Primary Phagocytosis of Neurons by Inflamed Microglia: Potential Roles in Neurodegeneration

Microglial phagocytosis of dead or dying neurons can be beneficial by preventing the release of damaging and/or pro-inflammatory intracellular components. However, there is now evidence that under certain conditions, such as inflammation, microglia can also phagocytose viable neurons, thus executing their death. Such phagocytic cell death may result from exposure of phosphatidylserine (PS) or other eat-me signals on otherwise viable neurons as a result of physiological activation or sub-toxic insult, and neuronal phagocytosis by activated microglia. In this review, we discuss the mechanisms of phagocytic cell death and its potential roles in Alzheimer’s Disease, Parkinson’s Disease, and Frontotemporal Dementia

Epigenetic control of microglial immune responses

Microglia, the major population of brain-resident macrophages, are now recognized as a heterogeneous population comprising several cell subtypes with different (so far mostly supposed) functions in health and disease. A number of studies have performed molecular characterization of these different microglial activation states over the last years making use of omics technologies, that is transcriptomics, proteomics and, less frequently, epigenomics profiling. These approaches offer the possibility to identify disease mechanisms, discover novel diagnostic biomarkers, and develop new therapeutic strategies. Here, we focus on epigenetic profiling as a means to understand microglial immune responses beyond what other omics methods can offer, that is, revealing past and present molecular responses, gene regulatory networks and potential future response trajectories, and defining cell subtype-specific disease relevance through mapping non-coding genetic variants. We review the current knowledge in the field regarding epigenetic regulation of microglial identity and function, provide an exemplary analysis that demonstrates the advantages of performing joint transcriptomic and epigenomic profiling of single microglial cells and discuss how comprehensive epigenetic analyses may enhance our understanding of microglial pathophysiology

The fluorescent pentameric oligothiophene pFTAA identifies filamentous tau in live neurons cultured from adult P301S tau mice.

Identification of fluorescent dyes that label the filamentous protein aggregates characteristic of neurodegenerative disease, such as β-amyloid and tau in Alzheimer's disease, in a live cell culture system has previously been a major hurdle. Here we show that pentameric formyl thiophene acetic acid (pFTAA) fulfills this function in living neurons cultured from adult P301S tau transgenic mice. Injection of pFTAA into 5-month-old P301S tau mice detected cortical and DRG neurons immunoreactive for AT100, an antibody that identifies solely filamentous tau, or MC1, an antibody that identifies a conformational change in tau that is commensurate with neurofibrillary tangle formation in Alzheimer's disease brains. In fixed cultures of dorsal root ganglion (DRG) neurons, pFTAA binding, which also identified AT100 or MC1+ve neurons, followed a single, saturable binding curve with a half saturation constant of 0.14 μM, the first reported measurement of a binding affinity of a beta-sheet reactive dye to primary neurons harboring filamentous tau. Treatment with formic acid, which solubilizes filamentous tau, extracted pFTAA, and prevented the re-binding of pFTAA and MC1 without perturbing expression of soluble tau, detected using an anti-human tau (HT7) antibody. In live cultures, pFTAA only identified DRG neurons that, after fixation, were AT100/MC1+ve, confirming that these forms of tau pre-exist in live neurons. The utility of pFTAA to discriminate between living neurons containing filamentous tau from other neurons is demonstrated by showing that more pFTAA+ve neurons die than pFTAA-ve neurons over 25 days. Since pFTAA identifies fibrillar tau and other misfolded proteins in living neurons in culture and in animal models of several neurodegenerative diseases, as well as in human brains, it will have considerable application in sorting out disease mechanisms and in identifying disease-modifying drugs that will ultimately help establish the mechanisms of neurodegeneration in human neurodegenerative diseases.This work was funded by grant number NC/L000741/1 from the National Council of the 3Rs (NC3Rs) and Alzheimer's Research UK (ARUK). KPR Nilsson is funded by an ERC Starting Independent Researcher Grant (Project: MUMID, number 260604).This is the final published version of the article. It was originally published in Frontiers in Neuroscience (Brelstaff J, Ossola B, Neher JJ, Klingstedt T, Nilsson KPR, Goedert M, Spillantini MG, Tolkovsky AM, Frontiers in Neuroscience, 2015, 9:184, doi:10.3389/fnins.2015.00184). The final version is available at http://dx.doi.org/10.3389/fnins.2015.0018

Endogenous but not sensory-driven activity controls migration, morphogenesis and survival of adult-born juxtaglomerular neurons in the mouse olfactory bulb.

The development and survival of adult-born neurons are believed to be driven by sensory signaling. Here, in vivo analyses of motility, morphology and Ca2+ signaling, as well as transcriptome analyses of adult-born juxtaglomerular cells with reduced endogenous excitability (via cell-specific overexpression of either Kv1.2 or Kir2.1 K+ channels), revealed a pronounced impairment of migration, morphogenesis, survival, and functional integration of these cells into the mouse olfactory bulb, accompanied by a reduction in cytosolic Ca2+ fluctuations, phosphorylation of CREB and pCREB-mediated gene expression. Moreover, K+ channel overexpression strongly downregulated genes involved in neuronal migration, differentiation, and morphogenesis and upregulated apoptosis-related genes, thus locking adult-born cells in an immature and vulnerable state. Surprisingly, cells deprived of sensory-driven activity developed normally. Together, the data reveal signaling pathways connecting the endogenous intermittent neuronal activity/Ca2+ fluctuations as well as enhanced Kv1.2/Kir2.1 K+ channel function to migration, maturation, and survival of adult-born neurons

