Structural reorganization of medullary dorsal horn astrocytes in a rat model of neuropathic pain.

Multiple studies have shown that astrocytes in the medullary dorsal horn (MDH) play an important role in the development of pathologic pain. However, little is known about the structural reorganization of the peripheral astrocytic processes (PAP), the main functional part of the astrocyte, in MDH in neuropathic state. For this, we investigated the structural relationship between PAP and their adjacent presynaptic axon terminals and postsynaptic dendrites in the superficial laminae of the MDH using electron microscopical immunohistochemistry for ezrin, a marker for PAP, and quantitative analysis in a rat model of neuropathic pain following chronic constriction injury of the infraorbital nerve (CCI-ION). We found that, compared to controls, in rats with CCI-ION, (1) the number, % area, surface density, and volume fraction of ezrin-positive (+) PAP, as well as the fraction of synaptic edge apposed by ezrin + PAP and the degree of its coverage of presynaptic axon terminals and postsynaptic dendrites increased significantly, (2) these effects were abolished by administration of the mGluR5 antagonist 2-methyl-6-(phenylethynyl) pyridine (MPEP). These findings indicate that PAP undergoes structural reorganization around the central synapses of sensory afferents following nerve injury, suggest that it may be mediated by mGluR5, and may represent the structural basis for enhancing astrocyte-neuron interaction in neuropathic pain.

Early and selective localization of tau filaments to glutamatergic subcellular domains within the human anterodorsal thalamus.

Widespread cortical accumulation of misfolded pathological tau proteins (ptau) in the form of paired helical filaments is a major hallmark of Alzheimer's disease. Subcellular localization of ptau at various stages of disease progression is likely to be informative of the cellular mechanisms involving its spread. Here, we found that the density of ptau within several distinct rostral thalamic nuclei in post-mortem human tissue (n = 25 cases) increased with the disease stage, with the anterodorsal nucleus (ADn) consistently being the most affected. In the ADn, ptau-positive elements were present already in the pre-cortical (Braak 0) stage. Tau pathology preferentially affected the calretinin-expressing subpopulation of glutamatergic neurons in the ADn. At the subcellular level, we detected ptau immunoreactivity in ADn cell bodies, dendrites, and in a specialized type of presynaptic terminal that expresses vesicular glutamate transporter 2 (vGLUT2) and likely originates from the mammillary body. The ptau-containing terminals displayed signs of degeneration, including endosomal/lysosomal organelles. In contrast, corticothalamic axon terminals lacked ptau. The data demonstrate the involvement of a specific cell population in ADn at the onset of the disease. The presence of ptau in subcortical glutamatergic presynaptic terminals supports hypotheses about the transsynaptic spread of tau selectively affecting specialized axonal pathways.

Neuropathologic Validation and Diagnostic Accuracy of Presynaptic Dopaminergic Imaging in the Diagnosis of Parkinsonism.

