The adenosine A2A receptor (A2AR) is a potential drug target for the treatment of Parkinson’s disease and other neurological disorders. In rodents, the therapeutic efficacy of A2AR modulation is improved by concomitant modulation of the metabotropic glutamate receptor 5 (mGluR5). To elucidate the anatomical substrate(s) through which these therapeutic benefits could be mediated, pre-embedding electron microscopy immunohistochemistry was used to conduct a detailed, quantitative ultrastructural analysis of A2AR localization in the primate basal ganglia and to assess the degree of A2AR/mGluR5 colocalization in the striatum. A2AR immunoreactivity was found at the highest levels in the striatum and external globus pallidus (GPe). However, the monkey, but not the rat, substantia nigra pars reticulata (SNr) also harbored a significant level of neuropil A2AR immunoreactivity. At the electron microscopic level, striatal A2AR labeling was most commonly localized in postsynaptic elements (58% ± 3% of labeled elements), whereas, in the GPe and SNr, the labeling was mainly presynaptic (71% ± 5%) or glial (27% ± 6%). In both striatal and pallidal structures, putative inhibitory and excitatory terminals displayed A2AR immunoreactivity. Striatal A2AR/mGluR5 colocalization was commonly found; 60–70% of A2AR-immunoreactive dendrites or spines in the monkey striatum coexpress mGluR5. These findings provide the first detailed account of the ultrastructural localization of A2AR in the primate basal ganglia and demonstrate that A2AR and mGluR5 are located to interact functionally in dendrites and spines of striatal neurons. Together, these data foster a deeper understanding of the substrates through which A2AR could regulate primate basal ganglia function and potentially mediate its therapeutic effects in parkinsonism.
mGluR5; Parkinson’s disease; primate; immunogold; globus pallidus; substantia nigra; putamen
An extensive literature shows that astrocytes exhibit metabotropic glutamate receptor 5 (mGluR5)–dependent increases in cytosolic calcium ions (Ca2+) in response to glutamatergic transmission and, in turn, modulate neuronal activity by their Ca2+-dependent release of gliotransmitters. These findings, based on studies of young rodents, have led to the concept of the tripartite synapse, in which astrocytes actively participate in neurotransmission. Using genomic analysis, immunoelectron microscopy, and two-photon microscopy of astrocytic Ca2+ signaling in vivo, we found that astrocytic expression of mGluR5 is developmentally regulated and is undetectable after postnatal week 3. In contrast, mGluR3, whose activation inhibits adenylate cyclase but not calcium signaling, was expressed in astrocytes at all developmental stages. Neuroglial signaling in the adult brain may therefore occur in a manner fundamentally distinct from that exhibited during development.
The demonstration that dopamine loss is the key pathological feature of Parkinson's disease (PD), and the subsequent introduction of levodopa have revolutionalized the field of PD therapeutics. This review will discuss the significant progress that has been made in the development of new pharmacological and surgical tools to treat PD motor symptoms since this major breakthrough in the 1960s. However, we will also highlight some of the challenges the field of PD therapeutics has been struggling with during the past decades. The lack of neuroprotective therapies and the limited treatment strategies for the nonmotor symptoms of the disease (ie, cognitive impairments, autonomic dysfunctions, psychiatric disorders, etc.) are among the most pressing issues to be addressed in the years to come. It appears that the combination of early PD nonmotor symptoms with imaging of the nigrostriatal dopaminergic system offers a promising path toward the identification of PD biomarkers, which, once characterized, will set the stage for efficient use of neuroprotective agents that could slow down and alter the course of the disease.
dopamine; nonmotor symptoms; striatum; substantia nigra; deep brain stimulation; transplantation; dopamine; movement disorders; neurodegeneration/neuroprotection; neurology; basal ganglia; substantia nigra; deep brain stimulation; nigrostriatal; adenosine
Electrical and pharmacological stimulation methods are commonly used to study neuronal brain circuits in vivo, but are problematic, because electrical stimulation has limited specificity, while pharmacological activation has low temporal resolution. A recently developed alternative to these methods is the use of optogenetic techniques, based on the expression of light sensitive channel proteins in neurons. While optogenetics have been applied in in vitro preparations and in in vivo studies in rodents, their use to study brain function in nonhuman primates has been limited to the cerebral cortex. Here, we characterize the effects of channelrhodopsin-2 (ChR2) transfection in subcortical areas, i.e., the putamen, the external globus pallidus (GPe) and the ventrolateral thalamus (VL) of rhesus monkeys. Lentiviral vectors containing the ChR2 sequence under control of the elongation factor 1α promoter (pLenti-EF1α -hChR2(H134R)-eYFP-WPRE, titer 109 particles/ml) were deposited in GPe, putamen and VL. Four weeks later, a probe combining a conventional electrode and an optic fiber was introduced in the previously injected brain areas. We found light-evoked responses in 31.5% and 32.7% of all recorded neurons in the striatum and thalamus, respectively, but only in 2.5% of recorded GPe neurons. As expected, most responses were time-locked increases in firing, but decreases or mixed responses were also seen, presumably via ChR2-mediated activation of local inhibitory connections. Light and electron microscopic analyses revealed robust expression of ChR2 on the plasma membrane of cell somas, dendrites, spines and terminals in the striatum and VL. This study demonstrates that optogenetic experiments targeting the striatum and basal ganglia-related thalamic nuclei can be successfully achieved in monkeys. Our results indicate important differences of the type and magnitude of responses in each structure. Experimental conditions such as the vector used, the number and rate of injections, or the light stimulation conditions have to be optimized for each structure studied.
