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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.

Background: Elevated levels of specific proinflammatory molecules in the cerebrospinal fluid (CSF) have been associated with disability progression, enhanced neurodegeneration and higher incidence of mood disorders in people with multiple sclerosis (MS). Studies in animal models of MS suggest that preventive exercise may play an immunomodulatory activity, with beneficial effects on both motor deficits and behavioral alterations. Here we explored the impact of lifestyle physical activity on clinical presentation and associated central inflammation in a large group of newly diagnosed patients with MS. Furthermore, we addressed the causal link between exercise-mediated immunomodulation and mood symptoms in the animal setting. Methods: A cross-sectional study was conducted on 235 relapsing-remitting MS patients at the time of the diagnosis. Patients were divided into 3 groups (“sedentary”, “lifestyle physical activity” and “exercise”) according to the level of physical activity in the six months preceding the evaluation. Patients underwent clinical, neuropsychological and psychiatric evaluation, magnetic resonance imaging and lumbar puncture for diagnostic purposes. The CSF levels of proinflammatory and anti-inflammatory cytokines were analyzed and compared with a group of 80 individuals with non-inflammatory and non-degenerative diseases. Behavioral and electrophysiological studies were carried out in control mice receiving intracerebral injection of IL-2 or vehicle. Behavior was also assessed in mice with experimental autoimmune encephalomyelitis (EAE), animal model of MS, reared in standard (sedentary group) or running wheel-equipped (exercise group) cages. Results: In exercising MS patients, depression and anxiety were reduced compared to sedentary patients. The CSF levels of the interleukin-2 and 6 (IL-2, IL-6) were increased in MS patients compared with control individuals. In MS subjects exercise was associated with normalized CSF levels of IL-2. In EAE mice exercise started before disease onset reduced both behavioral alterations and striatal IL-2 expression. Notably, a causal role of IL-2 in mood disorders was shown. IL-2 administration in control healthy mice induced anxious- and depressive-like behaviors and impaired type-1 cannabinoid (CB1) receptor-mediated neurotransmission at GABAergic synapses, mimicking EAE-induced synaptic dysfunction. Conclusions: Our results indicate an immunomodulatory effect of exercise in MS patients, associated with reduced CSF expression of IL-2, which might result in reduced mood disorders. These data suggest that exercise in the early stages may act as a disease-modifying therapy in MS although further longitudinal studies are needed to clarify this issue.

Growing data from human and animal studies indicate the beneficial effects of exercise on several clinical outcomes in patients with multiple sclerosis (MS), an autoimmune, demyelinating disease, suggesting that it may slow down the disease progression, by reducing brain damage. However, the mechanisms involved are still elusive.Aim of this study was to address the effects of voluntary running wheel in a toxic-demyelinating model of MS, in which demyelination and brain inflammation occur in response to cuprizone (CPZ) treatment. Mice were housed in standard or wheel-equipped cages starting from the day of CPZ or normal chow feeding for three or six weeks and evaluated for weight changes, locomotor skills and neuromuscular functions over the course of the experimental design. Biochemical, molecular biology and immunohistochemical analyses were performed.Exercise prevented early weight loss caused by CPZ, indicating improved wellness in these mice. Both neuromuscular function and motor coordination were significantly enhanced by exercise in CPZ-treated mice. Moreover, exercise induced an early protection against axonal damage and the loss of the myelin associated proteins, myelin basic protein (MBP) and 2′,3′-Cyclic-nucleotide 3′-phosphodiesterase (CNPase), in the striatum and the corpus callosum, in coincidence of a strongly attenuated microglia activation in both brain areas. Further, during the late phase of the treatment, exercise in CPZ mice reduced the recruitment of new OLs compared to sedentary CPZ mice, likely due to the precocious protection against myelin damage.Overall, these results suggest that life-style interventions can be effective against the demyelinating-inflammatory processes occurring in the brains of MS patients.

