Spatial-temporal patterns of reactive oxygen species release from microglia and synaptic impairment
Final Report Abstract
Oxidative stress is a known hallmark of Alzheimer’s Disease (AD) but the underlying mechanisms of how reactive oxygen species (ROS) are generated and trigger synaptic dysfunction and neuronal death remain elusive. Microglia are known to contribute to AD pathology. They can be neuroprotective or acquire neurotoxic phenotypes. This research project aimed to identify the effect of microglial production of ROS on synaptic depression following complement receptor stimulation and co-activation with a hypoxic insult. The second objective was to identify the receptor responsible for Aβ induced synaptic impairments on microglia. As a last step, microglia ROS production and synaptic depression was to be investigated in the 5xFAD mouse model of AD. To investigate these research objectives genetically encoded redox indicators (such as the redox sensitive green fluorescent protein roGFP) were used to quantify neuronal redox states in brain tissue. After successfully implementing 2-photon based ratiometric imaging techniques in the lab several stimulation protocols were tried to activate microglia and neuronal redox states and synaptic activities were measured simultaneously. Surprisingly, in these first datasets no stimulation could effectively induce notable redox changes in neurons. As a next step, redox imaging was performed in the 5xFAD mouse model of AD which is known for its massive overproduction of Aβ, plaque depositions, microglia activation and synaptic alterations as well as memory deficits. While oxidative stress and microglia are extensively researched studies on how microglia interact with neuronal oxidative stress have not been conducted due to a lack of imaging tools. Using different roGFP redox sensors we describe imaging strategies to simultaneously image microglia and neuronal redox states in live and paraformaldehyde fixed brain tissue. By employing these tools, we found that microglia closely associate and engulf highly oxidized dystrophic neurites around amyloid plaques in 5xFAD mice. Long-term in vivo 2-photon imaging revealed a highly dynamic spatial-temporal neuron-microglia interaction. We depleted microglia from 5xFAD mice using PLX3397 to study the impact on oxidative stress and neuronal survival. While oxidative stress was surprisingly exacerbated after microglia removal, we found less neuron loss in 5xFAD mice indicating neuroprotective as well as neurotoxic functions of microglia. We found that increased levels of ROS correlate with elevated LAMP1 levels indicating that lysosome accumulation in dystrophic neurites is directly linked to oxidative stress. This data indicates that oxidative stress around amyloid plaques might not be induced by microglia but is of neuronal origin instead. Identifying the individual molecular pathways for the different phenotypes of microglia will be crucial in order to find novel molecular targets to selectively manipulate neurotoxic functions and not the neuroprotective ones.
Publications
- Neuron Activity Dependent Redox Compartmentation Revealed with a Second Generation Red-Shifted Ratiometric Sensor. ACS Chem Neurosci. 2020 Sep2;11(17):2666-2678
Saranya Radhakrishnan, Jacob Norley, Stefan Wendt, Nathan LeRoy,Hana Hall, Stevie Norcross, Sara Doan, Jordan Snaider, Brian A MacVicar,Vikki M Weake, Libai Huang, Mathew Tantama
(See online at https://doi.org/10.1021/acschemneuro.0c00342)
