Quantitative Kinematic Characterization of Reaching Impairments in Mice After a Stroke

Background and Objective. Kinematic analysis of reaching movements is increasingly used to evaluate upper extremity function after cerebrovascular insults in humans and has also been applied to rodent models. Such analyses can require time-consuming frame-by-frame inspections and are affected by the experimenter's bias. In this study, we introduce a semi-automated algorithm for tracking forepaw movements in mice. This methodology allows us to calculate several kinematic measures for the quantitative assessment of performance in a skilled reaching task before and after a focal cortical stroke. Methods. Mice were trained to reach for food pellets with their preferred paw until asymptotic performance was achieved. Photothrombosis was then applied to induce a focal ischemic injury in the motor cortex, contralateral to the trained limb. Mice were tested again once a week for 30 days. A high frame rate camera was used to record the movements of the paw, which was painted with a nontoxic dye. An algorithm was then applied off-line to track the trajectories and to compute kinematic measures for motor performance evaluation. Results. The tracking algorithm proved to be fast, accurate, and robust. A number of kinematic measures were identified as sensitive indicators of poststroke modifications. Based on end-point measures, ischemic mice appeared to improve their motor performance after 2 weeks. However, kinematic analysis revealed the persistence of specific trajectory adjustments up to 30 days poststroke, indicating the use of compensatory strategies. Conclusions. These results support the use of kinematic analysis in mice as a tool for both detection of poststroke functional impairments and tracking of motor improvements following rehabilitation. Similar studies could be performed in parallel with human studies to exploit the translational value of this skilled reaching analysis

Pharmacological rescue of adult hippocampal neurogenesis in a mouse model of X-linked intellectual disability

Oligophrenin-1 (OPHN1) is a Rho GTPase activating protein whose mutations cause X-linked intellectual disability (XLID). How loss of function of Ophnl affects neuronal development is only partly understood. Here we have exploited adult hippocampal neurogenesis to dissect the steps of neuronal differentiation that are affected by Ophn1 deletion. We found that mice lacking Ophnl display a reduction in the number of newborn neurons in the dentate gyrus. A significant fraction of the Ophn1-deficient newly generated neurons failed to extend an axon towards CM, and showed an altered density of dendritic protrusions. Since Ophnl-deficient mice display overactivation of Rho-associated protein kinase (ROCK) and protein kinase A (PICA) signaling, we administered a clinically approved ROCK/PICA inhibitor (fasudil) to correct the neurogenesis defects. While administration of fasudil was not effective in rescuing axon formation, the same treatment completely restored spine density to control levels, and enhanced the long-term survival of adult-born neurons in mice lacking Ophn1. These results identify specific neurodevelopmental steps that are impacted by Ophn1 deletion, and indicate that they may be at least partially corrected by pharmacological treatment. (C) 2017 The Authors. Published by Elsevier Inc

The Chemokine CCL2 Mediates the Seizure-enhancing Effects of Systemic Inflammation

Epilepsy is a chronic disorder characterized by spontaneous recurrent seizures. Brain inflammation is increasingly recognized as a critical factor for seizure precipitation, but the molecular mediators of such proconvulsant effects are only partly understood. The chemokine CCL2 is one of the most elevated inflammatory mediators in patients with pharmacoresistent epilepsy, but its contribution to seizure generation remains unexplored. Here, we show, for the first time, a crucial role for CCL2 and its receptor CCR2 in seizure control. We imposed a systemic inflammatory challenge via lipopolysaccharide (LPS) administration in mice with mesial temporal lobe epilepsy. We found that LPS dramatically increased seizure frequency and upregulated the expression of many inflammatory proteins, including CCL2. To test the proconvulsant role of CCL2, we administered systemically either a CCL2 transcription inhibitor (bindarit) or a selective antagonist of the CCR2 receptor (RS102895). We found that interference with CCL2 signaling potently suppressed LPS-induced seizures. Intracerebral administration of anti-CCL2 antibodies also abrogated LPS-mediated seizure enhancement in chronically epileptic animals. Our results reveal that CCL2 is a key mediator in the molecular pathways that link peripheral inflammation with neuronal hyperexcitability

