D2 Receptor Binding Sites Research Articles

Maternal Cannabis Use Alters Ventral Striatal Dopamine D2 Gene Regulation in the Offspring

Prenatal cannabis exposure has been linked to addiction vulnerability, but the neurobiology underlying this risk is unknown. Striatal dopamine and opioid-related genes were studied in human fetal subjects exposed to cannabis (as well as cigarettes and alcohol). Cannabis-related gene disturbances observed in the human fetus were subsequently characterized with an animal model of prenatal Δ-9-tetrahydrocannabinol (THC) (.15 mg/kg) exposure. Prenatal cannabis exposure decreased dopamine receptor D2 (DRD2) messenger RNA expression in the human ventral striatum (nucleus accumbens [NAc]), a key brain reward region. No significant alterations were observed for the other genes in cannabis-exposed subjects. Maternal cigarette use was associated with reduced NAc prodynorphin messenger RNA expression, and alcohol exposure induced broad alterations primarily in the dorsal striatum of most genes. To explore the mechanisms underlying the cannabis-associated disturbances, we exposed pregnant rats to THC and examined the epigenetic regulation of the NAc Drd2 gene in their offspring at postnatal day 2, comparable to the human fetal period studied, and in adulthood. Chromatin immunoprecipitation of the adult NAc revealed increased 2meH3K9 repressive mark and decreased 3meH3K4 and RNA polymerase II at the Drd2 gene locus in the THC-exposed offspring. Decreased Drd2 expression was accompanied by reduced dopamine D2 receptor (D(2)R) binding sites and increased sensitivity to opiate reward in adulthood. These data suggest that maternal cannabis use alters developmental regulation of mesolimbic D(2)R in offspring through epigenetic mechanisms that regulate histone lysine methylation, and the ensuing reduction of D(2)R might contribute to addiction vulnerability later in life.

Patterns of neurotransmitter receptor distributions following cortical spreading depression

Spreading depression (SD), a self-propagating depolarization of neurons and glia, is believed to play a role in different neurological disorders including migraine aura and acute brain ischaemia. Initiation and propagation of SD modulate excitability of neuronal network. A brief period of excitation heralds SD which is immediately followed first by prolonged nerve cell depression and later by an excitatory phase. The aim of the present study was to characterize local and remote transmitter receptor changes after propagation of cortical SD. Quantitative receptor autoradiography was used to asses 16 transmitter receptor types in combined striatum-hippocampus–cortex slices of the rat 1 h after induction of cortical SD. In neocortical tissues, local increases of glutamate NMDA, AMPA, and kainate receptor binding sites were observed. In addition to up-regulation of ionotropic glutamate receptors, receptor binding sites of GABAA, muscarinic M1 and M2, adrenergic α1 and α2, and serotonergic 5-HT2 receptors were increased in the hippocampus. Cortical SD also upregulated NMDA, AMPA, kainate, GABAA, serotonergic 5-HT2, adrenergic α2 and dopaminergic D1 receptor binding sites in the striatum. These findings indicate selective changes in several receptors binding sites both in cortical and subcortical regions by SD which may explain delayed excitatory phase after SD. Mapping of receptor changes by cortical SD increases our understanding of the mechanism of SD action in associated neurological disorders.

99mTc]TRODAT-1/[ 123I]IBZM SPECT studies of the dopaminergic system in Tourette syndrome

The good clinical effectiveness of dopamine depleter and receptor antagonists on tics suggests dopaminergic hyperactivity in Tourette syndrome (TS). In this case-control study of 10 TS patients and 15 age-matched healthy controls, we evaluated (i) presynaptic and postsynaptic striatal dopaminergic function using [ 99mTc]TRODAT-1/[ 123I]IBZM single photon emission computed tomography (SPECT) and (ii) correlations between dopamine transporter (DAT)/D2 receptor binding sites and tic severity scores. Patients 1–5 were pretreated with haloperidol and were drug free for at least 3 months before SPECT imaging. Patients 6–10 were drug-naïve. We found no significant difference in DAT and D2 receptor binding sites between TS patients and healthy controls nor any association between striatal DAT or D2 receptor binding sites and tic severity assessed using the Modified Rush Videotape Rating Scale. Our findings provided no direct evidence of abnormally available striatal DAT or dopamine D2 receptors in TS. However, functional abnormalities of the dopaminergic system, e.g., alterations in the synaptic release of endogenous dopamine, cannot be completely ruled out.

