Involvement of ryanodine receptor type 3 in dopamine release from the striatum: evidence from mutant mice lacking this receptor

Although it is known that ryanodine receptor type 3 is expressed in the striatum, the function of this receptor has not been elucidated. Therefore, we examined whether caffeine- and ryanodine-induced dopamine release in striatal slices is affected in mice lacking ryanodine receptor type 3. Pretreatment with thapsigargin, an inhibitor of the Ca(2+) ATPase pump of the endoplasmic reticulum, abolished caffeine- or ryanodine-induced dopamine release in slices from normal mice. Dopamine concentration in the striatum and KCl-induced dopamine release were unaffected by a ryanodine receptor type 3 deficiency. Ryanodine-induced dopamine release was significantly attenuated in mice lacking ryanodine receptor type 3, whereas caffeine-induced dopamine release was partially attenuated. Caffeine produced a similar hyper-motor activity in both wild and homozygous mice. The present results suggest the involvement of ryanodine receptor type 3 in dopamine release from the striatum.     

Effect of Chronic Angiotensin-Converting Enzyme Inhibition on Striatal Dopamine Content in the MPTP-Treated Mouse

We have previously shown that chronic treatment with the angiotensin-converting enzyme inhibitor perindopril increased striatal dopamine levels by 2.5-fold in normal Sprague-Dawley rats, possibly via modulation of the striatal opioid or tachykinin levels. In the present study, we investigated if this effect of perindopril persists in an animal model of Parkinson's disease, the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated mouse. C57BL/6 mice were treated with the neurotoxin (30 mg/kg/day intraperitoneally) for 4 days and then left for 3 weeks to allow the degeneration of striatal dopaminergic terminals. At this time, the mice exhibited a 40% decrease in striatal dopamine content and an accompanying 46% increase in dopamine D2 receptor levels compared with control untreated mice. The dopamine content returned to control levels, and the increase in dopamine D2 receptor levels was attenuated in mice treated with perindopril (5 mg/kg/day orally for 7 days) 2 weeks after the last dose of MPTP. When the angiotensin-converting enzyme inhibitor was administered (5 mg/kg/day for 7 days) immediately after the cessation of the MPTP treatment, there was no reversal of the effect of the neurotoxin in decreasing striatal dopamine content. Our results demonstrate that perindopril is an effective agent in increasing striatal dopamine content in an animal model of Parkinson's disease.     

Aging of the dopaminergic system and motor behavior in mice intoxicated with the parkinsonian toxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine

J. Neurochem. (2012) 122, 1032-1046. ABSTRACT: 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) intoxication of mice is a standard model of Parkinson's disease (PD). However, it does not reproduce functionally PD. Given the occurrence of PD during aging, symptoms might only be detected in MPTP-intoxicated mice after aging. To address this, mice injected with MPTP at 2.5?months were followed up to a maximum age of 21?months. There was no loss of dopamine cells with aging in control mice; moreover, the initial post-MPTP intoxication decrease in dopamine cell was no longer significant at 21?months. With aging, striatal dopamine level remained constant, but concentrations of the dopamine metabolites dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) were markedly reduced in both groups. There was also a late impairment of fine motor skills. After MPTP intoxication, hyperactivity was immediately detected and it became greater than in control mice from 14?months of age; fine motor skills were also more impaired; both these symptoms were correlated with striatal dopamine, DOPAC and HVA concentrations. In bothgroups, neither motor symptoms nor dopamine changes worsened with age. These findings do not support the notion that PD develops with age in mice after MPTP intoxication and that the motor deficits seen are because of an aging process.     