Endogenous but Not Sensory-Driven Activity Controls Migration, Morphogenesis and Survival of Adult-Born Juxtaglomerular Neurons in the Mouse Olfactory Bulb

The development and survival of adult-born neurons are believed to be driven by sensory signaling. Here, in vivo analyses of motility, morphology and Ca2+ signaling, as well as transcriptome analyses of adult-born juxtaglomerular cells with reduced endogenous excitability (via cell-specific overexpression of either Kv1.2 or Kir2.1 K+ channels), revealed a pronounced impairment of migration, morphogenesis, survival, and functional integration of these cells into the mouse olfactory bulb, accompanied by a reduction in cytosolic Ca2+ fluctuations, phosphorylation of CREB and pCREB-mediated gene expression. Moreover, K+ channel overexpression strongly downregulated genes involved in neuronal migration, differentiation, and morphogenesis and upregulated apoptosis-related genes, thus locking adult-born cells in an immature and vulnerable state. Surprisingly, cells deprived of sensory-driven activity developed normally. Together, the data reveal signaling pathways connecting the endogenous intermittent neuronal activity/Ca2+ fluctuations as well as enhanced Kv1.2/Kir2.1 K+ channel function to migration, maturation, and survival of adult-born neurons

Electrotonic Signals along Intracellular Membranes May Interconnect Dendritic Spines and Nucleus

Synapses on dendritic spines of pyramidal neurons show a remarkable ability to induce phosphorylation of transcription factors at the nuclear level with a short latency, incompatible with a diffusion process from the dendritic spines to the nucleus. To account for these findings, we formulated a novel extension of the classical cable theory by considering the fact that the endoplasmic reticulum (ER) is an effective charge separator, forming an intrinsic compartment that extends from the spine to the nuclear membrane. We use realistic parameters to show that an electrotonic signal may be transmitted along the ER from the dendritic spines to the nucleus. We found that this type of signal transduction can additionally account for the remarkable ability of the cell nucleus to differentiate between depolarizing synaptic signals that originate from the dendritic spines and back-propagating action potentials. This study considers a novel computational role for dendritic spines, and sheds new light on how spines and ER may jointly create an additional level of processing within the single neuron

Alterations in voltage-sensing of the mitochondrial permeability transition pore in ANT1-deficient cells

The probability of mitochondrial permeability transition (mPT) pore opening is inversely related to the magnitude of the proton electrochemical gradient. The module conferring sensitivity of the pore to this gradient has not been identified. We investigated mPT's voltage-sensing properties elicited by calcimycin or H2O2 in human fibroblasts exhibiting partial or complete lack of ANT1 and in C2C12 myotubes with knocked-down ANT1 expression. mPT onset was assessed by measuring in situ mitochondrial volume using the 'thinness ratio' and the 'cobalt-calcein' technique. De-energization hastened calcimycin-induced swelling in control and partially-expressing ANT1 fibroblasts, but not in cells lacking ANT1, despite greater losses of mitochondrial membrane potential. Matrix Ca(2+) levels measured by X-rhod-1 or mitochondrially-targeted ratiometric biosensor 4mtD3cpv, or ADP-ATP exchange rates did not differ among cell types. ANT1-null fibroblasts were also resistant to H2O2-induced mitochondrial swelling. Permeabilized C2C12 myotubes with knocked-down ANT1 exhibited higher calcium uptake capacity and voltage-thresholds of mPT opening inferred from cytochrome c release, but intact cells showed no differences in calcimycin-induced onset of mPT, irrespective of energization and ANT1 expression, albeit the number of cells undergoing mPT increased less significantly upon chemically-induced hypoxia than control cells. We conclude that ANT1 confers sensitivity of the pore to the electrochemical gradient

Alterations in voltage-sensing of the mitochondrial permeability transition pore in ANT1-deficient cells

The probability of mitochondrial permeability transition (mPT) pore opening is inversely related to the magnitude of the proton electrochemical gradient. The module conferring sensitivity of the pore to this gradient has not been identified. We investigated mPT's voltage-sensing properties elicited by calcimycin or H2O2 in human fibroblasts exhibiting partial or complete lack of ANT1 and in C2C12 myotubes with knocked-down ANT1 expression. mPT onset was assessed by measuring in situ mitochondrial volume using the 'thinness ratio' and the 'cobalt-calcein' technique. De-energization hastened calcimycin-induced swelling in control and partially-expressing ANT1 fibroblasts, but not in cells lacking ANT1, despite greater losses of mitochondrial membrane potential. Matrix Ca(2+) levels measured by X-rhod-1 or mitochondrially-targeted ratiometric biosensor 4mtD3cpv, or ADP-ATP exchange rates did not differ among cell types. ANT1-null fibroblasts were also resistant to H2O2-induced mitochondrial swelling. Permeabilized C2C12 myotubes with knocked-down ANT1 exhibited higher calcium uptake capacity and voltage-thresholds of mPT opening inferred from cytochrome c release, but intact cells showed no differences in calcimycin-induced onset of mPT, irrespective of energization and ANT1 expression, albeit the number of cells undergoing mPT increased less significantly upon chemically-induced hypoxia than control cells. We conclude that ANT1 confers sensitivity of the pore to the electrochemical gradient