There is an increasing amount of literature data showing the positive effects on preclinical anti-Parkinsonian rodent models with selective positive allosteric modulators of metabotropic glutamate receptor 4 (mGlu4).1 However, most of the data generated utilize compounds that have not been optimized for drug-like properties and, as a consequence, they exhibit poor pharmacokinetic properties and thus do not cross the blood-brain barrier. Herein, we report on a series of N-4-(2,5-dioxopyrrolidin-1-yl)-phenylpicolinamides with improved PK properties with excellent potency and selectivity as well as improved brain exposure in rodents. Finally, ML182 was shown to be orally active in the haloperidol induced catalepsy model, a well-established anti-Parkinsonian model.
metabotropic glutamate receptors; mGlu4; positive allosteric modulators; Parkinson’s disease; haloperidol-induced catalepsy; structure-activity relationship (SAR); oral efficacy; brain penetration
The striatum receives glutamatergic inputs from two main thalamostriatal systems that originate either from the centre median/parafascicular complex (CM/PF-striatal system) or the rostral intralaminar, midline, associative and relay thalamic nuclei (non-CM/PF-striatal system). These dual thalamostriatal systems display striking differences in their anatomical and, most likely, functional organization. The CM/PF-striatal system is topographically organized, and integrated within functionally segregated basal ganglia-thalamostriatal circuits that process sensorimotor, associative and limbic information. CM/PF neurons are highly responsive to attention-related sensory stimuli, suggesting that the CM/PF-striatal system, through its strong connections with cholinergic interneurons, may play a role in basal ganglia-mediated learning, behavioral switching and reinforcement. In light of evidence for prominent CM/PF neuronal loss in Parkinson’s disease, we propose that the significant CM-striatal system degeneration, combined with the severe nigrostriatal dopamine loss in sensorimotor striatal regions, may alter normal automatic actions, and shift the processing of basal ganglia-thalamocortical motor programs towards goal-directed behaviors.
Centromedian; Parafascicular; set shifting; striatum; learning; cholinergic interneuron
The Biogenesis of Lysosome-Related Organelles Complex 1 (BLOC-1) is a protein complex containing the schizophrenia susceptibility factor dysbindin, which is encoded by the gene DTNBP1. However, mechanisms engaged by dysbindin defining schizophrenia susceptibility pathways have not been quantitatively elucidated. Here, we discovered prevalent and novel cellular roles of the BLOC-1 complex in neuronal cells by performing large-scale Stable Isotopic Labeling of Cells in Culture quantitative proteomics (SILAC) combined with genetic analyses in dysbindin-null mice (Mus musculus) and the genome of schizophrenia patients. We identified 24 proteins that associate with the BLOC-1 complex many of which were altered in content/distribution in cells or tissues deficient in BLOC-1. New findings include BLOC-1 interactions with the COG complex, a Golgi apparatus tether, and antioxidant enzymes peroxiredoxins 1-2. Importantly, loci encoding eight of the 24 proteins are affected by genomic copy number variation in schizophrenia patients. Thus, our quantitative proteomic studies expand the functional repertoire of the BLOC-1 complex and provide insight into putative molecular pathways of schizophrenia susceptibility.
