Role of aqueous extract of saffron in ameliorating effect of sofosbuvir on the cerebellar cortex in rat.

Sofosbuvir is a promising antiviral drug against chronic hepatitis C virus. Although it is characterized by its high efficacy, its adverse effects on nervous tissue are still unclear. Saffron is known for its neuroprotective property. This is a biochemical, histological and immunohistochemical study of the effect of sofosbuvir on the cerebellar cortex of rat and the possible ameliorating role of saffron's aqueous extract. Twenty-four adult male Wistar albino rats were equally divided into four groups; control, saffron extract-treated, sofosbuvir-treated (41.1 mg/kg/day for 6 weeks) and group concomitantly treated with saffron extract and sofosbuvir. Sofosbuvir-treated group recorded a significant increase in cerebellar malondialdehyde level coupling with a significant decrease in tissue glutathione and superoxide dismutase. Light microscopy revealed reduced number of Purkinje cells. The granular layer depicted many granular cells and Bergmann astrocytes with nuclear and cytoplasmic alterations. Electron microscopy revealed disorganized molecular layer with disarranged myelinated axons and disrupted mitochondria. Few shrunken Purkinje cells showed electron-dense cytoplasm and rarefied nuclei, indistinct nuclear envelope and dilated perinuclear space, areas of vacuolated cytoplasm, fragmented rough endoplasmic reticulum and few dark mitochondria. Some axons with tiny mitochondria were detected. A significant upregulation in immunohistochemical expression of GFAP-positive astrocytes was recorded. Concomitant administration of saffron extract significantly improved all studied parameters. Saffron extract is beneficial in ameliorating sofosbuvir-induced cerebellar morphological changes mainly through its antioxidant and neuroprotective properties.

3D RNA-seq: a powerful and flexible tool for rapid and accurate differential expression and alternative splicing analysis of RNA-seq data for biologists.

RNA-sequencing (RNA-seq) analysis of gene expression and alternative splicing should be routine and robust but is often a bottleneck for biologists because of different and complex analysis programs and reliance on specialized bioinformatics skills. We have developed the '3D RNA-seq' App, an R shiny App and web-based pipeline for the comprehensive analysis of RNA-seq data from any organism. It represents an easy-to-use, flexible and powerful tool for analysis of both gene and transcript-level gene expression to identify differential gene/transcript expression, differential alternative splicing and differential transcript usage (3D) as well as isoform switching from RNA-seq data. 3D RNA-seq integrates state-of-the-art differential expression analysis tools and adopts best practice for RNA-seq analysis. The program is designed to be run by biologists with minimal bioinformatics experience (or by bioinformaticians) allowing lab scientists to analyse their RNA-seq data. It achieves this by operating through a user-friendly graphical interface which automates the data flow through the programs in the pipeline. The comprehensive analysis performed by 3D RNA-seq is extremely rapid and accurate, can handle complex experimental designs, allows user setting of statistical parameters, visualizes the results through graphics and tables, and generates publication quality figures such as heat-maps, expression profiles and GO enrichment plots. The utility of 3D RNA-seq is illustrated by analysis of data from a time-series of cold-treated Arabidopsis plants and from dexamethasone-treated male and female mouse cortex and hypothalamus data identifying dexamethasone-induced sex- and brain region-specific differential gene expression and alternative splicing.

The possible neuroprotective role of grape seed extract on the histopathological changes of the cerebellar cortex of rats prenatally exposed to Valproic Acid: animal model of autism.

