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Abstract Frailty is a common age-related clinical syndrome characterized by a decline in the function of multiple organ systems, increased vulnerability to stressors, and a huge socio-economic burden. Despite recent research efforts, the physiopathological mechanisms underlying frailty remain elusive and biomarkers able to predate its occurrence in the early stages are still lacking. Beyond its physical component, cognitive decline represents a critical domain of frailty associated with higher risk of adverse health outcomes. We measured by High-Performance Liquid Chromatography (HPLC) a pool of serum amino acids including L-glutamate, L-aspartate, glycine, and D-serine, as well as their precursors L-glutamine, L-asparagine, and L-serine in a cohort of elderly subjects encompassing the entire continuum from fitness to frailty. These amino acids are known to orchestrate excitatory and inhibitory neurotransmission, and in turn, to play a key role as intermediates of energy homeostasis and in liver, kidney, muscle, and immune system metabolism. To comprehensively assess frailty, we employed both the Edmonton Frail Scale (EFS), as a practical tool to capture the multidimensionality of frailty, and the frailty phenotype, as a measure of physical function. We found that D-serine and D-/Total serine ratio were independent predictors of EFS but not of physical frailty. Furthermore, higher levels of glycine, glycine/L-serine and D-/Total serine were associated with worse cognition and depressive symptoms in the frail group. These findings suggest that changes in peripheral glycine and serine enantiomers homeostasis may represent a novel biochemical correlate of frailty.

IntroductionSchizophrenia (SCZ) and autism spectrum disorder (ASD) are neurodevelopmental diseases characterized by different psychopathological manifestations and divergent clinical trajectories. Various alterations at glutamatergic synapses have been reported in both disorders, including abnormal NMDA and metabotropic receptor signaling.MethodsWe conducted a bicentric study to assess the blood serum levels of NMDA receptors-related glutamatergic amino acids and their precursors, including L-glutamate, L-glutamine, D-aspartate, L-aspartate, L-asparagine, D-serine, L-serine and glycine, in ASD, SCZ patients and their respective control subjects. Specifically, the SCZ patients were subdivided into treatment-resistant and non-treatment-resistant SCZ patients, based on their responsivity to conventional antipsychotics.ResultsD-serine and D-aspartate serum reductions were found in SCZ patients compared to controls. Conversely, no significant differences between cases and controls were found in amino acid concentrations in the two ASD cohorts analyzed.DiscussionThis result further encourages future research to evaluate the predictive role of selected D-amino acids as peripheral markers for SCZ pathophysiology and diagnosis.

We recently described increased D- and L-serine concentrations in the striatum of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated monkeys, the post-mortem caudate-putamen of human Parkinson's disease (PD) brains and the cerebrospinal fluid (CSF) of de novo living PD patients. However, data regarding blood D- and L-serine levels in PD are scarce. Here, we investigated whether the serum profile of D- and L-serine, as well as the other glutamate N-methyl-D-aspartate ionotropic receptor (NMDAR)-related amino acids, (i) differs between PD patients and healthy controls (HC) and (ii) correlates with clinical-demographic features and levodopa equivalent daily dose (LEDD) in PD. Eighty-three consecutive PD patients and forty-one HC were enrolled. PD cohort underwent an extensive clinical characterization. Serum levels of D- and L-serine, L-glutamate, L-glutamine, L-aspartate, L-asparagine and glycine were determined using High Performance Liquid Chromatography.In age- and sex-adjusted analyses, no differences emerged in the serum levels of D-serine, L-serine and other NMDAR-related amino acids between PD and HC. However, we found that D-serine and D−/Total serine ratio positively correlated with age in PD but not in HC, and also with PD age at onset. Moreover, we found that higher LEDD correlated with lower levels of D-serine and the other excitatory amino acids. Following these results, the addition of LEDD as covariate in the analyses disclosed a selective significant increase of D-serine in PD compared to HC (Δ ≈ 38%). Overall, these findings suggest that serum D-serine and D−/Total serine may represent a valuable biochemical signature of PD.