BACKGROUND AND OBJECTIVES: Degeneration of the presynaptic nigrostriatal dopaminergic system is one of the main biological features of Parkinson disease (PD), multiple system atrophy (MSA), progressive supranuclear palsy (PSP), and corticobasal degeneration (CBD), which can be measured using single-photon emission CT imaging for diagnostic purposes. Despite its widespread use in clinical practice and research, the diagnostic properties of presynaptic nigrostriatal dopaminergic (DAT) imaging in parkinsonism have never been evaluated against the diagnostic gold standard of neuropathology. The aim of this study was to evaluate the diagnostic parameters of DAT imaging compared with pathologic diagnosis in patients with parkinsonism. METHODS: Retrospective cohort study of patients with DAT imaging for the investigation of a clinically uncertain parkinsonism with brain donation between 2010 and 2021 to the Queen Square Brain Bank (London). Patients with DAT imaging for investigation of pure ataxia or dementia syndromes without parkinsonism were excluded. Those with a pathologic diagnosis of PD, MSA, PSP, or CBD were considered presynaptic dopaminergic parkinsonism, and other pathologies were considered postsynaptic for the analysis. DAT imaging was performed in routine clinical practice and visually classified by hospital nuclear medicine specialists as normal or abnormal. The results were correlated with neuropathologic diagnosis to calculate diagnostic accuracy parameters for the diagnosis of presynaptic dopaminergic parkinsonism. RESULTS: All of 47 patients with PD, 41 of 42 with MSA, 68 of 73 with PSP, and 6 of 10 with CBD (sensitivity 100%, 97.6%, 93.2%, and 60%, respectively) had abnormal presynaptic dopaminergic imaging. Eight of 17 patients with presumed postsynaptic parkinsonism had abnormal scans (specificity 52.9%). DISCUSSION: DAT imaging has very high sensitivity and negative predictive value for the diagnosis of presynaptic dopaminergic parkinsonism, particularly for PD. However, patients with CBD, and to a lesser extent PSP (of various phenotypes) and MSA (with predominant ataxia), can show normal DAT imaging. A range of other neurodegenerative disorders may have abnormal DAT scans with low specificity in the differential diagnosis of parkinsonism. DAT imaging is a useful diagnostic tool in the differential diagnosis of parkinsonism, although clinicians should be aware of its diagnostic properties and limitations. CLASSIFICATION OF EVIDENCE: This study provides Class II evidence that DAT imaging does not accurately distinguish between presynaptic dopaminergic parkinsonism and non-presynaptic dopaminergic parkinsonism.

Synaptopathy: presynaptic convergence in frontotemporal dementia and amyotrophic lateral sclerosis.

Frontotemporal dementia and amyotrophic lateral sclerosis are common forms of neurodegenerative disease that share overlapping genetics and pathologies. Crucially, no significantly disease-modifying treatments are available for either disease. Identifying the earliest changes that initiate neuronal dysfunction is important for designing effective intervention therapeutics. The genes mutated in genetic forms of frontotemporal dementia and amyotrophic lateral sclerosis have diverse cellular functions, and multiple disease mechanisms have been proposed for both. Identification of a convergent disease mechanism in frontotemporal dementia and amyotrophic lateral sclerosis would focus research for a targetable pathway, which could potentially effectively treat all forms of frontotemporal dementia and amyotrophic lateral sclerosis (both familial and sporadic). Synaptopathies are diseases resulting from physiological dysfunction of synapses, and define the earliest stages in multiple neuronal diseases, with synapse loss a key feature in dementia. At the presynapse, the process of synaptic vesicle recruitment, fusion and recycling is necessary for activity-dependent neurotransmitter release. The unique distal location of the presynaptic terminal means the tight spatio-temporal control of presynaptic homeostasis is dependent on efficient local protein translation and degradation. Recently, numerous publications have shown that mutations associated with frontotemporal dementia and amyotrophic lateral sclerosis present with synaptopathy characterized by presynaptic dysfunction. This review will describe the complex local signalling and membrane trafficking events that occur at the presynapse to facilitate neurotransmission and will summarize recent publications linking frontotemporal dementia/amyotrophic lateral sclerosis genetic mutations to presynaptic function. This evidence indicates that presynaptic synaptopathy is an early and convergent event in frontotemporal dementia and amyotrophic lateral sclerosis and illustrates the need for further research in this area, to identify potential therapeutic targets with the ability to impact this convergent pathomechanism.

Axonal ER Ca2+ Release Selectively Enhances Activity-Independent Glutamate Release in a Huntington Disease Model.

