The distribution of Cdh20 mRNA demarcates somatotopic subregions and subpopulations of spiny projection neurons in the rat dorsolateral striatum.

The dorsolateral striatum (DLS) of rodents is functionally subdivided into somatotopic subregions that represent each body part along both the dorsoventral and anteroposterior (A-P) axes and play crucial roles in sensorimotor functions via corticostriatal pathways. However, little is known about the spatial gene expression patterns and heterogeneity of spiny projection neurons (SPNs) within somatotopic subregions. Here, we show that the cell adhesion molecule gene Cdh20, which encodes a Type II cadherin, is expressed in discrete subregions covering the inner orofacial area and part of the forelimb area in the ventral domain of the DLS (v-DLS) in rats. Cdh20-expressing cells were localized in the v-DLS at the intermediate level of the striatum along the A-P axis and could be classified as direct-pathway SPNs or indirect-pathway SPNs. Unexpectedly, comprehensive analysis revealed that Cdh20 is expressed in SPNs in the rat DLS but not in the mouse DLS or the ferret putamen (Pu). Our observations reveal that Cdh20 expression demarcates somatotopic subregions and subpopulations of SPNs specifically in the rat DLS and suggest divergent regulation of genes differentially expressed in the v-DLS and Pu among mammals.

Structures of α-synuclein filaments from multiple system atrophy.

Synucleinopathies, which include multiple system atrophy (MSA), Parkinson's disease, Parkinson's disease with dementia and dementia with Lewy bodies (DLB), are human neurodegenerative diseases1. Existing treatments are at best symptomatic. These diseases are characterized by the presence of, and believed to be caused by the formation of, filamentous inclusions of α-synuclein in brain cells2,3. However, the structures of α-synuclein filaments from the human brain are unknown. Here, using cryo-electron microscopy, we show that α-synuclein inclusions from the brains of individuals with MSA are made of two types of filament, each of which consists of two different protofilaments. In each type of filament, non-proteinaceous molecules are present at the interface of the two protofilaments. Using two-dimensional class averaging, we show that α-synuclein filaments from the brains of individuals with MSA differ from those of individuals with DLB, which suggests that distinct conformers or strains characterize specific synucleinopathies. As is the case with tau assemblies4-9, the structures of α-synuclein filaments extracted from the brains of individuals with MSA differ from those formed in vitro using recombinant proteins, which has implications for understanding the mechanisms of aggregate propagation and neurodegeneration in the human brain. These findings have diagnostic and potential therapeutic relevance, especially because of the unmet clinical need to be able to image filamentous α-synuclein inclusions in the human brain.

Thalamic degeneration in MPTP-treated Parkinsonian monkeys: impact upon glutamatergic innervation of striatal cholinergic interneurons.

In both Parkinson's disease (PD) patients and MPTP-treated non-human primates, there is a profound neuronal degeneration of the intralaminar centromedian/parafascicular (CM/Pf) thalamic complex. Although this thalamic pathology has long been established in PD (and other neurodegenerative disorders), the impact of CM/Pf cell loss on the integrity of the thalamo-striatal glutamatergic system and its regulatory functions upon striatal neurons remain unknown. In the striatum, cholinergic interneurons (ChIs) are important constituents of the striatal microcircuitry and represent one of the main targets of CM/Pf-striatal projections. Using light and electron microscopy approaches, we have analyzed the potential impact of CM/Pf neuronal loss on the anatomy of the synaptic connections between thalamic terminals (vGluT2-positive) and ChIs neurons in the striatum of parkinsonian monkeys treated chronically with MPTP. The following conclusions can be drawn from our observations: (1) as reported in PD patients, and in our previous monkey study, CM/Pf neurons undergo profound degeneration in monkeys chronically treated with low doses of MPTP. (2) In the caudate (head and body) nucleus of parkinsonian monkeys, there is an increased density of ChIs. (3) Despite the robust loss of CM/Pf neurons, no significant change was found in the density of thalamostriatal (vGluT2-positive) terminals, and in the prevalence of vGluT2-positive terminals in contact with ChIs in parkinsonian monkeys. These findings provide new information about the state of thalamic innervation of the striatum in parkinsonian monkeys with CM/Pf degeneration, and bring up an additional level of intricacy to the consequences of thalamic pathology upon the functional microcircuitry of the thalamostriatal system in parkinsonism. Future studies are needed to assess the importance of CM/Pf neuronal loss, and its potential consequences on the neuroplastic changes induced in the synaptic organization of the thalamostriatal system, in the development of early cognitive impairments in PD.