Abstract Beyond motor neuron degeneration, homozygous mutations in the survival motor neuron 1 (SMN1) gene cause multiorgan and metabolic defects in patients with spinal muscular atrophy (SMA). However, the precise biochemical features of these alterations and the age of onset in the brain and peripheral organs remain unclear. Using untargeted NMR-based metabolomics in SMA mice, we identify cerebral and hepatic abnormalities related to energy homeostasis pathways and amino acid metabolism, emerging already at postnatal day 3 (P3) in the liver. Through HPLC, we find that SMN deficiency induces a drop in cerebral norepinephrine levels in overt symptomatic SMA mice at P11, affecting the mRNA and protein expression of key genes regulating monoamine metabolism, including aromatic L-amino acid decarboxylase (AADC), dopamine beta-hydroxylase (DβH) and monoamine oxidase A (MAO-A). In support of the translational value of our preclinical observations, we also discovered that SMN upregulation increases cerebrospinal fluid norepinephrine concentration in Nusinersen-treated SMA1 patients. Our findings highlight a previously unrecognized harmful influence of low SMN levels on the expression of critical enzymes involved in monoamine metabolism, suggesting that SMN-inducing therapies may modulate catecholamine neurotransmission. These results may also be relevant for setting therapeutic approaches to counteract peripheral metabolic defects in SMA.

Physiology and behavior are structured temporally to anticipate daily cycles of light and dark, ensuring fitness and survival. Neuromodulatory systems in the brain—including those involving serotonin and dopamine—exhibit daily oscillations in neural activity and help shape circadian rhythms. Disrupted neuromodulation can cause circadian abnormalities that are thought to underlie several neuropsychiatric disorders, including bipolar mania and schizophrenia, for which a mechanistic understanding is still lacking. Here, we show that genetically depleting serotonin in Tph2 knockout mice promotes manic-like behaviors and disrupts daily oscillations of the dopamine biosynthetic enzyme tyrosine hydroxylase (TH) in midbrain dopaminergic nuclei. Specifically, while TH mRNA and protein levels in the Substantia Nigra (SN) and Ventral Tegmental Area (VTA) of wild-type mice doubled between the light and dark phase, TH levels were high throughout the day in Tph2 knockout mice, suggesting a hyperdopaminergic state. Analysis of TH expression in striatal terminal fields also showed blunted rhythms. Additionally, we found low abundance and blunted rhythmicity of the neuropeptide cholecystokinin (Cck) in the VTA of knockout mice, a neuropeptide whose downregulation has been implicated in manic-like states in both rodents and humans. Altogether, our results point to a previously unappreciated serotonergic control of circadian dopamine signaling and propose serotonergic dysfunction as an upstream mechanism underlying dopaminergic deregulation and ultimately maladaptive behaviors.

Nuzzo, Russo, Errico, D’Amico et al. investigate neuroinflammation in forty-eight pediatric spinal muscular atrophy patients before and after Nusinersen treatment. They find signatures of neuroinflammation that are specifically associated with severe disease and show that Nusinersen therapy has neuro-immunomodulatory effects.

L-serine generated in astrocytes plays a pivotal role in modulating essential neurometabolic processes, while its enantiomer, D-serine, specifically regulates NMDA receptor (NMDAR) signalling. Despite their physiological relevance in modulating cerebral activity, serine enantiomers metabolism in Parkinson's disease (PD) remains elusive. Using High-Performance Liquid Chromatography (HPLC), we measured D- and L-serine levels along with other amino acids known to modulate NMDAR function, such as L-glutamate, L-aspartate, D-aspartate, and glycine, in the post-mortem caudate putamen (CPu) and superior frontal gyrus (SFG) of PD patients. Moreover, we examined these amino acids in the cerebrospinal fluid (CSF) of de novo living PD, Alzheimer's disease (AD), and amyotrophic lateral sclerosis (ALS) patients versus subjects with other neurological disorders (OND), used as control. We found higher D-serine and L-serine levels in the CPu of PD patients but not in the SFG, a cerebral region that, in contrast to the CPu, is not innervated by nigral dopaminergic terminals. We also highlighted a significant elevation of both serine enantiomers in the CSF samples from PD but not in those of AD and ALS patients, compared with control subjects. By contrast, none or only minor changes were found in the amount of other NMDAR modulating amino acids. Our findings identify D-serine and L-serine level upregulation as a biochemical signature associated with nigrostriatal dopaminergic degeneration in PD.

