Assessment of the effects of plaques and glia on synaptic transmission in App knock-in models of Alzheimer’s disease

Alzheimer’s disease (AD) is a neurodegenerative disorder associated with the formation of extracellular amyloid plaques and intracellular neurofibrillary tangles, coupled to extensive synaptic denervation and gliosis. The role of microglia in disease progression has become a central focus for AD research with studies showing that they cluster around amyloid plaques and some indications that plaque-associated microglia phagocytose synapses in these regions. The present work aimed to explore if this is a protective mechanism to limit plaque-associated toxicity by examining the hippocampus of AppNL-F and AppNL-G-F using patch-clamp electrophysiology and immunohistochemistry. This work also explored AppNL-F/Trem2R47H mice to determine how a compromised microglial response affects synaptic activity. I found that AppNL-G-F mice exhibit heavier plaque pathology with age and a larger number of smaller-sized plaques, similar to what I observed in AppNL-F/Trem2R47H mice in comparison to AppNL-F mice. AppNL-G-F mice also showed increased microglial density compared to wild-types, regardless of age, and no changes in astrocyte density. In comparison, the presence of the Trem2R47H mutation in AppNL-F mice led to reduced microglial density in the SLM region, where the plaque load in these mice was the heaviest, and also reversed the increased astrocytic density which was observed in AppNL-F mice. In terms of electrophysiological changes, I found alterations in the kinetics of the postsynaptic response and changes in the failure rate in both models. Exploring electrophysiological changes in plaque-associated regions revealed that both AppNL-G- F and AppNL-F mice exhibit lower amplitudes in plaque compared to no-plaque conditions, while this effect was lost in AppNL-F/Trem2R47H mice. Furthermore, both AppNL-F and AppNL-F/Trem2R47H mice showed enhanced release probability in plaque conditions. Altogether, these results illustrate the dual effects of plaque presence around axons which are driven by the effects of soluble Aβ and microglia on the presynapse and postsynapse, respectively

Neuroprotective actions of leptin facilitated through balancing mitochondrial morphology and improving mitochondrial function

Authors would like to acknowledge ARUK for supporting this research. YC is Chinese Scholarship recipient. The University of St Andrews is a charity registered in Scotland: No SC013532Mitochondrial dysfunction has a recognised role in the progression of Alzheimer's disease (AD) pathophysiology. Cerebral perfusion becomes increasingly inefficient throughout ageing, leading to unbalanced mitochondrial dynamics. This effect is exaggerated by amyloid β (Aβ) and phosphorylated tau, two hallmark proteins of AD pathology. A neuroprotective role for the adipose‐derived hormone, leptin, has been demonstrated in neuronal cells. However, its effects with relation to mitochondrial function in AD remain largely unknown. To address this question, we have used both a glucose‐serum deprived (CGSD) model of ischaemic stroke in SH‐SY5Y cells and a Aβ1‐42‐treatment model of AD in differentiated hippocampal cells. Using a combination of JC‐1 and MitoRed staining techniques, we show that leptin prevents depolarisation of the mitochondrial membrane and excessive mitochondrial fragmentation induced by both CGSD and Aβ1‐42. Thereafter, we used ELISAs and a number of activity assays to reveal the biochemical underpinnings of these processes. Specifically, leptin was seen to inhibit upregulation of the mitochondrial fission protein Fis1 and downregulation of the mitochondrial fusion protein, Mfn2. Furthermore, leptin was seen to upregulate the expression and activity of the antioxidant enzyme, monoamine oxidase B. Herein we provide the first demonstration that leptin is sufficient to protect against aberrant mitochondrial dynamics and resulting loss of function induced by both CGSD and Aβ1‐42. We conclude that the established neuroprotective actions of leptin may be facilitated through regulation of mitochondrial dynamics.Publisher PDFPeer reviewe

Upregulation of Trem2 expression occurs exclusively on microglial contact with plaques