Action potential (AP)-independent (miniature) neurotransmission occurs at all chemical synapses but remains poorly understood, particularly in pathologic contexts. Axonal endoplasmic reticulum (ER) Ca2+ stores are thought to influence miniature neurotransmission, and aberrant ER Ca2+ handling is implicated in progression of Huntington disease (HD). Here, we report elevated mEPSC frequencies in recordings from YAC128 mouse (HD-model) neurons (from cortical cultures and striatum-containing brain slices, both from male and female animals). Pharmacological experiments suggest that this is mediated indirectly by enhanced tonic ER Ca2+ release. Calcium imaging, using an axon-localized sensor, revealed slow AP-independent ER Ca2+ release waves in both YAC128 and WT cultures. These Ca2+ waves occurred at similar frequencies in both genotypes but spread less extensively and were of lower amplitude in YAC128 axons, consistent with axonal ER Ca2+ store depletion. Surprisingly, basal cytosolic Ca2+ levels were lower in YAC128 boutons and YAC128 mEPSCs were less sensitive to intracellular Ca2+ chelation. Together, these data suggest that elevated miniature glutamate release in YAC128 cultures is associated with axonal ER Ca2+ depletion but not directly mediated by ER Ca2+ release into the cytoplasm. In contrast to increased mEPSC frequencies, cultured YAC128 cortical neurons showed less frequent AP-dependent (spontaneous) Ca2+ events in soma and axons, although evoked glutamate release detected by an intensity-based glutamate-sensing fluorescence reporter in brain slices was similar between genotypes. Our results indicate that axonal ER dysfunction selectively elevates miniature glutamate release from cortical terminals in HD. This, together with reduced spontaneous cortical neuron firing, may cause a shift from activity-dependent to -independent glutamate release in HD, with potential implications for fidelity and plasticity of cortical excitatory signaling.SIGNIFICANCE STATEMENT Miniature neurotransmitter release persists at all chemical neuronal synapses in the absence of action potential firing but remains poorly understood, particularly in disease states. We show enhanced miniature glutamate release from cortical neurons in the YAC128 mouse Huntington disease model. This effect is mediated by axonal ER Ca2+ store depletion, but is not obviously due to elevated ER-to-cytosol Ca2+ release. Conversely, YAC128 cortical pyramidal neurons fired fewer action potentials and evoked cortical glutamate release was similar between WT an YAC128 preparations, indicating axonal ER depletion selectively enhances miniature glutamate release in YAC128 mice. These results extend our understanding of action potential independent neurotransmission and highlight a potential involvement of elevated miniature glutamate release in Huntington disease pathology.

Inhibitory synapse loss and accumulation of amyloid beta in inhibitory presynaptic terminals in Alzheimer's disease.

BACKGROUND AND PURPOSE: Synapse degeneration in Alzheimer's disease (AD) correlates strongly with cognitive decline. There is well-established excitatory synapse loss in AD with known contributions of pathological amyloid beta (Aß) to excitatory synapse dysfunction and loss. Despite clear changes in circuit excitability in AD and model systems, relatively little is known about pathology in inhibitory synapses. METHODS: Here human postmortem brain samples (n = 5 control, 10 AD cases) from temporal and occipital cortices were examined to investigate whether inhibitory synapses and neurons are lost in AD and whether Aß may contribute to inhibitory synapse degeneration. Inhibitory neurons were counted in all six cortical layers using stereology software, and array tomography was used to examine synapse density and the accumulation of Aß in synaptic terminals. RESULTS: Differing inhibitory neuron densities were observed in the different cortical layers. The highest inhibitory neuron density was observed in layer 4 in both brain regions and the visual cortex had a higher inhibitory neuron density than the temporal cortex. There was significantly lower inhibitory neuron density in AD than in control cases in all six cortical layers. High-resolution array tomography imaging revealed plaque-associated loss of inhibitory synapses and accumulation of Aß in a small subset of inhibitory presynaptic terminals with the most accumulation near amyloid plaques. CONCLUSIONS: Inhibitory neuron and synapse loss in AD may contribute to disrupted excitatory/inhibitory balance and cognitive decline. Future work is warranted to determine whether targeting inhibitory synapse loss could be a useful therapeutic strategy.

The organization and development of cortical interneuron presynaptic circuits are area specific.