Longitudinal Evaluation of theHdh(CAG)150Knock-In Murine Model of Huntington's Disease

Several murine genetic models of Huntington's disease (HD) have been developed. Murine genetic models are crucial for identifying mechanisms of neurodegeneration in HD and for preclinical evaluation of possible therapies for HD. Longitudinal analysis of mutant phenotypes is necessary to validate models and to identify appropriate periods for analysis of early events in the pathogenesis of neurodegeneration. Here we report longitudinal characterization of the murine Hdh(CAG)150 knock-in model of HD. A series of behavioral tests at five different time points (20, 40, 50, 70, and 100 weeks) demonstrates an age-dependent, late-onset behavioral phenotype with significant motor abnormalities at 70 and 100 weeks of age. Pathological analysis demonstrated loss of striatal dopamine D1 and D2 receptor binding sites at 70 and 100 weeks of age, and stereological analysis showed significant loss of striatal neuron number at 100 weeks. Late-onset behavioral abnormalities, decrease in striatal dopamine receptors, and diminished striatal neuron number observed in this mouse model recapitulate key features of HD. The Hdh(CAG)150 knock-in mouse is a valid model to evaluate early events in the pathogenesis of neurodegeneration in HD.

Hyperdopaminergia and altered locomotor activity in GABAB1‐deficient mice

GABAB1-/- mice, which are devoid of functional GABAB receptors, consistently exhibit marked hyperlocomotion when exposed to a novel environment. Telemetry recordings now revealed that, in a familiar environment, GABAB1-/- mice display an altered pattern of circadian activity but no hyperlocomotion. This indicates that hyperlocomotion is only triggered when GABAB1-/- mice are aroused by novelty. In microdialysis experiments, GABAB1-/- mice exhibited a 2-fold increased extracellular level of dopamine in the striatum. Following D-amphetamine administration, GABAB1-/- mice released less dopamine than wild-type mice, indicative of a reduced cytoplasmic dopamine pool. The hyperdopaminergic state of GABAB1-/- mice is accompanied by molecular changes, including reduced levels of tyrosine hydroxylase mRNA, D1 receptor binding-sites and Ser40 phosphorylation of tyrosine hydroxylase. Tyrosine hydroxylase activity, tissue dopamine content and dopamine metabolism do not appear to be measurably altered. Pharmacological and electrophysiological experiments support that the hyperdopaminergic state of GABAB1-/- mice is not severe enough to inactivate dopamine D2 receptors and to disrupt D2-mediated feedback inhibition of tyrosine hydroxylase activity. The data support that loss of GABAB activity results in a sustained moderate hyperdopaminergic state, which is phenotypically revealed by contextual hyperlocomotor activity. Importantly, the presence of an inhibitory GABA tone on the dopaminergic system mediated by GABAB receptors provides an opportunity for therapeutic intervention.

Effects of Chronic Alcohol and Repeated Deprivations on Dopamine D1 and D2 Receptor Levels in the Extended Amygdala of Inbred Alcohol‐Preferring Rats

Dopaminergic (DA) activity in the extended amygdala (EA) has been known to play a pivotal role in mediating drug and alcohol addiction. Alterations of DA activity within the EA after chronic exposure to alcohol or substances of abuse are considered a major mechanism for the development of alcoholism and addiction. To date, it is not clear how different patterns of chronic alcohol drinking affect DA receptor levels. Therefore, the current studies investigated the effects of chronic ethanol consumption, with or without deprivations, on D1 and D2 receptor densities within the EA. Inbred alcohol-preferring (iP) rats were divided into 3 groups with the following treatments: (1) water for 14 weeks; (2) continuous alcohol (C-Alc) for 14 weeks [24-hour concurrent access to 15 and 30% (v/v) ethanol]; or (3) repeatedly deprived of alcohol (RD-Alc) (24-hour concurrent access to 15 and 30% ethanol for 6 weeks, followed by 2 cycles of 2 weeks of deprivation of and 2 weeks of reexposure to ethanol access). At the end of 14 weeks, the rats were killed for autoradiographic labeling of D1 and D2 receptors. Compared with the water control group, both the C-Alc and the RD-Alc groups displayed increases in D1 receptor binding density in the anterior region of the Acb core, whereas the RD-Alc group displayed additional increases in D1 receptor binding density in anterior regions of the lateral and intercalated nuclei of the amygdala. Additionally, both C-Alc and RD-Alc rats displayed increases in D2 receptor binding density in anterior regions of the Acb shell and core, whereas RD-Alc rats displayed additional increases in D2 receptor binding density in the dorsal striatum. The results of this study indicate that 14-week extended alcohol drinking with continuous chronic or repeated deprivations increase binding sites of D1 and D2 receptors in specific regions of the EA with greater sensitivity in the anterior regions. The repeated deprivation has greater effect on altering D1 and D2 receptor binding sites in the Acb, dorsal striatum, and subamygdala regions. The current result indicates that the two drinking paradigms may have common as well as differential mechanisms on alteration of dopamine receptor-binding sites in specific regions of the EA.