Neuroadaptations to hyperdopaminergia in dopamine D3 receptor-deficient mice

The dopamine D3 receptor (D3R) has been implicated in schizophrenia, drug addiction, depression and Parkinson's disease. The D3R is localized post-synaptically on nucleus accumbens neurons, but is also an autoreceptor on dopaminergic neurons in the mesencephalon. Its functional role as autoreceptor is highly debated, but supported by the elevated basal extracellular dopamine levels found in D3R-deficient mice. To investigate the functional role of the D3R in vivo, we used mice with a targeted disruption of the D3R gene. We found a higher basal level of grooming in D3R-deficient mice, compared to their wild-type littermates. This behavior, which is under the control of D1R stimulation, may be related to an increased dopaminergic tone, since no changes in the gene expression of dopamine D1 and D2 receptors were noticed in the striatum of these mice. D3R-deficient mice displayed other neuroadaptive changes, including decreased tyrosine hydroxylase, increased dopamine transporter mRNAs and increased dopamine reuptake in striatum. The level of tyrosine hydroxylase protein was unchanged in the striatum, as preprodynorphin and preproenkephalin gene expressions. All the changes identified in D3R-deficient mice cannot explain hyperdopaminergia, but, on the contrary, tend to attenuate this phenotype. These results support a distinct role for D2R and D3R as autoreceptors: the D2R is the release-regulating and firing rate-regulating autoreceptor, whereas the D3R may control basal dopamine levels in the striatum, by an unknown mechanism, which does not involve regulation of dopamine transporters or tyrosine hydroxylase. This hyperdopaminergia phenotype of D3R-deficient mice may explain their hyperactivity to drug-paired environmental cues.     

Aberrant striatal dopamine transmitter dynamics in brain-derived neurotrophic factor-deficient mice

Brain-derived neurotrophic factor (BDNF) modulates the synaptic transmission of several monoaminergic neuronal systems, including forebrain dopamine-containing neurons. Recent evidence shows a strong correlation between neuropsychiatric disorders and BDNF hypofunction. The aim of the present study was to characterize the effect of low endogenous levels of BDNF on dopamine system function in the caudate-putamen using heterozygous BDNF (BDNF(+/-) ) mice. Apparent extracellular dopamine levels in the caudate-putamen, determined by quantitative microdialysis, were significantly elevated in BDNF(+/-) mice compared with wildtype controls (12 vs. 5 nM, respectively). BDNF(+/-) mice also had a potentiated increase in dopamine levels following potassium (120 mM)-stimulation (10-fold) relative to wildtype controls (6-fold). Slice fast-scan cyclic voltammetry revealed that BDNF(+/-) mice had reductions in both electrically evoked dopamine release and dopamine uptake rates in the caudate-putamen. Superfusion of BDNF led to partial recovery of the electrically stimulated dopamine release response in BDNF(+/-) mice. Conversely, tissue accumulation of L-3,4-dihydroxyphenylalanine, extracellular levels of dopamine metabolites, and spontaneous locomotor activity were unaltered. Together, this study indicates that endogenous BDNF influences dopamine system homeostasis by regulating the release and uptake dynamics of pre-synaptic dopamine transmission.     

Estrogen Alters MPTP-Induced Neurotoxicity in Female Mice: Effects on Striatal Dopamine Concentrations and Release

Abstract: The effects of estrogen on MPTP-induced neurotoxicity of the nigrostriatal dopaminergic system were examined in C57Bl and CD-1 mice. Ovariectomized mice with and without estrogen were treated with MPTP or its vehicle. The effects of these treatments on striatal dopamine concentrations and l -DOPA-stimulated dopamine and l -3,4-dihydroxyphenylacetic acid (DOPAC) release in vitro were determined. Dopamine concentrations of C57Bl mice receiving estrogen before MPTP were significantly greater than those of non-estrogen-treated MPTP mice as well as estrogen-treated mice receiving the MPTP vehicle. Dopamine concentrations of the CD-1 mice did not differ with these treatments. l -DOPA-evoked dopamine release values of estrogen-treated C57Bl mice were significantly increased compared with non-estrogen-treated mice. No such differences were observed in the MPTP-treated C57Bl mice. DOPAC release rates were similar to that of dopamine in these C57Bl mice. In the CD-1 mice estrogen also produced a significant increase in l -DOPA-evoked dopamine release; however, this response was unaltered by MPTP treatment. A significant increase in l -DOPA-evoked DOPAC output was obtained only for estrogen-treated CD-1 mice. Both strains show very similar responses to the estrogen treatment, but differential responses of dopamine release to l -DOPA between the C57Bl and CD-1 mice were obtained with regard to the interactive effects of estrogen and MPTP. Our results suggest that in addition to its role as modulator, estrogen may also function as a neuroprotectant against MPTP neurotoxicity of the nigrostriatal dopaminergic system in the C57Bl mouse.     