The positron emission tomography (PET) tracer 2β-carbomethoxy-3β-(4-chlorophenyl)-8-(2-[18F]-fluoroethyl)-nortropane (18F-FECNT) is a highly specific ligand for dopamine transporter (DAT) that yields higher peak striatum-to-cerebellum ratios and offers more favorable kinetics than most 18F-radiolabeled DAT ligands currently available. The goal of this study is to validate the use of 18F-FECNT as a PET radiotracer to assess the degree of striatal dopamine terminals denervation and midbrain dopaminergic cell loss in MPTP-treated parkinsonian monkeys. Three rhesus monkeys received weekly injections of MPTP (0.2-0.5 mg/kg) for 21 weeks, which resulted in the progressive development of a moderate level of parkinsonism. We carried out 18F-FECNT PET at baseline (twice; ten weeks apart) and at week 21 post-MPTP injections. Postmortem stereological cell counts of dopaminergic neurons in the ventral midbrain, and intensity measurements of DAT and tyrosine hydroxylase (TH) immunoreactivity in the striatum were performed and correlated with striatal and ventral midbrain PET data. Three additional monkeys were used as controls for midbrain dopaminergic cell counts, and striatal DAT or TH immunoreactivity measurements. The correlation and coefficient of variance between 18F-FECNT test-retest specific uptake ratios were 0.99 (R2) and 2.65%, respectively. The 18F-FECNT binding potential of the ventral midbrain and striatal regions was tightly correlated with postmortem stereological cell counts of nigral dopaminergic neurons (R2 = 0.91), and striatal DAT (R2 = 0.83) or TH (R2 = 0.88) immunoreactivity intensity measurements. These findings demonstrate that 18F-FECNT is a highly sensitive PET imaging ligand to quantify both striatal dopamine denervation and midbrain dopaminergic cell loss associated with parkinsonism.
Parkinson’s disease; brain imaging; transporter; dopamine; striatum; MPTP; monkey; primate; substantia nigra; midbrain; animal model
Degeneration of the dopaminergic nigrostriatal system and of noradrenergic neurons in the locus coeruleus are important pathological features of Parkinson’s disease. There is an urgent need to develop therapies that slow down the progression of neurodegeneration in Parkinson’s disease. In the present study, we tested whether the highly specific metabotropic glutamate receptor 5 antagonist, 3-[(2-methyl-1,3-thiazol-4-yl) ethynyl] pyridine, reduces dopaminergic and noradrenergic neuronal loss in monkeys rendered parkinsonian by chronic treatment with low doses of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine. Weekly intramuscular 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine injections (0.2–0.5 mg/kg body weight), in combination with daily administration of 3-[(2-methyl-1,3-thiazol-4-yl) ethynyl] pyridine or vehicle, were performed until the development of parkinsonian motor symptoms in either of the two experimental groups (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine/3-[(2-methyl-1,3-thiazol-4-yl) ethynyl] pyridine versus 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine/vehicle). After 21 weeks of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine treatment, all 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine/vehicle-treated animals displayed parkinsonian symptoms, whereas none of the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine/3-[(2-methyl-1,3-thiazol-4-yl) ethynyl] pyridine-treated monkeys were significantly affected. These behavioural observations were consistent with in vivo positron emission tomography dopamine transporter imaging data, and with post-mortem stereological counts of midbrain dopaminergic neurons, as well as striatal intensity measurements of dopamine transporter and tyrosine hydroxylase immunoreactivity, which were all significantly higher in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine/3-[(2-methyl-1,3-thiazol-4-yl) ethynyl] pyridine-treated animals than in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine/vehicle-treated monkeys. The 3-[(2-methyl-1,3-thiazol-4-yl) ethynyl] pyridine treatment also had a significant effect on the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced loss of norepinephrine neurons in the locus coeruleus and adjoining A5 and A7 noradrenaline cell groups. In 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine/vehicle-treated animals, almost 40% loss of tyrosine hydroxylase-positive norepinephrine neurons was found in locus coeruleus/A5/A7 noradrenaline cell groups, whereas the extent of neuronal loss was lower than 15% of control values in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine/3-[(2-methyl-1,3-thiazol-4-yl) ethynyl] pyridine-treated monkeys. Our data demonstrate that chronic treatment with the metabotropic glutamate receptor 5 antagonist, 3-[(2-methyl-1,3-thiazol-4-yl) ethynyl] pyridine, significantly reduces 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine toxicity towards dopaminergic and noradrenergic cell groups in non-human primates. This suggests that the use of metabotropic glutamate receptor 5 antagonists may be a useful strategy to reduce degeneration of catecholaminergic neurons in Parkinson’s disease.
