Microglia not only gobble up detritus from dead cells, they also have a penchant for chowing down on live ones. At a Keystone joint symposia—Neurodegenerative Diseases: New Insights and Therapeutic Opportunities and Neural Environment in Disease: Glial Responses and Neuroinflammation—held June 16–21 in Keystone, Colorado, researchers described how microglia use different receptors to sense neurons that are stressed, dying, or behaving oddly. Microglia start eating their synapses, or even the entire cell. Sometimes the purging seems justified, scientists reported, for example when microglia use the TAM receptors Axl and Mer to clean up apoptotic debris, plaques, and overactive neurons. Other times, microglia take it too far, killing off functional neurons and synapses and impairing cognition. Researchers proposed strategies to block these overzealous microglia, which run amok when neurons are most stressed.
- Stress, or aberrant neuronal activity, may bait microglia early in disease.
- Microglial TAM receptors mediate removal of apoptotic neurons.
- Microglia over-prune synapses early in Huntington’s disease.
Microglial pruning of neurons is crucial during development, when these cells sculpt fledgling synaptic circuits. Their appetite for neurons is helpful during infections or injury, when they dispatch infected neurons or mop up apoptotic debris. However, if the immune cells nibble neurons that are merely stressed, they may contribute to neurodegeneration.
For example, scientists had previously reported that P2Y6—a purinoreceptor primarily known for its role in neuropathic pain—recognizes uridine 5'diphosphate (UDP), which is released from injured or stressed neurons (Koizumi et al., 2007). By blocking P2Y6 with the small molecule antagonist MRS2578, they spared neurons from an untimely death following intracerebral injection of lipopolysaccharide into rats. In culture, MRS2578 protected neurons exposed to Aβ from being eaten by overzealous microglia (Neher et al., 2014). At Keystone, Guy Brown of the University of Cambridge, U.K., extended this line of investigation to models of neurodegeneration and aging. He found that PY26 deficiency spared mice from neuronal loss and memory deficits induced by injection of Aβ oligomers, by tau pathology, or by the aging process itself.
Yajing Xu, University College London, wondered how sparing dying neurons from microglial engulfment could be beneficial, given that those neurons would remain dysfunctional or perhaps die soon. Brown replied that neurons releasing UDP might only be stressed, and not destined to die at that time. Similarly, Youtong Huang of the Salk Institute, San Diego, questioned whether deleting or blocking P2Y6 might lead to a build-up of apoptotic cells. Brown said that he has not checked for an accumulation of apoptotic debris in his models, but hypothesized that P2Y6 primarily facilitates microglial engulfment of viable cells—a process dubbed “phagoptosis” (for review, see Brown and Neher, 2012).
In her talk, Huang, a graduate student in Greg Lemke’s lab at Salk, reported how microglia use another set of receptors to keep the brain clear of apoptotic detritus in AD models. Lemke previously discovered that Tryo3, Axl, and Mer, collectively known as the TAM receptors, enable microglial engulfment of sick or dying neurons (see Fourgeaud et al., 2016, for review; Lemke 2017). From the microglial surface, the receptors recognize phosphatidylserine, a membrane phospholipid that flips from the inner to outer membrane when neurons are stressed or apoptotic. Preventing phosphatidylserine exposure on the cell surface protected virally infected neurons from being culled by microglia (Tufail et al., 2017). Other researchers have found TAM receptors on the surface of myeloid cells surrounding plaques in AD mouse models (Feb 2015 news).
At Keystone, Huang reported that in 9.5-month-old APP/PS1 mice, microglia surrounding plaques expressed gobs of Axl and Mer. While these cells always make some Mer, only activated microglia turn on Axl. Huang said that microglia mingling with plaques had 30-fold higher levels of Axl than those farther away, in keeping with the 25-fold upregulation of Axl previously reported in disease-associated microglia (DAM). Huang showed additional data implying, essentially, that microglia expressing Axl/Mer, their ligands, phosphatyldserine, and Aβ all mingle together around dystrophic neurites.
Huang believes that microglial TAM receptor expression is an attempt to limit AD pathogenesis. In agreement with this idea, she found that the survival of APP/PS1 mice plummeted when Axl and Mer were knocked out. Forty percent of the animals died of seizures in their first year of life. Seizures occur in AD. The surviving 1-year-old APP/PS1 mice lacking Axl and Mer had worse memory loss on a contextual fear conditioning test than APP/PS1 controls. Huang also observed a build-up of cleaved caspase-3 puncta in the hippocampus, indicative of uncleared apoptotic cells.