Dopaminergic transmission in STOP null mice

Neuroleptics are thought to exert their anti-psychotic effects by counteracting a hyper-dopaminergic transmission. Here, we have examined the dopaminergic status of STOP (stable tubule only polypeptide) null mice, which lack a microtubule-stabilizing protein and which display neuroleptic-sensitive behavioural disorders. Dopamine transmission was investigated using both behavioural analysis and measurements of dopamine efflux in different conditions. Compared to wild-type mice in basal conditions or following mild stress, STOP null mice showed a hyper-locomotor activity, which was erased by neuroleptic treatment, and an increased locomotor reactivity to amphetamine. Such a behavioural profile is indicative of an increased dopaminergic transmission. In STOP null mice, the basal dopamine concentrations, measured by quantitative microdialysis, were normal in both the nucleus accumbens and the striatum. When measured by electrochemical techniques, the dopamine efflux evoked by electrical stimulations mimicking physiological stimuli was dramatically increased in the nucleus accumbens of STOP null mice, apparently due to an increased dopamine release, whereas dopaminergic uptake and auto-inhibition mechanisms were normal. In contrast, dopamine effluxes were slightly diminished in the striatum. Together with previous results, the present study indicates the association in STOP null mice of hippocampal hypo-glutamatergy and of limbic hyper-dopaminergy. Such neurotransmission defects are thought to be central to mental diseases such as schizophrenia.     

MsrA knockout mouse exhibits abnormal behavior and brain dopamine levels

Oxidative stress can cause methionine oxidation that has been implicated in various proteins malfunctions, if not adequately reduced by the methionine sulfoxide reductase system. Recent evidence has found oxidized methionine residues in neurodegenerative conditions. Previously, we have described elevated levels of brain pathologies and an abnormal walking pattern in the methionine sulfoxide reductase A knockout (MsrA(-/-)) mouse. Here we show that MsrA(-/-) mice have compromised complex task learning capabilities relative to wild-type mice. Likewise, MsrA(-/-) mice exhibit lower locomotor activity and altered gait that exacerbated with age. Furthermore, MsrA(-/-) mice were less responsive to amphetamine treatment. Consequently, brain dopamine levels were determined. Surprisingly, relative to wild-type mice, MsrA(-/-) brains contained significantly higher levels of dopamine up to 12 months of age, while lower levels of dopamine were observed at 16 months of age. Moreover, striatal regions of MsrA(-/-) mice showed an increase of dopamine release parallel to observed dopamine levels. Similarly, the expression pattern of tyrosine hydroxylase activating protein correlated with the age-dependent dopamine levels. Thus, it is suggested that dopamine regulation and signaling pathways are impaired in MsrA(-/-) mice, which may contribute to their abnormal behavior. These observations may be relevant to age-related neurological diseases associated with oxidative stress.     