substantia nigra; locus coeruleus; striatum; neuroprotection; noradrenaline
Changes in GABAergic transmission in the external and internal segments of the globus pallidus (GPe and GPi) contribute to the pathophysiology of the basal ganglia network in Parkinson’s disease. Because GABA-B receptors are involved in the modulation of GABAergic transmission in GPe and GPi, it is possible that changes in the functions or localization of these receptors contribute to the changes in GABAergic transmission. To further examine this question, we investigated the anatomical localization of GABA-B receptors and the electrophysiologic effects of microinjections of GABA-B receptor ligands in GPe and GPi of MPTP-treated (parkinsonian) monkeys. We found that the pattern of cellular and ultrastructural localization of the GABA-BR1 subunit of the GABA-B receptor in GPe and GPi was not significantly altered in parkinsonian monkeys. However, the magnitude of reduction in firing rate of GPe and GPi neurons produced by microinjections of the GABA-B receptor agonist baclofen was larger in MPTP-treated animals than in normal monkeys. Injections of the GABA-B receptor antagonist CGP55845A were more effective in reducing the firing rate of GPi neurons in parkinsonian monkeys than in normal animals. In addition, the injections of baclofen in GPe and GPi, or of CGP55845A in GPi lead to a significant increase in the proportion of spikes in rebound bursts in parkinsonian animals, but not in normal monkeys. Thus, despite the lack of changes in the localization of GABA-BR1 subunits in the pallidum, GABA-B receptor-mediated effects are altered in the GPe and GPi of parkinsonian monkeys. These changes in GABA-B receptors function may contribute to bursting activities in the parkinsonian state.
External segment of the globus pallidus; internal segment of the globus pallidus; Parkinson’s disease; GABAergic transmission; ultrastructural localization; nonhuman primate; in vivo electrophysiology
Although the existence of prominent connections between the intralaminar thalamic nuclei and the basal ganglia have long been established, the limited knowledge of the functional relevance of this network has considerably hampered progress in our understanding of the neural mechanisms by which the thalamostriatal system integrates and regulates the basal ganglia circuitry. In this brief commentary, we will address this gap of knowledge through a discussion of the key points of a symposium entitled “Thalamic Contributions to Basal Ganglia-Related Behavioral Switching and Reinforcement” that will be presented at the 2011 Society for Neuroscience meeting. Recent anatomical and physiological data that support the role of the thalamostriatal system in action selection, attentional shifting and reinforcement will be discussed. We will also address the possibility that degeneration of the thalamostriatal system could underlie some of the deficits in redirection of attention in response to salient stimuli seen in Parkinson’s disease.
Centre median; parafascicular nucleus; superior colliculus; striatum; thalamostriatal; interneuron; acetylcholine; attention; Parkinson’s disease
A novel vesicle transport mechanism is described that requires dysbindin-associated complexes for cargo targeting from neuronal cell bodies to neurites and nerve terminals. The results suggest that mistargeting of specific vesicular cargoes may underlie, in part, the molecular pathogenesis of schizophrenia.
Dysbindin assembles into the biogenesis of lysosome-related organelles complex 1 (BLOC-1), which interacts with the adaptor protein complex 3 (AP-3), mediating a common endosome-trafficking route. Deficiencies in AP-3 and BLOC-1 affect synaptic vesicle composition. However, whether AP-3-BLOC-1–dependent sorting events that control synapse membrane protein content take place in cell bodies upstream of nerve terminals remains unknown. We tested this hypothesis by analyzing the targeting of phosphatidylinositol-4-kinase type II α (PI4KIIα), a membrane protein present in presynaptic and postsynaptic compartments. PI4KIIα copurified with BLOC-1 and AP-3 in neuronal cells. These interactions translated into a decreased PI4KIIα content in the dentate gyrus of dysbindin-null BLOC-1 deficiency and AP-3–null mice. Reduction of PI4KIIα in the dentate reflects a failure to traffic from the cell body. PI4KIIα was targeted to processes in wild-type primary cultured cortical neurons and PC12 cells but failed to reach neurites in cells lacking either AP-3 or BLOC-1. Similarly, disruption of an AP-3–sorting motif in PI4KIIα impaired its sorting into processes of PC12 and primary cultured cortical neuronal cells. Our findings indicate a novel vesicle transport mechanism requiring BLOC-1 and AP-3 complexes for cargo sorting from neuronal cell bodies to neurites and nerve terminals.