Interferon-γ (IFN-γ) has been implicated in the pathogenesis of multiple sclerosis (MS) and in its animal model, experimental autoimmune encephalomyelitis (EAE). The type-1 cannabinoid receptors (CB1Rs) are heavily involved in MS pathophysiology, and a growing body of evidence suggests that mood disturbances reflect specific effects of proinflammatory cytokines on neuronal activity. Here, we investigated whether IFN-γ could exert a role in the anxiety- and depressive-like behavior observed in mice with EAE, and in the modulation of CB1Rs. Anxiety and depression in fact are often diagnosed in MS, and have already been shown to depend on cannabinoid system. We performed biochemical, behavioral and electrophysiological experiments to assess the role of IFN-γ on mood control and on synaptic transmission in mice. Intracerebroventricular delivery of IFN-γ caused a depressive- and anxiety-like behavior in mice, associated with the selective dysfunction of CB1Rs controlling GABA transmission in the striatum. EAE induction was associated with increased striatal expression of IFN-γ, and with CB1R transmission deficits, which were rescued by pharmacological blockade of IFN-γ. IFN-γ was unable to replicate the effects of EAE on excitatory and inhibitory transmission in the striatum, but mimicked the effects of EAE on CB1R function in this brain area. Overall these results indicate that IFN-γ exerts a relevant control on mood, through the modulation of CB1R function. A better understanding of the biological pathways underling the psychological disorders during neuroinflammatory conditions is crucial for developing effective therapeutic strategies.

Mood disturbances are frequent in patients with multiple sclerosis (MS), even in non-disabled patients and in the remitting stages of the disease. It is still largely unknown how the pathophysiological process on MS causes anxiety and depression, but the dopaminergic system is likely involved.Aim of the present study was to investigate depressive-like behavior in mice with experimental autoimmune encephalomyelitis (EAE), a model of MS, and its possible link to dopaminergic neurotransmission. Behavioral, amperometric and biochemical experiments were performed to determine the role of inflammation in mood control in EAE. First, we assessed the independence of mood alterations from motor disability during the acute phase of the disease, by showing a depressive-like behavior in EAE mice with mild clinical score and preserved motor skills (mild-EAE). Second, we linked such behavioral changes to the selective increased striatal expression of interleukin-1beta (IL-1β) in a context of mild inflammation and to dopaminergic system alterations. Indeed, in the striatum of EAE mice, we observed an impairment of dopamine (DA) neurotransmission, since DA release was reduced and signaling through DA D1- and D2-like receptors was unbalanced.In conclusion, the present study provides first evidence of the link between the depressive-like behavior and the alteration of dopaminergic system in EAE mice, raising the possibility that IL-1β driven dysfunction of dopaminergic signaling might play a role in mood disturbances also in MS patients.

A significant proportion of multiple sclerosis (MS) patients have functionally relevant cerebellar deficits, which significantly contribute to disability. Although clinical and experimental studies have been conducted to understand the pathophysiology of cerebellar dysfunction in MS, no electrophysiological and morphological studies have investigated potential alterations of synaptic connections of cerebellar Purkinje cells (PC). For this reason we analyzed cerebellar PC GABAergic connectivity in mice with MOG(35–55)-induced experimental autoimmune encephalomyelitis (EAE), a mouse model of MS. We observed a strong reduction in the frequency of the spontaneous inhibitory post-synaptic currents (IPSCs) recorded from PCs during the symptomatic phase of the disease, and in presence of prominent microglia activation not only in the white matter (WM) but also in the molecular layer (ML). The massive GABAergic innervation on PCs from basket and stellate cells was reduced and associated to a decrease of the number of these inhibitory interneurons. On the contrary no significant loss of the PCs could be detected. Incubation of interleukin-1beta (IL-1β) was sufficient to mimic the electrophysiological alterations observed in EAE mice.We thus suggest that microglia and pro-inflammatory cytokines, together with a degeneration of basket and stellate cells and their synaptic terminals, contribute to impair GABAergic transmission on PCs during EAE. Our results support a growing body of evidence that GABAergic signaling is compromised in EAE and in MS, and show a selective susceptibility to neuronal and synaptic degeneration of cerebellar inhibitory interneurons.

Transient receptor potential vanilloid 1 (TRPV1) channels are involved in several inflammatory diseases. However, their action is still controversial, and both pro-inflammatory and anti-inflammatory roles have been described. We used a strain of TRPV1-KO mice to characterize the role of these channels in experimental autoimmune encephalomyelitis (EAE), which models multiple sclerosis (MS) in mice.EAE mice showed higher lethality in the peak phase of the disease and a better recovery of the surviving animals in the chronic stages, compared to their wild-type (WT) counterparts. By means of whole-cell patch clamp experiments in corticostriatal brain slices, we found that the absence of TRPV1 channels exacerbated the defect of glutamate transmission occurring in the peak phase of EAE, and attenuated the alterations of GABA synapses in the chronic phase of EAE, thus paralleling the dual effects of TRPV1-KO on the motor deficits of EAE mice. Furthermore, in slices from non-EAE mice, we found that genetic or pharmacological blockade of TRPV1 channels enhanced the synaptic effects of tumor necrosis factor α (TNF-α) on glutamate-mediated excitatory postsynaptic currents, and prevented the action of interleukin 1β (IL-1β) on GABAergic inhibitory postsynaptic currents.Together, our results suggest that TRPV1 channels contrast TNF-α-mediated synaptic deficits in the peak phase of EAE and, in the chronic stages, enhance IL-1β-induced GABAergic defects. The opposing interplay with the synaptic actions of the two major pro-inflammatory cytokines might explain the bimodal effects of TRPV1 ablation on the motor deficits of EAE, and suggests that the inflammatory milieu determines whether TRPV1 channels exert preferentially aversive or protective effects on neurons during neuroinflammatory diseases.

