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Fragile X Syndrome (FXS) is a genetic neurodevelopmental disorder that causes autism and intellectual disabilities: exhibiting hyperactivity, elevated anxiety, and impaired cognitive/sensory processing. These deficits result from mutations in the X-linked gene Fragile X messenger ribonucleoprotein 1 (Fmr1). Fmr1-knock- out (KO) mouse models have shown consistency with observations in humans, displaying seizures and sensory processing deficits. Utilizing Fmr1-KO mice to identify a potential treatment for these symptoms, we administered a drug called 2-chloro-4-((2,5-dimethyl-1-(4-trifluoromethoxy)phenyl)-1H-imidazole-4-yl) ethynyl)pyridine (CTEP) to KO mice and measured behavioral changes. CTEP in- hibits metabotropic glutamate receptor pathways, which are upregulated in FXS. Two types of experiments were run: open field test (OFT) and elevated plus maze (EPM), commonly used to study anxiety and hyperactivity. Experimental mice with higher anxiety depict decreased exploration and more time spent near the arena’s wall or closed arms. We found that CTEP reduces the distance traveled in the OFT across both wild-type (WT) and KO groups, suggesting reduced locomotion. There were no statistically significant differences in time spent in EPM closed arms between WT and KO mice, indicating no treatment of anxiety. These results suggest that more effective intervention is needed to target anxiety deficits related to FXS.

[Background] Developing biological based approaches for preventing suicide has become a priority. In recent years, there has been a surge in studies investigating the role of the glutamatergic system in suicide, although it remains unclear., [Methods] We evaluated changes in the gene expression of the metabotropic glutamate receptor 5 (mGluR5) and its scaffolding proteins Homer1a and p11 in the dorsolateral prefrontal cortex (DLPFC), amygdala (AMY), and hippocampus (HIP) of 28 suicide decedents (S) (with no clinical psychiatric history or treatment with anxiolytics or antidepressants) and 26 controls (C) by real-time PCR (qPCR). Indeed, we measured BDNF gene expression and VGluT1 and VGAT immunoreactivities in the HIP by qPCR and immunohistochemistry, respectively. Cases and controls matched for age (C: 48.6 ± 11.6 years; S: 46.9 ± 14.5 years) and postmortem interval (PMI; C: 20.1 ± 13h; S: 16.9 ± 5h)., [Results] In DLPFC, S had lower p11 gene expression levels, but no differences were found in mGluR5 or Homer1a. In the AMY and HIP, mGluR5 and Homer1a were increased, p11 and BDNF were reduced. In the HIP, there were less VGAT-ir and more VGluT1-ir., [Limitations] Future studies are necessary to evaluate protein levels, and determine the cell types and potential compensatory mechanisms in a larger sample including S diagnosed with psychiatric disorders, females and different ethnicities., [Conclusions] This study identified significant alterations in mGluR5, Homer1a, p11, BDNF and excitatory/inhibitory balance in corticolimbic brain areas of S. These results further characterize the biological basis of suicide, contributing to the identification of potential biomarkers for suicide prevention.

Background Perturbed functional coupling between the metabotropic glutamate receptor-5 (mGluR5) and N-methyl-D-aspartate (NMDA) receptor-mediated excitatory glutamatergic neurotransmission may contribute to the pathophysiology of psychiatric disorders such as schizophrenia. We aimed to establish the functional interaction between mGluR5 and NMDA receptors in brain of mice with genetic ablation of the mGluR5. Methods We first measured the brain glutamate levels with magnetic resonance spectroscopy (MRS) in mGluR5 knockout (KO) and wild-type (WT) mice. Then, we assessed brain glucose metabolism with [18F]fluorodeoxyglucose ([18F]FDG) positron emission tomography before and after the acute administration of an NMDA antagonist, MK-801 (0.5 mg/kg), in the same mGluR5 KO and WT mice. Results Between-group comparisons showed no significant differences in [18F]FDG standardized uptake values (SUVs) in brain of mGluR5 KO and WT mice at baseline, but widespread reductions in mGluR5 KO mice compared to WT mice after MK-801 administration (p < 0.05). The baseline glutamate levels did not differ significantly between the two groups. However, there were significant negative correlations between baseline prefrontal glutamate levels and regional [18F]FDG SUVs in mGluR5 KO mice (p < 0.05), but no such correlations in WT mice. Fisher’s Z-transformation analysis revealed significant between-group differences in these correlations (p < 0.05). Conclusions This is the first multimodal neuroimaging study in mGluR5 KO mice and the first report on the association between cerebral glucose metabolism and glutamate levels in living rodents. The results indicate that mGluR5 KO mice respond to NMDA antagonism with reduced cerebral glucose metabolism, suggesting that mGluR5 transmission normally moderates the net effects of NMDA receptor antagonism on neuronal activity. The negative correlation between glutam

Electronic cigarettes (e-cigs) use has been dramatically increased recently, especially among youths. Previous studies from our laboratory showed that chronic exposure to e-cigs, containing 24 mg/mL nicotine, was associated with dysregulation of glutamate transporters and neurotransmitter levels in the brain of a mouse model. In this study, we evaluated the effect of three months continuous exposure to e-cig vapor (JUUL pods), containing a high nicotine concentration, on the expression of glutamate receptors and transporters in drug reward brain regions such as the nucleus accumbens (NAc) core (NAc-core), NAc shell (NAc-shell) and hippocampus (HIP) in female C57BL/6 mice. Three months exposure to mint- or mango-flavored JUUL (containing 5% nicotine, 59 mg/mL) induced upregulation of metabotropic glutamate receptor 1 (mGluR1) and postsynaptic density protein 95 (phosphorylated and total PSD95) expression, and downregulation of mGluR5 and glutamate transporter 1 (GLT-1) in the NAc-shell. In addition, three months exposure to JUUL was associated with upregulation of mGluR5 and GLT-1 expression in the HIP. These findings demonstrated that three-month exposure to e-cig vapor containing high nicotine concentrations induced differential effects on the glutamatergic system in the NAc and HIP, suggesting dysregulation of glutamatergic system activity in mesolimbic brain regions.

Purpose: To determine how the low-affinity (Z)-isomer of the radiotracer [11C]ABP688 affects binding potential values in vivo in humans. Methods: High-resolution [11C]ABP688 PET scans were acquired on 74 healthy volunteers (25 male, 49 female, mean age 20 ± 3.0). The relative contents of (E)- and (Z)-isomers were determined prior to injection using analytical high-performance liquid chromatography [rt(E) = 10 min, rt(Z) = 8.5 min]. Mean binding potential [BPND = fND * (Bavail/KD)] values were calculated in the striatum, limbic regions, and prefrontal cortex using the simplified reference tissue model with cerebellar grey matter as reference. Results: Mean ± SD (E)-isomer content in [11C]ABP688 production was 92 ± 3.8% (range 78–97%). Percent (E)-isomer was positively correlated with BPND in the striatum (ρ = 0.28, p = 0.015) and limbic regions (ρ = 0.25, p = 0.036). In multiple regression analysis, sex (β = 0.39, p = 0.001) and (E)-isomer content (β = 0.23, p = 0.040) were significant predictors of BPND. Conclusions: Even modest levels of (Z)-[11C]ABP688 can reduce estimates of tracer binding in vivo. Future studies should use production methods that enrich levels of (E)-[11C]ABP688, report tracer isomer ratios, and account for this factor in their analyses.