M₅ muscarinic receptors mediate striatal dopamine activation by ventral tegmental morphine and pedunculopontine stimulation in mice

Opiates, like other addictive drugs, elevate forebrain dopamine levels and are thought to do so mainly by inhibiting GABA neurons near the ventral tegmental area (VTA), in turn leading to a disinhibition of dopamine neurons. However, cholinergic inputs from the laterodorsal (LDT) and pedunculopontine (PPT) tegmental nucleus to the VTA and substantia nigra (SN) importantly contribute, as either LDT or PPT lesions strongly attenuate morphine-induced forebrain dopamine elevations. Pharmacological blockade of muscarinic acetylcholine receptors in the VTA or SN has similar effects. M₅ muscarinic receptors are the only muscarinic receptor subtype associated with VTA and SN dopamine neurons. Here we tested the contribution of M₅ muscarinic receptors to morphine-induced dopamine elevations by measuring nucleus accumbens dopamine efflux in response to intra-VTA morphine infusion using in vivo chronoamperometry. Intra-VTA morphine increased nucleus accumbens dopamine efflux in urethane-anesthetized wildtype mice starting at 10 min after infusion. These increases were absent in M₅ knockout mice and were similarly blocked by pre-treatment with VTA scopolamine in wildtype mice. Furthermore, in wildtype mice electrical stimulation of the PPT evoked an initial, short-lasting increase in striatal dopamine efflux, followed 5 min later by a second prolonged increase in dopamine efflux. In M₅ knockout mice, or following systemic pre-treatment with scopolamine in wildtype mice, the prolonged increase in striatal dopamine efflux was absent. The time course of increased accumbal dopamine efflux in wildtype mice following VTA morphine was consistent with both the prolonged M₅-mediated excitation of striatal dopamine efflux following PPT electrical stimulation and accumbal dopamine efflux following LDT electrical stimulation. Therefore, M₅ receptors appear critical for prolonged PPT excitation of dopamine efflux and for dopamine efflux induced by intra-VTA morphine.     

Correlated alterations in serotonergic and dopaminergic modulations at the hippocampal mossy fiber synapse in mice lacking dysbindin

Dysbindin-1 (dystrobrevin-binding protein 1, DTNBP1) is one of the promising schizophrenia susceptibility genes. Dysbindin protein is abundantly expressed in synaptic regions of the hippocampus, including the terminal field of the mossy fibers, and this hippocampal expression of dysbindin is strongly reduced in patients with schizophrenia. In the present study, we examined the functional role of dysbindin in hippocampal mossy fiber-CA3 synaptic transmission and its modulation using the sandy mouse, a spontaneous mutant with deletion in the dysbindin gene. Electrophysiological recordings were made in hippocampal slices prepared from adult male sandy mice and their wild-type littermates. Basic properties of the mossy fiber synaptic transmission in the mutant mice were generally normal except for slightly reduced frequency facilitation. Serotonin and dopamine, two major neuromodulators implicated in the pathophysiology of schizophrenia, can potentiate mossy fiber synaptic transmission probably via an increase in cAMP levels. Synaptic potentiation induced by serotonin and dopamine was very variable in magnitude in the mutant mice, with some mice showing prominent enhancement as compared with the wild-type mice. In addition, the magnitude of potentiation induced by these monoamines significantly correlated with each other in the mutant mice, indicating that a subpopulation of sandy mice has marked hypersensitivity to both serotonin and dopamine. While direct activation of the cAMP cascade by forskolin induced robust synaptic potentiation in both wild-type and mutant mice, this forskolin-induced potentaition correlated in magnitude with the serotonin-induced potentiation only in the mutant mice, suggesting a possible change in coupling of receptor activation to downstream signaling. These results suggest that the dysbindin deficiency could be an essential genetic factor that causes synaptic hypersensitivity to dopamine and serotonin. The altered monoaminergic modulation at the mossy fiber synapse could be a candidate pathophysiological basis for impairment of hippocampus-dependent brain functions in schizophrenia.     