Characterization of highly proliferative secondary tumor clusters along host blood vessels in malignant glioma.

The aim of the present study was to investigate the extensive invasion of tumor cells into normal brain tissue, a life­threatening feature of malignant gliomas. How invasive tumor cells migrate into normal brain tissue and form a secondary tumor structure remains to be elucidated. In the present study, the morphological and phenotypic changes of glioma cells during invasion in a C6 glioma model were investigated. C6 glioma cells were stereotactically injected into the right putamen region of adult Sprague­Dawley rats. The brain tissue sections were then subjected to hematoxylin and eosin, immunohistochemical or immunofluorescent staining. High magnification views of the tissue sections revealed that C6 cells formed tumor spheroids following implantation and marked invasion was observed shortly after spheroid formation. In the later stages of invasion, certain tumor cells invaded the perivascular space and formed small tumor clusters. These small tumor clusters exhibited certain common features, including tumor cell multilayers surrounding an arteriole, which occurred up to several millimeters away from the primary tumor mass; a high proliferation rate; and similar gene expression profiles to the primary tumor. In conclusion, the present study revealed that invading tumor cells are capable of forming highly proliferative cell clusters along arterioles near the tumor margin, which may be a possible cause of the recurrence of malignant glioma.

A novel, fully automated, observer-independent program for semiquantifying striatal 123I-FP-CIT uptake / Un nuevo programa, totalmente automático e independiente del observador para semi-cuantificar la captación estriatal de 123I-FP-CIT

Objective: To describe and validate a novel, fully automated program specifically designed for the semi- quantification of striatal 123I-FP-CIT uptake using volumes of interest (VOI) analysis. Material and methods: The proposed algorithm is based on a template that mimics the striatal 123 I-FP-CIT uptake in a healthy subjects, derived from defined anatomical VOIs available from WFU PickAtlas. Four SPECT studies of the anthropomorphic Alderson phantom filled with variable radioactive concentrations were acquired for the experimental validation. Experimental SPECT images were spatially normalized with respect to the previously created template. The binary VOIs corresponding to left caudate and puta- men and right caudate and putamen, which were used to construct the template, were projected onto the experimental images to obtain the counts for these regions. To minimize the partial volume effect, a percentage of the voxels in these regions (threshold), rather than all of them, was used. A binary occipital VOI was used to quantify the non-specific uptake. Experimental binding potentials (BPs) were calculated from the counts in these regions. True BPs were calculated from aliquots taken from the solutions used to fill the phantom. Results: There were statistically significant differences in the experimental BP values (p < 0.002) accord- ing to the percentage of voxels used. A highly significant correlation was achieved between true and experimental BP values, regardless of the percentage of voxels included for quantification. Conclusions: Our novel, observer-independent program automatically performs the semiquantification of striatal 123I-FP-CIT uptake in experimental studies (AU)

Objetivos: Describir y validar un nuevo software, totalmente automático, específicamente diseñado para semicuantificar la captación estriatal de 123I-FP-CIT usando volúmenes de interés (VOIs). Material y métodos: El algoritmo propuesto se basa en una plantilla que remeda la captación estriatal de 123I-FP-CIT en un sujeto sano, obtenida a partir de VOIs anatómicos definidos en WFU PickAtlas. Para la validación experimental de este algoritmo se adquirieron 4 estudios SPECT del maniquí antropomórfico Alderson llenado con concentraciones radioactivas variables. Las imágenes SPECT experimentales se normalizaron espacialmente respecto a la plantilla creada. Los VOIs binarios correspondientes a núcleo caudado y putámen derechos e izquierdos, utilizados para disen ̃ar la plantilla, se proyectaron sobre las imágenes experimentales para obtener las cuentas en estas regiones. Para minimizar los efectos de volumen parcial se utilizó un porcentaje de vóxeles, en vez de utilizar todos los vóxeles contenidos en estos VOIs. Se ha utilizado un VOI binario situado en región occipital para cuantificar la unión no específica. Los potenciales de unión (BPs) experimentales se calcularon a partir de las cuentas obtenidas en estas regiones. Los BPs reales se calcularon a partir de alícuotas tomadas de las soluciones utilizadas para llenar el maniquí. Resultados: Hubo diferencias estadísticamente significativas en los BPs experimentales en función del porcentaje de vóxeles utilizados para la cuantificación (p < 0.002). Se alcanzó una alta correlación entre los BPs reales y los experimentales, independientemente del porcentaje de vóxeles utilizados para la cuantificación. Conclusiones: Este nuevo programa automático e independiente del observador realiza la semicuantificación de la captación estriatal de 123I-FP-CIT en estudios experimentales (AU)

