Periodic Reporting for period 4 - MICROGLIA-CIRCUIT (Microglia action towards neuronal circuit formation and function in health and disease)
Reporting period: 2021-11-01 to 2022-10-31
The brain is assembled from thousands of cell types, which are functionally organized in neuronal circuits to collect, encode and process environmental information. Classical studies have provided important insights into the necessity of correct neuron wiring as otherwise the signal processing is impaired resulting in behavioral dysfunction. Microglia traditionally classified as immune-responsive cells are embedded within the neuronal network and have been shown to be involved in cell removal and synapse refinement. How microglia interact with other cell types and sense changes during neuronal circuit development, maintenance and degeneration is still an open question. The goal of this project is to provide an answer. We will elucidate the key molecular and genetic principles underlying microglial phenotype during neuronal circuit formation, identify the circuit components with which microglia are interacting with at a spatial-temporal resolution, pin-point how neuronal activity alters microglial function, and finally differentiate human induced pluripotent stem cells (hiPSCs) into microglia to develop a strategy to address microglia-associated disease genes in neurodevelopmental diseases.
The knowledge is fundamental for the society because microglia are frequently found in a phagocytic-associated state in post-mortem human brain of neurodevelopmental and neurodegenerative diseases. Also, an initial sign of the disease onset is the activation of the immune system that precedes synapse impairment and neuronal cell death making microglia a prime candidate. Furthermore, genome-wide association studies have identified gene mutation that are linked to microglia. Thus, it is critical to obtain insights into microglia-neuron interaction.
The overall objectives cover to identify the different phenotypic states of microglia on the morphological and functional level. Then, we are interfering with input signals to the brain and identify the functional consequences on the microglia-to-neuronal communication. Finally, we will translate our observations to the human microglia-neuron model system.
In this project, we found that microglia enable reinstatement of juvenile-like neuronal plasticity in the adult brain by removing the perineuronal nets (PNN) after repeated anesthetic ketamine exposure. Strikingly, we recapitulated the ketamine effect with non-invasive, selective 60-Hz light entrainment. To provide insights into the microglia function, we investigated the highly branched 3D-morphology. We realized that traditionally strategies to analyze morphology are insufficient to distinguish microglia across brain regions, sex, development and disease progression. Thus, we development MorphOMICs, an algorithm that allows a high-throughput, minimally biased, and consistent way to capture context-specific and sex-dependent microglia morphology signatures during different stages of development and disease progression. We applied this algorithm to a self-build library of 41’886 3D-microglial reconstructions and established for seven brain regions a reference atlas that can be expanded unlimitedly map how microglia morphologically adapt in a condition-of-interest. This provides a first critical step to establish a morphology-to-function relationship.
Furthermore, we build a foundation for AAV-mediated retinal microglial targeting in vivo and provide novel insights into environment-dependent microglial transduction, which has not been described previously and will be highly relevant for future studies.
Finally, we identified that hiPSC-derived retinal organoid differentiation also results in the occurrence of microglia, however, they preferentially enrich in mesenchymal-rich regions and rarely infiltrate the neuronal tissue suggesting that certain signaling molecules are involved.
In conclusion, our results indicate a surprising complex role of microglia in the neuronal network that influence not only the neuronal network during development and degeneration but also in adulthood.
The knowledge is fundamental for the society because microglia are frequently found in a phagocytic-associated state in post-mortem human brain of neurodevelopmental and neurodegenerative diseases. Also, an initial sign of the disease onset is the activation of the immune system that precedes synapse impairment and neuronal cell death making microglia a prime candidate. Furthermore, genome-wide association studies have identified gene mutation that are linked to microglia. Thus, it is critical to obtain insights into microglia-neuron interaction.
The overall objectives cover to identify the different phenotypic states of microglia on the morphological and functional level. Then, we are interfering with input signals to the brain and identify the functional consequences on the microglia-to-neuronal communication. Finally, we will translate our observations to the human microglia-neuron model system.