Autism Spectrum Disorder (ASD) is a heterogeneous neurodevelopmental disease characterized by defect in verbal and nonverbal communications. As, the cerebellum has the greatest number of neurons and synapses in the central nervous system so, the cerebellum has emerged as one of the target brain areas affected in autism. The aim of this work was to study the biochemical, immunohistochemical and ultrastructural characteristics of autism and the possible neuroprotective role of grape seed extract. In this study 28 male pups were divided into Control groups; Group I (saline), Group II (GSE 400 mg/kg), Group III (VPA 500 mg/kg) and Group IV (VPA and GSE). Cerebellar hemispheres were dissected out and prepared to determine the oxidative stress markers, histological, immunohistochemical and morphometric study were done. A significant elevation in oxidative stress markers in off spring of VPA treated rats in comparison to control group was detected. A significant decrease in the Purkinje cell count and nuclear size were observed. Numerous shrunken cells with hyperchromatic nuclei and ultrastructural degeneration of cytoplasmic organelles were detected. A significant rise in the area percentage of GFAP-positive immune stained cells in comparison to that of the control groups was seen. Strikingly, GSE revealed significant improvement in the oxidative stress markers and then the histological and morphometric picture of the cerebellum. GSE has neuroprotective effect on the cerebellum of VPA treated rats through its potent antioxidant effect.

Conversion of Graded Presynaptic Climbing Fiber Activity into Graded Postsynaptic Ca2+ Signals by Purkinje Cell Dendrites.

The brain must make sense of external stimuli to generate relevant behavior. We used a combination of in vivo approaches to investigate how the cerebellum processes sensory-related information. We found that the inferior olive encodes contexts of sensory-associated external cues in a graded manner, apparent in the presynaptic activity of their axonal projections (climbing fibers) in the cerebellar cortex. Individual climbing fibers were broadly responsive to different sensory modalities but relayed sensory-related information to the cortex in a lobule-dependent manner. Purkinje cell dendrites faithfully transformed this climbing fiber activity into dendrite-wide Ca2+ signals without a direct contribution from the mossy fiber pathway. These results demonstrate that the size of climbing-fiber-evoked Ca2+ signals in Purkinje cell dendrites is largely determined by the firing level of climbing fibers. This coding scheme emphasizes the overwhelming role of the inferior olive in generating salient signals useful for instructing plasticity and learning.

Remarkable increases of α1-antichymotrypsin in brain tissues of rodents during prion infection.

α1-Antichymotrypsin (α1-ACT) belongs to a kind of acute-phase inflammatory protein. Recently, such protein has been proved exist in the amyloid deposits which is the hallmark of Alzheimer's disease, but limitedly reported in prion disease. To estimate the change of α1-ACT during prion infection, the levels of α1-ACT in the brain tissues of scrapie agents 263K-, 139A- and ME7-infected rodents were analyzed, respectively. Results shown that α1-ACT levels were significantly increased in the brain tissues of the three kinds of scrapie-infected rodents, displaying a time-dependent manner during prion infection. Immunohistochemistry assays revealed the increased α1-ACT mainly accumulated in some cerebral regions of rodents infected with prion, such as cortex, thalamus and cerebellum. Immunofluorescent assays illustrated ubiquitously localization of α1-ACT with GFAP positive astrocytes, Iba1-positive microglia and NeuN-positive neurons. Moreover, double-stained immunofluorescent assays and immunohistochemistry assays using series of brain slices demonstrated close morphological colocalization of α1-ACT signals with that of PrP and PrPSc in the brain slices of 263K-infected hamster. However, co-immunoprecipitation does not identify any detectable molecular interaction between the endogenous α1-ACT and PrP either in the brain homogenates of 263K-infected hamsters or in the lysates of prion-infected cultured cells. Our data here imply that brain α1-ACT is increased abnormally in various scrapie-infected rodent models. Direct molecular interaction between α1-ACT and PrP seems not to be essential for the morphological colocalization of those two proteins in the brain tissues of prion infection.

Perinatal thiamine restriction affects central GABA and glutamate concentrations and motor behavior of adult rat offspring.

The purposes of the present study were to investigate the effects of perinatal thiamine deficiency, from the 11th day of gestation until the 5th day of lactation, on motor behavior and neurochemical parameters in adult rat offspring, using 3-month-old, adult, male Wistar rats. All rats were submitted to motor tests, using the rotarod and paw print tasks. After behavioral tests, their thalamus, cerebellum and spinal cord were dissected for glutamate and GABA quantifications by high performance liquid chromatography. The thiamine-restricted mothers (RM) group showed a significant reduction of time spent on the rotarod at 25 rpm and an increase in hind-base width. A significant decrease of glutamate concentration in the cerebellum and an increase of GABA concentrations in the thalamus were also observed. For the offspring from control mothers (CM) group there were significant correlations between thalamic GABA concentrations and both rotarod performance and average hind-base width. In addition, for rats from the RM group a significant correlation between stride length and cerebellar GABA concentration was found. These results show that the deficiency of thiamine during an early developmental period affects certain motor behavior parameters and GABA and glutamate levels in specific brain areas. Hence, a thiamine deficiency episode during an early developmental period can induce motor impairments and excitatory and inhibitory neurotransmitter changes that are persistent and detectable in later periods of life.