Neuroplastic Changes Following Brain Ischemia and their Contribution to Stroke Recovery: Novel Approaches in Neurorehabilitation

Ischemic damage to the brain triggers substantial reorganization of spared areas and pathways, which is associated with limited, spontaneous restoration of function. A better understanding of this plastic remodeling is crucial to develop more effective strategies for stroke rehabilitation. In this review article, we discuss advances in the comprehension of post-stroke network reorganization in patients and animal models. We first focus on rodent studies that have shed light on the mechanisms underlying neuronal remodeling in the perilesional area and contralesional hemisphere after motor cortex infarcts. Analysis of electrophysiological data has demonstrated brain-wide alterations in functional connectivity in both hemispheres, well beyond the infarcted area. We then illustrate the potential use of non-invasive brain stimulation (NIBS) techniques to boost recovery. We finally discuss rehabilitative protocols based on robotic devices as a tool to promote endogenous plasticity and functional restoration

Electrophysiology of glioma: a Rho GTPase-activating protein reduces tumor growth and spares neuron structure and function

Background. Glioblastomas are the most aggressive type of brain tumor. A successful treatment should aim at halting tumor growth and protecting neuronal cells to prevent functional deficits and cognitive deterioration. Here, we exploited a Rho GTPase-activating bacterial protein toxin, cytotoxic necrotizing factor 1 (CNF1), to interfere with glioma cell growth in vitro and vivo. We also investigated whether this toxin spares neuron structure and function in peritumoral areas. Methods. We performed a microarray transcriptomic and in-depth proteomic analysis to characterize the molecular changes triggered by CNF1 in glioma cells. We also examined tumor cell senescence and growth in vehicle-and CNF1-treated glioma-bearing mice. Electrophysiological and morphological techniques were used to investigate neuronal alterations in peritumoral cortical areas. Results. Administration of CNF1 triggered molecular and morphological hallmarks of senescence in mouse and human glioma cells in vitro. CNF1 treatment in vivo induced glioma cell senescence and potently reduced tumor volumes. In peritumoral areas of glioma-bearing mice, neurons showed a shrunken dendritic arbor and severe functional alterations such as increased spontaneous activity and reduced visual responsiveness. CNF1 treatment enhanced dendritic length and improved several physiological properties of pyramidal neurons, demonstrating functional preservation of the cortical network. Conclusions. Our findings demonstrate that CNF1 reduces glioma volume while at the same time maintaining the physiological and structural properties of peritumoral neurons. These data indicate a promising strategy for the development of more effective antiglioma therapies

Re-Assembled Botulinum Neurotoxin Inhibits CNS Functions without Systemic Toxicity

The therapeutic potential of botulinum neurotoxin type A (BoNT/A) has recently been widely recognized. BoNT/A acts to silence synaptic transmission via specific proteolytic cleavage of an essential neuronal protein, SNAP25. The advantages of BoNT/A-mediated synaptic silencing include very long duration, high potency and localized action. However, there is a fear of possible side-effects of BoNT/A due to its diffusible nature which may lead to neuromuscular blockade away from the injection site. We recently developed a “protein-stapling” technology which allows re-assembly of BoNT/A from two separate fragments. This technology allowed, for the first time, safe production of this popular neuronal silencing agent. Here we evaluated the re-assembled toxin in several CNS assays and assessed its systemic effects in an animal model. Our results show that the re-assembled toxin is potent in inhibiting CNS function at 1 nM concentration but surprisingly does not exhibit systemic toxicity after intraperitoneal injection even at 200 ng/kg dose. This shows that the re-assembled toxin represents a uniquely safe tool for neuroscience research and future medical applications

Epithelioid variant of pleomorphic liposarcoma as potential mimic of metastatic carcinoma