Schizophrenia is a severe neuropsychiatric disorder, arising in adolescence and early adulthood and characterised by hallucinations, delusions, blunted affect and disorganised thought patterns. One of the most enduring features of schizophrenia and psychosis are structural brain deficits, whose pathophysiological mechanism is unknown. Accumulating evidence indicates that inflammation both peripherally and centrally in the form of increased activation of the brain’s immune cells, microglia, may be a potential cause of structural deficits in psychosis. The evidence is multi-faceted ranging from mouse models that demonstrate increased numbers of microglia, to clinical studies of patients with schizophrenia showing increased pro- inflammatory molecules within peripheral blood. However, there are still many questions that remain unanswered, including whether inflammation varies across stages of psychosis, whether it is related to structural brain deficits and symptomatology and how inflammation identified in schizophrenia relates to other candidate pathways implicated in psychosis. In this thesis a multi-disciplinary approach was adopted, considered appropriate to tackling the complexity of these questions. Firstly, to determine whether inflammation was associated with other candidate pathways implicated in psychosis, we conducted an animal study, utilising an mGluR5 KO mouse model of psychosis. Glutamate has been shown to influence neuroinflammation, with cellular studies demonstrating that mGluR5 can regulate microglial numbers and activation. At the time of conducting the study, there were no satisfactory mGluR5 PET ligands that enabled in vivo monitoring within the clinical population. The mGluR5 KO mouse had been shown to display neuropsychiatric endophenotypes related to schizophrenia and thereby offered an alternative approach to gaining further insight into the role of mGluR5 in neuroinflammation and how this may impact symptoms associated with psychosis. Our aim

Cognitive abilities are critical for human functioning and flourishing yet unfortunately dysfunction of cognition is a common symptom in developmental, psychiatric and neurodegenerative disorders. Observational studies in humans have demonstrated that variation in cognitive abilities is associated with genetic and environmental factors but are unable to easily examine gene-environment interactions or make causal claims regarding the contribution of each factor. Rodent models enable the investigation of causal effects in cognition as they make feasible the necessary genetic and environmental manipulations. This thesis examined a genetic mouse model of cognitive impairment housed under differing environmental conditions. These animals were cognitively phenotyped using touchscreen operant chambers (touchscreens), a testing method that offers a broader and more consistent assay of cognitive testing in rodents than traditional behavioural tasks. Environmental factors such as exercise and education are known to be associated with improved cognitive performance in humans. A plausibly analogous environmental manipulation to examine these factors is environmental enrichment (EE), which entails any positive modification of the ‘standard housing’ (SH) conditions in which laboratory animals are typically held, usually involving increased opportunity for cognitive stimulation and physical activity. EE has been reported to enhance baseline performance of wild-type animals on traditional cognitive behavioural tasks. We used touchscreens to analyse the effects of EE on a battery of cognitive tasks, hypothesising that EE would enhance the performance of mice. Our hypothesis was partially supported in that EE induced enhancements in cognitive flexibility as observed in pairwise discrimination and reversal learning improvements. However, no other significant effects of EE on cognitive performance were observed. Genetic variation was examined using metabotropic glutamate receptor 5 kno

L-DOPA is the main pharmacological therapy for Parkinson's disease. However, long-term exposure to L-DOPA induces involuntary movements termed dyskinesia. Clinical trials show that dyskinesia is attenuated by metabotropic glutamate receptor type 5 (mGluR5) antagonists. Further, the onset of dyskinesia is delayed by nicotine and mGluR5 expression is lower in smokers than in non-smokers. However, the mechanisms by which mGluR5 modulates dyskinesia and how mGluR5 and nicotine interact have not been established. To address these issues, we studied the role of mGluR5 in D1R-containing neurons in dyskinesia and examined whether nicotine reduces dyskinesia via mGluR5. In the aphakia mouse model of Parkinson's disease, we selectively knocked down mGluR5 in D1R-containing neurons (aphakia-mGluR5KD-D1). We found that genetic downregulation of mGluR5 decreased dyskinesia in aphakia mice. Although chronic nicotine increased the therapeutic effect of L-DOPA in both aphakia and aphakia-mGluR5KD-D1 mice, it caused a robust reduction in dyskinesia only in aphakia, and not in aphakia-mGluR5KD-D1 mice. Downregulating mGluR5 or nicotine treatment after L-DOPA decreased ERK and histone 3 activation, and FosB expression. Combining nicotine and mGluR5 knockdown did not have an added antidyskinetic effect, indicating that the effect of nicotine might be mediated by downregulation of mGluR5 expression. Treatment of aphakia-mGluR5KD-D1 mice with a negative allosteric modulator did not further modify dyskinesia, suggesting that mGluR5 in non-D1R-containing neurons does not play a role in its development. In conclusion, this work suggests that mGluR5 antagonists reduce dyskinesia by mainly affecting D1R-containing neurons and that the effect of nicotine on dyskinetic signs in aphakia mice is likely via mGluR5.

This thesis examined the role of mGlu5 and NMDA receptors in the extinction and reinstatement of alcohol seeking. The first series of experiments (Chapter 3) examined the effect of systemic antagonism of mGlu5 and NMDA receptors on the instrumental extinction of alcohol seeking. These experiments demonstrated that antagonism of mGluR5 with MTEP prior to extinction, impaired the between session extinction of alcohol seeking in a time and dose dependent manner. Conversely, NMDA receptor antagonism alone (MK-801 and Ifenprodil), had no effect on the extinction of alcohol seeking. Furthermore, the findings revealed that MTEP acts, at least in part, via a state dependent mechanism as there was no deficit in the acquisition or expression of extinction when animals were trained and tested under the effects of MTEP. The final experiments reported in Chapter 3, provided novel evidence that co-antagonism of mGlu5 and NMDA receptors impaired both the within and between session extinction of instrumental alcohol seeking. The second series of experiments (Chapter 4) examined the role of mGluR5 in the context-induced reinstatement, and reacquisition of alcohol seeking. These experiments provided mixed evidence as to the role of mGluR5 in these recovery phenomena. While there was, evidence in Experiment 9 that MTEP impaired context-induced reinstatement, this was not replicated in Experiment 10 or 11. Finally, there was evidence that MTEP potentiates the reacquisition of alcohol seeking. However, there was no influence on the break point or overall performance in a progressive ratio test of motivation. Together, the findings from this thesis provide novel evidence for the role of mGluR5 in the instrumental extinction of alcohol seeking. They suggest a functional interaction between mGlu5 and NMDA receptors in the processes, which underpin the extinction of alcohol seeking. Finally, the mixed results for the role of mGluR5 in context-induced reinstatement and the finding that MTEP

Long-term levodopa (l-dopa) treatment in patients with Parkinson ́s disease (PD) is associated with the development of motor complications (ie, motor fluctuations and dyskinesias). The principal etiopathogenic factors are the degree of nigro-striatal dopaminergic loss and the duration and dose of l-dopa treatment. In this review article we concentrate on analysis of the mechanisms underlying l-dopa–induced dyskinesias, a phenomenon that causes disability in a proportion of patients and that has not benefited from major therapeutic advances. Thus, we discuss the main neurotransmitters, receptors, and pathways that have been thought to play a role in l-dopa–induced dyskinesias from the perspective of basic neuroscience studies. Some important advances in deciphering the molecular pathways involved in these abnormal movements have occurred in recent years to reveal potential targets that could be used for therapeutic purposes. However, it has not been an easy road because there have been a plethora of components involved in the generation of these undesired movements, even bypassing the traditional and well-accepted dopamine receptor activation, as recently revealed by optogenetics. Here, we attempt to unify the available data with the hope of guiding and fostering future research in the field of striatal activation and abnormal movement generation. © 2017 International Parkinson and Movement Disorder Society.