Cerebellar compensation is a reliable model of lesion-induced plasticity occurring through profound synaptic and neurochemical modifications in cortical and sub-cortical regions. As the recovery from cerebellar deficits progresses, the firstly enhanced glutamate striatal transmission is then normalized. The time course of cerebellar compensation and the concomitant striatal modifications might be influenced by protocols of environmental enrichment (EE) differently timed in respect to cerebellar lesion. In the present study, we analyzed the effects of different EE protocols on postural and locomotor behaviors (by means of a neurological rating scale), and on striatal synaptic activity (by means of recordings of spontaneous glutamate-mediated excitatory postsynaptic currents (sEPSCs)) and on morphological correlates (by means of density and dendritic length of Fast Spiking (FS) interneurons) following hemicerebellectomy (HCb) in rats. Cerebellar motor deficits were reduced faster in the enriched animals in comparison to standard housed HCbed rats. The beneficial influence of EE was higher in the animals enriched before the HCb than in rats enriched only after the lesion. In parallel, the HCb-induced increase in striatal sEPSCs was not observed in rats enriched before HCb and attenuated in rats enriched after HCb. Furthermore, the EE prevented the shrinkage of dendritic arborization of FS striatal interneurons. Also this effect was more marked in animals enriched before than after the HCb. The exposure to EE exerted either neuro-protective or therapeutic actions on the cerebellar deficits. The experience-dependent changes of the synaptic and neuronal connectivity observed in the striatal neurons may represent one of the mechanisms through which the enrichment facilitates functional compensation following the cerebellar damage.

Voluntary exercise is beneficial in models of primarily neurodegenerative disorders. Whether exercise also affects inflammatory neurodegeneration is unknown. In the present study, we evaluated the clinical, synaptic and neuropathological effects of voluntary wheel running in mice with myelin oligodendrocyte glycoprotein (MOG)-induced experimental autoimmune encephalomyelitis (EAE), a model of multiple sclerosis. Exercising EAE mice exhibited less severe neurological deficits compared to control EAE animals. The sensitivity of striatal GABA synapses to the stimulation of cannabinoid CB1 receptors was dramatically downregulated following EAE induction, and was rescued by exercise in EAE mice with access to a running wheel. Finally, we found that exercise was able to contrast dendritic spine loss induced by EAE in striatal neurons, although the degree of inflammatory response was similar in the two experimental groups.Our work suggests that life style and experiences can impact the clinical course of inflammatory neurodegenerative diseases by affecting their synaptic bases.

Synaptic rearrangements in the peri-infarct regions are believed to contribute to the partial recovery of function that takes place after stroke. Here, we performed neurophysiological recordings from single neurons of rats with permanent occlusion of the middle cerebral artery (pMCAO) during the resolution of their neurological deficits. Our results show that complex and dynamic changes of glutamate transmission in the peri-infarct area parallel the recovery from brain infarct. We have observed that frequency and duration of spontaneous glutamate-mediated synaptic events were markedly increased in striatal neurons during the early phase of the recovery (3 days after pMCAO), due to potentiation of both NMDA (N-methyl-d-aspartate) and non-NMDA receptor-mediated transmission. In the late phase of recovery (7 days after pMCAO), glutamate transmission was still enhanced because of a selective facilitation of non-NMDA receptor-mediated transmission. Spiny projection neurons but not aspiny interneurons underwent detectable changes of synaptic excitability in the striatum following pMCAO, indicating that the process of neuronal adaptation after focal brain ischemia is cell-type-specific. Our results provide a synaptic correlate of the long-lasting brain hyperexcitability mediating recovery described with noninvasive neurophysiological approaches.