Parvalbumin and somatostatin inhibitory interneurons gate information flow in discrete cortical areas that compute sensory and cognitive functions. Despite the considerable differences between areas, individual interneuron subtypes are genetically invariant and are thought to form canonical circuits regardless of which area they are embedded in. Here, we investigate whether this is achieved through selective and systematic variations in their afferent connectivity during development. To this end, we examined the development of their inputs within distinct cortical areas. We find that interneuron afferents show little evidence of being globally stereotyped. Rather, each subtype displays characteristic regional connectivity and distinct developmental dynamics by which this connectivity is achieved. Moreover, afferents dynamically regulated during development are disrupted by early sensory deprivation and in a model of fragile X syndrome. These data provide a comprehensive map of interneuron afferents across cortical areas and reveal the logic by which these circuits are established during development.

Disassembly and rewiring of a mature converging excitatory circuit following injury.

Specificity and timing of synapse disassembly in the CNS are essential to learning how individual circuits react to neurodegeneration of the postsynaptic neuron. In sensory systems such as the mammalian retina, synaptic connections of second-order neurons are known to remodel and reconnect in the face of sensory cell loss. Here we analyzed whether degenerating third-order neurons can remodel their local presynaptic connectivity. We injured adult retinal ganglion cells by transiently elevating intraocular pressure. We show that loss of presynaptic structures occurs before postsynaptic density proteins and accounts for impaired transmission from presynaptic neurons, despite no evidence of presynaptic cell loss, axon terminal shrinkage, or reduced functional input. Loss of synapses is biased among converging presynaptic neuron types, with preferential loss of the major excitatory cone-driven partner and increased connectivity with rod-driven presynaptic partners, demonstrating that this adult neural circuit is capable of structural plasticity while undergoing neurodegeneration.

Oxytocin in the anterior cingulate cortex attenuates neuropathic pain and emotional anxiety by inhibiting presynaptic long-term potentiation.

Oxytocin is a well-known neurohypophysial hormone that plays an important role in behavioral anxiety and nociception. Two major forms of long-term potentiation, presynaptic LTP (pre-LTP) and postsynaptic LTP (post-LTP), have been characterized in the anterior cingulate cortex (ACC). Both pre-LTP and post-LTP contribute to chronic-pain-related anxiety and behavioral sensitization. The roles of oxytocin in the ACC have not been studied. Here, we find that microinjections of oxytocin into the ACC attenuate nociceptive responses and anxiety-like behavioral responses in animals with neuropathic pain. Application of oxytocin selectively blocks the maintenance of pre-LTP but not post-LTP. In addition, oxytocin enhances inhibitory transmission and excites ACC interneurons. Similar results are obtained by using selective optical stimulation of oxytocin-containing projecting terminals in the ACC in animals with neuropathic pain. Our results demonstrate that oxytocin acts on central synapses and reduces chronic-pain-induced anxiety by reducing pre-LTP.

Automated in vivo drug screen in zebrafish identifies synapse-stabilising drugs with relevance to spinal muscular atrophy.

Synapses are particularly vulnerable in many neurodegenerative diseases and often the first to degenerate, for example in the motor neuron disease spinal muscular atrophy (SMA). Compounds that can counteract synaptic destabilisation are rare. Here, we describe an automated screening paradigm in zebrafish for small-molecule compounds that stabilize the neuromuscular synapse in vivo. We make use of a mutant for the axonal C-type lectin chondrolectin (chodl), one of the main genes dysregulated in SMA. In chodl-/- mutants, neuromuscular synapses that are formed at the first synaptic site by growing axons are not fully mature, causing axons to stall, thereby impeding further axon growth beyond that synaptic site. This makes axon length a convenient read-out for synapse stability. We screened 982 small-molecule compounds in chodl chodl-/- mutants and found four that strongly rescued motor axon length. Aberrant presynaptic neuromuscular synapse morphology was also corrected. The most-effective compound, the adenosine uptake inhibitor drug dipyridamole, also rescued axon growth defects in the UBA1-dependent zebrafish model of SMA. Hence, we describe an automated screening pipeline that can detect compounds with relevance to SMA. This versatile platform can be used for drug and genetic screens, with wider relevance to synapse formation and stabilisation.

In vivo aggregation of presynaptic alpha-synuclein is not influenced by its phosphorylation at serine-129.