Effect of Qingxinkaiqiao compound on cortical mRNA expression of the apoptosis-related genes Bcl-2, BAX, caspase-3, and Aß in an Alzheimer's disease rat model.

OBJECTIVE: To investigate the effects of Qingxinkaiqiao (QK) compound in a rat model of Alzheimer's disease induced with ß-amyloid (Aß) 1-40. METHODS: Fifty-six three months, male, Sprague-Dawley rats were randomly divided into seven groups: blank control group, surgery group, model group, low-dose QK group, middle-dose QK group, high-dose QK group, and Aricept (donepezil hydrochloride) group, with eight rats in each group. Apart from the control and surgery groups, an Alzheimer's disease model was established in all groups by bilateral hippocampal injection of Aß 1-40. The surgery group received an injection of the same volume of physiological saline. Two days after model establishment, rats from the drug groups were administered the corresponding drugs; the control group and model group were administered an equal volume of physiological saline for 14 days. After treatment, real-time quantitative polymerase chain reaction, immunohistochemistry, and western blot assay were employed to confirm mRNA and protein expressions of Bcl-2, Bax, caspase-3, and Aß, respectively. CONCLUSION: QK treatment resulted in significantly up-regulated Bcl-2 expression, down-regulated Bax, caspase-3, and Aß expression, and reduced numbers of apoptotic cells in the cortex.

A bird's eye view of anisatin induced convulsive seizures in brain by a (1)H NMR based metabolic approach.

Anisatin is the main convulsant component in plants of the genus Illicium, many of which are important spices or folk medicines. The neurotoxicity of anisatin has been widely investigated, mainly focusing on its action on the γ-amino butyrate (GABA) system; however, little is known about the metabolic alterations that it causes. In this study, a NMR-based metabolomic approach was performed on the extracts of cortexes and cerebellums of mice administered with anisatin to explore the metabolic events associated with its intoxication. Orthogonal partial least squares-discriminant analysis (OPLS-DA) revealed many differential metabolites that indicated metabolic disturbance in neurotransmission and neuromodulation (GABA, glutamate, glutamine, and taurine), stress of reactive oxygen species (ROS) (ascorbate, phosphatidylcholine, choline, and ethanolamine), energy metabolism (NAD(+)i.e., nicotinamide-adenine dinucleotide, lactate, citrate, fumarate, creatine/phosphocreatine, and creatinine), amino acid metabolism (leucine, isoleucine, valine, phenylalanine, tyrosine, tryptophan, alanine, threonine, and glycine) and nucleic acid metabolism (NAD(+), nicotinamide/niacinamide, adenosine, and guanosine). This pilot metabolomic study on anisatin intoxication should help to develop a holistic view of convulsive seizures induced by anisatin, and provide a better understanding of the mechanisms.

Brainstem neurons survive the identical ischemic stress that kills higher neurons: insight to the persistent vegetative state.