Amyloid PET imaging with [11C]PIB enabled detection of Aβ for the first time in vivo. However, [11C]PIB is a small molecule that binds only the insoluble Aβ plaque. Rather, the soluble Aβ aggregates are considered the cause of Alzheimer’s disease (AD). As such, a more sensitive and specific PET tracer is needed for tracking longitudinal AD pathology. Soluble Aβ aggregates likely interact with the metabotropic glutamate receptor 5 (mGluR5) to cause neurotoxic effects. However, with [11C]ABP688 PET we were unable to detect aberrant mGluR5 binding in AD mouse models, although we find elevated mGluR5 protein levels with immunoblotting. Antibodies are highly specific large molecules that can bind specifically to soluble Aβ aggregates, thus they can be a good marker for AD pathology. Unfortunately, due to their large size they cannot cross the blood-brain barrier (BBB). However, it is possible to shuttle antibodies into the brain by taking advantage of endogenous transporter systems on the BBB. By creating bispecific antibodies binding both to soluble Aβ aggregates and to the transferrin receptor (BBB target), we successfully transported the antibody into the brain and could visually detect soluble Aβ aggregates with PET. Recombinant expression further improved and optimized antibody design, creating smaller bispecific antibody-based constructs that had better pharmacokinetic properties allowing for earlier PET scanning (1 day instead of 3), and more sensitive signal. Lastly, using TCO-tetrazine click chemistry, we indirectly labeled our antibodies with fluorine-18, and could successfully perform PET already 11 h post-injection with a fluorine-18 labeled antibody.

Alzheimer's disease (AD) is characterized by aggregation of amyloid beta (Aβ) into insoluble plaques. Intermediates, Aβ oligomers (Aβo), appear to be the mechanistic cause of disease. The de facto PET AD ligand, [(11)C]PIB, binds and visualizes Aβ plaque load, which does not correlate well with disease severity. Therefore, finding a dynamic target that changes with pathology progression in AD is of great interest. Aβo alter synaptic plasticity, inhibit long-term potentiation, and facilitate long-term depression; key mechanisms involved in memory and learning. In order to convey these neurotoxic effects, Aβo requires interaction with the metabotropic glutamate 5 receptor (mGluR5). The aim was to investigate in vivo mGluR5 changes in an Aβ pathology model using PET. Wild type C57/BL6 (wt) and AβPP transgenic mice (tg-ArcSwe), 4, 8, and 16 months old, were PET scanned with [(11)C]ABP688, which is highly specific to mGluR5, to investigate changes in mGluR5. Mouse brains were extracted postscan and mGluR5 and Aβ protofibril levels were assessed with immunoblotting and ELISA respectively. Receptor-dense brain regions (hippocampus, thalamus, and striatum) displayed higher [(11)C]ABP688 concentrations corresponding to mGluR5 expression pattern. Mice had similar uptake levels of [(11)C]ABP688 regardless of genotype or age. Immunoblotting revealed general decline in mGluR5 expression and elevated levels of mGluR5 in 16 months old tg-ArcSwe compared with wt mice. [(11)C]ABP688 could visualize mGluR5 in the mouse brain. In conclusion, mGluR5 levels were found to decrease with age and tended to be higher in tg-ArcSwe compared with wt mice, however these changes could not be quantified with PET.

Amyloid PET imaging with [11C]PIB enabled detection of Aβ for the first time in vivo. However, [11C]PIB is a small molecule that binds only the insoluble Aβ plaque. Rather, the soluble Aβ aggregates are considered the cause of Alzheimer’s disease (AD). As such, a more sensitive and specific PET tracer is needed for tracking longitudinal AD pathology. Soluble Aβ aggregates likely interact with the metabotropic glutamate receptor 5 (mGluR5) to cause neurotoxic effects. However, with [11C]ABP688 PET we were unable to detect aberrant mGluR5 binding in AD mouse models, although we find elevated mGluR5 protein levels with immunoblotting. Antibodies are highly specific large molecules that can bind specifically to soluble Aβ aggregates, thus they can be a good marker for AD pathology. Unfortunately, due to their large size they cannot cross the blood-brain barrier (BBB). However, it is possible to shuttle antibodies into the brain by taking advantage of endogenous transporter systems on the BBB. By creating bispecific antibodies binding both to soluble Aβ aggregates and to the transferrin receptor (BBB target), we successfully transported the antibody into the brain and could visually detect soluble Aβ aggregates with PET. Recombinant expression further improved and optimized antibody design, creating smaller bispecific antibody-based constructs that had better pharmacokinetic properties allowing for earlier PET scanning (1 day instead of 3), and more sensitive signal. Lastly, using TCO-tetrazine click chemistry, we indirectly labeled our antibodies with fluorine-18, and could successfully perform PET already 11 h post-injection with a fluorine-18 labeled antibody.

The metabotropic glutamate receptor 5 (mGluR5) represents the glutamate sensor of astrocytes and is thought to participate in the regulation of diverse astrocytic functions. The mGluR5 has the particularity to show distinct calcium patterns upon activation including sustained calcium mobilization or calcium oscillations. In our laboratory, we are convinced that these patterns differently impact on the mGluR5-mediated regulation of astrocytic functions. In this thesis, we highlight that mGluR5 activation associated with calcium oscillations protects against receptor desensitization while sustained calcium mobilization is associated with robust desensitization. In addition, we demonstrate that calcium oscillations induced by mGluR5 activation regulate the astrocytic glutamate transport while sustained calcium mobilization has no effect. We also add new insight into the role of PKC epsilon, by showing its implication in the regulation of mGluR5 calcium oscillations., (ART - Art et sciences de l'art) -- UCL, 2017

Fragile X syndrome is the most common hereditary cause of intellectual disability and autism. Currently, no targeted treatment is available. The identification of its causative gene, followed by the generation of animal models, has boosted research options. The past decades, preclinical research has focused on involved pathways as targets for therapeutic interventions. Targeting many of these pathways in animal models improved core features of the disease. These promising results were received with great enthusiasm, and led to clinical trials in patients with fragile X syndrome. The aim of this thesis was to develop and evaluate therapeutic intervention strategies for fragile X syndrome with the ultimate goal to advance care and treatment options for patients with the disorder. One aspect was to identify possible targets for therapeutic intervention using social behavior of _Fmr1_ knockout mice as outcome measure. In addition, new strategies were explored using combination therapy (chapter one - four). A second aspect was to investigate a potential target, the mGluR5, for its efficacy in clinical trials. The mGluR5 antagonist mavoglurant was studied in a multicenter randomized, double-blind, placebo-controlled trial (chapter five). A third aspect was to center patient care, in order to advance clinical expertise and enable clinical research. An expert center and clinical guidelines are indispensable in patient care and further development of treatment for fragile X syndrome (chapter seven). The findings were then placed in the light of current knowledge and future prospectives (chapter six and eight).

UnlabelledAltered function of the Gq-coupled, Group 1 metabotropic glutamate receptors, specifically mGlu5, is implicated in multiple mouse models of autism and intellectual disability. mGlu5 dysfunction has been most well characterized in the fragile X syndrome mouse model, the Fmr1 knock-out (KO) mouse, where pharmacological and genetic reduction of mGlu5 reverses many phenotypes. mGlu5 is less associated with its scaffolding protein Homer in Fmr1 KO mice, and restoration of mGlu5-Homer interactions by genetic deletion of a short, dominant negative of Homer, H1a, rescues many phenotypes of Fmr1 KO mice. These results suggested that disruption of mGlu5-Homer leads to phenotypes of FXS. To test this idea, we examined mice with a knockin mutation of mGlu5 (F1128R; mGlu5(R/R)) that abrogates binding to Homer. Although FMRP levels were normal, mGlu5(R/R) mice mimicked multiple phenotypes of Fmr1 KO mice, including reduced mGlu5 association with the postsynaptic density, enhanced constitutive mGlu5 signaling to protein synthesis, deficits in agonist-induced translational control, protein synthesis-independent LTD, neocortical hyperexcitability, audiogenic seizures, and altered behaviors, including anxiety and sensorimotor gating. These results reveal new roles for the Homer scaffolds in regulation of mGlu5 function and implicate a specific molecular mechanism in a complex brain disease.Significance statementAbnormal function of the metabotropic, or Gq-coupled, glutamate receptor 5 (mGlu5) has been implicated in neurodevelopmental disorders, including a genetic cause of intellectual disability and autism called fragile X syndrome. In brains of a mouse model of fragile X, mGlu5 is less associated with its binding partner Homer, a scaffolding protein that regulates mGlu5 localization to synapses and its ability to activate biochemical signaling pathways. Here we show that a mouse expressing a mutant mGlu5 that cannot bind to Homer is sufficient to mimic many of the biochem