Loss of dopaminergic midbrain neurons perturbs l-serine and d-serine homeostasis in the post-mortem caudate putamen (CPu) of Parkinson's disease (PD) patients. However, it is unclear whether the severity of dopaminergic nigrostriatal degeneration plays a role in deregulating serine enantiomers' metabolism. Here, through high-performance liquid chromatography (HPLC), we measured the levels of these amino acids in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated monkeys and MPTP-plus-probenecid (MPTPp)-treated mice to determine whether and how dopaminergic midbrain degeneration affects the levels of serine enantiomers in various basal ganglia subregions. In addition, in the same brain regions, we measured the levels of key neuroactive amino acids modulating glutamatergic neurotransmission, including l-glutamate, glycine, l-aspartate, d-aspartate, and their precursors l-glutamine, l-asparagine. In monkeys, MPTP treatment produced severe denervation of nigrostriatal dopaminergic fibers (⁓75%) and increased the levels of serine enantiomers in the rostral putamen (rPut), but not in the subthalamic nucleus, and the lateral and medial portion of the globus pallidus. Moreover, this neurotoxin significantly reduced the protein expression of the astrocytic serine transporter ASCT1 and the glycolytic enzyme GAPDH in the rPut of monkeys. Conversely, concentrations of d-serine and l-serine, as well as ASCT1 and GAPDH expression were unaffected in the striatum of MPTPp-treated mice, which showed only mild dopaminergic degeneration (⁓30%). These findings unveil a link between the severity of dopaminergic nigrostriatal degeneration and striatal serine enantiomers concentration, ASCT1 and GAPDH expression. We hypothesize that the up-regulation of d-serine and l-serine levels occurs as a secondary response within a homeostatic loop to support the metabolic and neurotransmission demands imposed by the degeneration of dopaminergic neurons.

Abstract The D-aspartate oxidase (DDO) gene encodes the enzyme responsible for the catabolism of D-aspartate, an atypical amino acid enriched in the mammalian brain and acting as an endogenous NMDA receptor agonist. Considering the key role of NMDA receptors in neurodevelopmental disorders, recent findings suggest a link between D-aspartate dysmetabolism and schizophrenia. To clarify the role of D-aspartate on brain development and functioning, we used a mouse model with constitutive Ddo overexpression and D-aspartate depletion. In these mice, we found reduced number of BrdU-positive dorsal pallium neurons during corticogenesis, and decreased cortical and striatal gray matter volume at adulthood. Brain abnormalities were associated with social recognition memory deficit at juvenile phase, suggesting that early D-aspartate occurrence influences neurodevelopmental related phenotypes. We corroborated this hypothesis by reporting the first clinical case of a young patient with severe intellectual disability, thought disorders and autism spectrum disorder symptomatology, harboring a duplication of a chromosome 6 region, including the entire DDO gene.

Abstract Schizophrenia is a disorder of synaptic plasticity and aberrant connectivity in which a major dysfunction in glutamate synapse has been suggested. However, a multi-level approach tackling diverse clusters of interacting molecules of the glutamate signaling in schizophrenia is still lacking. We investigated in the post-mortem dorsolateral prefrontal cortex (DLPFC) and hippocampus of schizophrenia patients and non-psychiatric controls, the levels of neuroactive d- and l-amino acids (l-glutamate, d-serine, glycine, l-aspartate, d-aspartate) by HPLC. Moreover, by quantitative RT-PCR and western blotting we analyzed, respectively, the mRNA and protein levels of pre- and post-synaptic key molecules involved in the glutamatergic synapse functioning, including glutamate receptors (NMDA, AMPA, metabotropic), their interacting scaffolding proteins (PSD-95, Homer1b/c), plasma membrane and vesicular glutamate transporters (EAAT1, EAAT2, VGluT1, VGluT2), enzymes involved either in glutamate-dependent GABA neurotransmitter synthesis (GAD65 and 67), or in post-synaptic NMDA receptor-mediated signaling (CAMKIIα) and the pre-synaptic marker Synapsin-1. Univariable analyses revealed that none of the investigated molecules was differently represented in the post-mortem DLPFC and hippocampus of schizophrenia patients, compared with controls. Nonetheless, multivariable hypothesis-driven analyses revealed that the presence of schizophrenia was significantly affected by variations in neuroactive amino acid levels and glutamate-related synaptic elements. Furthermore, a Machine Learning hypothesis-free unveiled other discriminative clusters of molecules, one in the DLPFC and another in the hippocampus. Overall, while confirming a key role of glutamatergic synapse in the molecular pathophysiology of schizophrenia, we reported molecular signatures encompassing elements of the glutamate synapse able to discriminate patients with schizophrenia and normal individuals.