Using spatial cell-type-enriched transcriptomics, we compare plaque-induced gene (PIG) expression in microglia touching plaques, neighboring plaques, and far from plaques in 18-month-old APPNLF/NLF knock-in mice with and without the Alzheimer’s disease risk mutation Trem2R47H/R47H. We report that, in AppNLF/NLF mice, expression of 35/55 PIGs, is exclusively upregulated in microglia that are touching plaques. In 7 PIGs including Trem2 this upregulation is prevented by the Trem2R47H/R47H mutation. Unlike in young mice, knockin of the Trem2R47H/R47H mutation does not significantly decrease the Trem2 expression but decreases protein levels by 20% in the absence of plaques. On plaques, despite the mutation preventing increased gene expression, TREM2 protein levels increased by 1.6-fold (compared to 3-fold with Trem2WT/WT) and microglial density increased 20-fold compared to 30-fold. Hence microglia must touch plaques before Trem2 gene expression is increased but small changes in protein expression can increase microglia density without a change in gene expression

Plaque contact and unimpaired Trem2 is required for the microglial response to amyloid pathology

Using spatial cell-type-enriched transcriptomics, we compare plaque-induced gene (PIG) expression in microglia-touching plaques, neighboring plaques, and far from plaques in an aged Alzheimer’s mouse model with late plaque development. In 18-month-old APPNL-F/NL-F knockin mice, with and without the Alzheimer’s disease risk mutation Trem2R47H/R47H, we report that expression of 38/55 PIGs have plaque-induced microglial upregulation, with a subset only upregulating in microglia directly contacting plaques. For seven PIGs, including Trem2, this upregulation is prevented in APPNL-F/NL-FTrem2R47H/R47H mice. These TREM2-dependent genes are all involved in phagocytic and degradative processes that we show correspond to a decrease in phagocytic markers and an increase in the density of small plaques in Trem2-mutated mice. Furthermore, despite the R47H mutation preventing increased Trem2 gene expression, TREM2 protein levels and microglial density are still marginally increased on plaques. Hence, both microglial contact with plaques and functioning TREM2 are necessary for microglia to respond appropriately to amyloid patholog

Knock-in models related to Alzheimer’s disease: synaptic transmission, plaques and the role of microglia

Funder: Cure Alzheimer's Fund; doi: http://dx.doi.org/10.13039/100007625Funder: UK Dementia Research Institute (GB)Funder: Censejo Nacional de Ciencia Tecnilogia (MX)Funder: Alzheimerfonden; doi: http://dx.doi.org/10.13039/501100008599Funder: Dolby Family FundAbstract: Background: Microglia are active modulators of Alzheimer’s disease but their role in relation to amyloid plaques and synaptic changes due to rising amyloid beta is unclear. We add novel findings concerning these relationships and investigate which of our previously reported results from transgenic mice can be validated in knock-in mice, in which overexpression and other artefacts of transgenic technology are avoided. Methods: AppNL-F and AppNL-G-F knock-in mice expressing humanised amyloid beta with mutations in App that cause familial Alzheimer’s disease were compared to wild type mice throughout life. In vitro approaches were used to understand microglial alterations at the genetic and protein levels and synaptic function and plasticity in CA1 hippocampal neurones, each in relationship to both age and stage of amyloid beta pathology. The contribution of microglia to neuronal function was further investigated by ablating microglia with CSF1R inhibitor PLX5622. Results: Both App knock-in lines showed increased glutamate release probability prior to detection of plaques. Consistent with results in transgenic mice, this persisted throughout life in AppNL-F mice but was not evident in AppNL-G-F with sparse plaques. Unlike transgenic mice, loss of spontaneous excitatory activity only occurred at the latest stages, while no change could be detected in spontaneous inhibitory synaptic transmission or magnitude of long-term potentiation. Also, in contrast to transgenic mice, the microglial response in both App knock-in lines was delayed until a moderate plaque load developed. Surviving PLX5266-depleted microglia tended to be CD68-positive. Partial microglial ablation led to aged but not young wild type animals mimicking the increased glutamate release probability in App knock-ins and exacerbated the App knock-in phenotype. Complete ablation was less effective in altering synaptic function, while neither treatment altered plaque load. Conclusions: Increased glutamate release probability is similar across knock-in and transgenic mouse models of Alzheimer’s disease, likely reflecting acute physiological effects of soluble amyloid beta. Microglia respond later to increased amyloid beta levels by proliferating and upregulating Cd68 and Trem2. Partial depletion of microglia suggests that, in wild type mice, alteration of surviving phagocytic microglia, rather than microglial loss, drives age-dependent effects on glutamate release that become exacerbated in Alzheimer’s disease