mTORC1 Signaling Pathway Mediates Chronic Stress-Induced Synapse Loss in the Hippocampus

Background: The mechanistic target of rapamycin complex 1 (mTORC1) signaling has served as a promising target for therapeutic intervention of major depressive disorder (MDD), but the mTORC1 signaling underlying MDD has not been well elucidated. In the present study, we investigated whether mTORC1 signaling pathway mediates synapse loss induced by chronic stress in the hippocampus.Methods: Chronic restraint stress-induced depression-like behaviors were tested by behavior tests (sucrose preference test, forced swim test and tail suspension test). Synaptic proteins and alternations of phosphorylation levels of mTORC1 signaling-associated molecules were measured using Western blotting. In addition, mRNA changes of immediate early genes (IEGs) and glutamate receptors were measured by RT-PCR. Rapamycin was used to explore the role of mTORC1 signaling in the antidepressant effects of fluoxetine.Results: After successfully establishing the chronic restraint stress paradigm, we observed that the mRNA levels of some IEGs were significantly changed, indicating the activation of neurons and protein synthesis alterations. Then, there was a significant downregulation of glutamate receptors and postsynaptic density protein 95 at protein and mRNA levels. Additionally, synaptic fractionation assay revealed that chronic stress induced synapse loss in the dorsal and ventral hippocampus. Furthermore, these effects were associated with the mTORC1 signaling pathway-mediated protein synthesis, and subsequently the phosphorylation of associated downstream signaling targets was reduced after chronic stress. Finally, we found that intracerebroventricular infusion of rapamycin simulated depression-like behavior and also blocked the antidepressant effects of fluoxetine.Conclusion: Overall, our study suggests that mTORC1 signaling pathway plays a critical role in mediating synapse loss induced by chronic stress, and has part in the behavioral effects of antidepressant treatment.

Friend or Foe? The Varied Faces of Homeostatic Synaptic Plasticity in Neurodegenerative Disease

Homeostatic synaptic plasticity (HSP) regulates synaptic strength both pre- and postsynaptically to ensure stability and efficient information transfer in neural networks. A number of neurological diseases have been associated with deficits in HSP, particularly diseases characterised by episodic network instability such as migraine and epilepsy. Recently, it has become apparent that HSP also plays a role in many neurodegenerative diseases. In this mini review, we present an overview of the evidence linking HSP to each of the major neurodegenerative diseases, finding that HSP changes in each disease appear to belong to one of three broad functional categories: (1) deficits in HSP at degenerating synapses that contribute to pathogenesis or progression; (2) HSP induced in a heterosynaptic or cell non-autonomous manner to support the function of networks of which the degenerating synapses or cells are part; and (3) induction of HSP within the degenerating population of synapses to preserve function and to resist the impact of synapse loss. Understanding the varied manifestations of HSP in neurodegeneration will not only aid understanding mechanisms of disease but could also inspire much-needed novel approaches to therapy.

Microglia Mediate Early Corticostriatal Synapse Loss and Cognitive Dysfunction in Huntington’s Disease Through Complement-Dependent Mechanisms

Huntington’s disease (HD) is a devastating monogenic neurodegenerative disease characterized by early, selective pathology in the basal ganglia despite the ubiquitous expression of mutant huntingtin. The molecular mechanisms underlying this region-specific neuronal degeneration and how these relate to the development of early cognitive phenotypes are poorly understood. Here, we show that there is selective loss of synaptic connections between the cortex and striatum in postmortem tissue from HD patients that is associated with the increased activation and localization of complement proteins, innate immune molecules, to markers of these synaptic elements. We also find that levels of these secreted innate immune molecules are elevated in the CSF of premanifest HD patients and correlate with established measures of disease burden.In preclinical genetic models of HD we show that complement proteins mediate the selective elimination of corticostriatal synapses at an early stage in disease pathogenesis marking them for removal by microglia, the brain’s resident macrophage population. This process requires mutant huntingtin to be expressed in both cortical and striatal neurons and inhibition of this complement-dependent elimination mechanism through administration of a therapeutically relevant C1q function blocking antibody or genetic ablation of a complement receptor on microglia, prevented synapse loss, increased excitatory input to the striatum and rescued the early development of visual discrimination learning and cognitive flexibility deficits in these models. Together, our findings implicate microglia and the complement cascade in the selective, early degeneration of corticostriatal synapses and the development of cognitive deficits in presymptomatic HD, and also provide new preclinical data to support complement as a therapeutic target for early intervention.

Stroke subtype-dependent synapse elimination by reactive gliosis in mice

The pathological role of reactive gliosis in CNS repair remains controversial. In this study, using murine ischemic and hemorrhagic stroke models, we demonstrated that microglia/macrophages and astrocytes are differentially involved in engulfing synapses in the reactive gliosis region. By specifically deleting MEGF10 and MERTK phagocytic receptors, we determined that inhibiting phagocytosis of microglia/macrophages or astrocytes in ischemic stroke improved neurobehavioral outcomes and attenuated brain damage. In hemorrhagic stroke, inhibiting phagocytosis of microglia/macrophages but not astrocytes improved neurobehavioral outcomes. Single-cell RNA sequencing revealed that phagocytosis related biological processes and pathways were downregulated in astrocytes of the hemorrhagic brain compared to the ischemic brain. Together, these findings suggest that reactive microgliosis and astrogliosis play individual roles in mediating synapse engulfment in pathologically distinct murine stroke models and preventing this process could rescue synapse loss.