In this project, we found that microglia enable reinstatement of juvenile-like neuronal plasticity in the adult brain by removing the perineuronal nets (PNN) after repeated anesthetic ketamine exposure. Strikingly, we recapitulated the ketamine effect with non-invasive, selective 60-Hz light entrainment. To provide insights into the microglia function, we investigated the highly branched 3D-morphology. We realized that traditionally strategies to analyze morphology are insufficient to distinguish microglia across brain regions, sex, development and disease progression. Thus, we development MorphOMICs, an algorithm that allows a high-throughput, minimally biased, and consistent way to capture context-specific and sex-dependent microglia morphology signatures during different stages of development and disease progression. We applied this algorithm to a self-build library of 41’886 3D-microglial reconstructions and established for seven brain regions a reference atlas that can be expanded unlimitedly map how microglia morphologically adapt in a condition-of-interest. This provides a first critical step to establish a morphology-to-function relationship.
Furthermore, we build a foundation for AAV-mediated retinal microglial targeting in vivo and provide novel insights into environment-dependent microglial transduction, which has not been described previously and will be highly relevant for future studies.
Finally, we identified that hiPSC-derived retinal organoid differentiation also results in the occurrence of microglia, however, they preferentially enrich in mesenchymal-rich regions and rarely infiltrate the neuronal tissue suggesting that certain signaling molecules are involved.
In conclusion, our results indicate a surprising complex role of microglia in the neuronal network that influence not only the neuronal network during development and degeneration but also in adulthood.
We established a new methodology called MorphOMICs, an algorithm that allows a high-throughput, minimally biased, and consistent way to capture context-specific and sex-dependent microglia morphology signatures during different stages of development and disease progression. This technology combines biology, applied mathematics (topology), statistics, and machine learning strategies. We then applied this algorithm to our dataset of 41,886 3D-traced microglial cells at different phenotypic states across development, adulthood, and degenerative disease across 7 brain regions (olfactory bulb, frontal cortex, substantia nigra, dentate gyrus of the hippocampus, cerebellum, cochlear nucleus, primary somatosensory cortex). This study has been published in Nature neuroscience in 2022 with the following press release. The code for performing the analysis is shared on github. We uploaded the microglial reconstructions to the NeuroMorpho.org database, a centrally curated inventory of digitally reconstructed neurons and glia. This work was already recently commented.
We found that ketamine anaesthesia in mice stimulates microglia to remove structures involved in juvenile synaptic plasticity. This study has been published in Cell reports in 2021 with the following press release. The paper raised interest in various communities and received special mentioning at the Alzforum, Lucid, and Faculty Opinion. We also wrote a STAR Protocols to describe the reported method in more detail.
We adapted a virus capsid to increase the transduction efficiency of microglia in the retina. This paper has been published in Molecular Therapy – Methods and Clinical Development in 2021 and was a follow up of an extensive review about strategies to transduce microglia with viral vectors published in Neuroscience Letters.
Finally, we show that during retinal organoid differentiation microglia occur intrinsically during the differentiation process, however they preferentially enrich in the mesenchymal niche and rarely infiltrate the neuronal environment. This paper has been published in iScience in 2022 with a follow-up STAR Protocols that describes how to functional characterize microglia in culture.
We found that ketamine anaesthesia in mice stimulates microglia to remove structures involved in juvenile synaptic plasticity. This study has been published in Cell reports in 2021 with the following press release. The paper raised interest in various communities and received special mentioning at the Alzforum, Lucid, and Faculty Opinion. We also wrote a STAR Protocols to describe the reported method in more detail.
We adapted a virus capsid to increase the transduction efficiency of microglia in the retina. This paper has been published in Molecular Therapy – Methods and Clinical Development in 2021 and was a follow up of an extensive review about strategies to transduce microglia with viral vectors published in Neuroscience Letters.
Finally, we show that during retinal organoid differentiation microglia occur intrinsically during the differentiation process, however they preferentially enrich in the mesenchymal niche and rarely infiltrate the neuronal environment. This paper has been published in iScience in 2022 with a follow-up STAR Protocols that describes how to functional characterize microglia in culture.
We are exploring the option for a start-up that takes advantage of the 60-Hz light entrainment and establish it as a medical therapeutic intervention.