Allopregnanolone Reinstates Tyrosine Hydroxylase Immunoreactive Neurons and Motor Performance in an MPTP-Lesioned Mouse Model of Parkinson's Disease

Restorative/protective therapies to restore dopamine neurons in the substantia nigra pars compacta (SNpc) are greatly needed to effectively change the debilitating course of Parkinson''s disease. In this study, we tested the therapeutic potential of a neurogenic neurosteroid, allopregnanolone, in the restoration of the components of the nigrostriatal pathway in MPTP-lesioned mice by measuring striatal dopamine levels, total and tyrosine hydroxylase immunoreactive neuron numbers and BrdU-positive cells in the SNpc. An acute treatment (once/week for two weeks) with allopregnanolone restored the number of tyrosine hydroxylase-positive and total cell numbers in the SNpc of MPTP-lesioned mice, even though this did not increase striatal dopamine. It was also noted that MPTP treated mice to which allopregnanolone was administered had an increase in BrdU-positive cells in the SNpc. The effects of allopregnanolone in MPTP-lesioned mice were more apparent in mice that underwent behavioral tests. Interestingly, mice treated with allopregnanolone after MPTP lesion were able to perform at levels similar to that of non-lesioned control mice in a rotarod test. These data demonstrate that allopregnanolone promotes the restoration of tyrosine hydroxylase immunoreactive neurons and total cells in the nigrostriatal tract, improves the motor performance in MPTP-treated mice, and may serve as a therapeutic strategy for Parkinson''s disease.     

Dopamine release is severely compromised in the R6/2 mouse model of Huntington's disease

Recently, alterations in dopamine signaling have been implicated in Huntington's disease. In this work, dopamine release and uptake was measured in striatal slices from the R6/2 transgenic mouse model of Huntington's disease using fast-scan cyclic voltammetry at carbon-fiber microelectrodes. Dopamine release in brain slices from 6-week-old R6/2 mice is substantially reduced (53% of wild type), while dopamine uptake is unaffected. In agreement with this, R6/2 mice injected with the dopamine uptake inhibitor cocaine exhibited a blunted motor activity response (54% of wild type). At 10 weeks of age, an even more dramatic motor activity decrease in response to cocaine injection (21% of wild type) was observed. Moreover, the pre-drug activity of 10-week-old R6/2 mice was significantly reduced (by 37%) compared with 6-week-old R6/2 mice. Striatal dopamine release decreased with age, indicating that progressive alterations in dopaminergic pathways may affect motor activity. The inhibition constants of cocaine and methamphetamine (METH) determined in brain slices differed little between genotype or age group, suggesting that the decreased responses to cocaine and METH arise from compromised dopamine release rather than differences in uptake or drug action. Collectively, these data demonstrate (i) a reduction in the ability of dopamine terminals to release dopamine and (ii) the importance of this attenuation of release on the motor symptoms of Huntington's disease.     

Region-specific enhancement of basal extracellular and cocaine-evoked dopamine levels following constitutive deletion of the Serotonin(1B) receptor

The behavioral effects of cocaine are enhanced following constitutive deletion of the serotonin(1B) receptor. The neural substrates mediating the enhanced response to cocaine are unknown. The present studies determined whether basal dopamine dynamics or cocaine-evoked dopamine levels are altered in projection areas of mesostriatal or mesoaccumbens dopamine neurons following serotonin(1B) receptor deletion. Male wild-type and serotonin(1B) knockout mice were implanted with microdialysis guide cannulas aimed at the dorsal striatum or nucleus accumbens. The zero net flux method of quantitative microdialysis was used to quantify basal extracellular dopamine concentrations (DA(ext)) and the extraction fraction of dopamine (E(d)), which provides an index of dopamine uptake. Conventional microdialysis techniques were used to quantify cocaine (0, 5.0, and 20.0 mg/kg)-evoked dopamine overflow. Basal DA(ext) and E(d) did not differ in striatum of wild-type and knockout mice. Similarly, cocaine-stimulated dopamine overflow did not differ between genotype. The basal E(d) did not differ in the nucleus accumbens of wild-type and knockout mice. However, DA(ext) was significantly elevated in the nucleus accumbens of knockout mice. Cocaine-evoked dopamine overflow (nM) was also enhanced in the nucleus accumbens of knockout mice. However, the cocaine-induced increase in dopamine levels, relative to basal values, did not differ between genotype. These data demonstrate that deletion of the serotonin(1B) receptor is associated with increases in basal DA(ext) in the nucleus accumbens. This increase is not associated with an alteration in E(d), suggesting increased basal dopamine release in these animals. It is hypothesized that these alterations in presynaptic neuronal activity are a compensatory response to constitutive deletion of the serotonin(1B) receptor and may contribute to the enhanced behavioral effects of psychostimulants observed in knockout mice.     