Altered function of the Gq-coupled, Group 1 metabotropic glutamate receptors, specifically mGlu5, is implicated in multiple mouse models of autism and intellectual disability. mGlu5 dysfunction has been most well characterized in the fragile X syndrome mouse model, the Fmr1 knock-out (KO) mouse, where pharmacological and genetic reduction of mGlu5 reverses many phenotypes. mGlu5 is less associated with its scaffolding protein Homer in Fmr1 KO mice, and restoration of mGlu5-Homer interactions by genetic deletion of a short, dominant negative of Homer, H1a, rescues many phenotypes of Fmr1 KO mice. These results suggested that disruption of mGlu5-Homer leads to phenotypes of FXS. To test this idea, we examined mice with a knockin mutation of mGlu5 (F1128R; mGlu5(R/R)) that abrogates binding to Homer. Although FMRP levels were normal, mGlu5(R/R) mice mimicked multiple phenotypes of Fmr1 KO mice, including reduced mGlu5 association with the postsynaptic density, enhanced constitutive mGlu5 signaling to protein synthesis, deficits in agonist-induced translational control, protein synthesis-independent LTD, neocortical hyperexcitability, audiogenic seizures, and altered behaviors, including anxiety and sensorimotor gating. These results reveal new roles for the Homer scaffolds in regulation of mGlu5 function and implicate a specific molecular mechanism in a complex brain disease.Significance statementAbnormal function of the metabotropic, or Gq-coupled, glutamate receptor 5 (mGlu5) has been implicated in neurodevelopmental disorders, including a genetic cause of intellectual disability and autism called fragile X syndrome. In brains of a mouse model of fragile X, mGlu5 is less associated with its binding partner Homer, a scaffolding protein that regulates mGlu5 localization to synapses and its ability to activate biochemical signaling pathways. Here we show that a mouse expressing a mutant mGlu5 that cannot bind to Homer is sufficient to mimic many of the biochemical, neur

Metabotropic glutamate receptor subtype 5 (mGluR5)-targeting therapeutics display a favourable profile of effects in preclinical paradigms, particularly for the treatment of cognitive dysfunction. It however remains unclear whether mGluR5 plays a causal or epiphenomenal role in the pathophysiology of schizophrenia. It is important to understand the aspects of mGluR5 that might be altered within the pathological condition, as this could have implications for novel drugs targeted at these systems in the patient. This thesis therefore aimed to deconstruct [at least some] of the complexities of mGluR5 in schizophrenia, with a focus on potential dysregulation of mGluR5, predominately utilising human tissues (postmortem brain tissues and DNA samples derived from schizophrenia patients). Considering many symptoms associated with schizophrenia overlap with other closely related neuropsychiatric disorders, the first study in this thesis (Chapter 2) aimed to address whether mGluR5 alterations were specific to a particular set of symptoms (such as depressive symptoms or psychotic symptoms, or a combination thereof). To do this, mGluR5 binding was performed in postmortem samples from the anterior cingulate cortex, a region that modulates cognitive and emotional processes, of subjects with schizophrenia, major depression (with or without psychosis) or bipolar disorder. The results revealed that mGluR5 binding was unaltered across all neuropsychiatric pathologies, providing preliminary evidence that mGluR5 is not affected, at least in this region. Binding studies provide valuable insight into the ability of a receptor to interact with ligands and are thus useful for determining the therapeutic utility of a receptor; however this method is unable to detect wider changes, such as altered regulation, which may be underlying the observed pathologies. Building on from this work, Chapter 3 aimed to explore whether dysregulation was present within the mGluR5 system in schizophrenia. Pos

Metabotropic glutamate receptor subtype 5 (mGluR5)-targeting therapeutics display a favourable profile of effects in preclinical paradigms, particularly for the treatment of cognitive dysfunction. It however remains unclear whether mGluR5 plays a causal or epiphenomenal role in the pathophysiology of schizophrenia. It is important to understand the aspects of mGluR5 that might be altered within the pathological condition, as this could have implications for novel drugs targeted at these systems in the patient. This thesis therefore aimed to deconstruct [at least some] of the complexities of mGluR5 in schizophrenia, with a focus on potential dysregulation of mGluR5, predominately utilising human tissues (postmortem brain tissues and DNA samples derived from schizophrenia patients). Considering many symptoms associated with schizophrenia overlap with other closely related neuropsychiatric disorders, the first study in this thesis (Chapter 2) aimed to address whether mGluR5 alterations were specific to a particular set of symptoms (such as depressive symptoms or psychotic symptoms, or a combination thereof). To do this, mGluR5 binding was performed in postmortem samples from the anterior cingulate cortex, a region that modulates cognitive and emotional processes, of subjects with schizophrenia, major depression (with or without psychosis) or bipolar disorder. The results revealed that mGluR5 binding was unaltered across all neuropsychiatric pathologies, providing preliminary evidence that mGluR5 is not affected, at least in this region. Binding studies provide valuable insight into the ability of a receptor to interact with ligands and are thus useful for determining the therapeutic utility of a receptor; however this method is unable to detect wider changes, such as altered regulation, which may be underlying the observed pathologies. Building on from this work, Chapter 3 aimed to explore whether dysregulation was present within the mGluR5 system in schizophrenia. Pos

Metabotropic glutamate receptor subtype 5 (mGluR5)-targeting therapeutics display a favourable profile of effects in preclinical paradigms, particularly for the treatment of cognitive dysfunction. It however remains unclear whether mGluR5 plays a causal or epiphenomenal role in the pathophysiology of schizophrenia. It is important to understand the aspects of mGluR5 that might be altered within the pathological condition, as this could have implications for novel drugs targeted at these systems in the patient. This thesis therefore aimed to deconstruct [at least some] of the complexities of mGluR5 in schizophrenia, with a focus on potential dysregulation of mGluR5, predominately utilising human tissues (postmortem brain tissues and DNA samples derived from schizophrenia patients). Considering many symptoms associated with schizophrenia overlap with other closely related neuropsychiatric disorders, the first study in this thesis (Chapter 2) aimed to address whether mGluR5 alterations were specific to a particular set of symptoms (such as depressive symptoms or psychotic symptoms, or a combination thereof). To do this, mGluR5 binding was performed in postmortem samples from the anterior cingulate cortex, a region that modulates cognitive and emotional processes, of subjects with schizophrenia, major depression (with or without psychosis) or bipolar disorder. The results revealed that mGluR5 binding was unaltered across all neuropsychiatric pathologies, providing preliminary evidence that mGluR5 is not affected, at least in this region. Binding studies provide valuable insight into the ability of a receptor to interact with ligands and are thus useful for determining the therapeutic utility of a receptor; however this method is unable to detect wider changes, such as altered regulation, which may be underlying the observed pathologies. Building on from this work, Chapter 3 aimed to explore whether dysregulation was present within the mGluR5 system in schizophrenia. Pos

A premutation (PM) expansion (55-200 CGG) in the fragile X mental retardation gene 1 causes elevated messenger RNA and reduced fragile X mental retardation gene 1 protein. Young PM carriers can develop characteristic physical features and mild cognitive disabilities. In addition, individuals with PM, particularly male carriers, are at high risk to develop fragile X-associated tremor/ataxia syndrome (FXTAS) with aging. Human postmortem FXTAS brains show extensive white matter disease in the cerebellum and the presence of intranuclear inclusions throughout the brain, although their etiologic significance is unknown. In the current work, expression levels of the metabotropic glutamate (Glu) receptor 5 and the Glu transporter excitatory amino acid transporter 1, examined by reverse transcription polymerase chain reaction and western blot analyses, were found to be reduced in the postmortem cerebellum of PM carriers with FXTAS compared with age matched controls, with higher CGG repeat number having greater reductions in both proteins. These data suggests a dysregulation of Glu signaling in PM carriers, which would likely contribute to the development and severity of FXTAS.

