from brain membranes. The predictive utility of the CoMFA model was evaluated using several high-affinity dopamine agonists and m- and p-tyramine, two compounds with a single hydroxyl group on the aromatic ring. Predictions were fairly accurate for all compounds but the two tyramines.