Dopamine-independent locomotor actions of amphetamines in a novel acute mouse model of Parkinson disease

Brain dopamine is critically involved in movement control, and its deficiency is the primary cause of motor symptoms in Parkinson disease. Here we report development of an animal model of acute severe dopamine deficiency by using mice lacking the dopamine transporter. In the absence of transporter-mediated recycling mechanisms, dopamine levels become entirely dependent on de novo synthesis. Acute pharmacological inhibition of dopamine synthesis in these mice induces transient elimination of striatal dopamine accompanied by the development of a striking behavioral phenotype manifested as severe akinesia, rigidity, tremor, and ptosis. This phenotype can be reversed by administration of the dopamine precursor, L-DOPA, or by nonselective dopamine agonists. Surprisingly, several amphetamine derivatives were also effective in reversing these behavioral abnormalities in a dopamine-independent manner. Identification of dopamine transporter- and dopamine-independent locomotor actions of amphetamines suggests a novel paradigm in the search for prospective anti-Parkinsonian drugs.     

Adaptive Changes in Postsynaptic Dopamine Receptors Despite Unaltered Dopamine Dynamics in Mice Lacking Monoamine Oxidase B

Monoamine oxidase (MAO) B is considered a key enzyme in dopamine metabolism. The present studies, conducted in MAO B knockout mice, show that lack of MAO B does not alter extracellular levels of dopamine in striatum. Similarly, the synthesis, storage, uptake, and release of dopamine are also unaltered. However, autoradiography revealed a significant up-regulation of the D2-like dopamine receptors in the striatum of MAO B knockout mice. Mutant mice also exhibit a functional supersensitivity of D1-dopamine receptors in the nucleus accumbens. Thus, the agonist SKF 38,393-induced c-Fos immunoreactivity was significantly increased in knockout mice as compared with wild-type controls. In view of the apparently normal basal dopamine dynamics observed in MAO B knockout mice, we hypothesize that a dopamine-independent mechanism underlies adaptations in dopamine receptor function that occur as a consequence of MAO B depletion. Finally, these findings suggest that chronic administration of MAO inhibitors, as occurs in the treatment of Parkinson's disease and depression, may be associated with an increased responsiveness of CNS neurons to dopamine receptor ligands.     

Methamphetamine Increases Locomotion and Dopamine Transporter Activity in Dopamine D5 Receptor-Deficient Mice

Dopamine regulates the psychomotor stimulant activities of amphetamine-like substances in the brain. The effects of dopamine are mediated through five known dopamine receptor subtypes in mammals. The functional relevance of D5 dopamine receptors in the central nervous system is not well understood. To determine the functional relevance of D5 dopamine receptors, we created D5 dopamine receptor-deficient mice and then used these mice to assess the roles of D5 dopamine receptors in the behavioral response to methamphetamine. Interestingly, D5 dopamine receptor-deficient mice displayed increased ambulation in response to methamphetamine. Furthermore, dopamine transporter threonine phosphorylation levels, which regulate amphetamine-induced dopamine release, were elevated in D5 dopamine receptor-deficient mice. The increase in methamphetamine-induced locomotor activity was eliminated by pretreatment with the dopamine transporter blocker GBR12909. Taken together, these results suggest that dopamine transporter activity and threonine phosphorylation levels are regulated by D5 dopamine receptors.     