Background Pharmacological modulation of metabotropic glutamate receptor 5 (mGluR5) is of marked interest as a novel therapeutic mechanism to treat schizophrenia and major depression. However, the status of mGluR5 in the pathophysiology of these disorders remains unknown. Discussion The majority of studies in the schizophrenia post-mortem brain indicate that total mGluR5 expression is unaltered. However, close examination of the literature suggests that these findings are superficial, and in actuality, a number of critical factors have not yet been considered; alterations may be highly dependent on brain region, neuronal population or molecular organisation in specific cellular compartments. A number of genetic knockout studies (mGluR5, Norbin, Homer1 etc.) continue to lend support to a role of mGluR5 in the pathology of schizophrenia, providing impetus to explore the regulation of mGluR5 beyond total mGluR5 protein and mRNA levels. With regards to major depression, preliminary evidence to date shows a reduction in total mGluR5 protein and mRNA levels; however, as in schizophrenia, there are no studies examining mGluR5 function or regulation in the pathological state. A comprehensive understanding of mGluR5 regulation in major depression, particularly in comparison to schizophrenia, is crucial as this has extensive implications for mGluR5 targeting novel therapeutics, especially considering that opposing modulation of mGluR5 is of therapeutic interest for these two disorders. Summary Despite the complexities, examinations of post-mortem human brain provide valuable insights into the pathologies of these inherently human disorders. It is important, especially with regards to the identification of novel therapeutic drug targets, to have an in depth understanding of the pathophysiologies of these disorders. We posit that brain region- and cell type-specific alterations exist in mGluR5 in schizophrenia and depression, with evidence pointing towards altered regulation of

Background Pharmacological modulation of metabotropic glutamate receptor 5 (mGluR5) is of marked interest as a novel therapeutic mechanism to treat schizophrenia and major depression. However, the status of mGluR5 in the pathophysiology of these disorders remains unknown. Discussion The majority of studies in the schizophrenia post-mortem brain indicate that total mGluR5 expression is unaltered. However, close examination of the literature suggests that these findings are superficial, and in actuality, a number of critical factors have not yet been considered; alterations may be highly dependent on brain region, neuronal population or molecular organisation in specific cellular compartments. A number of genetic knockout studies (mGluR5, Norbin, Homer1 etc.) continue to lend support to a role of mGluR5 in the pathology of schizophrenia, providing impetus to explore the regulation of mGluR5 beyond total mGluR5 protein and mRNA levels. With regards to major depression, preliminary evidence to date shows a reduction in total mGluR5 protein and mRNA levels; however, as in schizophrenia, there are no studies examining mGluR5 function or regulation in the pathological state. A comprehensive understanding of mGluR5 regulation in major depression, particularly in comparison to schizophrenia, is crucial as this has extensive implications for mGluR5 targeting novel therapeutics, especially considering that opposing modulation of mGluR5 is of therapeutic interest for these two disorders. Summary Despite the complexities, examinations of post-mortem human brain provide valuable insights into the pathologies of these inherently human disorders. It is important, especially with regards to the identification of novel therapeutic drug targets, to have an in depth understanding of the pathophysiologies of these disorders. We posit that brain region- and cell type-specific alterations exist in mGluR5 in schizophrenia and depression, with evidence pointing towards altered regulation of

Background Pharmacological modulation of metabotropic glutamate receptor 5 (mGluR5) is of marked interest as a novel therapeutic mechanism to treat schizophrenia and major depression. However, the status of mGluR5 in the pathophysiology of these disorders remains unknown. Discussion The majority of studies in the schizophrenia post-mortem brain indicate that total mGluR5 expression is unaltered. However, close examination of the literature suggests that these findings are superficial, and in actuality, a number of critical factors have not yet been considered; alterations may be highly dependent on brain region, neuronal population or molecular organisation in specific cellular compartments. A number of genetic knockout studies (mGluR5, Norbin, Homer1 etc.) continue to lend support to a role of mGluR5 in the pathology of schizophrenia, providing impetus to explore the regulation of mGluR5 beyond total mGluR5 protein and mRNA levels. With regards to major depression, preliminary evidence to date shows a reduction in total mGluR5 protein and mRNA levels; however, as in schizophrenia, there are no studies examining mGluR5 function or regulation in the pathological state. A comprehensive understanding of mGluR5 regulation in major depression, particularly in comparison to schizophrenia, is crucial as this has extensive implications for mGluR5 targeting novel therapeutics, especially considering that opposing modulation of mGluR5 is of therapeutic interest for these two disorders. Summary Despite the complexities, examinations of post-mortem human brain provide valuable insights into the pathologies of these inherently human disorders. It is important, especially with regards to the identification of novel therapeutic drug targets, to have an in depth understanding of the pathophysiologies of these disorders. We posit that brain region- and cell type-specific alterations exist in mGluR5 in schizophrenia and depression, with evidence pointing towards altered regulation of

Background: Novel experiences activate the brains reward system in a manner similar to drugs of abuse, and high levels of novelty-seeking and sensation-seeking behavior have been associated with increased susceptibility to alcohol and drug abuse. Here, we show that metabotropic glutamate receptor 5 (mGluR5) signaling on dopaminoceptive neurons is necessary for both novelty-seeking behavior and the abstinence-induced escalation of alcohol drinking. less thanbrgreater than less thanbrgreater thanMethods: Mice harboring a transgene expressing microRNA hairpins against mGluR5 messenger RNA under the control of the D1 dopamine receptor gene promoter (mGluR5KD-D1) were tested in a battery of behavioral tests measuring learning abilities, anxiety levels, reactions to novelty, operant sensation seeking, and alcohol sensitivity. In addition, we have developed a method to assess long-term patterns of alcohol drinking in mice housed in groups using the IntelliCage system. less thanbrgreater than less thanbrgreater thanResults: mGluR5KD-D1 mice showed no behavioral deficits and exhibited normal anxiety-like behaviors and learning abilities. However, mGluR5KD-D1 animals showed reduced locomotor activity when placed in a novel environment, and exhibited decreased interaction with a novel object. Moreover, unlike control animals, mutant mice did not perform instrumental responses under the operant sensation-seeking paradigm, although they learned to respond for food normally. When mGluR5KD-D1 mice were provided access to alcohol, they showed similar patterns of consumption as wild-type animals. However, mutant mice did not escalate their alcohol consumption after a period of forced abstinence, but control mice almost doubled their intake. less thanbrgreater than less thanbrgreater thanConclusions: These data identify mGluR5 receptors on D1-expressing neurons as a common molecular substrate of novelty-seeking behaviors and behaviors associated with alcohol abuse., Funding Agencies|Polish Ministry of Science and Higher Education|N405 143238|Institute of Pharmacology of Polish Academy of Sciences||Department of Toxicology of Faculty of Pharmacy of Jagiellonian University Collegium Medicum||European Research Council||Swedish Research Council||German Bundesministerium fur Bildung und Forschung (National Genome Research Network Plus)|FKZ: 01GS08152