Regional differences in human ependymal and subventricular zone cytoarchitecture are unchanged in neuropsychiatric disease.

Much work has focused on the possible contribution of adult hippocampal neurogenesis to neuropsychiatric diseases. The hippocampal subgranular zone and the other stem cell-containing neurogenic niche, the subventricular zone (SVZ), share several cytological features and are regulated by some of the same molecular mechanisms. However, very little is known about the SVZ in neuropsychiatric disorders. This is important since it surrounds the lateral ventricles and in schizophrenia ventricular enlargement frequently follows forebrain nuclei shrinkage. Also, adult neurogenesis has been implicated in pharmacotherapy for affective disorders and many of the molecules associated with neuropsychiatric disorders affect SVZ biology. To assess the neurogenic niche, we examined material from 60 humans (Stanley Collection) and characterized the cytoarchitecture of the SVZ and ependymal layer in age-, sex- and post mortem interval-matched controls, and patients diagnosed with schizophrenia, bipolar illness, and depression (n = 15 each). There is a paucity of post mortem brains available for study in these diseases, so to maximize the number of possible parameters examined here, we quantified individual sections rather than a large series. Previous work showed that multiple sclerosis is associated with increased width of the hypocellular gap, a cell-sparse region that typifies the human SVZ. Statistically there were no differences between disease groups and controls in the width of the hypocellular gap or in the density of cells in the hypocellular gap. Because ventricular enlargement in schizophrenia may disrupt ependymal cells, we quantified them, but observed no difference between diagnostic groups and controls. There are significant differences in the prevalence of neuropsychiatric illness between the sexes. Therefore, we looked for male versus female differences, but did not observe any in the parameters quantified. We next turned to a finer spatial resolution and asked if there were differences amongst the disease groups in dorsal ventral subdivisions of the SVZ. Similar to when we treated the SVZ as a whole, we did not find such differences. However, compared to the dorsal SVZ, the ventral SVZ had a wider hypocellular gap and more ependymal cells in all four groups. In contrast, cell density was similar in dorsal ventral subregions of the SVZ hypocellular gap. These results show that though there are regional differences in the SVZ in humans, neuropsychiatric disorders do not seem to alter several fundamental histological features of this adult neurogenic zone.

Mitochondria in the striatum of subjects with schizophrenia.

OBJECTIVES: Schizophrenia is a severe mental illness that manifests pathology in many brain regions, including the striatum. Among the abnormalities in schizophrenia are those related to mitochondria. The present study sought to determine whether the number of mitochondria was affected at the level of the synapse. METHODS: Human postmortem striatum from schizophrenia subjects and controls was examined at the ultrastructural level. The density of mitochondria and synapses were tabulated using stereology. RESULTS: There were similar overall numbers of mitochondria in the caudate nucleus and putamen of schizophrenia subjects vs. controls, but a differential distribution of existing mitochondria. Schizophrenia subjects had 26?30% fewer mitochondria per synapse compared to controls. This may contribute to the pathophysiology of the illness, may be a medication effect, or an adaptive response to normalize the high number of striatal synapses we have previously found. The higher density of mitochondria in dendrites in the caudate nucleus in certain subgroups of schizophrenia vs. controls (>34%) may be related to more synaptic inputs. CONCLUSIONS: The role of mitochondria in the various symptoms of schizophrenia is still unclear. A comparison of schizophrenia subjects with differing symptoms or treatment response might shed light on whether differences in mitochondrial density are abnormal or adaptive.