Global ischemia caused by heart attack, pulmonary failure, near-drowning or traumatic brain injury often damages the higher brain but not the brainstem, leading to a 'persistent vegetative state' where the patient is awake but not aware. Approximately 30,000 U.S. patients are held captive in this condition but not a single research study has addressed how the lower brain is preferentially protected in these people. In the higher brain, ischemia elicits a profound anoxic depolarization (AD) causing neuronal dysfunction and vasoconstriction within minutes. Might brainstem nuclei generate less damaging AD and so be more resilient? Here we compared resistance to acute injury induced from simulated ischemia by 'higher' hippocampal and striatal neurons versus brainstem neurons in live slices from rat and mouse. Light transmittance (LT) imaging in response to 10 minutes of oxygen/glucose deprivation (OGD) revealed immediate and acutely damaging AD propagating through gray matter of neocortex, hippocampus, striatum, thalamus and cerebellar cortex. In adjacent brainstem nuclei, OGD-evoked AD caused little tissue injury. Whole-cell patch recordings from hippocampal and striatal neurons under OGD revealed sudden membrane potential loss that did not recover. In contrast brainstem neurons from locus ceruleus and mesencephalic nucleus as well as from sensory and motor nuclei only slowly depolarized and then repolarized post-OGD. Two-photon microscopy confirmed non-recoverable swelling and dendritic beading of hippocampal neurons during OGD, while mesencephalic neurons in midbrain appeared uninjured. All of the above responses were mimicked by bath exposure to 100 µM ouabain which inhibits the Na+/K+ pump or to 1-10 nM palytoxin which converts the pump into an open cationic channel. Therefore during ischemia the Na+/K+ pump of higher neurons fails quickly and extensively compared to naturally resilient hypothalamic and brainstem neurons. The selective survival of lower brain regions that maintain vital functions will support the persistent vegetative state.

Ameliorative effect of Pimpinella anisum oil on immunohistochemical and ultrastuctural changes of cerebellum of albino rats induced by aspartame.

The study aims to investigate the protective effect of Pimpinella anisum oil on aspartame (ASP) which resulted in cerebellar changes. The rats were divided into four equal groups: Group 1: (control group): served as control animals. Group 2: control P. anisum oil received .5 mL/kg/d/b wt. once daily. Group 3 (ASP group): received daily 250 mg/kg/b wt. of ASP dissolved in distilled water and given orally to the animals by intra-gastric tube for 2 months. Group 4: received .5 mL/kg/b wt. of prophylactic P. anisum oil once daily, followed by ASP after 2 h for 2 months. The histopathological approach revealed marked changes in the Purkinje cells, myleinated nerve fibers and granular cells of ASP-treated animals. Some of these cells appeared with deeply stained cytoplasm. Ultrastructural examination showed Purkinje cells with dilated rough endoplasmic reticulum and condensed mitochondria. Granular cells appeared with less c nuclei and surrounded by dissolution of most Mossy rosettes structures. Most myelinated nerve fibers showed thickening of myelinated sheath and others showed splitting of their myelin sheath. The histopathological, immunohistochemical and ultrastructural alterations were much less observed in concomitant use of P. anisum oil with ASP. Cerebellar cortex is considered target areas of ASP neurotoxicity, while P. anisum oil, when used in combination with ASP displays a protective action against neurotoxicity.

On the origin of tremor in Parkinson's disease.

The exact origin of tremor in Parkinson's disease remains unknown. We explain why the existing data converge on the basal ganglia-thalamo-cortical loop as a tremor generator and consider a conductance-based model of subthalamo-pallidal circuits embedded into a simplified representation of the basal ganglia-thalamo-cortical circuit to investigate the dynamics of this loop. We show how variation of the strength of dopamine-modulated connections in the basal ganglia-thalamo-cortical loop (representing the decreasing dopamine level in Parkinson's disease) leads to the occurrence of tremor-like burst firing. These tremor-like oscillations are suppressed when the connections are modulated back to represent a higher dopamine level (as it would be the case in dopaminergic therapy), as well as when the basal ganglia-thalamo-cortical loop is broken (as would be the case for ablative anti-parkinsonian surgeries). Thus, the proposed model provides an explanation for the basal ganglia-thalamo-cortical loop mechanism of tremor generation. The strengthening of the loop leads to tremor oscillations, while the weakening or disconnection of the loop suppresses them. The loop origin of parkinsonian tremor also suggests that new tremor-suppression therapies may have anatomical targets in different cortical and subcortical areas as long as they are within the basal ganglia-thalamo-cortical loop.

Heterogeneity of parvalbumin expression in the avian cerebellar cortex and comparisons with zebrin II.