Glutamatergic agents have been conceptualized as powerful, fast-acting alternatives to monoaminergic-based antidepressants. NMDA receptor antagonists such as ketamine or MK-801 are therapeutically effective, but their clinical use is hampered by psychotomimetic effects, accompanied by neurotoxicity in the retrosplenial and cingulate cortex. Antagonists of metabotropic mGlu5 receptors like MPEP elicit both robust antidepressant and anxiolytic effects; however, the underlying mechanisms are yet unknown. mGlu5 receptors closely interact with NMDA receptors, but whether MPEP induces neurotoxicity similar to NMDA receptor antagonists has not been elucidated. We show here using c-Fos brain mapping that MPEP administration results in a restricted activation of distinct stress-related brain areas, including the bed nucleus of stria terminalis (BNST), central nucleus of the amygdala, and paraventricular nucleus of the hypothalamus (PVNH), in a pattern similar to that induced by classical antidepressants and anxio-lytics. Unlike the NMDA antagonist MK-801, MPEP does not injure the adult retrosplenial cortex, in which it fails to induce heat shock protein 70 (Hsp70). Moreover, MPEP does not elicit to the same extent as MK-801 apoptosis in cortical areas at perinatal stages, as revealed by caspase 3 expression. These data identify new cellular targets for the anxiolytic and antidepressant effect of MPEP, indicating also in addition that in contrast to MK-801, it lacks the cortical neurotoxicity associated with psychotomimetic side-effects. (C) 2012 Elsevier Ltd. All rights reserved.

Glutamatergic agents have been conceptualized as powerful, fast-acting alternatives to monoaminergic-based antidepressants. NMDA receptor antagonists such as ketamine or MK-801 are therapeutically effective, but their clinical use is hampered by psychotomimetic effects, accompanied by neurotoxicity in the retrosplenial and cingulate cortex. Antagonists of metabotropic mGlu5 receptors like MPEP elicit both robust antidepressant and anxiolytic effects; however, the underlying mechanisms are yet unknown. mGlu5 receptors closely interact with NMDA receptors, but whether MPEP induces neurotoxicity similar to NMDA receptor antagonists has not been elucidated. We show here using c-Fos brain mapping that MPEP administration results in a restricted activation of distinct stress-related brain areas, including the bed nucleus of stria terminalis (BNST), central nucleus of the amygdala, and paraventricular nucleus of the hypothalamus (PVNH), in a pattern similar to that induced by classical antidepressants and anxio-lytics. Unlike the NMDA antagonist MK-801, MPEP does not injure the adult retrosplenial cortex, in which it fails to induce heat shock protein 70 (Hsp70). Moreover, MPEP does not elicit to the same extent as MK-801 apoptosis in cortical areas at perinatal stages, as revealed by caspase 3 expression. These data identify new cellular targets for the anxiolytic and antidepressant effect of MPEP, indicating also in addition that in contrast to MK-801, it lacks the cortical neurotoxicity associated with psychotomimetic side-effects. (C) 2012 Elsevier Ltd. All rights reserved.

of a poster that presented at the Society of Biological Psychiatry 67th Annual Meeting.

of a poster that presented at the Society of Biological Psychiatry 67th Annual Meeting.

of a poster that presented at the Society of Biological Psychiatry 67th Annual Meeting.

Prepulse inhibition (PPI) may useful for exploring the proposed shared neurobiology between idiopathic autism and autism caused by FXS. We compared PPI in four groups: typically developing controls (n = 18), FXS and autism (FXS+A; n = 15), FXS without autism spectrum disorder (FXS-A; n = 17), and idiopathic autism (IA; n = 15). Relative to controls, the FXS+A (p < 0.002) and FXS-A (p < 0.003) groups had impaired PPI. The FXS+A (p < 0.01) and FXS-A (p < 0.03) groups had lower PPI than the IA group. Prolonged startle latency was seen in the IA group. The differing PPI profiles seen in the FXS+A and IA indicates these groups may not share a common neurobiological abnormality of sensorimotor gating.

Prepulse inhibition (PPI) may useful for exploring the proposed shared neurobiology between idiopathic autism and autism caused by FXS. We compared PPI in four groups: typically developing controls (n = 18), FXS and autism (FXS+A; n = 15), FXS without autism spectrum disorder (FXS-A; n = 17), and idiopathic autism (IA; n = 15). Relative to controls, the FXS+A (p < 0.002) and FXS-A (p < 0.003) groups had impaired PPI. The FXS+A (p < 0.01) and FXS-A (p < 0.03) groups had lower PPI than the IA group. Prolonged startle latency was seen in the IA group. The differing PPI profiles seen in the FXS+A and IA indicates these groups may not share a common neurobiological abnormality of sensorimotor gating.

One of the main difficulties of addiction treatment is the high risk of relapse even after a longabstinence and fully detoxification. Therefore, discovering the underlying molecular principlesof relapse is essential. The metabotropic glutamate receptor, mGluR5, is considered to beinvolved in this aspect. One of the brain structures expressing mGluR5 is the striatum, an areawith well-established role in addiction which is largely composed of medium-sized spinyneurons (MSNs). These neurons are basically divided into two major subpopulationscharacterized based on their projections and protein properties. It is known that the mGluR5receptor is expressed on both subpopulations of MSNs. Consequently, it can be used to establishthe proportional contribution of each of MSNs subpopulations in relapse to addiction. In ourconstellation, we have generated a mouse line designed to have a selective mGluR5 knock-downin one of these subpopulations – the dopamine D1 receptor (D1R) expressing neurons. It hashowever been unclear if the expression of the transgene is indeed limited to only D1R-expressingneurons. By immunofluorescence technique, I here show that the construct is expressed only inMSNs and is restricted to the D1R-expressing cell population in the striatum. Thus the transgenicmouse line is a good tool for the study of mGluR5 selectively in D1R expressing neurons.

One of the main difficulties of addiction treatment is the high risk of relapse even after a longabstinence and fully detoxification. Therefore, discovering the underlying molecular principlesof relapse is essential. The metabotropic glutamate receptor, mGluR5, is considered to beinvolved in this aspect. One of the brain structures expressing mGluR5 is the striatum, an areawith well-established role in addiction which is largely composed of medium-sized spinyneurons (MSNs). These neurons are basically divided into two major subpopulationscharacterized based on their projections and protein properties. It is known that the mGluR5receptor is expressed on both subpopulations of MSNs. Consequently, it can be used to establishthe proportional contribution of each of MSNs subpopulations in relapse to addiction. In ourconstellation, we have generated a mouse line designed to have a selective mGluR5 knock-downin one of these subpopulations – the dopamine D1 receptor (D1R) expressing neurons. It hashowever been unclear if the expression of the transgene is indeed limited to only D1R-expressingneurons. By immunofluorescence technique, I here show that the construct is expressed only inMSNs and is restricted to the D1R-expressing cell population in the striatum. Thus the transgenicmouse line is a good tool for the study of mGluR5 selectively in D1R expressing neurons.