Lrrk2 localization in the primate basal ganglia and thalamus: a light and electron microscopic analysis in monkeys.

The Leucine Rich Repeat Kinase-2 (LRRK2) gene is a common mutation target in Parkinson's disease (PD), but the cellular mechanisms by which such mutations underlie the pathophysiology of PD remain poorly understood. Thus, to better characterize the neuronal target sites of LRRK2 mutations in the primate brain, we studied the cellular and ultrastructural localization of Lrrk2 immunoreactivity in the monkey basal ganglia. As previously described, the monkey striatum was the most enriched basal ganglia structure in Lrrk2 labeling. Both projection neurons and parvalbumin-containing GABAergic interneurons displayed Lrrk2 immunoreactivity. At the electron microscopic level, striatal Lrrk2 labeling was associated predominantly with dendritic shafts and subsets of putative glutamatergic axon terminals. At the pallidal level, moderate cellular Lrrk2 immunostaining was found in the external globus pallidus (GPe), while neurons in the internal globus pallidus (GPi) were devoid of Lrrk2 immunoreactivity. Strong labeling was associated with cholinergic neurons in the nucleus basalis of Meynert. Midbrain dopaminergic neurons in the primate substantia nigra pars compacta (SNc) and ventral tegmental area harbored a significant level of Lrrk2 labeling, while neurons in the subthalamic nucleus were lightly immunostained. Most thalamic nuclei were enriched in Lrrk2 immunoreactivity, except for the centromedian nucleus that was completely devoid of labeling. Thus, Lrrk2 protein is widely distributed in the monkey basal ganglia, suggesting that gene mutations in PD may result in multifarious pathophysiological effects that could impact various target sites in the functional circuitry of the primate basal ganglia.

Dopamine D1 and N-methyl-D-aspartate receptors and extracellular signal-regulated kinase mediate neuronal morphological changes induced by repeated cocaine administration.

The development of drug addiction involves persistent cellular and molecular changes in the CNS. The brain dopamine and glutamate systems play key roles in mediating drug-induced neuroadaptation. Changes in dendritic morphology in medium spiny neurons (MSNs) in the nucleus accumbens (NAc) and caudate putamen (CPu) accompany drug-induced enduring behavioral and molecular changes. We have investigated the potential involvement of dopamine D1 and D2 receptors, the N-methyl-D-aspartate (NMDA) receptor, and the extracellular signal-regulated kinase (ERK) in dendritic morphological changes induced by repeated cocaine administration. We show that either a genetic mutation or pharmacological blockade of dopamine D1 receptors attenuated cocaine-induced changes in both dendritic branching and spine density of MSNs in the shell of the NAc and CPu. In contrast, antagonism of dopamine D2 receptors had no obvious effect on changes in dendritic branching but had a partial effect on changes in spine density of MSNs in these brain regions following repeated cocaine injections. Pharmacological inhibition of either NMDA receptors or ERK attenuated cocaine-induced changes in both dendritic branching and spine density of MSNs in the shell of the NAc and CPu. These results suggest that dopamine D1 and NMDA receptors and ERK contribute significantly to neuronal morphological changes induced by repeated exposure to cocaine.

Focal white matter changes in spasmodic dysphonia: a combined diffusion tensor imaging and neuropathological study.

Spasmodic dysphonia is a neurological disorder characterized by involuntary spasms in the laryngeal muscles during speech production. Although the clinical symptoms are well characterized, the pathophysiology of this voice disorder is unknown. We describe here, for the first time to our knowledge, disorder-specific brain abnormalities in these patients as determined by a combined approach of diffusion tensor imaging (DTI) and postmortem histopathology. We used DTI to identify brain changes and to target those brain regions for neuropathological examination. DTI showed right-sided decrease of fractional anisotropy in the genu of the internal capsule and bilateral increase of overall water diffusivity in the white matter along the corticobulbar/corticospinal tract in 20 spasmodic dysphonia patients compared to 20 healthy subjects. In addition, water diffusivity was bilaterally increased in the lentiform nucleus, ventral thalamus and cerebellar white and grey matter in the patients. These brain changes were substantiated with focal histopathological abnormalities presented as a loss of axonal density and myelin content in the right genu of the internal capsule and clusters of mineral depositions, containing calcium, phosphorus and iron, in the parenchyma and vessel walls of the posterior limb of the internal capsule, putamen, globus pallidus and cerebellum in the postmortem brain tissue from one patient compared to three controls. The specificity of these brain abnormalities is confirmed by their localization, limited only to the corticobulbar/corticospinal tract and its main input/output structures. We also found positive correlation between the diffusivity changes and clinical symptoms of spasmodic dysphonia (r = 0.509, P = 0.037). These brain abnormalities may alter the central control of voluntary voice production and, therefore, may underlie the pathophysiology of this disorder.