Abnormal aggregation of the α-synuclein protein is a key molecular feature of Parkinson's disease and other neurodegenerative diseases. The precise mechanisms that trigger α-synuclein aggregation are unclear, and it is not known what role aggregation plays in disease pathogenesis. Here we use an in vivo zebrafish model to express several different forms of human α-synuclein and measure its aggregation in presynaptic terminals. We show that human α-synuclein tagged with GFP can be expressed in zebrafish neurons, localizing normally to presynaptic terminals and undergoing phosphorylation at serine-129, as in mammalian neurons. The visual advantages of the zebrafish system allow for dynamic in vivo imaging to study α-synuclein, including the use of fluorescence recovery after photobleaching (FRAP) techniques to probe protein mobility. These experiments reveal three distinct terminal pools of α-synuclein with varying mobility, likely representing different subpopulations of aggregated and non-aggregated protein. Human α-synuclein is phosphorylated by an endogenous zebrafish Polo-like kinase activity, and there is a heterogeneous population of neurons containing either very little or extensive phosphorylation throughout the axonal arbor. Both pharmacological and genetic manipulations of serine-129 show that phosphorylation of α-synuclein at this site does not significantly affect its mobility. This suggests that serine-129 phosphorylation alone does not promote α-synuclein aggregation. Together our results show that human α-synuclein can be expressed and measured quantitatively in zebrafish, and that disease-relevant post-translational modifications occur within neurons. The zebrafish model provides a powerful in vivo system for measuring and manipulating α-synuclein function and aggregation, and for developing new treatments for neurodegenerative disease.

Preserved noradrenergic function in Parkinson's disease patients with rest tremor.

Noradrenergic neurotransmission may play an important role in tremor modulation through its innervation of key structures of the central tremor circuits. Here, Parkinson's disease (PD) patients with (PDT+) or without (PDT-) rest tremor had 11C-methylreboxetine(11C-MeNER) positron emission tomography (PET) to test the hypothesis that noradrenaline terminal function was relatively preserved in PDT+ compared to PDT-. METHODS: Sixty-five PD patients and 28 healthy controls (HC) were scanned with 11C-MeNER PET. Patients were categorized as PDT+ if subscores in UPDRS-III item 3 or MDS-UPDRS-III item 17 was ≥2; remaining were categorized as PDT-. Simplified reference tissue model 2 distribution volume ratios (DVR) for 11C-MeNER were calculated for thalamus, dorsal and median raphe, locus coeruleus (LC) and red nucleus using time activity curves (TACs) obtained from volumes of interest (VOI). Data were statistically interrogated with a general linear mixed model using 'region', and 'group' as factors and the interaction of 'region x group' was examined. RESULTS: Tremor positive PD patients had a significantly higher mean 11C-MeNER DVR compared to PDT- in LC and thalamus. The PDT+ mean LC DVR was similar to that of HC. PDT+ mean 11C-MeNER DVRs were significantly lower than HC in the dorsal raphe while the PDT- group showed significantly lower mean 11C-MeNER DVR across all regions compared to HC. CONCLUSION: While both PD T+ and PD T- groups showed a significant loss of noradrenaline terminal function compared to controls, noradrenergic neurons were relatively preserved in PDT+ in LC and thalamus. The greater loss of noradrenergic transporters in PDT- in LC and thalamus compared with PDT+ is in line with earlier in-vitro studies and could potentially contribute to their tremor negative phenotype.

Genetic deletion of Polo-like kinase 2 reduces alpha-synuclein serine-129 phosphorylation in presynaptic terminals but not Lewy bodies.