Motor Dysfunction in Type 5 Adenylyl Cyclase-null Mice

Various neurotransmitters, such as dopamine, stimulate adenylyl cyclase to produce cAMP, which regulates neuronal functions. Genetic disruption of the type 5 adenylyl cyclase isoform led to a major loss of adenylyl cyclase activity in a striatum-specific manner with a small increase in the expression of a few other adenylyl cyclase isoforms. D1 dopaminergic agonist-stimulated adenylyl cyclase activity was attenuated, and this was accompanied by a decrease in the expression of the D1 dopaminergic receptor and G(s)alpha. D2 dopaminergic agonist-mediated inhibition of adenylyl cyclase activity was also blunted. Type 5 adenylyl cyclase-null mice exhibited Parkinsonian-like motor dysfunction, i.e. abnormal coordination and bradykinesia detected by Rotarod and pole test, respectively, and to a lesser extent locomotor impairment was detected by open field tests. Selective D1 or D2 dopaminergic stimulation improved some of these disorders in this mouse model, suggesting the partial compensation of each dopaminergic receptor signal through the stimulation of remnant adenylyl cyclase isoforms. These findings extend our knowledge of the role of an effector enzyme isoform in regulating receptor signaling and neuronal functions and imply that this isoform provides a site of convergence of both D1 and D2 dopaminergic signals and balances various motor functions.

Substituted 3-amino and/or 3-aminomethyl-3,4-dihydro-2H-1-benzopyrans: synthesis and biological activity

A series of new 3-amino, 3-aminomethyl-5-alkoxy-3,4-dihydro-2H-1-benzopyran and 5′-alkoxy-3′,4′-dihydrospiro[piperazine-2,3′(2′H)-benzopyran] derivatives was prepared and evaluated for affinity at 5-HT1A, 5-HT2A and D2 receptors. Two of the compounds (1f and 2b) can be considered as potent and selective 5-HT2A ligands. One compound (1g) demonstrated high affinity for 5-HT1A and D2 receptor binding sites and one compound (1d) proved to be a mixed 5-HT1A/5-HT2A ligand.

Characterization of [ 3H]mazindol binding sites in cultured monkey amniotic epithelial cells

Our previous studies showed that monkey amniotic epithelial cells (MAEC) synthesize and release catecholamines and possess D1 and D2 dopamine (DA) receptors (Elwan, M.A., Ishii, T., Ono, F. and Sakuragawa, N., Evidence for the presence of dopamine D1 receptor mRNA and binding sites in monkey amniotic epithelial cells. Neurosci. Lett., 262 (1999) 9–12; Elwan, M.A., Ishii, T. and Sakuragawa, N., Detection of dopamine D2 receptor mRNA and binding sites in monkey amniotic epithelial cells. J. Neurosci. Res., 56 (1999) 316–322; Elwan, M.A., Thangavel, R., Ono, F. and Sakuragawa, N., Synthesis and release of catecholamines by cultured monkey amniotic epithelial cells. J. Neurosci. Res., 53 (1998) 107–113). In the present study we tested the presence of DA transporter (DAT) in MAEC using radioligand binding experiments. Saturation studies showed that [ 3H]mazindol binds to a high affinity site with K D and B max values of 7.85±1.25 nM and 123.22±18.34 fmol/mg protein, respectively. Competition studies indicated that selective DAT inhibitors are potent displacers of [ 3H]mazindol binding, compared to inhibitors of other types of transporters. The rank order of potency of the competing drugs is consistent with the pharmacology of DAT. These results provide, for the first time, clear evidence that MAEC natively possess DAT binding sites and suggest that MAEC may provide a potential primate cell model to study DA release and uptake processes and to explore new drugs active at this site.