High levels of free D-aspartate (D-Asp) are present in vertebrate testis during post-natal development, coinciding with the onset of testosterone production, which suggests that this atypical amino acid might participate in the regulation of hormone biosynthesis. To elucidate the unknown role of D-Asp on testicular function, we investigated steroidogenesis and spermatogenesis in a one-month-old knockin mouse model with the constitutive depletion of D-Asp levels due to the targeted overexpression of D-aspartate oxidase (DDO), which catalyzes the deaminative oxidation of D-Asp to generate the corresponding α-keto acid, oxaloacetate, hydrogen peroxide, and ammonium ions. In the Ddo knockin mice, we found a dramatic reduction in testicular D-Asp levels, accompanied by a significant decrease in the serum testosterone levels and testicular 17β-HSD, the enzyme involved in testosterone biosynthesis. Additionally, in the testes of these Ddo knockin mice, the expression of PCNA and SYCP3 proteins decreased, suggesting alterations in spermatogenesis-related processes, as well as an increase in the cytosolic cytochrome c protein levels and TUNEL-positive cell number, which indicate an increase in apoptosis. To further investigate the histological and morphometric testicular alterations in Ddo knockin mice, we analyzed the expression and localization of prolyl endopeptidase (PREP) and disheveled-associated activator of morphogenesis 1 (DAAM1), two proteins involved in cytoskeletal organization. Our results showed that the testicular levels of DAAM1 and PREP in Ddo knockin mice were different from those in wild-type animals, suggesting that the deficiency of D-Asp is associated with overall cytoskeletal disorganization. Our findings confirmed that physiological D-Asp influences testosterone biosynthesis and plays a crucial role in germ cell proliferation and differentiation, which are required for successful reproduction.

Introduction: Schizophrenia (SCZ) and autism spectrum disorder (ASD) are neurodevelopmental diseases characterized by different psychopathological manifestations and divergent clinical trajectories. Various alterations at glutamatergic synapses have been reported in both disorders, including abnormal NMDA and metabotropic receptor signaling. Methods: We conducted a bicentric study to assess the blood serum levels of NMDA receptors-related glutamatergic amino acids and their precursors, including L-glutamate, L-glutamine, D-aspartate, L-aspartate, L-asparagine, Dserine, L-serine and glycine, in ASD, SCZ patients and their respective control subjects. Specifically, the SCZ patients were subdivided into treatment-resistant and non-treatment-resistant SCZ patients, based on their responsivity to conventional antipsychotics. Results: D-serine and D-aspartate serum reductions were found in SCZ patients compared to controls. Conversely, no significant differences between cases and controls were found in amino acid concentrations in the two ASD cohorts analyzed. Discussion: This result further encourages future research to evaluate the predictive role of selected D-amino acids as peripheral markers for SCZ pathophysiology and diagnosis. [ABSTRACT FROM AUTHOR]

Physiology and behavior are structured temporally to anticipate daily cycles of light and dark, ensuring fitness and survival. Neuromodulatory systems in the brain—including those involving serotonin and dopamine—exhibit daily oscillations in neural activity and help shape circadian rhythms. Disrupted neuromodulation can cause circadian abnormalities that are thought to underlie several neuropsychiatric disorders, including bipolar mania and schizophrenia, for which a mechanistic understanding is still lacking. Here, we show that genetically depleting serotonin in Tph2 knockout mice promotes manic-like behaviors and disrupts daily oscillations of the dopamine biosynthetic enzyme tyrosine hydroxylase (TH) in midbrain dopaminergic nuclei. Specifically, while TH mRNA and protein levels in the Substantia Nigra (SN) and Ventral Tegmental Area (VTA) of wild-type mice doubled between the light and dark phase, TH levels were high throughout the day in Tph2 knockout mice, suggesting a hyperdopaminergic state. Analysis of TH expression in striatal terminal fields also showed blunted rhythms. Additionally, we found low abundance and blunted rhythmicity of the neuropeptide cholecystokinin (Cck) in the VTA of knockout mice, a neuropeptide whose downregulation has been implicated in manic-like states in both rodents and humans. Altogether, our results point to a previously unappreciated serotonergic control of circadian dopamine signaling and propose serotonergic dysfunction as an upstream mechanism underlying dopaminergic deregulation and ultimately maladaptive behaviors. [ABSTRACT FROM AUTHOR]