Phosphorylation of alpha-synuclein at serine-129 is an important marker of pathologically relevant, aggregated forms of the protein in several important human diseases, including Parkinson's disease, Dementia with Lewy bodies, and Multiple system atrophy. Although several kinases have been shown to be capable of phosphorylating alpha-synuclein in various model systems, the identity of the kinase that phosphorylates alpha-synuclein in the Lewy body remains unknown. One member of the Polo-like kinase family, PLK2, is a strong candidate for being the Lewy body kinase. To examine this possibility, we have used a combination of approaches, including biochemical, immunohistochemical, and in vivo multiphoton imaging techniques to study the consequences of PLK2 genetic deletion on alpha-synuclein phosphorylation in both the presynaptic terminal and preformed fibril-induced Lewy body pathology in mouse cortex. We find that PLK2 deletion reduces presynaptic terminal alpha-synuclein serine-129 phosphorylation, but has no effect on Lewy body phosphorylation levels. Serine-129 mutation to the phosphomimetic alanine or the unphosphorylatable analog aspartate does not change the rate of cell death of Lewy inclusion-bearing neurons in our in vivo multiphoton imaging paradigm, but PLK2 deletion does slow the rate of neuronal death. Our data indicate that inhibition of PLK2 represents a promising avenue for developing new therapeutics, but that the mechanism of neuroprotection by PLK2 inhibition is not likely due to reducing alpha-synuclein serine-129 phosphorylation and that the true Lewy body kinase still awaits discovery.

Activity-Dependent Remodeling of Synaptic Protein Organization Revealed by High Throughput Analysis of STED Nanoscopy Images.

The organization of proteins in the apposed nanodomains of pre- and postsynaptic compartments is thought to play a pivotal role in synaptic strength and plasticity. As such, the alignment between pre- and postsynaptic proteins may regulate, for example, the rate of presynaptic release or the strength of postsynaptic signaling. However, the analysis of these structures has mainly been restricted to subsets of synapses, providing a limited view of the diversity of synaptic protein cluster remodeling during synaptic plasticity. To characterize changes in the organization of synaptic nanodomains during synaptic plasticity over a large population of synapses, we combined STimulated Emission Depletion (STED) nanoscopy with a Python-based statistical object distance analysis (pySODA), in dissociated cultured hippocampal circuits exposed to treatments driving different forms of synaptic plasticity. The nanoscale organization, characterized in terms of coupling properties, of presynaptic (Bassoon, RIM1/2) and postsynaptic (PSD95, Homer1c) scaffold proteins was differently altered in response to plasticity-inducing stimuli. For the Bassoon - PSD95 pair, treatments driving synaptic potentiation caused an increase in their coupling probability, whereas a stimulus driving synaptic depression had an opposite effect. To enrich the characterization of the synaptic cluster remodeling at the population level, we applied unsupervised machine learning approaches to include selected morphological features into a multidimensional analysis. This combined analysis revealed a large diversity of synaptic protein cluster subtypes exhibiting differential activity-dependent remodeling, yet with common features depending on the expected direction of plasticity. The expanded palette of synaptic features revealed by our unbiased approach should provide a basis to further explore the widely diverse molecular mechanisms of synaptic plasticity.

Brain γ-Tocopherol Levels Are Associated with Presynaptic Protein Levels in Elderly Human Midfrontal Cortex.

BACKGROUND: Higher vitamin E intake has been widely related to lower risks of cognitive decline and dementia. Animal models suggest that this relationship might be (partially) explained by the protection of vitamin E against presynaptic protein oxidation. OBJECTIVE: In this cross-sectional study, we aimed to examine the associations between brain tocopherols and presynaptic protein levels in elderly humans. METHODS: We examined associations of α- and γ-tocopherol brain levels with presynaptic protein levels in 113 deceased participants (age 88.5±6.0 years, 45 (40%) female) from the prospective Memory and Aging project. Three distinct presynaptic proteins, a SNARE protein composite, a synaptotagmin synaptophysin composite and the protein-protein interaction between synaptosomal-associated protein 25 (SNAP-25), and syntaxin were measured in two cortical brain regions. Linear regression models assessed associations of brain tocopherols with presynaptic protein levels. RESULTS: Higher brain γ-tocopherol levels were associated with higher levels of the SNARE protein composite, complexin-I, complexin-II, the synaptotagmin synaptophysin composite, and septin-5 in the midfrontal cortex (B(SE) = 0.272 to 0.412 (0.084 to 0.091), p < 0.001 to 0.003). When additionally adjusted for global Alzheimer's disease pathology, cerebral infarcts, and Lewy body disease pathology, these associations remained largely similar. No associations were found between α-tocopherol and presynaptic protein levels. CONCLUSION: In this cross-sectional study, we found higher brain γ-tocopherol levels were associated with presynaptic protein levels in the midfrontal cortex. These results are consistent with a proposed role of vitamin E to maintain presynaptic protein levels.