The cerebellar cortex has a fundamental parasagittal organization that is reflected in the physiological responses of Purkinje cells, afferent and efferent connections, and the expression of several molecular markers. The most thoroughly studied of these molecular markers is zebrin II (ZII; a.k.a. aldolase C). ZII is differentially expressed in Purkinje cells, resulting in a pattern of sagittal stripes of high expression interdigitated with stripes of little or no expression. In this study, we examined the expression of the calcium binding protein parvalbumin (PV) in the cerebellum of several avian species (pigeons, hummingbirds, zebra finches) and compared it to the expression of ZII. We found that PV immunoreactivity was distributed across the cerebellar cortex such that there were sagittal stripes of PV immunopositive (PV+) Purkinje cells alternating with PV immunonegative (PV-) Purkinje cells. Although most Purkinje cells in the anterior lobe were PV+, there were several thin (i.e. only a few Purkinje cells wide) PV- stripes spanning the folia. In the posterior lobe, PV+ and PV- stripes were also apparent, but the PV- stripes were much wider than in the anterior lobe. In sections processed for both ZII and PV, the expression was generally complementary: PV+ stripes were ZII-, and vice-versa. This complementary expression was most apparent in folia II-IV and VIII-IXcd. The complementary expression was not, however, absolute; some Purkinje cells co-expressed PV and ZII whereas others lacked both. These novel findings relate to the complex neurochemical organization of the cerebellum, and are likely important to issues regarding cerebellar plasticity.

Quantitative organization of GABAergic synapses in the molecular layer of the mouse cerebellar cortex.

In the cerebellar cortex, interneurons of the molecular layer (stellate and basket cells) provide GABAergic input to Purkinje cells, as well as to each other and possibly to other interneurons. GABAergic inhibition in the molecular layer has mainly been investigated at the interneuron to Purkinje cell synapse. In this study, we used complementary subtractive strategies to quantitatively assess the ratio of GABAergic synapses on Purkinje cell dendrites versus those on interneurons. We generated a mouse model in which the GABAA receptor alpha1 subunit (GABAARalpha1) was selectively removed from Purkinje cells using the Cre/loxP system. Deletion of the alpha1 subunit resulted in a complete loss of GABAAR aggregates from Purkinje cells, allowing us to determine the density of GABAAR clusters in interneurons. In a complementary approach, we determined the density of GABA synapses impinging on Purkinje cells using alpha-dystroglycan as a specific marker of inhibitory postsynaptic sites. Combining these inverse approaches, we found that synapses received by interneurons represent approximately 40% of all GABAergic synapses in the molecular layer. Notably, this proportion was stable during postnatal development, indicating synchronized synaptogenesis. Based on the pure quantity of GABAergic synapses onto interneurons, we propose that mutual inhibition must play an important, yet largely neglected, computational role in the cerebellar cortex.

Distribution of the SNAP25 and SNAP23 synaptosomal-associated protein isoforms in rat cerebellar cortex.

Synaptosome-associated protein of 25 kDa (SNAP25) is a component of the fusion complex that mediates synaptic vesicle exocytosis, regulates calcium dynamics and neuronal plasticity. Despite its crucial role in vesicle release, SNAP25 is not distributed homogenously within the brain. It seems to be virtually absent in mature inhibitory terminals and is observed in a subtype of excitatory neurons defined by the expression of vesicular glutamate transporter 1 (VGluT1). Since a complementary distribution of VGluT1 and VGluT2 in excitatory synapses is correlated with different probabilities of release (Pr), we evaluated whether SNAP25 localization is associated with specific synaptic properties. In the cerebellum, climbing fiber (CF) and parallel fiber (PF) inputs, which impinge onto the same Purkinje cell (PC), have very different functional properties. In the cerebellum of adult rats, using confocal and electron microscopy, we observed that VGluT2-positive CFs, characterized by a high Pr, only weakly express SNAP25, while VGluT1-positive PFs that show a low Pr abundantly express SNAP25. Moreover, SNAP25 was less profuse in the VGluT2-positive rosettes of mossy fibers (MFs) and was almost absent in inhibitory terminals. We extended our analysis to the SNAP23 homolog; this is expressed at different levels in both gamma-aminobutyric acid-containing terminals (GABAergic) and glutamatergic terminals of the cerebellar cortex. In conclusion, the preferential localization of SNAP25 in specific synaptic boutons suggests a correlation between SNAP25 and the Pr. This evidence supports the hypothesis that SNAP25 has a modulatory role in shaping synaptic responses.