Introduction: Plasticity at corticostriatal synapses is a key substrate for a variety of brain functions – including motor control, learning and reward processing – and is often disrupted in disease conditions. Despite intense research pointing toward a dynamic interplay between glutamate, dopamine (DA), and serotonin (5-HT) neurotransmission, their precise circuit and synaptic mechanisms regulating their role in striatal plasticity are still unclear. Here, we analyze the role of serotonergic raphe-striatal innervation in the regulation of DA-dependent corticostriatal plasticity. Methods: Mice (males and females, 2–6 months of age) were housed in standard plexiglass cages at constant temperature (22 ± 1°C) and maintained on a 12/12h light/dark cycle with food and demineralized water ad libitum. In the present study, we used a knock-in mouse line in which the green fluorescent protein reporter gene (GFP) replaced the I Tph2 exon (Tph2GFP mice), allowing selective expression of GFP in the whole 5-HT system, highlighting both somata and neuritis of serotonergic neurons. Heterozygous, Tph2+/GFP, mice were intercrossed to obtain experimental cohorts, which included Wild-type (Tph2+/+), Heterozygous (Tph2+/GFP), and Mutant serotonin-depleted (Tph2GFP/GFP) animals. Results: Using male and female mice, carrying on different Tph2 gene dosages, we show that Tph2 gene modulation results in sex-specific corticostriatal abnormalities, encompassing the abnormal amplitude of spontaneous glutamatergic transmission and the loss of Long Term Potentiation (LTP) in Tph2GFP/GFP mice of both sexes, while this form of plasticity is normally expressed in control mice (Tph2+/+). Once LTP is induced, only the Tph2+/GFP female mice present a loss of synaptic depotentiation. Conclusion: We showed a relevant role of the interaction between dopaminergic and serotonergic systems in controlling striatal synaptic plasticity. Overall, our data unveil that 5-HT plays a primary role in regulating DA-dependent corticostriatal plasticity in a sex-related manner and propose altered 5-HT levels as a critical determinant of disease-associated plasticity defects.

Intrathecal delivery of Nusinersen–an antisense oligonucleotide that promotes survival motor neuron (SMN) protein induction–is an approved therapy for spinal muscular atrophy (SMA). Here, we employed nuclear magnetic resonance (NMR) spectroscopy to longitudinally characterize the unknown metabolic effects of Nusinersen in the cerebrospinal fluid (CSF) of SMA patients across disease severity. Modulation of amino acid metabolism is a common denominator of biochemical changes induced by Nusinersen, with distinct downstream metabolic effects according to disease severity. In severe SMA1 patients, Nusinersen stimulates energy-related glucose metabolism. In intermediate SMA2 patients, Nusinersen effects are also related to energy homeostasis but involve ketone body and fatty acid biosynthesis. In milder SMA3 patients, Nusinersen mainly modulates amino acid metabolism. Moreover, Nusinersen modifies the CSF metabolome of a more severe clinical group towards the profile of untreated SMA patients with milder disease. These findings reveal disease severity-specific neurometabolic signatures of Nusinersen treatment, suggesting a selective modulation of peripheral organ metabolism by this CNS-directed therapy in severe SMA patients.

Levodopa (L-DOPA) treatment is the main gold-standard therapy for Parkinson disease (PD). Besides good antiparkinsonian effects, prolonged use of this drug is associated to the development of involuntary movements known as L-DOPA-induced dyskinesia (LID). L-DOPA-induced dyskinesia is linked to a sensitization of dopamine (DA) D1 receptors located on spiny projection neurons (SPNs) of the dorsal striatum. Several evidences have shown that the emergence of LID can be related to striatal D1/cAMP/PKA/DARPP-32 and extracellular signal-regulated kinases (ERK1/2) pathway overactivation associated to aberrant N-methyl-d-aspartate (NMDA) receptor function. In addition, within striatum, ERK1/2 is also able to modulate in a D1 receptor-dependent manner the activity of the mammalian target of rapamycin complex 1 (mTORC1) pathway under DA depletion and L-DOPA therapy. Consistently, increased mTORC1 signaling appears during chronic administration of L-DOPA and shows a high correlation with the severity of dyskinesia. Furthermore, the abnormal activation of the D1/PKA/DARPP-32 cascade is paralleled by increased phosphorylation of the GluA1 subunit of the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor at the PKA Ser845 site. The GluA1 promotes excitatory AMPA receptor-mediated transmission and may be implicated in the alterations found at the corticostriatal synapses of dyskinetic animals. In our study, we investigated the role of mTORC1 pathway activation in modulating bidirectional striatal synaptic plasticity in L-DOPA-treated parkinsonian rats. Inhibition of mTORC1 by coadministration of rapamycin to L-DOPA was able to limit the magnitude of LID expression, accounting for a therapeutic effect of this drug. In particular, behavioral data showed that, in L-DOPA-treated rats, rapamycin administration induced a selective decrease of distinct components of abnormal involuntary movements (i.e., axial and orolingual dyskinesia). Furthermore, ex vivo patch clamp and intracellular recordings of SPNs revealed that pharmacological inhibition of mTORC1 also resulted associated with a physiological bidirectional plasticity, when compared to dyskinetic rats treated with L-DOPA alone. This study uncovers the important role of mTORC1 inhibition to prevent the loss of striatal bidirectional plasticity under chronic L-DOPA treatment in rodent models of PD.