Synaptic differences in the patch matrix compartments of subjects with schizophrenia: a postmortem ultrastructural study of the striatum.

The striatum processes motor, cognitive, and limbic circuitry. Striatal patch and matrix compartments are organized differently in many aspects including connectivity. Abnormalities in either compartment could have different functional consequences. The present study compares the synaptic organization in the patches and matrix in subjects with schizophrenia (SZ, n = 14) versus normal controls (NC, n = 8). Postmortem striatal tissue was processed for calbindin immunocytochemistry to identify the patch versus matrix compartments, prepared for electron microscopy, and analyzed using stereology. Several synaptic changes were observed in the SZ subjects vs. NCs including a higher density of cortical-type synapses in the putamen patch (44% higher) and in the caudate matrix (36% higher) in SZ cases on typical antipsychotic drugs. These changes appeared to be normalized rather than caused by treatment. The abnormal connectivity may represent a failure of normal synaptic pruning and may play a role in limbic or cognitive dysfunction in schizophrenia.

Activity-regulated cytoskeleton-associated protein Arc is targeted to dendrites and coexpressed with mu-opioid receptors in postnatal rat caudate-putamen nucleus.

Dendritic expression of the activity-regulated cytoskeleton-associated protein (Arc) is dramatically enhanced by increased synaptic activity in adult brain. We used immunocytochemical electron microscopy to determine whether the subcellular localization of Arc in developing dendrites corresponds to the peak period of synaptogenesis in the postnatal rat caudate-putamen nucleus (CPN). The distribution was compared with that of mu-opioid receptors (MORs), whose localization in dendritic spines closely parallels excitatory synapse formation during postnatal development (Wang et al. [2003] Neuroscience 118:695-708). Sections were processed for immunocytochemical detection of antisera against Arc or MORs at the beginning (postnatal day 15; P15) and the end (P30) of the peak period of synaptogenesis in rat CPN. At P15, immunolabeling for Arc showed a punctate distribution in the cytoplasm of dendritic shafts, some of which was associated with polyribosomes. In some spiny dendrites, Arc immunoreactivity was more intensely localized in putative spines than in their parental dendrites, whereas, in other spiny dendrites, Arc labeling was restricted in the shafts. Many dendritic shafts and spines also showed immunoreactivity for MORs, although dually labeled spines were less numerous than the shafts. At P30, the proportion of singly and dually labeled spines significantly increased from 2.0% to 7.5% and from 9.5% to 21%, respectively. Arc labeling in spines was more detectable beneath the postsynaptic density or at extrasynaptic sites on the plasma membrane. Our results suggest a correlation between Arc expression in dendritic spines during postnatal development and the onset of synaptogenesis in opioid-responsive neurons in the rat CPN.

Ultrastructural localization of delta-opioid receptors in the rat caudate-putamen nucleus during postnatal development: relation to synaptogenesis.