Blood harmane is correlated with cerebellar metabolism in essential tremor: a pilot study.

BACKGROUND: On proton magnetic resonance spectroscopic imaging ((1)H MRSI), there is a decrease in cerebellar N-acetylaspartate/total creatine (NAA/tCr) in essential tremor (ET), signifying cerebellar neuronal dysfunction or degeneration. Harmane, which is present in the human diet, is a potent tremor-producing neurotoxin. Blood harmane concentrations seem to be elevated in ET. OBJECTIVES: To assess in patients with ET whether blood harmane concentration is correlated with cerebellar NAA/tCR, a neuroimaging measure of neuronal dysfunction or degeneration. METHODS: Twelve patients with ET underwent (1)H MRSI. The major neuroanatomic structure of interest was the cerebellar cortex. Secondary regions were the central cerebellar white matter, cerebellar vermis, thalamus, and basal ganglia. Blood concentrations of harmane and another neurotoxin, lead, were also assessed. RESULTS: Mean +/- SD cerebellar NAA/tCR was 1.52 +/- 0.41. In a linear regression model that adjusted for age and gender, log blood harmane concentration was a predictor of cerebellar NAA/tCR (beta = -0.41, p = 0.009); every 1 g(-10)/mL unit increase in log blood harmane concentration was associated with a 0.41 unit decrease in cerebellar NAA/tCR. The association between blood harmane concentration and brain NAA/tCR only occurred in the cerebellar cortex; it was not observed in secondary brain regions of interest. Furthermore, the association was specific to harmane and not another neurotoxin, lead. CONCLUSION: This study provides additional support for the emerging link between harmane, a neurotoxin, and ET. Further studies are warranted to address whether cerebellar harmane concentrations are associated with cerebellar pathology in postmortem studies of the ET brain.

Grape seed flavanols, but not Port wine, prevent ethanol-induced neuronal lipofuscin formation.

Lipofuscin is an end-product of lipid peroxidation which dramatically increases following ethanol consumption, as we have shown in hippocampal and cerebellar neurons. In this work, we corroborated observations indicating that supplementation of ethanol with 200 mg/l of grape seed flavanols prevents increased lipofuscin formation, an action that has been ascribed to the antioxidant properties of the flavanols. Because wine is an alcoholic beverage naturally rich in flavanols, we decided to study the effect of chronic ingestion of Port wine (PW), which also contains 20% ethanol and approximately 200 mg/l of flavanol oligomers, upon lipofuscin accumulation in the hippocampal CA1 and CA3 pyramidal neurons and in the cerebellar Purkinje cells. Six months old rats were fed with PW and results were compared with those obtained in ethanol-treated groups and pair-fed controls. After 6 months of treatment, the volume of lipofuscin per neuron was estimated using unbiased stereological methods. Treatment with PW resulted in an increase of lipofuscin in all neuronal populations studied when compared to controls and to rats treated with ethanol supplemented with flavanols. No differences were observed when comparisons were made with ethanol drinking rats. We conclude that PW, despite containing 20% ethanol and flavanols, does not prevent ethanol-induced lipofuscin formation as previously found in animals drinking ethanol plus flavanols. The reduced antioxidant capacity of PW might depend on the type and amount of flavanols present and on its content in sugars.

Transcriptional profiling of depolarization-dependent phenotypic alterations in primary cultures of developing granule neurons.