Apparent dopamine D1 and D2 receptors in the weaver mutant mouse: receptor binding and coupling to adenylyl cyclase.

Weaver mutant mice have a selective degeneration of the nigrostriatal dopamine pathway arising between 7-21 days after birth. The goal of this study was to investigate the effects of this mutation on different parameters of the nigrostriatal and mesolimbic dopamine system: apparent D1 and D2 receptor binding sites as well as their signal transduction pathway. Using quantitative autoradiography of ligands for dopamine D1, D2 receptors and the dopamine uptake site, we found a significant loss in apparent D1 receptor binding sites throughout the neostriatum, significant increase of apparent D2 receptor binding in the dorsal aspect of the neostriatum, and almost complete loss of DA uptake sites in these regions of the weaver mouse. In contrast to the neostriatum, the density of dopamine receptors and uptake sites in the nucleus accumbens of the weaver mouse did not differ from controls. Despite alterations in the binding of apparent D1 and D2 receptors, there was no significant difference in either basal, DA stimulated or GTPgammaS stimulated cAMP production. These findings suggest the down-regulation of apparent D1 receptor binding sites reported in this model, probably does not reflect an important physiological mechanism through which these animals compensate for loss of dopamine innervation.

Detection of dopamine D2 receptor mRNA and binding sites in monkey amniotic epithelial cells

Previous results from our laboratory showed that monkey amniotic epithelial cells (MAEC) possess the catecholamine synthesizing enzymes and have the capacity to synthesize and release CA. Recently, we also reported that these cells express dopamine D1 receptor mRNA and binding sites. This study was designed to investigate the presence of dopamine D2 receptors in MAEC. Using RT-PCR, we found that MAEC express dopamine D2 receptor mRNA that is having 98% homology with human dopamine D2 receptors. Radioligand saturation binding studies showed a 3H-YM-09151-2 high-affinity binding site with a KD of 0.293 ± 0.06 nM and Bmax of 180.69 ± 11.61 fmol/mg protein. Competition experiments with a variety of displacing drugs demonstrated that D2 antagonists potently compete with 3H-YM-09151-2 binding, whereas D1 antagonists displayed a weaker competition for the binding sites. The rank order of potency of these compounds in competing with 3H-YM-09151-2 for binding sites was consistent with the pharmacology of the dopamine D2 receptors. All competition curves were better fitted to a one-site model with a Hill coefficient around unity, indicating that 3H-YM-09151-2 is labeling a single population of receptors. These results provide, for the first time, a compelling evidence that MAEC natively express dopamine D2 receptor mRNA and binding sites, and they suggest that monkey amniotic epithelial cells (MAEC) could represent a source of primate dopamine receptors without the need for transformation or cloning procedures using nonprimate cells, as generally happens. J. Neurosci. Res. 56:316–322, 1999. © 1999 Wiley-Liss, Inc.

Paradoxical Locomotor Behavior of Dopamine D1 Receptor Transgenic Mice

The behavioral effects of augmenting dopamine D1 receptor expression in the brain were investigated in mice incorporating additional copies of the mouse D1 receptor gene. Two transgenic lines showed increases in brain D1 receptor binding sites, which were greatest in extrastriatal regions. The full D1 agonist SKF 81297, when administered systemically to control animals, stimulated a dose-dependent increase in locomotor activity. In contrast, in D1 receptor overexpressing transgenic mice, this drug caused a marked suppression of locomotion due to a decrease in the frequency of movement initiation. Amphetamine and cocaine induced comparable locomotor activation in both transgenic animals and their control littermates. In the transgenic animals, D1 agonist-induced rearing and climbing behaviors were suppressed. However, on rotarod testing, the agonist-treated transgenic and control mice performed comparably, indicating that sensorimotor coordination was unaffected. These studies demonstrate that altering the levels of D1 receptor expression reverses the effects of D1 agonism on locomotor initiation and rearing.

Detection of dopamine D2 receptor mRNA and binding sites in monkey amniotic epithelial cells.