Schizophrenia (SCZ) is a polygenic severe mental illness. Genome-wide association studies (GWAS) have detected genomic variants associated with this psychiatric disorder and pathway analyses have indicated immune system and dopamine signaling as core components of risk in dorsolateral-prefrontal cortex (DLPFC) and hippocampus, but the mechanistic links remain unknown. The RasGRP1 gene, encoding for a guanine nucleotide exchange factor, is implicated in dopamine signaling and immune response. RasGRP1 has been identified as a candidate risk gene for SCZ and autoimmune disease, therefore representing a possible point of convergence between mechanisms involving the nervous and the immune system. Here, we investigated RasGRP1 mRNA and protein expression in post-mortem DLPFC and hippocampus of SCZ patients and healthy controls, along with RasGRP1 protein content in the serum of an independent cohort of SCZ patients and control subjects. Differences in RasGRP1 expression between SCZ patients and controls were detected both in DLPFC and peripheral blood of samples analyzed. Our results indicate RasGRP1 may mediate risk for SCZ by involving DLPFC and peripheral blood, thus encouraging further studies to explore its possible role as a biomarker of the disease and/or a target for new medication.

Schizophrenia (SCZ) is a mental illness characterized by aberrant synaptic plasticity and connectivity. A large bulk of evidence suggests genetic and functional links between postsynaptic abnormalities and SCZ. Here, we performed quantitative PCR and Western blotting analysis in the dorsolateral prefrontal cortex (DLPFC) and hippocampus of SCZ patients to investigate the mRNA and protein expression of three key spine shapers: the actin-binding protein cyclase-associated protein 2 (CAP2), the sheddase a disintegrin and metalloproteinase 10 (ADAM10), and the synapse-associated protein 97 (SAP97). Our analysis of the SCZ post-mortem brain indicated increased DLG1 mRNA in DLPFC and decreased CAP2 mRNA in the hippocampus of SCZ patients, compared to non-psychiatric control subjects, while the ADAM10 transcript was unaffected. Conversely, no differences in CAP2, SAP97, and ADAM10 protein levels were detected between SCZ and control individuals in both brain regions. To assess whether DLG1 and CAP2 transcript alterations were selective for SCZ, we also measured their expression in the superior frontal gyrus of patients affected by neurodegenerative disorders, like Parkinson’s and Alzheimer’s disease. Interestingly, also in Parkinson’s disease patients, we found a selective reduction of CAP2 mRNA levels relative to controls but unaltered protein levels. Taken together, we reported for the first time altered CAP2 expression in the brain of patients with psychiatric and neurological disorders, thus suggesting that aberrant expression of this gene may contribute to synaptic dysfunction in these neuropathologies.