Reducing voltage-dependent potassium channel Kv3.4 levels ameliorates synapse loss in a mouse model of Alzheimers disease

Synapse loss is associated with cognitive decline in Alzheimers disease (AD) and owing to their plastic nature, synapses are an ideal target for therapeutic intervention. Oligomeric amyloid beta (Ab) around amyloid plaques is known to contribute to synapse loss in mouse models and is associated with synapse loss in human AD brain tissue, but the mechanisms leading from Ab; to synapse loss remain unclear. Recent data suggest that the fast-activating and -inactivating voltage-gated potassium channel subtype 3.4 (Kv3.4) may play a role in Ab-mediated neurotoxicity. Here, we tested whether this channel could also be involved in Ab synaptotoxicity. Using adeno-associated virus and CRISPR (clustered regularly interspaced short palindromic repeats) technology, we reduced Kv3.4 expression in neurons of the somatosensory cortex of APP/PS1 mice. These mice express human familial AD associated mutations in amyloid precursor protein and presenilin 1 and develop amyloid plaques and plaque-associated synapse loss similar to that observed in AD brain. We observe that reducing Kv3.4 levels ameliorates dendritic spine loss and changes spine morphology compared to control virus. In support of translational relevance, Kv3.4 protein was observed in human AD and control brain and is associated with synapses in human iPSC-derived cortical neurons. Interestingly, we observe a decrease in Kv3.4 expression in iPSC derived cortical neurons when they are challenged with human Alzheimers disease derived brain homogenate. These results suggest that approaches to reduce Kv3.4 expression and/or function could be protective against Ab-induced synaptic alterations.

Unbiased high-content screening reveals Aβ- and tau-independent synaptotoxic activities in human brain homogenates from Alzheimer’s patients and high-pathology controls

Alzheimer’s disease (AD) is tightly correlated with synapse loss in vulnerable brain regions. It is assumed that specific molecular entities such as Aβ and tau cause synapse loss in AD, yet unbiased screens for synaptotoxic activities have not been performed. Here, we performed size exclusion chromatography on soluble human brain homogenates from AD cases, high pathology non-demented controls, and low pathology age-matched controls using our novel high content primary cultured neuron-based screening assay. Both presynaptic and postsynaptic toxicities were elevated in homogenates from AD cases and high pathology non-demented controls to a similar extent, with more modest synaptotoxic activities in homogenates from low pathology normal controls. Surprisingly, synaptotoxic activities were found in size fractions peaking between the 17–44 kDa size standards that did not match well with Aβ and tau immunoreactive species in these homogenates. The fractions containing previously identified high molecular weight soluble amyloid beta aggregates/”oligomers” were non-toxic in this assay. Furthermore, immunodepletion of Aβ and tau did not reduce synaptotoxic activity. This result contrasts with previous findings involving the same methods applied to 3xTg-AD mouse brain extracts. The nature of the synaptotoxic species has not been identified. Overall, our data indicates one or more potential Aβ and tau independent synaptotoxic activities in human AD brain homogenates. This result aligns well with the key role of synaptic loss in the early cognitive decline and may provide new insight into AD pathophysiology.

Mechanical overstimulation causes acute injury and synapse loss followed by fast recovery in lateral-line neuromasts of larval zebrafish

Excess noise damages sensory hair cells, resulting in loss of synaptic connections with auditory nerves and, in some cases, hair-cell death. The cellular mechanisms underlying mechanically induced hair-cell damage and subsequent repair are not completely understood. Hair cells in neuromasts of larval zebrafish are structurally and functionally comparable to mammalian hair cells but undergo robust regeneration following ototoxic damage. We therefore developed a model for mechanically induced hair-cell damage in this highly tractable system. Free swimming larvae exposed to strong water wave stimulus for 2 hours displayed mechanical injury to neuromasts, including afferent neurite retraction, damaged hair bundles, and reduced mechanotransduction. Synapse loss was observed in apparently intact exposed neuromasts, and this loss was exacerbated by inhibiting glutamate uptake. Mechanical damage also elicited an inflammatory response and macrophage recruitment. Remarkably, neuromast hair-cell morphology and mechanotransduction recovered within hours following exposure, suggesting severely damaged neuromasts undergo repair. Our results indicate functional changes and synapse loss in mechanically damaged lateral-line neuromasts that share key features of damage observed in noise-exposed mammalian ear. Yet, unlike the mammalian ear, mechanical damage to neuromasts is rapidly reversible.

URMC-099 Prophylaxis Repairs Hippocampal Vascular Damage in an Orthopedic Model of Delirium Superimposed on Dementia

Systemic perturbations can drive a neuroimmune cascade after surgical trauma, including affecting the blood-brain barrier (BBB), activating microglia, and contributing to cognitive deficits such as delirium. Delirium superimposed on dementia (DSD) is a particularly debilitating complication that renders the brain further vulnerable to neuroinflammation and neurodegeneration, albeit these molecular mechanisms remain poorly understood. Here we have used an orthopedic model of tibial fracture/fixation in APPSwDI/mNos2-/- AD (CVN-AD) mice to investigate relevant pathogenetic mechanisms underlying DSD. We conducted the present study in 6 months-old CVN-AD mice, an age at which we speculated amyloid-β pathology had not saturated BBB and neuroimmune functioning. We found that URMC-099, our brain-penetrant anti-inflammatory neuroprotective drug, prevented inflammatory endothelial activation, synapse loss, and microglial activation in our DSD model. Taken together, our data link post-surgical endothelial activation, microglial MafB immunoreactivity, and synapse loss as key substrates for DSD, all of which can be reversed by URMC-099.