Future therapies for diseases associated with altered dopaminergic signaling, including Parkinson's disease, schizophrenia and drug addiction or drug dependence may substantially build on the existence of intramembrane receptor-receptor interactions within dopamine receptor containing receptor mosaics (RM; dimeric or high-order receptor oligomers) where it is believed that the dopamine D-2 receptor may operate as the 'hub receptor' within these complexes. The constitutive adenosine A(2A)/dopamine D-2 RM, located in the dorsal striatopallidal GABA neurons, are of particular interest in view of the demonstrated antagonistic A(2A)/D-2 interaction within these heteromers; an interaction that led to the suggestion and later demonstration that A(2A) antagonists could be used as novel anti-Parkinsonian drugs. Based on the likely existence of A(2A)/D-2/mGluR5 RM located both extrasynaptically on striato-pallidal GABA neurons and on cortico-striatal glutamate terminals, multiple receptor-receptor interactions within this RM involving synergism between A(2A)/mGluR5 to counteract D-2 signaling, has led to the proposal of using combined mGluR5 and A(2A) antagonists as a future anti-Parkinsonian treatment. Based on the same RM in the ventral striato-pallidal GABA pathways, novel strategies for the treatment of schizophrenia, building on the idea that A(2A) agonists and/or mGluR5 agonists will help reduce the increased dopaminergic signaling associated with this disease, have been suggested. Such treatment may ensure the proper glutamatergic drive from the mediodorsal thalamic nucleus to the prefrontal cortex, one which is believed to be reduced in schizophrenia due to a dominance of D-2-like signaling in the ventral striatum. Recently, A(2A) receptors also have been shown to counteract the locomotor and sensitizing actions of cocaine and increases in A(2A) receptors have also been observed in the nucleus accumbens after extended cocaine self-administration, probably representing, authorCount :15

Endogenous cannabinoids (endocannabinoids) mediate retrograde signals for short- and long-term suppression of transmitter release at synapses of striatal medium spiny (MS) neurons. An endocannabinoid, 2-arachidonoyl-glycerol (2-AG), is synthesized from diacylglycerol (DAG) after membrane depolarization and Gq-coupled receptor activation. To understand 2-AG-mediated retrograde signaling in the striatum, we determined precise subcellular distributions of the synthetic enzyme of 2-AG, DAG lipase-alpha (DAGLalpha), and its upstream metabotropic glutamate receptor 5 (mGluR5) and muscarinic acetylcholine receptor 1 (M1). DAGLalpha, mGluR5, and M1 were all richly distributed on the somatodendritic surface of MS neurons, but their subcellular distributions were different. Although mGluR5 and DAGLalpha levels were highest in spines and accumulated in the perisynaptic region, M1 level was lowest in spines and was rather excluded from the mGluR5-rich perisynaptic region. These subcellular arrangements suggest that mGluR5 and M1 might differentially affect endocannabinoid-mediated, depolarization-induced suppression of inhibition (DSI) and depolarization-induced suppression of excitation (DSE) in MS neurons. Indeed, mGluR5 activation enhanced both DSI and DSE, whereas M1 activation enhanced DSI only. Importantly, DSI, DSE, and receptor-driven endocannabinoid-mediated suppression were all abolished by the DAG lipase inhibitor tetrahydrolipstatin, indicating 2-AG as the major endocannabinoid mediating retrograde suppression at excitatory and inhibitory synapses of MS neurons. Accordingly, CB1 cannabinoid receptor, the main target of 2-AG, was present at high levels on GABAergic axon terminals of MS neurons and parvalbumin-positive interneurons and at low levels on excitatory corticostriatal afferents. Thus, endocannabinoid signaling molecules are arranged to modulate the excitability of the MS neuron effectively depending on cortical activity and cholinergic tone as measured by m

Although metabotropic glutamate receptor 5 (mGluR5) is essential for cocaine self-administration and drug-seeking behavior, there is limited knowledge of the cellular actions of this receptor in the nucleus accumbens (NAc). Although mGluR5 has the potential to regulate neurons directly, recent studies have shown the importance of mGluR5 in regulating Ca(2+) signaling in astrocytes and, as a consequence, the Ca(2+)-dependent release of excitatory transmitters from these glia. In this study, we demonstrate that activation of mGluR5 induces Ca(2+) oscillations in NAc astrocytes with the correlated appearance of NMDA receptor-dependent slow inward currents detected in medium spiny neurons (MSNs). Photolysis of caged Ca(2+) loaded specifically into astrocytes evoked slow inward currents demonstrating that Ca(2+) elevations in astrocytes are responsible for these excitatory events. Pharmacological evaluation of these glial-evoked NMDA currents shows that they are mediated by NR2B-containing NMDA receptors, whereas synaptic NMDA receptors rely on NR2A-containing receptors. Stimulation of glutamatergic afferents activates mGluR5-dependent astrocytic Ca(2+) oscillations and gliotransmission that is sustained for minutes beyond the initial stimulus. Because gliotransmission is mediated by NMDA receptors, depolarized membrane potentials exhibited during up-states augment excitation provided by gliotransmission, which drives bursts of MSN action potentials. Because the predominant mGluR5-dependent action of glutamatergic afferents is to cause the sustained activation of astrocytes, which in turn excite MSNs through extrasynaptic NMDA receptors, our results raise the potential for gliotransmission being involved in prolonged mGluR5-dependent adaptation in the NAc.

Endogenous cannabinoids (endocannabinoids) mediate retrograde signals for short- and long-term suppression of transmitter release at synapses of striatal medium spiny (MS) neurons. An endocannabinoid, 2-arachidonoyl-glycerol (2-AG), is synthesized from diacylglycerol (DAG) after membrane depolarization and Gq-coupled receptor activation. To understand 2-AG-mediated retrograde signaling in the striatum, we determined precise subcellular distributions of the synthetic enzyme of 2-AG, DAG lipase-alpha (DAGLalpha), and its upstream metabotropic glutamate receptor 5 (mGluR5) and muscarinic acetylcholine receptor 1 (M1). DAGLalpha, mGluR5, and M1 were all richly distributed on the somatodendritic surface of MS neurons, but their subcellular distributions were different. Although mGluR5 and DAGLalpha levels were highest in spines and accumulated in the perisynaptic region, M1 level was lowest in spines and was rather excluded from the mGluR5-rich perisynaptic region. These subcellular arrangements suggest that mGluR5 and M1 might differentially affect endocannabinoid-mediated, depolarization-induced suppression of inhibition (DSI) and depolarization-induced suppression of excitation (DSE) in MS neurons. Indeed, mGluR5 activation enhanced both DSI and DSE, whereas M1 activation enhanced DSI only. Importantly, DSI, DSE, and receptor-driven endocannabinoid-mediated suppression were all abolished by the DAG lipase inhibitor tetrahydrolipstatin, indicating 2-AG as the major endocannabinoid mediating retrograde suppression at excitatory and inhibitory synapses of MS neurons. Accordingly, CB1 cannabinoid receptor, the main target of 2-AG, was present at high levels on GABAergic axon terminals of MS neurons and parvalbumin-positive interneurons and at low levels on excitatory corticostriatal afferents. Thus, endocannabinoid signaling molecules are arranged to modulate the excitability of the MS neuron effectively depending on cortical activity and cholinergic tone as measured by m

Endocannabinoids play central roles in retrograde signaling at a wide variety of synapses throughout the CNS. Although several molecular components of the endocannabinoid system have been identified recently, their precise location and contribution to retrograde synaptic signaling is essentially unknown. Here we show, by using two independent riboprobes, that principal cell populations of the hippocampus express high levels of diacylglycerol lipase alpha (DGL-alpha), the enzyme involved in generation of the endocannabinoid 2-arachidonoyl-glycerol (2-AG). Immunostaining with two independent antibodies against DGL-alpha revealed that this lipase was concentrated in heads of dendritic spines throughout the hippocampal formation. Furthermore, quantification of high-resolution immunoelectron microscopic data showed that this enzyme was highly compartmentalized into a wide perisynaptic annulus around the postsynaptic density of axospinous contacts but did not occur intrasynaptically. On the opposite side of the synapse, the axon terminals forming these excitatory contacts were found to be equipped with presynaptic CB1 cannabinoid receptors. This precise anatomical positioning suggests that 2-AG produced by DGL-alpha on spine heads may be involved in retrograde synaptic signaling at glutamatergic synapses, whereas CB1 receptors located on the afferent terminals are in an ideal position to bind 2-AG and thereby adjust presynaptic glutamate release as a function of postsynaptic activity. We propose that this molecular composition of the endocannabinoid system may be a general feature of most glutamatergic synapses throughout the brain and may contribute to homosynaptic plasticity of excitatory synapses and to heterosynaptic plasticity between excitatory and inhibitory contacts.