Dopamine receptors (DARs) in the nucleus accumbens (NAc) are critical for cocaine's actions but the nature of adaptations in DAR function after repeated cocaine exposure remains controversial. This may be due in part to the fact that different methods used in previous studies measured different DAR pools. In the present study, we used a protein crosslinking assay to make the first measurements of DAR surface expression in the NAc of cocaine-experienced rats. Intracellular and total receptor levels were also quantified. Rats self-administered saline or cocaine for ten days. The entire NAc, or core and shell subregions, were collected one or 45 days later, when rats are known to exhibit low and high levels of cue-induced drug seeking, respectively. We found increased cell surface D1 DARs in the NAc shell on the first day after discontinuing cocaine self-administration (designated withdrawal day 1, or WD1) but this normalized by WD45. Decreased intracellular and surface D2 DAR levels were observed in the cocaine group. In shell, both measures decreased on WD1 and WD45. In core, decreased D2 DAR surface expression was only observed on WD45. Similarly, WD45 but not WD1 was associated with increased D3 DAR surface expression in the core. Taking into account many other studies, we suggest that decreased D2 DAR and increased D3 DAR surface expression on WD45 may contribute to enhanced cocaine-seeking after prolonged withdrawal, although this is likely to be a modulatory effect, in light of the mediating effect previously demonstrated for AMPA-type glutamate receptors.
cocaine; dopamine receptors; nucleus accumbens; receptor trafficking
Progressive loss of the ascending dopaminergic projection in the basal ganglia is a fundamental pathological feature of Parkinson’s disease. Studies in animals and humans have identified spatially segregated functional territories in the basal ganglia for the control of goal-directed and habitual actions. In patients with Parkinson’s disease the loss of dopamine is predominantly in the posterior putamen, a region of the basal ganglia associated with the control of habitual behaviour. These patients may therefore be forced into a progressive reliance on the goal-directed mode of action control that is mediated by comparatively preserved processing in the rostromedial striatum. Thus, many of their behavioural difficulties may reflect a loss of normal automatic control owing to distorting output signals from habitual control circuits, which impede the expression of goal-directed action.
The GABA transporters GAT-1 and GAT-3 are abundant in the external and internal segments of the globus pallidus (GPe and GPi, respectively). We have shown that pharmacological blockade of either of these transporters results in decreased neuronal firing, and in elevated levels of extracellular GABA in normal monkeys. We now studied whether the electrophysiologic and biochemical effects of local intra-pallidal injections of GAT-1 and GAT-3 blockers, or the subcellular localization of these transporters, are altered in monkeys rendered parkinsonian by the administration of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). The subcellular localization of the transporters in GPe and GPi, studied with electron microscopy immunoperoxidase, was similar to that found in normal animals: i.e., GAT-3 immunoreactivity was mostly confined to glial processes, while GAT-1 labeling was expressed in unmyelinated axons and glial processes. A combined injection/recording device was used to record extracellular activity of single neurons in GPe and GPi, before, during and after administration of small volumes (1 μl) of either the GAT-1 inhibitor, SKF-89976A hydrochloride (720 ng), or the GAT-3 inhibitor, (S)-SNAP-5114 (500 ng). In GPe, the effects of GAT-1 or GAT-3 blockade were similar to those seen in normal monkeys. However, unlike the findings in the normal state, the firing of most neurons was not affected by blockade of either transporter in GPi. These results suggest that, after dopaminergic depletion, the functions of GABA transporters are altered in GPi; without major changes in their subcellular localization.
Parkinsonism; globus pallidus external segment; globus pallidus internal segment; GABAergic transmission; single unit extracellular recording; electron microscope; subcellular distribution
Striatal spine loss is a key pathological feature of Parkinson's disease (PD). Knowing that striatal glutamatergic afferents target dendritic spines, these data appear difficult to reconcile with evidence for an increased expression of the vesicular glutamate transporter 1 (vGluT1) in the striatum of PD patients and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated monkeys, as well as in some electrophysiological studies showing overactivity of the corticostriatal glutamatergic system in models of parkinsonism. To address the possibility that structural changes in glutamatergic afferents may underlie these discrepancies, we undertook an ultrastructural analysis of vGluT1-positive (i.e., corticostriatal) and vGluT2-positive (i.e., mostly thalamostriatal) axo-spinous glutamatergic synapses using a 3D electron microscopic approach in normal and MPTP-treated monkeys. Three main conclusions can be drawn: 1) spines contacted by vGluT1-containing terminals have larger volume and harbor significantly larger postsynaptic densities (PSDs) than those contacted by vGluT2-immunoreactive boutons; 2) a subset of vGluT2-, but not vGluT1-immunoreactive, terminals display a pattern of multisynaptic connectivity in normal and MPTP-treated monkeys; and 3) VGluT1- and vGluT2-positive axo-spinous synapses undergo ultrastructural changes (larger spine volume, larger PSDs, increased PSD perforations, larger presynaptic terminal) indicative of increased synaptic activity in parkinsonian animals. Furthermore, spines contacted by cortical terminals display an increased volume of their spine apparatus in MPTP-treated monkeys, suggesting an increased protein synthesis at corticostriatal synapses. These findings demonstrate that corticostriatal and thalamostriatal glutamatergic axo-spinous synapses display significantly different ultrastructural features, and that both systems undergo complex morphological changes that could underlie the pathophysiology of corticostriatal and thalamostriatal systems in PD.