Brown noted that in addition to marking apoptotic cells, phosphatidylserine also decorates the cell surface in response to increases in neuronal activity. He wondered whether a failure to clear those hyperactive neurons in Axl/Mer knockout mice could lead to a buildup of seizure-provoking neurons, perhaps explaining why the animals tend to die young from seizures. Huang said the lab is investigating the idea, as well as the role of TAM receptors in clearance of Aβ plaques.
Beth Stevens of Children’s Hospital in Boston discussed another form of microglial regulation. Stevens previously reported that microglial pruning of synapses, a normal developmental process, switches into overdrive in AD models (Apr 2016 news; Shi et al., 2017). Others have spotted overzealous pruning in tauopathy mice (Jul 2018 news). Essentially, microglia target synapses adorned with complement proteins.
At Keystone, Stevens asked what triggers synapses to start dressing up in complement in the first place. She turned to models of Huntington’s disease to address this question, because neurons in the cortico-striatal circuit are particularly vulnerable to the disease. Using two mouse models—BACHD and Hdh-Q175—postdoc Daniel Wilton found a disconnection in the pre- and postsynaptic compartments in this circuit starting at 3 months of age. Complement proteins crowded the vulnerable circuits, and microglia in the region were loaded with neuronal synapses. Treating mice with an antibody that blocks C1q not only spared synapses but, according to preliminary data, it also restored functional synaptic firing in the cortico-striatal circuit. Some of this work was previously presented at the 2015 Society for Neuroscience annual meeting (Nov 2015 conference news).
Would saving these synapses help? Indeed, Stevens reported that in preliminary experiments done in collaboration with William Yang at the University of California, Los Angeles, 12-month-old C3-deficient BACHD mice managed to balance on a spinning rod better than their C3-expressing counterparts, though they were not quite as agile as wild-type. Similar to BACHD mice treated with a C1q antibody, C3 deficiency spared synapses in the cortico-striatal circuit.
What about the human disease? Using postmortem brain tissue from people at different stages, Stevens identified progressive synapse loss, as well as an uptick in complement proteins, in the striatum. Notably, this synapse loss was apparent even in people who had died in the early stages of HD. Stevens also reported preliminary CSF findings from two HD patient cohorts—HD Clarity and Enroll HD—in which some complement proteins, C3 and iC3b in particular, rose in CSF but not plasma during presymptomatic stages. That said, CSF complement levels overlapped significantly between the different disease stage groups.
Stevens stopped short of addressing exactly what causes the vulnerable neurons in the cortico-striatal circuit to wave the complement flag, but speculated that aberrant neuronal activity and phosphatidylserine could be involved.
In neurodegenerative disease, malfunctions in neuronal circuitry could be an early trigger of damaging microglial responses, Stevens said. In his talk, Paul Worley, Johns Hopkins University in Baltimore, described how the immediate early genes Arc and Neuronal Pentraxin 2 (NPTX2) might facilitate those initial changes to neuronal circuitry, at least in Alzheimer’s disease. Worley reviewed his body of published work on the two proteins, which are important in memory consolidation and maintaining the brain’s circuitry. In a nutshell, Arc is rapidly translated in dendrites, and serves to weaken inactive synapses, while NPTX2 quells hyperexcitability in pyramidal neurons by strengthening their connections with inhibitory interneurons (Xiao et al., 2017).
Worley proposed that NPTX2 could be required to protect neurons from hyperactivity caused by Aβ or other insults. Interestingly, Worley and colleagues reported that CSF levels of NPTX2 drop as AD worsens, and correlate with loss of functional connectivity, suggesting this synaptic regulator could make a useful biomarker for AD progression (see Soldan et al., 2019). At the Alzheimer’s Association International Conference, which just concluded in Los Angeles on July 18, several proteomic and fluid-based studies noted a decline of NPTX2 in the run-up to AD dementia, as well (Aug 2019 coference news).
Stevens noted that the early circuitry imbalances described by Worley are likely what instigates microglia hunger pangs early in neurodegenerative disease.—Jessica Shugart
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