We report a case of epithelioid variant of pleomorphic liposarcoma (EPL) found in the the infrapatellar fat pad of Hoffa of a 31-year old male. Histologically, the predominant population was formed by epithelioid cells with eosinophilic or clear cytoplasm admixed with rare pleomorphic lipoblasts. The immunohistochemical panel was not helpful in the diagnosis. FISH analysis using the locus-specific indicator CHOP (12q13) dual color break apart was applied to representative formalin-fixed, paraffin-embedded tissue sections. The result of FISH indicated a rearranged CHOP (DDIT3) gene and confirmed the diagnosis of EPL. The EPL should be differentiated from a metastatic carcinoma or other type of sarcoma. In these cases a clinicopathological correlation and an exhaustive sampling of the specimen for demonstration of lipogenic areas or pleomorphic lipoblasts is always necessary. FISH with demonstration of CHOP gene rearrangement is useful in providing specific ancillary information for the difficult differential diagnosis of this case

Obese mice exposed to psychosocial stress display cardiac and hippocampal dysfunction associated with local brain-derived neurotrophic factor depletion

Introduction: Obesity and psychosocial stress (PS) co-exist in individuals of Western society. Nevertheless, how PS impacts cardiac and hippocampal phenotype in obese subjects is still unknown. Nor is it clear whether changes in local brain-derived neurotrophic factor (BDNF) account, at least in part, for myocardial and behavioral abnormalities in obese experiencing PS. Methods: In adult male WT mice, obesity was induced via a high-fat diet (HFD). The resident-intruder paradigm was superimposed to trigger PS. In vivo left ventricular (LV) performance was evaluated by echocardiography and pressure-volume loops. Behaviour was indagated by elevated plus maze (EPM) and Y-maze. LV myocardium was assayed for apoptosis, fibrosis, vessel density and oxidative stress. Hippocampus was analyzed for volume, neurogenesis, GABAergic markers and astrogliosis. Cardiac and hippocampal BDNF and TrkB levels were measured by ELISA and WB. We investigated the pathogenetic role played by BDNF signaling in additional cardiac-selective TrkB (cTrkB) KO mice. Findings: When combined, obesity and PS jeopardized LV performance, causing prominent apoptosis, fibrosis, oxidative stress and remodeling of the larger coronary branches, along with lower BDNF and TrkB levels. HFD/PS weakened LV function similarly in WT and cTrkB KO mice. The latter exhibited elevated LV ROS emission already at baseline. Obesity/PS augmented anxiety-like behaviour and impaired spatial memory. These changes were coupled to reduced hippocampal volume, neurogenesis, local BDNF and TrkB content and augmented astrogliosis. Interpretation: PS and obesity synergistically deteriorate myocardial structure and function by depleting cardiac BDNF/TrkB content, leading to augmented oxidative stress. This comorbidity triggers behavioral deficits and induces hippocampal remodeling, potentially via lower BDNF and TrkB levels. Fund: J.A. was in part supported by Rotary Foundation Global Study Scholarship. G.K. was supported by T32 National Institute of Health (NIH) training grant under award number 1T32AG058527. S.C. was funded by American Heart Association Career Development Award (19CDA34760185). G.A.R.C. was funded by NIH (K01HL133368-01). APB was funded by a Grant from the Friuli Venezia Giulia Region entitled: “Heart failure as the Alzheimer disease of the heart; therapeutic and diagnostic opportunities”. M.C. was supported by PRONAT project (CNR). N.P. was funded by NIH (R01 HL136918) and by the Magic-That-Matters fund (JHU). V.L. was in part supported by institutional funds from Scuola Superiore Sant'Anna (Pisa, Italy), by the TIM-Telecom Italia (WHITE Lab, Pisa, Italy), by a research grant from Pastificio Attilio Mastromauro Granoro s.r.l. (Corato, Italy) and in part by ETHERNA project (Prog. n. 161/16, Fondazione Pisa, Italy). Funding source had no such involvement in study design, in the collection, analysis, interpretation of data, in the writing of the report; and in the decision to submit the paper for publication