Previous results from our laboratory showed that monkey amniotic epithelial cells (MAEC) possess the catecholamine synthesizing enzymes and have the capacity to synthesize and release CA. Recently, we also reported that these cells express dopamine D1 receptor mRNA and binding sites. This study was designed to investigate the presence of dopamine D2 receptors in MAEC. Using RT-PCR, we found that MAEC express dopamine D2 receptor mRNA that is having 98% homology with human dopamine D2 receptors. Radioligand saturation binding studies showed a 3H-YM-09151-2 high-affinity binding site with a K(D) of 0.293+/-0.06 nM and Bmax of 180.69+/-11.61 fmol/mg protein. Competition experiments with a variety of displacing drugs demonstrated that D2 antagonists potently compete with 3H-YM-09151-2 binding, whereas D1 antagonists displayed a weaker competition for the binding sites. The rank order of potency of these compounds in competing with 3H-YM-09151-2 for binding sites was consistent with the pharmacology of the dopamine D2 receptors. All competition curves were better fitted to a one-site model with a Hill coefficient around unity, indicating that 3H-YM-09151-2 is labeling a single population of receptors. These results provide, for the first time, a compelling evidence that MAEC natively express dopamine D2 receptor mRNA and binding sites, and they suggest that monkey amniotic epithelial cells (MAEC) could represent a source of primate dopamine receptors without the need for transformation or cloning procedures using nonprimate cells, as generally happens.

Alternative splicing of the D2 dopamine receptor messenger ribonucleic acid is modulated by activated sex steroid receptors in the MMQ prolactin cell line.

The two isoforms of the D2 dopamine receptor are generated by alternative splicing of the exon 6 of the premessenger RNA (pre-mRNA), changing the length of the third cytoplasmic loop involved in the coupling to G proteins. In the MMQ PRL cell line, sex steroid hormones modulated the proportion of the two D2 receptor isoforms. Under controlled culture conditions, 17beta-estradiol (E2) strongly favored the production of the long isoform of D2 mRNA over the short one, whereas both isoforms were equally abundant when culture medium was hormone depleted. In the presence of progesterone (P), E2 action was inhibited, and equal amounts of each D2 receptor isoform were produced in the cells. Hormone treatments never modified either the total amount of D2 receptor mRNA and D2 receptor binding sites or D2 receptor-mediated inhibition of adenylyl cyclase. Specific antagonists demonstrated that the activity of each hormone depended on their nuclear receptors. Inhibitors of gene transcription or translation also showed that their activity required protein synthesis. The expression of the short D2 receptor isoform was never prominent, even at the single cell level. Analysis of the intron sequence flanking alternative exon 6 showed that only the upstream intron presented two sequence tracts known to be targets for splicing factors. Taken together, these results provide converging evidence for a physiologically relevant mechanism by which sex steroid receptors could regulate the expression of a splicing factor favoring the production of the long dopamine D2 receptor isoform.

Changes in the Midbrain-Rostral Forebrain Dopamine Circuitry in the Cocaine-Exposed Primate Fetal Braina.

To ascertain cocaine's effects in the fetus, we developed a nonhuman primate model in which pregnant monkeys were administered cocaine (3 mg/kg im) or saline four times a day from day 20 through days 40-70 of a 165-day gestation. At the time of cesarean section, plasma levels of cocaine in fetal blood were 231 ± 70 ng/ml. Fetal brains were examined using immunocytochemistry, in situ hybridization, receptor autoradiography, and nuclease protection assay analysis. No differences were found in the expression of tyrosine hydroxylase and dopamine receptor mRNAs by days 40-45 of gestation. However, by day 60 the midbrain of monkeys exposed to cocaine had significantly reduced expression of tyrosine hydroxylase, the rate-limiting enzyme in dopamine synthesis. Moreover, dopamine D1 and D2 receptor mRNAs were significantly elevated in the rostral forebrain as were D1 and D2 receptor binding sites in days 60-70 cocaine-exposed fetuses. Cocaine treatment from day 20 to days 60 and 70 of gestation also significantly increased the mRNA concentrations of dynorphin and enkephalin in the rostral forebrain. These findings suggest that in utero cocaine exposure has profound effects on the developing dopamine neurocircuitry.