Passive Transfer of Sera from ALS Patients with Identified Mutations Evokes an Increased Synaptic Vesicle Number and Elevation of Calcium Levels in Motor Axon Terminals, Similar to Sera from Sporadic Patients.

Previously, we demonstrated increased calcium levels and synaptic vesicle densities in the motor axon terminals (MATs) of sporadic amyotrophic lateral sclerosis (ALS) patients. Such alterations could be conferred to mice with an intraperitoneal injection of sera from these patients or with purified immunoglobulin G. Later, we confirmed the presence of similar alterations in the superoxide dismutase 1 G93A transgenic mouse strain model of familial ALS. These consistent observations suggested that calcium plays a central role in the pathomechanism of ALS. This may be further reinforced by completing a similar analytical study of the MATs of ALS patients with identified mutations. However, due to the low yield of muscle biopsy samples containing MATs, and the low incidence of ALS patients with the identified mutations, these examinations are not technically feasible. Alternatively, a passive transfer of sera from ALS patients with known mutations was used, and the MATs of the inoculated mice were tested for alterations in their calcium homeostasis and synaptic activity. Patients with 11 different ALS-related mutations participated in the study. Intraperitoneal injection of sera from these patients on two consecutive days resulted in elevated intracellular calcium levels and increased vesicle densities in the MATs of mice, which is comparable to the effect of the passive transfer from sporadic patients. Our results support the idea that the pathomechanism underlying the identical manifestation of the disease with or without identified mutations is based on a common final pathway, in which increasing calcium levels play a central role.

Neuroprotective activity of ursodeoxycholic acid in CHMP2BIntron5 models of frontotemporal dementia.

Frontotemporal dementia (FTD) is one of the most prevalent forms of early-onset dementia. It represents part of the FTD-Amyotrophic Lateral Sclerosis (ALS) spectrum, a continuum of genetically and pathologically overlapping disorders. FTD-causing mutations in CHMP2B, a gene encoding a core component of the heteromeric ESCRT-III Complex, lead to perturbed endosomal-lysosomal and autophagic trafficking with impaired proteostasis. While CHMP2B mutations are rare, dysfunctional endosomal-lysosomal signalling is common across the FTD-ALS spectrum. Using our established Drosophila and mammalian models of CHMP2BIntron5 induced FTD we demonstrate that the FDA-approved compound Ursodeoxycholic Acid (UDCA) conveys neuroprotection, downstream of endosomal-lysosomal dysfunction in both Drosophila and primary mammalian neurons. UDCA exhibited a dose dependent rescue of neuronal structure and function in Drosophila pan-neuronally expressing CHMP2BIntron5. Rescue of CHMP2BIntron5 dependent dendritic collapse and apoptosis with UDCA in rat primary neurons was also observed. UDCA failed to ameliorate aberrant accumulation of endosomal and autophagic organelles or ubiquitinated neuronal inclusions in both models. We demonstrate the neuroprotective activity of UDCA downstream of endosomal-lysosomal and autophagic dysfunction, delineating the molecular mode of action of UDCA and highlighting its potential as a therapeutic for the treatment of FTD-ALS spectrum disorders.

A candidate gene study of intermediate histopathological phenotypes in HIV-associated neurocognitive disorders.