dopamine; glutamate; vesicular glutamate transporters; electron microscopy; dendritic spine; 3D-reconstruction
The transcription factors in the myocyte enhancer factor 2 (MEF2) family play important roles in cell survival by regulating nuclear gene expression. Here, we report that MEF2D is present in rodent neuronal mitochondria, where it can regulate the expression of a gene encoded within mitochondrial DNA (mtDNA). Immunocytochemical, immunoelectron microscopic, and biochemical analyses of rodent neuronal cells showed that a portion of MEF2D was targeted to mitochondria via an N-terminal motif and the chaperone protein mitochondrial heat shock protein 70 (mtHsp70). MEF2D bound to a MEF2 consensus site in the region of the mtDNA that contained the gene NADH dehydrogenase 6 (ND6), which encodes an essential component of the complex I enzyme of the oxidative phosphorylation system; MEF2D binding induced ND6 transcription. Blocking MEF2D function specifically in mitochondria decreased complex I activity, increased cellular H2O2 level, reduced ATP production, and sensitized neurons to stress-induced death. Toxins known to affect complex I preferentially disrupted MEF2D function in a mouse model of Parkinson disease (PD). In addition, mitochondrial MEF2D and ND6 levels were decreased in postmortem brain samples of patients with PD compared with age-matched controls. Thus, direct regulation of complex I by mitochondrial MEF2D underlies its neuroprotective effects, and dysregulation of this pathway may contribute to PD.
GABA transporter type 1 and 3 (GAT-1 and GAT-3, respectively) are the two main subtypes of GATs responsible for the regulation of extracellular GABA levels in the central nervous system. These transporters are widely expressed in neuronal (mainly GAT-1) and glial (mainly GAT-3) elements throughout the brain, but most data obtained so far relate to their role in the regulation of GABAA receptor-mediated postsynaptic tonic and phasic inhibition in the hippocampus, cerebral cortex and cerebellum. Taking into consideration the key role of GABAergic transmission within basal ganglia networks, and the importance for these systems to be properly balanced to mediate normal basal ganglia function, we analyzed in detail the localization and function of GAT-1 and GAT-3 in the globus pallidus of normal and Parkinsonian animals, in order to further understand the substrate and possible mechanisms by which GABA transporters may regulate basal ganglia outflow, and may become relevant targets for new therapeutic approaches for the treatment of basal ganglia-related disorders. In this review, we describe the general features of GATs in the basal ganglia, and give a detailed account of recent evidence that GAT-1 and GAT-3 regulation can have a major impact on the firing rate and pattern of basal ganglia neurons through pre- and post-synaptic GABAA- and GABAB-receptor-mediated effects.
GABA transporter; striatum; globus pallidus; substantia nigra; patch clamp recording
The basal ganglia receive cortical inputs through two main stations – the striatum and the subthalamic nucleus (STN). The information flowing along the corticostriatal system is transmitted to the basal ganglia circuitry via the “direct and indirect” striatofugal pathways, while information that flows through the STN is transmitted along the so-called “hyperdirect” pathway. The functional significance of this dual entry system is not clear. Although the corticostriatal system has been thoroughly characterized anatomically and electrophysiologically, such is not the case for the corticosubthalamic system. In order to provide further insights into the intricacy of this complex anatomical organization, this review examines and compares the anatomical and functional organization of the corticostriatal and corticosubthalamic systems, and highlights some key issues that must be addressed to better understand the mechanisms by which these two neural systems may interact to regulate basal ganglia functions and dysfunctions.