During development, delta-opioid receptors (DORs) in the rat caudate-putamen nucleus (CPN) appear later than mu-opioid receptors (MORs), whose developmental pattern specifically relates to synaptogenesis. We used electron microscopic immunocytochemistry to determine whether there are also age-related changes in subcellular localization of DORs in the rat CPN. Sections from postnatal day (P) 0-P30 and adult dorsomedial CPN were immunogold-silver labeled to examine the plasmalemmal and cytoplasmic distribution of these receptors. In addition, immunoperoxidase labeling was used to determine the numerical density of synapses relative to DOR-labeled profiles. Immunolabeling for DOR was undetectable at P0, light at P5, and dense from P10 onward. The labeling during P5-P10 was mainly localized in somatodendritic profiles but also was readily seen in axon terminals, most of which formed asymmetric synapses with dendrites. From P15, a few immunogold particles were seen in contact with postsynaptic densities in spines, and the proportion of these particles significantly increased in P30 and adult CPN. Other particles were localized in the cytoplasm of dendrites and terminals without significant age-related changes. Stereological analysis showed that compared with labeled dendritic shafts and spines, labeled axon terminals have a closer correlation with synapse formation. These results are in marked contrast with MORs, which show an age-related increase in association with dendritic plasma membrane and a good correlation in the developmental pattern of MOR-labeled spines with synapse formation (Wang et al. [2003] Neuroscience 118:695-708). Together, our results suggest receptor-type specific roles for endogenous opioids acting at both pre- and postsynaptic sides in the developing CPN.

Postnatal development of mu-opioid receptors in the rat caudate-putamen nucleus parallels asymmetric synapse formation.

The mu-opioid receptor (MOR) in the caudate-putamen nucleus (CPN) appears early during prenatal development, and shows a patch-like distribution throughout the postnatal period and adulthood. In the adult rat CPN, neurons in patch compartments receive glutamatergic excitatory input mainly from the cortex through synapses onto spines, many of which express MORs. Thus, MOR expression in spines may be related to corticostriatal synaptogenesis. We used electron microscopic immunocytochemistry to determine potential age-dependent changes in the distribution pattern of MOR during postnatal synaptogenesis in the rat CPN. Immunogold-silver labeling revealed that the dendritic plasmalemmal density of MOR at postnatal day (P) 0 was significantly lower than, but after P10 was similar to, that of adult. In contrast, such age-dependent changes were not observed in axon terminals. Stereological analysis of immunoperoxidase labeling for MOR showed a good correlation in the developmental numerical densities of synapses with MOR-labeled spines and those of total asymmetric axospinous synapses, linear correlation coefficient r=0.99. Synapses with MOR-labeled dendrites, however, had a low correlation with axodendritic synapses (r=0.61), and synapses with MOR-labeled terminals showed no correlation with axospinous and axodendritic synapses (r=0.19). These results provide ultrastructural evidence that the targeting of MOR on the plasma membrane of dendrites and spines parallels the peak period of synaptogenesis during the third postnatal week in the rat CPN. Thus, the postnatal spatiotemporal expression pattern of MOR appears to match the functional maturation of corticostriatal glutamate transmission.

The ultrastructural organization of the patch matrix compartments in the human striatum.

The mammalian striatum is a heterogeneous structure characterized by striosomes and matrix. The synaptic organization of the striatum has been described previously in various mammalian species including human; however, potential ultrastructural differences in striosomal organization have not been well studied. Samples (n = 7) of striatal tissue were obtained from the Maryland Brain Collection (mean age, 37.7 +/- 9.4 years; and mean PMI, 5.3 +/- 1.4 hours). Tissue was prepared for calbindin immunocytochemistry to identify striosomal (patch) and extrastriosomal matrix (matrix) compartments and subsequently prepared for electron microscopy. Synaptic density was determined, using stereologic methods, for all synapses combined and for various subsets of synapses such as asymmetric, symmetric, axospinous, axodendritic, and perforated in the patch and matrix of the caudate (CP, CM) and putamen (PP, PM). An ANOVA revealed significant between-group (CP, CM, PP, PM) differences (P < 0.05) for the following types of synapses: total combined, asymmetric, axospinous, and asymmetric axospinous. Each of these four types was significantly increased in density in the CP vs the PP, whereas the matrix (CM vs PM) showed no significant differences in density in these or other synapses. In the caudate (CP vs CM), the synaptic density of the types of synapses studied did not vary significantly between the patch and the matrix. In the putamen, the matrix (PM) had higher synaptic densities than that of the patches (PP) for total synapses, symmetric dendritic, and perforated. These data show that the patch and matrix compartments are heterogeneous at the ultrastructural level, imparting another level of complexity to the striatum-a fact that should be taken into consideration when studying diseases of this brain region at the electron microscopic level.