Rat cerebellar granule neurons cultured in medium supplemented with elevated KCl are extensively used as a model to examine the coupling between neural activity and Ca(2+)-dependent gene expression. Elevated (25 mM) KCl is believed to mimic endogenous neural activity because it promotes depolarization and Ca(+2)-dependent survival and some aspects of maturation. By comparison, at least half of the granule neurons grown in standard medium containing 5 mM KCl undergo apoptosis beginning approximately 4 days in vitro. However, accumulating evidence suggests that chronic depolarization induces phenotypic abnormalities whereas growth in chemically defined medium containing 5 mM KCl more closely resembles the constitutive phenotype. To examine this, oligonucleotide microarrays and RT-PCR of selected mRNAs were used to compare transcription profiles of cultures grown in 5 mM and 25 mM KCl. In some cases, N-methyl-D-aspartate (NMDA) which, like elevated KCl, promotes long-term survival was also tested. Robust changes in several gene groups were observed and indicated that growth in elevated KCl: induces expression of mRNAs that are not normally observed; represses expression of mRNAs that should be present; maintains expression of mRNAs that are markers of immature neurons. Supplementation of the growth medium with NMDA instead of elevated KCl produces similar abnormalities. Altogether, these data indicate that growth in 5 mM KCl more closely mimics survival and maturation of granule neurons in vivo and should therefore be adopted in future studies.

Screening cascade and development of potential Positron Emission Tomography radiotracers for mGluR5: in vitro and in vivo characterization.

PURPOSE: Use of mGluR5 receptor radiotracers to determine whether an in vitro binding assay is able to predict how good a radiotracer is likely to be in imaging receptor in the central nervous system (CNS) via positron emission tomography (PET). PROCEDURES: Saturation and equilibrium competition studies in rat and rhesus membranes were used to determine receptor concentrations and tracer affinities. In addition, specific binding of metabotropic receptor subtype 5 (mGluR5) radioligands in rhesus and rat brain sections was determined using a "no-wash protocol," and the in vivo binding signal in rats was determined using micro-PET. RESULTS: Affinity values were determined for a series of mGluR5 antagonists (1-5) and ranged from 0.1 to 11 nM in rat. A previously reported "no-wash protocol" was then employed to determine specific binding in tissue sections following a 20-min incubation, and the regional distribution of these mGluR5 radiotracers determined in rat brain via autoradiography. The analogs 1b, 2b, 3b, and 4b, but not 5b, displayed good signal-to-noise ratios under these conditions with high density of binding in caudate, cortex, and hippocampus and lower density in cerebellum. With this information it was predicted that 1c, 2c, 3b, and 4b would display measurable signal-to-noise ratios in vivo, and that the larger in vitro signals for 3b and 4b would translate to 3b and 4b yielding the best in vivo signals. These predictions were investigated using micro-PET imaging in rat. Compound 1c showed a rapid wash-in and rapid wash-out profile in rat brain. Compound 2c showed similar signal-to-noise ratio as 1b, but slower washout. Compounds 3b and 4b showed the best signal-to-noise ratio in vivo, while 5b did not provide a significant signal, as predicted. In vivo occupancy estimates for 2-methyl-6-(phenylethynyl)-pyridine (MPEP) following intravenous administration were determined using radiolabeled compounds 1c, 2c, and 3b; they were essentially the same and were on the order of 1 mg kg(-1) (ID(50)). CONCLUSIONS: An in vitro screen of several mGluR5 tracers was used to rapidly predict whether radiolabeled mGluR5 analogs would be useful as PET radiotracers. Results provided an extension to previously reported data. Two of the four radiotracers with the best in vitro "no-wash" results also showed the best potential as measured noninvasively using micro-PET.

Further evidence for altered cerebellar neuronal integrity in schizophrenia.

OBJECTIVE: The authors' goal was to investigate the distribution of metabolites and voxel composition in the pons and three cerebellar subregions and compare metabolite integral values and differences in voxel composition between patients with schizophrenia and healthy subjects. METHOD: Proton magnetic resonance spectroscopic imaging was used to study the cerebellum and pons of 14 patients with schizophrenia and 14 healthy comparison subjects. RESULTS: The voxel composition was not significantly different between the groups, but the patients with schizophrenia had significantly lower N-acetylaspartate levels in the cerebellar cortex and vermis. CONCLUSIONS: The lower integral value of N-acetylaspartate in the cerebellar cortex and the vermis of patients with schizophrenia supports the theory of a dysfunctional corticocerebellar-thalamic-cortical circuit in schizophrenia.