subthalamic nucleus; striatum; hyperdirect; Parkinson's disease; basal ganglia; cerebral cortex; monkey; deep brain stimulation
Striatal dopamine denervation is the pathological hallmark of Parkinson's disease (PD). Another major pathological change described in animal models and PD patients is a significant reduction in the density of dendritic spines on medium spiny striatal projection neurons. Simultaneously, the ultrastructural features of the neuronal synaptic elements at the remaining corticostriatal and thalamostriatal glutamatergic axo-spinous synapses undergo complex ultrastructural remodeling consistent with increased synaptic activity (Villalba and Smith, 2011). The concept of tripartite synapses (TS) was introduced a decade ago, according to which astrocytes process and exchange information with neuronal synaptic elements at glutamatergic synapses (Araque et al., 1999a). Although there has been compelling evidence that astrocytes are integral functional elements of tripartite glutamatergic synaptic complexes in the cerebral cortex and hippocampus, their exact functional role, degree of plasticity and preponderance in other CNS regions remain poorly understood. In this review, we discuss our recent findings showing that neuronal elements at cortical and thalamic glutamatergic synapses undergo significant plastic changes in the striatum of MPTP-treated parkinsonian monkeys. We also present new ultrastructural data that demonstrate a significant expansion of the astrocytic coverage of striatal TS synapses in the parkinsonian state, providing further evidence for ultrastructural compensatory changes that affect both neuronal and glial elements at TS. Together with our limited understanding of the mechanisms by which astrocytes respond to changes in neuronal activity and extracellular transmitter homeostasis, the role of both neuronal and glial components of excitatory synapses must be considered, if one hopes to take advantage of glia–neuronal communication knowledge to better understand the pathophysiology of striatal processing in parkinsonism, and develop new PD therapeutics.
glia; MPTP; non-human primates; tripartite synapses; astrocyte; striatum; corticostriatal; thalamostriatal
Oxytocin regulates partner preference formation and alloparental behavior in the socially monogamous prairie vole (Microtus ochrogaster) by activating oxytocin receptors in the nucleus accumbens of females. Mating facilitates partner preference formation, and oxytocin-immunoreactive fibers in the nucleus accumbens have been described in prairie voles. However, there has been no direct evidence of oxytocin release in the nucleus accumbens during sociosexual interactions, and the origin of the oxytocin fibers is unknown. Here we show for the first time that extracellular concentrations of oxytocin are increased in the nucleus accumbens of female prairie vole during unrestricted interactions with a male. We further show that the distribution of oxytocin-immunoreactive fibers in the nucleus accumbens is conserved in prairie voles, mice and rats, despite remarkable species differences in oxytocin receptor binding in the region. Using a combination of site-specific and peripheral infusions of the retrograde tracer, Fluorogold, we demonstrate that the nucleus accumbens oxytocin-immunoreactive fibers likely originate from paraventricular and supraoptic hypothalamic neurons. This distribution of retrogradely labeled neurons is consistent with the hypothesis that striatal oxytocin fibers arise from collaterals of magnocellular neurons of the neurohypophysial system. If correct, this may serve to coordinate peripheral and central release of oxytocin with appropriate behavioral responses associated with reproduction, including pair bonding after mating, and maternal responsiveness following parturition and during lactation.
pair bonding; nucleus accumbens; paraventricular nucleus; supraoptic nucleus; neurohypophysial peptides; alloparental behavior
Endosomal sorting mechanisms mediated by AP-3 and BLOC-1 are perturbed in Hermansky-Pudlak Syndrome, a human genetic condition characterized by albinism and prolonged bleeding (OMIM #203300). Additionally, mouse models defective in either one of these complexes possess defective synaptic vesicle biogenesis (Newell-Litwa et al., 2009). These synaptic vesicle phenotypes were presumed uniform throughout the brain. However, here we report that AP-3 and BLOC-1 differentially regulate the composition of pre-synaptic terminals in the striatum and dentate gyrus of the hippocampus. Quantitative immunoelectron microscopy demonstrated that the majority of AP-3 immunoreactivity in both wild type striatum and hippocampus localizes to pre-synaptic axonal compartments, where it regulates synaptic vesicle size. In the striatum, loss of AP-3 (Ap3dmh/mh) resulted in decreased synaptic vesicle size. In contrast, loss of AP-3 in the dentate gyrus increased synaptic vesicle size, thus suggesting anatomically specific AP-3-regulatory mechanisms. Loss-of-function alleles of BLOC-1, Pldnpa/pa and Mutedmu/mu, revealed that this complex acts as a brain-region specific regulator of AP-3. In fact, BLOC-1 deficiencies selectively reduced AP-3 and AP-3 cargo immunoreactivity in pre-synaptic compartments within the dentate gyrus both at the light and/or electron microscopy level. However, the striatum did not exhibit these BLOC-1-null phenotypes. Our results demonstrate that distinct brain regions differentially regulate AP-3-dependent synaptic vesicle biogenesis. We propose that anatomically restricted mechanisms within the brain diversify the biogenesis and composition of synaptic vesicles.