Nigral and pallidal inputs to functionally segregated thalamostriatal neurons in the centromedian/parafascicular intralaminar nuclear complex in monkey.

In primates, thalamostriatal projections from the centromedian (CM) and parafascicular (Pf) nuclei are strong and organized according to a strict pattern of functional connectivity with various regions of the striatal complex. In turn, the CM/Pf complex receives a substantial innervation from the internal globus pallidus (GPi). In this study, we demonstrate that the substantia nigra pars reticulata (SNr) also provides a massive input to Pf in monkeys. These pallidothalamic and nigrothalamic projections provide routes whereby information can flow in functional loops between the basal ganglia and the intralaminar nuclear group. To understand better the anatomical organization and the degree of functional specificity of these loops, we combined retrograde and anterograde labeling methods from functionally defined regions of the striatum and GPi/SNr to determine the relationships between thalamostriatal neurons and basal ganglia afferents. Together with previous studies, our data suggest the existence of tightly connected functional circuits between the basal ganglia and the CM/Pf in primates: 1) A "sensorimotor" circuit links together the medial two-thirds of CM, the postcommissural putamen, and the ventrolateral part of the caudal GPi; 2) a "limbic" circuit involves the rostral one-third of Pf, the ventral striatum, and the rostromedial pole of GPi; and 3) an "associative"circuit exists between the caudal two-thirds of Pf, the caudate nucleus, and the SNr. An additional "associative" circuit that involves the caudate-receiving territory of GPi (dorsal one-third), the dorsolateral Pf (Pfdl), and the precommissural putamen was also disclosed. In conclusion, findings of this study provide additional evidence for the high degree of functional specificity of the thalamostriatal system through which CM/Pf may provide attention-specific sensory information important for conditional responses to the primate striatum.

Impaired preprodynorphin, but not preproenkephalin, mRNA induction in the striatum of mGluR1 mutant mice in response to acute administration of the full dopamine D(1) agonist SKF-82958.

Metabotropic glutamate receptor 1 (mGluR1) is highly expressed in striatonigral projection neurons of rat striatum. To define the role of mGluR1 in the regulation of striatal gene expression, the responsiveness of the three neuropeptide gene expression to a single injection of the dopamine D(1) agonist SKF-82958 was compared between mGluR1 mutant and wild-type control mice. We found that acute injection of SKF-82958 increased preprodynorphin (PPD), substance P (SP), and preproenkephalin (PPE) mRNAs in the dorsal and ventral striatum of mutant and wild-type mice in a dose-dependent manner (0.125, 0.5, and 2 mg/kg, i.p.) as revealed by quantitative in situ hybridization. However, the induction of PPD mRNA in both the dorsal and ventral striatum of mGluR1 minus sign/minus sign mice was significantly less than that of wild-type +/+ mice in response to the two higher doses of SKF-82958. In contrast to PPD, SP and PPE in the dorsal and ventral striatum of mGluR1 mutant mice were elevated to a similar level as that of wild-type mice. There were no differences in basal levels and distribution patterns of all three mRNAs between the two genotypes of mice treated with saline. These results indicate that mGluR1 selectively participates in striatonigral PPD induction in response to D(1) receptor stimulation.

Dopamine D2 receptors are present in prefrontal cortical afferents and their targets in patches of the rat caudate-putamen nucleus.

Glutamatergic neurons within the deep layers of the prefrontal cortex and dopaminergic neurons of the substantia nigra pars compacta preferentially terminate in patch-like regions within the caudate putamen nucleus (CPN). Activation of dopamine D2 receptors is known to potently modulate striatal glutamatergic transmission and may play a role in reward-based motor learning. To determine the cellular substrate for D2-mediated regulation of prefrontal corticostriatal transmission in striatal patches, we combined anterograde transport of biotinylated dextran amine (BDA) with immunogold-silver labeling of a D2 receptor antipeptide antiserum in rat brain. Injections centered in deep layers of the dorsal part of the anterior cingulate cortex, one of the prefrontal cortical regions, produced varicose axonal BDA labeling in a patch-like distribution in the dorsomedial CPN. Electron microscopy showed that in these patch compartments, BDA labeling was present exclusively in axons and terminals (total number = 581), 9% of which contained detectable D2-like immunoreactivity. Thirty percent of the BDA-labeled terminals formed asymmetric excitatory synapses with dendritic spine heads, and the remainder were without recognizable junctions. The recipient spines were unlabeled or contained immunogold-silver particles for D2 receptors. A few of the D2-labeled spines also received convergent, often nonsynaptic contact from D2-labeled terminals resembling dopaminergic afferents. In addition, the corticostriatal terminals often apposed spiny and nonspiny neuronal profiles that contained D2 labeling. These results suggest that dopamine D2 receptors are strategically positioned for presynaptic and postsynaptic modulation of prefrontal corticostriatal excitation of spiny neurons in striatal patches. The findings have direct implications for D2-mediated control of reward-related motor learning.