Exogenous, but not endogenous nociceptin modulates mesolimbic dopamine release in mice

The effect of nociceptin (an endogenous ligand of the ORL1 receptor) on mesolimbic dopamine release and simultaneous horizontal locomotion was studied in freely moving mice undergoing microdialysis of the nucleus accumbens. Intracerebroventricular (i.c.v.) administration of nociceptin (7 nmol) induced a long-lasting suppression of mesolimbic dopamine release and horizontal locomotion in wild-type but not ORL1 knockout mice. I.c.v. administration of the recently reported peptide nociceptin antagonist [Nphe1, Arg14, Lys15] nociceptin-NH(2) (known also as UFP-101, 5 nmol) completely abolished the suppressive effect of nociceptin on mesolimbic dopamine release. However, UFP-101 administration alone induced a mild and lasting suppression of mesolimbic dopamine release in both wild-type and ORL1 knockout mice that was magnified in ORL1 knockout mice by coadministration of nociceptin. UFP-101 administration alone suppressed locomotion in both genotypes. These results confirm that the suppressive action of nociceptin on mesolimbic dopamine release is mediated entirely by the ORL1 receptor, and that UFP-101 effectively antagonizes this action. However, the lack of a stimulatory effect of UFP-101 in wild-type mice indicates that despite being sensitive to exogenous nociceptin action, basal mesolimbic dopaminergic activity is not determined by endogenous nociceptin in mice.     

Behavioral, cellular and molecular consequences of the dopamine transporter gene inactivation

Mice lacking the the plasma membrane dopamine transporter (DAT), following gene inactivation or knock out, show an increase in their spontaneous locomotor activity that is of the same magnitude than in normal mice treated with amphetamine or cocaine, known to increase levels of dopamine in the basal ganglia. Many adaptive responses have occurred in these animals than could not compensate for the hyper activity of the dopamine system. Surprisingly, while intracellular dopamine levels were of only 5%, extracellular dopamine levels were increased by 300%. We investigated the regulation of tyrosine hydroxylase (TH), the rate limiting enzyme of dopamine synthesis, and found that this enzyme is regulated at the levels of mRNA, protein, trafficking as well as in its regional, cellular and subcellular organization. Our results establish not only the central importance of the transporter as the key element controlling dopamine levels in the brain, but also its role in the behavioral and biochemical action of amphetamine, cocaine and morphine. In addition, these mice have provided key elements leading to possible clinical and social implications for illnesses such as Parkinson disease, attention deficit disorder and drug addiction.     

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.     

多巴胺D5受体转基因小鼠的建立

目的建立人多巴胺D5受体突变基因F173 L（D5F173L）,S390G（D5S390G）及正常人D5基因（hD5 WT）的转基因小鼠,利用该转基因动物模型来研究D5受体在原发性高血压中的发病机制。方法利用显微注射的技术将插入CMV启动子下游的D5F173L,D5S390G及hD5 WT基因的转基因载体注射入C57BL/6小鼠体内,建立多巴胺D5F173L,D5S390G及hD5 WT的转基因小鼠。通过PCR鉴定转基因小鼠的基因型。利用Western Blotting方法鉴定该受体蛋白在肾脏的表达情况。使用智能无创血压测量仪测量转基因小鼠的血压值。结果分别建立了多巴胺D5F173L,D5S390G及hD5 WT转基因C57BL/6小鼠,Western Blotting方法鉴定结果显示,与非转基因C57BL/6小鼠比较,D5转基因小鼠D5受体在肾脏有较高的表达。3-6月龄D5 F173L转基因小鼠的收缩压、舒张压和平均动脉血压均明显高于多巴胺D5S390G及hD5 WT转基因小鼠（n=6-8,P〈0.05）。结论多巴胺D5受体在原发性高血压发病中具有重要作用,但作用机理还有待于进一步研究。