N-methyl-d-aspartate receptor (NMDAR) subunit composition strictly commands receptor function and pharmacological responses. Changes in NMDAR subunit composition have been documented in brain disorders such as Parkinson's disease (PD) and levodopa (L-DOPA)-induced dyskinesias (LIDs), where an increase of NMDAR GluN2A/GluN2B subunit ratio at striatal synapses has been observed. A therapeutic approach aimed at rebalancing NMDAR synaptic composition represents a valuable strategy for PD and LIDs. To this, the comprehension of the molecular mechanisms regulating the synaptic localization of different NMDAR subtypes is required.We have recently demonstrated that Rabphilin 3A (Rph3A) is a new binding partner of NMDARs containing the GluN2A subunit and that it plays a crucial function in the synaptic stabilization of these receptors. Considering that protein-protein interactions govern the synaptic retention of NMDARs, the purpose of this work was to analyse the role of Rph3A and Rph3A/NMDAR complex in PD and LIDs, and to modulate Rph3A/GluN2A interaction to counteract the aberrant motor behaviour associated to chronic L-DOPA administration. Thus, an array of biochemical, immunohistochemical and pharmacological tools together with electron microscopy were applied in this study. Here we found that Rph3A is localized at the striatal postsynaptic density where it interacts with GluN2A. Notably, Rph3A expression at the synapse and its interaction with GluN2A-containing NMDARs were increased in parkinsonian rats displaying a dyskinetic profile. Acute treatment of dyskinetic animals with a cell-permeable peptide able to interfere with Rph3A/GluN2A binding significantly reduced their abnormal motor behaviour.Altogether, our findings indicate that Rph3A activity is linked to the aberrant synaptic localization of GluN2A-expressing NMDARs characterizing LIDs. Thus, we suggest that Rph3A/GluN2A complex could represent an innovative therapeutic target for those pathological conditions where NMDAR composition is significantly altered.

NMDA receptor: Enzyme breaks down ion channel activator in schizophrenic brain Altered metabolism of an amino acid activator of ion channels in the brain could explain dysfunctional nerve signaling in schizophrenia. Researchers in Italy led by Alessandro Usiello from Ceinge Biotecnologie Avanzate and Loredano Pollegioni from the University of Insubria measured the levels of two amino acids—D-aspartate and D-serine—in post-mortem tissues taken from two brain regions of patients with and without schizophrenia. Both amino acids activate the N-methyl D-aspartate receptor, which is known to be less active in people with schizophrenia. The researchers found a mild increase in D-serine levels but a major decrease in D-aspartate in the schizophrenia patients’ dorsolateral prefrontal cortex (DLPFC), a memory and reasoning part of the brain, but not in the hippocampus. They also documented a greater activity of the enzyme responsible for D-aspartate breakdown in the DLPFC.

We performed ultra-deep methylation analysis at single molecule level of the promoter region of developmentally regulated D-Aspartate oxidase (Ddo), as a model gene, during brain development and embryonic stem cell neural differentiation. Single molecule methylation analysis enabled us to establish the effective epiallele composition within mixed or pure brain cell populations. In this framework, an epiallele is defined as a specific combination of methylated CpG within Ddo locus and can represent the epigenetic haplotype revealing a cell-to-cell methylation heterogeneity. Using this approach, we found a high degree of polymorphism of methylated alleles (epipolymorphism) evolving in a remarkably conserved fashion during brain development. The different sets of epialleles mark stage, brain areas, and cell type and unravel the possible role of specific CpGs in favoring or inhibiting local methylation. Undifferentiated embryonic stem cells showed non-organized distribution of epialleles that apparently originated by stochastic methylation events on individual CpGs. Upon neural differentiation, despite detecting no changes in average methylation, we observed that the epiallele distribution was profoundly different, gradually shifting toward organized patterns specific to the glial or neuronal cell types. Our findings provide a deep view of gene methylation heterogeneity in brain cell populations promising to furnish innovative ways to unravel mechanisms underlying methylation patterns generation and alteration in brain diseases.

Rhes is one of the most interesting genes regulated by thyroid hormones that, through the inhibition of the striatal cAMP/PKA pathway, acts as a modulator of dopamine neurotransmission. Rhes mRNA is expressed at high levels in the dorsal striatum, with a medial-to-lateral expression gradient reflecting that of both dopamine D2 and adenosine A2A receptors. Rhes transcript is also present in the hippocampus, cerebral cortex, olfactory tubercle and bulb, substantia nigra pars compacta (SNc) and ventral tegmental area of the rodent brain. In line with Rhes-dependent regulation of dopaminergic transmission, data showed that lack of Rhes enhanced cocaine- and amphetamine-induced motor stimulation in mice. Previous studies showed that pharmacological depletion of dopamine significantly reduces Rhes mRNA levels in rodents, non-human primates and Parkinson’s disease (PD) patients, suggesting a link between dopaminergic innervation and physiological Rhes mRNA expression. Rhes protein binds to and activates striatal mTORC1, and modulates L-DOPA-induced dyskinesia in PD rodent models. Finally, Rhes is involved in the survival of mouse midbrain dopaminergic neurons of SNc, thus pointing towards a Rhes-dependent modulation of autophagy and mitophagy processes, and encouraging further investigations about mechanisms underlying dysfunctions of the nigrostriatal system.