Ultrastructural localization of the CB1 cannabinoid receptor in mu-opioid receptor patches of the rat Caudate putamen nucleus.

Cannabinoids and opioids are widely consumed drugs of abuse that produce motor depression, in part via respective activation of the cannabinoid subtype 1 receptor (CB1R) and the mu-opioid receptor (muOR), in the striatal circuitry originating in the caudate putamen nucleus (CPN). Thus, the CB1R and muOR may show similar targeting in the CPN. To test this hypothesis, we examined the electron microscopic immunocytochemical labeling of CB1R and muOR in CPN patches of rat brain. Of the CB1R-labeled profiles, 34% (588) were dendrites, presumably arising from spiny as well as aspiny-type somata, which also contained CB1R immunoreactivity. In dendrites, CB1R often was localized to nonsynaptic and synaptic plasma membranes, particularly near asymmetric excitatory-type junctions. Almost one-half of the CB1R-labeled dendrites contained muOR immunoreactivity, whereas only 20% of all muOR-labeled dendrites expressed CB1R. Axons and axon terminals as well as abundant glial processes also showed plasmalemmal CB1R and were mainly without muOR immunoreactivity. Many CB1R-labeled axon terminals were small and without recognizable synaptic junctions, but a few also formed asymmetric, or more rarely symmetric, synapses. The CB1R-labeled glial processes were often perivascular or perisynaptic, surrounding asymmetric excitatory-type axospinous synapses. Our results show that in CPN patches CB1R and muOR are targeted strategically to some of the same postsynaptic neurons, which may account for certain similarities in motor function. Furthermore, they also provide evidence that CB1R may play a major role in the modulation of presynaptic transmitter release and glial functions that are unaffected in large part by opioids active at muOR in CPN.

Ultrastructural localization of nitrotyrosine within the caudate-putamen nucleus and the globus pallidus of normal rat brain.

Nitration of protein tyrosine residues by nitric oxide (NO)-derived reactive species results in the production of stable nitrotyrosine (NT) moieties that are immunochemically detectable in many regions of normal brain and enriched in those areas containing constitutive nitric oxide synthase (cNOS). These include the caudate-putamen nucleus (CPN) and the globus pallidus, which receives major inhibitory input from the CPN. To determine the functional sites for NT production in these critical motor nuclei, we examined the electron microscopic immunocytochemical localization of NT and cNOS in rat brain. In the CPN, NT was localized to the somata and dendrites of cNOS-containing interneurons and spiny neurons, some of which received input from cNOS-labeled terminals. The NT immunoreactivity was most prevalent on outer mitochondrial membranes and nearby segments of the plasma membranes in dendrites and within asymmetric synapses on dendritic spines. In the CPN and globus pallidus, there was also a prominent labeling of NT in astrocytic processes, small axons, and tubulovesicles and/or synaptic vesicles in axon terminals. These terminals formed mainly asymmetric synapses in the CPN and inhibitory-type synapses in the globus pallidus where they often apposed cNOS-containing terminals that also formed asymmetric, excitatory-type synapses. Our results suggest that NT is generated by mechanisms requiring the dual actions of excitatory transmitters and NO derived either from interneurons in the CPN or from excitatory afferents in the globus pallidus. The findings also implicate NT in the physiological actions of NO within the striatal circuitry and, particularly, in striatopallidal neurons severely affected in Huntington's disease.