Synapse; Synaptic; dentate gyrus; striatum; presynaptic regulation; Membrane
Electrophysiological and pharmacological studies have demonstrated that alpha-1 adrenergic receptor (α1AR) activation facilitates dopamine (DA) transmission in the striatum and ventral midbrain. However, because little is known about the localization of α1ARs in dopaminergic regions, the substrate(s) and mechanism(s) underlying this facilitation of DA signaling are poorly understood. To address this issue, we used light and electron microscopy immunoperoxidase labeling to examine the cellular and ultrastructural distribution of α1ARs in the caudate putamen, nucleus accumbens, ventral tegmental area, and substantia nigra in the rat. Analysis at the light microscopic level revealed α1AR immunoreactivity mainly in neuropil, with occasional staining in cell bodies. At the electron microscopic level, α1AR immunoreactivity was found primarily in presynaptic elements, with scarce postsynaptic labeling. Unmyelinated axons and about 30–50% terminals forming asymmetric synapses contained the majority of presynaptic labeling in the striatum and midbrain, while in the midbrain a subset of terminals forming symmetric synapses also displayed immunoreactivity. Postsynaptic labeling was scarce in both striatal and ventral midbrain regions. On the other hand, only 3–6% of spines displayed α1AR immunoreactivity in the caudate putamen and nucleus accumbens,. These data suggest that the facilitation of dopaminergic transmission by α1ARs in the mesostriatal system is probably achieved primarily by pre-synaptic regulation of glutamate and GABA release.
norepinephrine; Parkinson’s disease; addiction; dopamine; axon; electron microscopy
Although we have gained significant knowledge in the anatomy and microcircuitry of the thalamostriatal system over the last decades, the exact function(s) of these complex networks remain(s) poorly understood. It is now clear that the thalamostriatal system is not a unique entity, but consists of multiple neural systems that originate from a wide variety of thalamic nuclei and terminate in functionally segregated striatal territories. The primary source of thalamostriatal projections is the caudal intralaminar nuclear group which, in primates, comprises the centromedian and parafascicular nuclei (CM/Pf). These two nuclei provide massive, functionally organized glutamatergic inputs to the whole striatal complex. There are several anatomical and physiological features that distinguish this system from other thalamostriatal projections. Although all glutamatergic thalamostriatal neurons express vGluT2 and release glutamate as neurotransmitter, CM/Pf neurons target preferentially the dendritic shafts of striatal projection neurons, whereas all other thalamic inputs are almost exclusively confined to the head of dendritic spines. This anatomic arrangement suggests that transmission of input from sources other than CM/Pf to the striatal neurons is likely regulated by dopaminergic afferents in the same manner as cortical inputs, while the CM/Pf axo-dendritic synapses do not display any particular relationships with dopaminergic terminals. A better understanding of the role of these systems in the functional circuitry of the basal ganglia relies on future research of the physiology and pathophysiology of these networks in normal and pathological basal ganglia conditions. Although much remains to be known about the role of these systems, recent electrophysiological studies from awake monkeys have provided convincing evidence that the CM/Pf-striatal system is the entrance for attention-related stimuli to the basal ganglia circuits. However, the processing and transmission of this information likely involves intrinsic GABAergic and cholinergic striatal networks, thereby setting the stage for complex physiological responses of striatal output neurons to CM/Pf activation. Finally, another exciting development that will surely generate significant interest towards the thalamostriatal systems in years to come is the possibility that CM/Pf may be a potential surgical target for movement disorders, most particularly Tourette syndrome and Parkinson's disease. Although the available clinical evidence is encouraging, these procedures remain empirical at this stage because of the limited understanding of the thalamostriatal systems.
Parkinson's disease; thalamus; striatum; glutamate transporter; synaptic plasticity
Striatal spine loss is a key pathological feature of human Parkinson’s disease (PD) that can be induced after complete degeneration of the nigrostriatal dopaminergic system in rodent models of parkinsonism. In line with these observations, our findings reveal a significant (30–50%) reduction in spine density in both the caudate nucleus and putamen of severely DA-depleted striata of MPTP-treated monkeys; the sensorimotor post-commissural putamen being the most severely affected region for both dopamine depletion and spine loss. Using MPTP-treated monkeys with complete or partial striatal dopamine (DA) denervation, we also demonstrate that striatal spine loss is an early pathological feature of parkinsonism, which progresses along a positive rostrocaudal and mediolateral gradient in parallel with the extent of striatal dopamine denervation. Quantitative electron microscopy immunocytochemistry for D1 dopamine receptor (D1) in the striatum of control and severely DA-depleted animals revealed that both D1-immunoreactive and immunonegative spines are lost in the putamen of MPTP-treated monkeys.
These data demonstrate that striatal spine loss in MPTP-treated monkeys is an early pathological event of parkinsonism, tightly correlated with the degree of nigrostriatal dopamine denervation that likely affects both direct and indirect striatofugal pathways.