Huntington's disease (HD) is a genetic neurodegenerative disorder characterized by motor abnormalities and cognitive impairment. The irreversible succinate dehydrogenase (SD) inhibitor 3-nitropropionic acid (3NP) causes neurodegeration in the striatum resembling HD when administered to rodents or primates.Using corticostriatal brain slice preparations, we analyzed the pattern of gene expression following 3NP application utilizing cDNA microarrays. Acute 3NP treatment modulates the expression of several genes involved in dopaminergic and glutamatergic signaling in corticostriatal brain slices, and unbalances the downstream serine/threonine protein kinase and phosphatase network affecting the dopamine- and cAMP-regulated phosphoprotein of 32 kDa (DARPP-32). Our data provide new information about the molecular events possibly underlying neurodegeneration induced by this mitochondrial toxin.

Although experimental studies have widely shown that the pharmacological blockade of ionotropic glutamate receptors reduces ischemic damage, clinical trials with classical AMPA and NMDA glutamate receptor antagonists have provided negative results. To address the involvement of ionotropic glutamate receptors in ischemic damage, corticostriatal brain slices were prepared from adult rats. Extracellular recordings were performed in the striatum after stimulation of the glutamatergic corticostriatal fibres. In vitro ischemia was induced for a 10-min period by omitting oxygen and glucose from the external medium. Under control conditions, ischemia produced an irreversible loss of the corticostriatal field potential amplitude, AP5, a competitive NMDA receptor antagonist, induced a slight rescue of the potential, while ifenprodil, a positive modulator of the proton sensor of the NMDA receptors, allowed a complete recovery from the ischemic insult. Similar neuroprotection was achieved by utilizing either CNQX, a broad spectrum AMPA receptors antagonist, or Joro spider toxin, a selective blocker of calcium permeable AMPA receptors. Interestingly, while CNZX also fully suppressed physiological excitatory transmission, Joro spider toxin was ineffective on this parameter. Finally, lamotrigine and remacemide, two antiepileptic drugs that differentially affect excitatory transmission, exerted neuroprotective effects against ischemia. Noticeably, a combination of low concentrations of these two drugs exerted a stronger neuroprotection than a single drug given in isolation. Thus, it might be possible to reach a neuroprotective action by utilizing doses of these compounds low enough to avoid side effects. Our experimental data still support the idea that a negative modulation of excitatory transmission can be neuroprotective against ischemia. In addition, our findings support the concept that it is possible to produce a significant neuroprotective action in the absence of a relevant interference with normal synaptic transmission.

The irreversible loss of the dopamine-mediated control of striatal function is considered the functional substrate of the motor symptoms of Parkinson's disease. This pathological event causes a complex rearrangement of neuronal activity which involves specific dopamine-regulated cellular functions and, secondarily, several other cellular properties and transmitter systems. In the present study, we applied recently developed cDNA microarray technology to investigate the genetic correlates of the alterations produced by 6-hydroxydopamine-induced dopamine denervation in the nucleus striatum. We found that chronic dopamine denervation caused the modulation of 50 different genes involved in several cellular functions. In particular, products of the genes modulated by this experimental manipulation are involved both in the intracellular transduction of dopamine signal and in the regulation of glutamate transmission in striatal neurons, providing some information on the possible neuronal events which lead to the reorganization of glutamate transmission in the striatum of parkinsonian rats.

In the present study we have characterized a rat model of manganese (Mn) intoxication leading to behavioral disinhibition in the absence of major motor alterations. These behavioral changes were associated with significantly increased brain Mn levels but were uncoupled to anatomical lesions of the striatum or to morphological and cytochemical changes of the nigrostriatal dopaminergic pathway. The analysis of this model at cellular level showed an enhanced dopaminergic inhibitory control of the corticostriatal excitatory transmission via presynaptic D2-like dopamine (DA) receptors in slices obtained from Mn-treated rats. Conversely, the use of agonists acting on presynaptic purinergic, muscarinic, and glutamatergic metabotropic receptors revealed a normal sensitivity. Moreover, membrane responses recorded from single dopaminergic neurons following activation of D2 DA autoreceptors were also unchanged following Mn intoxication. Thus, our findings indicate a selective involvement of the D2-like DA receptors located on glutamatergic corticostriatal terminals in this pathological condition and suggest that the behavioral symptoms described in the “early” clinical phase of manganism may be caused by an abnormal dopaminergic inhibitory control on corticostriatal inputs. The identification of the synaptic mechanism underlying the “early” phase of Mn intoxication might have a critical importance to understand the causes of the progression of this pathological condition towards an “established” phase characterized by motor abnormalities and anatomical lesions of the basal ganglia.