Modulation of Dopamine D<sub>2</sub> Receptor Expression by an NMDA Receptor Antagonist in Rat Brain

The expression of dopamine D2 receptor mRNA was studied in rat brain following micro-injection of a competitive N-methyl D-aspartate (NMDA) receptor antagonist at the prefrontal cortex. Male Sprague-Dawley rats cannulated bilaterally into the medial prefrontal cortex were injected with a competitive NMDA receptor antagonist (+/-)-3-(2-carboxypiperazin-4-yl)-propyl-1-phosphonic acid (CPP). The levels of mRNA for NMDA-R1 and dopamine D2 receptors were measured by reverse transcriptase-polymerase chain reaction (RT-PCR), and D2 receptor density was quantified by [3H]spiperone binding in the cortex and striatum of these animals. In the prefrontal cortex, the levels of NMDA-R1 receptor mRNA showed significant decrease in CPP-treated animals compared to control animals. However, NMDA-R1 mRNA levels in striatum remained unchanged in any of the experimental groups. The D2 receptor mRNA levels and [3H]spiroperidol binding in prefrontal cortical membranes showed no significant difference between the CPP-treated and control groups of animals. In the striatum, a significant increase in striatal dopamine D2 receptor mRNA levels was shown in animals treated with CPP. The increase in D2 mRNA level was correlated with an increase in the D2 receptor binding sites in the striatal membranes. These results suggest a possible interaction between prefrontal cortical NMDA receptors and striatal dopamine receptors.

Mitochondrial toxins in models of neurodegenerative diseases. II: elevated zif268 transcription and independent temporal regulation of striatal D1 and D2 receptor mRNAs and D1 and D2 receptor-binding sites in C57BL/6 mice during MPTP treatment

Sporadic Parkinson's disease (PD) may arise from a defect in complex I of the mitochondrial electron transport chain (ETC), transmitted through mitochondrial DNA mutations. The N-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) model of experimental PD is believed to arise from loss of complex I activity in dopamine (DA) neurons after accumulation of MPP+, a potent complex I inhibitor and the two electron monoamine oxidase B oxidation product of MPTP. Acute MPP+ infusion into striatum, possibly mimicking the in vivo situation after MPTP treatment, increases release of DA and production of hydroxyl radical ( ⋅OH). We treated C57BL/6 mice with MPTP and followed the expression of the immediate-early gene zif268 in striatum as a marker of DA synaptic activity, determined the pharmacology of its activation during MPTP toxicity, and assayed the time course of MPTP effects on striatal DA transporter (DAT), and D1 and D2 DA receptor-binding sites and their mRNAs. MPTP (24 mg/kg b.i.d. for 4 doses) increased striatal zif268 expression, with peak effects observed 24 h after starting MPTP. Increased striatal zif268 was dependent mainly on DA D1 and to a lesser extent on non-NMDA glutamate receptors and was not altered by inhibition of nitric oxide synthase (NOS). Our MPTP schedule resulted in a loss of about one-third of nigral DA neurons. We observed with [ 3H]mazindol autoradiography that loss of striatal DAT sites after starting MPTP was heterogenous and greatest in centromedial striatum, reached a maximum at 48 h and showed a slight recovery at 2 weeks. Striatal D1 and D2 receptor-binding sites (measured with [ 3H]SCH23390 and [ 3H]spiperone binding, respectively) and mRNA levels for D1 and D2 receptors (determined with quantitative in situ hybridization) were altered after MPTP treatment in temporally independent manners. MPTP toxicity to the nigrostriatal system likely induces substantial striatal DA release in vivo and stimulates transcription of at least one major IEG, zif268, in striatal neurons. Increased striatal zif268 expression after MPTP appears to derive mainly from DA released onto D1 receptors, not by a NO-dependent process which has been described in striatal neurons in vitro. The rapid loss of striatal DA terminals after MPTP treatment alters D1 and D2 receptor sites independently of changes in their mRNA levels. Increased D1 and D2 gene transcription in this model may depend on re-innervation by DA terminals of striatal neurons and likely is not related to the increased zif268 transcription observed after MPTP.

Alterations in the turnover rate of dopamine D1 but not D2 receptors in the adult rat neostriatum after a neonatal dopamine denervation

