Single-cell resolution analysis of the crosstalk between chemogenically activated astrocytes and microglia

Abstract

25Astrocytes are the most common glial cell type in the brain, yet, it is still not clear how their 26activation affects the transcriptome of other brain cells such as microglia and neurons. 27Engineered G protein-coupled receptors called Designer Receptors Exclusively Activated by 28Designer Drugs (DREADDS) make it possible to selectively activate specific cell types, such as 29 neurons and astrocytes. By combining the selective activation of astrocytes with single cell RNA 30 sequencing, we were able to study transcriptional changes that occur in response to the 31 activation of astrocytes at the single cell level. Interestingly, our data shows that long-term 32 activation of astrocytes in healthy mice results in dramatic alteration in the transcriptome of 33 astrocytes and microglia. Genes that were differentially expressed in these Gq-DREADD-34 activated astrocytes were involved in neurogenesis and low density lipoprotein particle biology, 35while those in the microglia were involved in the response to lipoproteins, and the migration and 36 chemotaxis of immune cells. Furthermore, network analysis showed that Gq-DREADD-37 mediated activation in astrocytes resulted in an upregulation of genes involved in the G protein-38 coupled receptor signaling pathway and calcium ion homeostasis. This confirmed the activation 39 of astrocytes through the expressed DREADDS. Our findings show the importance of 40considering the transcriptomic alteration in microglia and neurons after the activation of 41 astrocytes in in vivo models. Therefore, our data will serve as a resource for the broader 42 neuroscience community.

A perspective on astrocyte regulation of neural circuit function and animal behavior

“…Clearly, there is an urgent need for new and improved actuators that allow precise control over astrocytes’ spatial and temporal dynamics and specific signaling pathways. Thorough characterization, including transcriptomic and proteomic profiling, will be necessary to develop a mechanistic understanding of precisely how their activation influences astrocytes, neural circuits, and animal behavior, either directly or indirectly ( Figueiredo et al, 2014 ; Octeau et al, 2019 ; Philtjens et al, 2021 ; Yu et al, 2020 ). Such experiments should also consider how the observed effects depend on stimulation parameters and the approach itself (e.g., light-induced heating-mediated effects; Courtney et al, 2021 ; Owen et al, 2019 ; Schmidt & Oheim, 2020 ).…”

Impairment of hippocampal astrocyte-mediated striatal dopamine release and locomotion in Alzheimer’s disease

“…Chronic DREADD activation of astrocytes has been shown to induce transcriptomic changes in adjacent microglia, notably with the upregulation of a set of Alzheimer’s disease-associated genes that are associated to a microglial phenotype which is protective against AD pathology 31,32 . Our results extend this finding by showing that, functionally, the result of chronic astrocytic DREADD stimulation is associated to an increase in microglial density and especially in a microglial phenotype that more efficiently responds to and limits AD pathology.…”

The DREADDful Hurdles and Opportunities of the Chronic Chemogenetic Toolbox

“…A literature review reveals that only a handful of studies (unbiasedly) unveiled the transcriptomic/proteomic profile upon chemogenetic activation, as summarized in Table 2 . These studies focused on CNO-induced DREADD modulation of neurons [ 24 , 81 , 82 , 83 , 84 , 85 ] or astrocytes [ 86 , 87 , 88 ], and reported corresponding molecular effects on the neurons [ 24 , 81 , 82 , 84 ] or astrocytes [ 86 , 87 , 89 ] themselves, and/or on neighboring endothelial cells [ 85 ] or microglia [ 87 ]. Briefly, studies depicting neuronal alterations exclusively related to DREADD activation, reported an activity-dependent upregulation of several genes in the BDNF-TrkB signaling pathway [ 81 , 82 ] or an upregulation of multiple immediately early genes, JUNB interaction partners and a possible involvement of PKA signaling pathway [ 84 ].…”

“…Briefly, studies depicting neuronal alterations exclusively related to DREADD activation, reported an activity-dependent upregulation of several genes in the BDNF-TrkB signaling pathway [ 81 , 82 ] or an upregulation of multiple immediately early genes, JUNB interaction partners and a possible involvement of PKA signaling pathway [ 84 ]. Similarly, studies focusing on the astrocytic alterations described altered signaling pathways associated with neuroinflammatory responses [ 89 ], GPCR signaling [ 87 ], and calcium ion homeostasis [ 87 ] or biological functions related to immune responses, regulation of transcription, and translation and cell proliferation/growth [ 89 ]. Moreover, astrocytic activation also led to an upregulation of Thbs1 , which is involved in synapse formation and function [ 86 , 89 ].…”

“…Umpierre et al [ 97 ] elegantly described altered calcium signaling in microglia upon chemogenetic modulation of neuronal activity. Moreover, Philtjens et al [ 87 ] used DREADDs in astrocytes and did not only observe changes in the astrocytes, but also in neighboring microglia. Unfortunately, they could not report on the effect on nearby neurons, as glial cells were enriched and neurons depleted in their dissociation protocol.…”