Endocannabinoids play central roles in retrograde signaling at a wide variety of synapses throughout the CNS. Although several molecular components of the endocannabinoid system have been identified recently, their precise location and contribution to retrograde synaptic signaling is essentially unknown. Here we show, by using two independent riboprobes, that principal cell populations of the hippocampus express high levels of diacylglycerol lipase alpha (DGL-alpha), the enzyme involved in generation of the endocannabinoid 2-arachidonoyl-glycerol (2-AG). Immunostaining with two independent antibodies against DGL-alpha revealed that this lipase was concentrated in heads of dendritic spines throughout the hippocampal formation. Furthermore, quantification of high-resolution immunoelectron microscopic data showed that this enzyme was highly compartmentalized into a wide perisynaptic annulus around the postsynaptic density of axospinous contacts but did not occur intrasynaptically. On the opposite side of the synapse, the axon terminals forming these excitatory contacts were found to be equipped with presynaptic CB1 cannabinoid receptors. This precise anatomical positioning suggests that 2-AG produced by DGL-alpha on spine heads may be involved in retrograde synaptic signaling at glutamatergic synapses, whereas CB1 receptors located on the afferent terminals are in an ideal position to bind 2-AG and thereby adjust presynaptic glutamate release as a function of postsynaptic activity. We propose that this molecular composition of the endocannabinoid system may be a general feature of most glutamatergic synapses throughout the brain and may contribute to homosynaptic plasticity of excitatory synapses and to heterosynaptic plasticity between excitatory and inhibitory contacts.

Endocannabinoids play central roles in retrograde signaling at a wide variety of synapses throughout the CNS. Although several molecular components of the endocannabinoid system have been identified recently, their precise location and contribution to retrograde synaptic signaling is essentially unknown. Here we show, by using two independent riboprobes, that principal cell populations of the hippocampus express high levels of diacylglycerol lipase alpha (DGL-alpha), the enzyme involved in generation of the endocannabinoid 2-arachidonoyl-glycerol (2-AG). Immunostaining with two independent antibodies against DGL-alpha revealed that this lipase was concentrated in heads of dendritic spines throughout the hippocampal formation. Furthermore, quantification of high-resolution immunoelectron microscopic data showed that this enzyme was highly compartmentalized into a wide perisynaptic annulus around the postsynaptic density of axospinous contacts but did not occur intrasynaptically. On the opposite side of the synapse, the axon terminals forming these excitatory contacts were found to be equipped with presynaptic CB1 cannabinoid receptors. This precise anatomical positioning suggests that 2-AG produced by DGL-alpha on spine heads may be involved in retrograde synaptic signaling at glutamatergic synapses, whereas CB1 receptors located on the afferent terminals are in an ideal position to bind 2-AG and thereby adjust presynaptic glutamate release as a function of postsynaptic activity. We propose that this molecular composition of the endocannabinoid system may be a general feature of most glutamatergic synapses throughout the brain and may contribute to homosynaptic plasticity of excitatory synapses and to heterosynaptic plasticity between excitatory and inhibitory contacts., Journal Article, Research Support, N.I.H. Extramural, Research Support, Non-U.S. Gov't, info:eu-repo/semantics/published

Endocannabinoids play central roles in retrograde signaling at a wide variety of synapses throughout the CNS. Although several molecular components of the endocannabinoid system have been identified recently, their precise location and contribution to retrograde synaptic signaling is essentially unknown. Here we show, by using two independent riboprobes, that principal cell populations of the hippocampus express high levels of diacylglycerol lipase alpha (DGL-alpha), the enzyme involved in generation of the endocannabinoid 2-arachidonoyl-glycerol (2-AG). Immunostaining with two independent antibodies against DGL-alpha revealed that this lipase was concentrated in heads of dendritic spines throughout the hippocampal formation. Furthermore, quantification of high-resolution immunoelectron microscopic data showed that this enzyme was highly compartmentalized into a wide perisynaptic annulus around the postsynaptic density of axospinous contacts but did not occur intrasynaptically. On the opposite side of the synapse, the axon terminals forming these excitatory contacts were found to be equipped with presynaptic CB1 cannabinoid receptors. This precise anatomical positioning suggests that 2-AG produced by DGL-alpha on spine heads may be involved in retrograde synaptic signaling at glutamatergic synapses, whereas CB1 receptors located on the afferent terminals are in an ideal position to bind 2-AG and thereby adjust presynaptic glutamate release as a function of postsynaptic activity. We propose that this molecular composition of the endocannabinoid system may be a general feature of most glutamatergic synapses throughout the brain and may contribute to homosynaptic plasticity of excitatory synapses and to heterosynaptic plasticity between excitatory and inhibitory contacts.

Endocannabinoids play central roles in retrograde signaling at a wide variety of synapses throughout the CNS. Although several molecular components of the endocannabinoid system have been identified recently, their precise location and contribution to retrograde synaptic signaling is essentially unknown. Here we show, by using two independent riboprobes, that principal cell populations of the hippocampus express high levels of diacylglycerol lipase alpha (DGL-alpha), the enzyme involved in generation of the endocannabinoid 2-arachidonoyl-glycerol (2-AG). Immunostaining with two independent antibodies against DGL-alpha revealed that this lipase was concentrated in heads of dendritic spines throughout the hippocampal formation. Furthermore, quantification of high-resolution immunoelectron microscopic data showed that this enzyme was highly compartmentalized into a wide perisynaptic annulus around the postsynaptic density of axospinous contacts but did not occur intrasynaptically. On the opposite side of the synapse, the axon terminals forming these excitatory contacts were found to be equipped with presynaptic CB1 cannabinoid receptors. This precise anatomical positioning suggests that 2-AG produced by DGL-alpha on spine heads may be involved in retrograde synaptic signaling at glutamatergic synapses, whereas CB1 receptors located on the afferent terminals are in an ideal position to bind 2-AG and thereby adjust presynaptic glutamate release as a function of postsynaptic activity. We propose that this molecular composition of the endocannabinoid system may be a general feature of most glutamatergic synapses throughout the brain and may contribute to homosynaptic plasticity of excitatory synapses and to heterosynaptic plasticity between excitatory and inhibitory contacts., Journal Article, Research Support, N.I.H. Extramural, Research Support, Non-U.S. Gov't, info:eu-repo/semantics/published

Slow Ca2(+) oscillations caused by release from intracellular stores have been observed in neurons in the lamprey spinal cord. These oscillations are triggered by activation of metabotropic glutamate receptors on the cell surface. The pathway leading from receptor activation to the inositol triphosphate-mediated release of Ca2(+) from the endoplasmatic reticulum has been modelled in order to facilitate further understanding of the nature of these oscillations. The model generates Ca2(+) oscillations with a frequency range of 0.01-0.09 Hz. A prediction of the model is that the frequency will increase with a stronger extracellular glutamate signal., QC 20111019

Slow Ca2+ oscillations caused by release from intracellular stores have been observed in neurons in the lamprey spinal cord. These oscillations are triggered by activation of metabotropic glutamate receptors on the cell surface. The pathway leading from receptor activation to the inositol triphosphate-mediated release of Ca2+ from the endoplasmatic reticulum has been modelled in order to facilitate further understanding of the nature of these oscillations. The model generates Ca2+ oscillations with a frequency range of 0.01–0.09 Hz. A prediction of the model is that the frequency will increase with a stronger extracellular glutamate signal.