HIV-associated neurocognitive disorders (HAND) describe a spectrum of neuropsychological impairment caused by HIV-1 infection. While the sequence of cellular and physiological events that lead to HAND remains obscure, it likely involves chronic neuroinflammation. Host genetic markers that increase the risk for HAND have been reported, but replication of such studies is lacking, possibly due to inconsistent application of a behavioral phenotype across studies. In the current study, we used histopathologic phenotypes in order to validate putative risk alleles for HAND. The National NeuroAIDS Tissue Consortium, a longitudinal study of the neurologic manifestations of HIV. Data and specimens were obtained from 175 HIV-infected adults. After determining several potential covariates of neurocognitive functioning, we quantified levels of six histopathological markers in the frontal lobe in association with neurocognitive functioning: SYP, MAP 2, HLA-DR, Iba1, GFAP, and ß-amyloid. We then determined alleles of 15 candidate genes for their associations with neurocognitive functioning and histopathological markers. Finally, we identified the most plausible causal pathway based on our data using a multi-stage linear regression-based mediation analysis approach. None of the genetic markers were associated with neurocognitive functioning. Of the histopathological markers, only MAP 2 and SYP were associated with neurocognitive functioning; however, MAP 2 and SYP did not vary as a function of genotype. Mediation analysis suggests a causal pathway in which presynaptic degeneration (SYP) leads to somatodendritic degeneration (MAP 2) and ultimately neurocognitive impairment. This study did not support the role of host genotype in the histopathology underlying HAND. The findings lend further support for synaptodendritic degeneration as the proximal underlying neuropathological substrate of HAND.

Perisynaptic Schwann cells phagocytose nerve terminal debris in a mouse model of Guillain-Barré syndrome.

In mouse models of acute motor axonal neuropathy, anti-ganglioside antibodies (AGAbs) bind to motor axons, notably the distal nerve, and activate the complement cascade. While complement activation is well studied in this model, the role of inflammatory cells is unknown. Herein we aimed to investigate the contribution of phagocytic cells including macrophages, neutrophils and perisynaptic Schwann cells (pSCs) to distal nerve pathology. To observe this, we first created a subacute injury model of sufficient duration to allow inflammatory cell recruitment. Mice were injected intraperitoneally with an anti-GD1b monoclonal antibody that binds strongly to mouse motor nerve axons. Subsequently, mice received normal human serum as a source of complement. Dosing was titrated to allow humane survival of mice over a period of 3 days, yet still induce the characteristic neurological impairment. Behaviour and pathology were assessed in vivo using whole-body plethysmography and post-sacrifice by immunofluorescence and flow cytometry. ex vivo nerve-muscle preparations were used to investigate the acute phagocytic role of pSCs following distal nerve injury. Following complement activation at distal intramuscular nerve sites in the diaphragm macrophage localisation or numbers are not altered, nor do they shift to a pro- or anti-inflammatory phenotype. Similarly, neutrophils are not significantly recruited. Instead, ex vivo nerve-muscle preparations exposed to AGAb plus complement reveal that pSCs rapidly become phagocytic and engulf axonal debris. These data suggest that pSCs, rather than inflammatory cells, are the major cellular vehicle for axonal debris clearance following distal nerve injury, in contrast to larger nerve bundles where macrophage-mediated clearance predominates.

Presynaptic Caytaxin prevents apoptosis via deactivating DAPK1 in the acute phase of cerebral ischemic stroke.

Death-associated protein kinase 1 (DAPK1) is a key protein that mediates neuronal death in ischemic stroke. Although the substrates of DAPK1 and molecular signal in stroke have been gradually discovered, the modulation of DAPK1 itself is still unclear. Here we first reveal that Caytaxin, a brain-specific member of BCL2/adenovirus E1B -interacting protein (BNIP-2), increases and interacts with DAPK1 as early as 2 h after middle cerebral artery occlusion (MCAO) in the penumbra area of mouse brain. Furthermore, Caytaxin binds to DAPK1 at the presynaptic site and inhibits DAPK1 catalytic activity. Silencing Caytaxin by Caytaxin shRNA (Sh-Caytaxin) enhances DAPK1 activity, deteriorates neuronal apoptosis and brain injuries both in vivo and in vitro. Thus, elevating presynaptic Caytaxin could prevent neuronal apoptosis by inhibiting DAPK1 activation in the acute stage of ischemic stroke. Caytaxin may physiologically protect neuronal cells and represent a potential prevention and therapeutic target in the early phase of cerebral ischemic stroke.