Adult rats that were treated with intracerebral ventricular injection of 6-hydroxydopamine (6-OHDA) as neonates exhibit a profound loss of nigrostriatal dopamine innervation in addition to a variety of other neurochemical and anatomical changes, including alterations in the number of neostriatal D1 and D2 receptor binding sites. In the present study, the turnover of neostriatal dopamine D1 and D2 receptors was measured in rats previously treated with 6-OHDA or ascorbic acid vehicle as neonates at various time intervals after peripheral N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline (EEDQ, 10 mg/kg) administration. Dopamine receptors were labelled with [ 3H]SCH23390 (D1) and [ 3H]raclopride (D2), while the degree of dopamine denervation was assessed by the measurement of neostriatal dopamine, homovanillic acid and dihydroxyphenylacetic acid content. Two days after acute EEDQ treatment, the maximal binding capacity ( B max) of [ 3H]SCH23390 and [ 3H]raclopride binding was significantly decreased to 58 and 32% of control values, respectively, without any significant alteration in their equilibrium dissociation constants ( K d). A time-dependent increase in the density of [ 3H]SCH23390 and [ 3H]raclopride binding was observed in both treatment groups following a single dose of EEDQ. The rate of recovery of D1 receptors was significantly slower in the 6-OHDA-lesioned animals as compared to controls with a half-life of 103 compared to 53 h, respectively. No differences were observed in the rate of recovery of D2 receptors in these two treatment groups. These data are consistent with the findings of decreased expression of D1 receptors in neonatal 6-OHDA-lesioned rats owing to decreased receptor synthesis, and further suggest that in this model the up-regulation of D2 receptors is a result of a post-transcriptional mechanism, such as an increased rate of post-synthetic maturation.

A Neural Substrate of Hyperactivity in Borna Disease: Changes in Brain Dopamine Receptors

Rats experimentally infected with the neurotropic RNA virus, Borna disease virus, have a hyperactive movement disorder. Because locomotor activity is modulated by the nucleus accumbens (N. Acc.) dopamine (DA) system, high-affinity DA uptake, DA D1, D2, and D3 receptor binding sites were examined in N. Acc. subregions of normal and infected rats by quantitative receptor autoradiography. The N. Acc. of infected rats had decreased mazindol and D2 and D3 radioligand binding in the core and decreased D3 radioligand binding in rostral subregions. The abnormalities observed in the N. Acc. DA system of infected rats may offer insights into the potential viral pathogenesis of psychiatric conditions with a dopaminergic substrate such as schizophrenia and affective disorders.

Development of striatal dopaminergic function. I. Pre- and postnatal development of mRNAs and binding sites for striatal D1 (D1a) and D2 (D2a) receptors.

Quantitative receptor autoradiography with iodinated ligands, quantitative in situ hybridization histochemistry and reverse transcriptase-polymerase chain reaction (RT-PCR) were used to describe the prenatal and early postnatal ontogeny (embryonic day 14 to postnatal day 7, or E14 to P7) of striatal D1 and D2 dopamine receptor binding sites and mRNA levels, respectively, in relation to the development of dopaminergic nigrostriatal innervation D1 dopamine receptor, measured by [125I]SCH23982 binding, and dopamine transporter binding sites, measured by [125I]RTI-55 binding, were present in low amounts beginning on E14 (2-3% and 0.3-0.6% of adult values, respectively) and increased slowly during the prenatal period. D2 receptor binding sites, measured with [125I]spiperone, were also detected on E14 but in higher relative quantities (17% of adult values) than D1 receptor and dopamine transporter binding sites at the same age. Other than abrupt declines in the late prenatal period for D1 and D2 receptor binding sites, all three binding sites increased throughout development and increased maximally between P7 and adulthood. On P5, both D1 and D2 receptors were functionally coupled to their respective G proteins, based on GTP-induced decreases in affinity of dopamine for [125I]SCH23982 and [125I]spiperone binding. D1 receptor mRNA was present in E14 striatal anlage, increased prenatally, declined on P0, then increased to a peak on P5, after which it declined to its lowest levels (20% of peak values) in the adult. In contrast, D2 receptor mRNA levels were presented also on E14, increased to a peak on P0, declined until P5, and increased thereafter to adulthood. Anatomically, nigrostriatal innervation and D1 and D2 receptor mRNA levels increased from the medial to lateral striatal quadrants. In contrast, D1 and D2 receptor binding site ontogency exhibited fairly homogenous distributions from E18 to P7. D1 and D2 receptor mRNAs appear to be expressed early in prenatal development before there is any significant dopaminergic innervation. In contrast, the majority of D1 and D2 receptor binding activity, representing expressed receptor proteins, develops in the postnatal period and correlates well with the increase in dopaminergic innervation. Intrinsic genetic programming is more likely to be responsible for D1 and D2 receptor gene transcription in striatal neuroblasts and newly born neurons, while factors derived from ingrowing dopaminergic afferents may direct post-transcriptional dopamine receptor development. The dissociation between the ontogeny of dopamine receptor binding sites and mRNAs suggests that the developmental regulation of D1 and D2 receptor synthesis is independent of D1 and D2 receptor gene transcription.