In situ and transcriptomic identification of microglia in synapse-rich regions of the developing zebrafish brain

Silva, Nicholas J.; Dorman, Leah C.; Vainchtein, Ilia D.; Horneck, Nadine C.; Molofsky, Anna V.

doi:10.1038/s41467-021-26206-x

Cited by 44 publications

(54 citation statements)

References 74 publications

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“…markers (Lange et al, 2020). Lastly, we identified OT24 as microglial due to expression of mpeg1.1, slc7a7, and ctsba (Figure 3C), the latter a lysosomal gene found to be enriched in tectal populations of microglia (Silva et al, 2021; see Supplementary Table 5 for all glial markers).…”

Section: The Larval Optic Tectum Contains Mature and Developing Neuro...mentioning

confidence: 88%

Single-Cell RNA Sequencing Characterizes the Molecular Heterogeneity of the Larval Zebrafish Optic Tectum

Martin

Babbitt

Pickens

et al. 2022

Front. Mol. Neurosci.

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The optic tectum (OT) is a multilaminated midbrain structure that acts as the primary retinorecipient in the zebrafish brain. Homologous to the mammalian superior colliculus, the OT is responsible for the reception and integration of stimuli, followed by elicitation of salient behavioral responses. While the OT has been the focus of functional experiments for decades, less is known concerning specific cell types, microcircuitry, and their individual functions within the OT. Recent efforts have contributed substantially to the knowledge of tectal cell types; however, a comprehensive cell catalog is incomplete. Here we contribute to this growing effort by applying single-cell RNA Sequencing (scRNA-seq) to characterize the transcriptomic profiles of tectal cells labeled by the transgenic enhancer trap line y304Et(cfos:Gal4;UAS:Kaede). We sequenced 13,320 cells, a 4X cellular coverage, and identified 25 putative OT cell populations. Within those cells, we identified several mature and developing neuronal populations, as well as non-neuronal cell types including oligodendrocytes and microglia. Although most mature neurons demonstrate GABAergic activity, several glutamatergic populations are present, as well as one glycinergic population. We also conducted Gene Ontology analysis to identify enriched biological processes, and computed RNA velocity to infer current and future transcriptional cell states. Finally, we conducted in situ hybridization to validate our bioinformatic analyses and spatially map select clusters. In conclusion, the larval zebrafish OT is a complex structure containing at least 25 transcriptionally distinct cell populations. To our knowledge, this is the first time scRNA-seq has been applied to explore the OT alone and in depth.

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Section: The Larval Optic Tectum Contains Mature and Developing Neuro...mentioning

confidence: 88%

Single-Cell RNA Sequencing Characterizes the Molecular Heterogeneity of the Larval Zebrafish Optic Tectum

Martin

Babbitt

Pickens

et al. 2022

Front. Mol. Neurosci.

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“…In addition to oligodendrocytes, we identified several populations enriched for radial glial markers, notably clusters OT2, OT6, and OT10 (Figure 3C). Lastly, we identified OT24 as microglial, due to expression of mpeg1.1, slc7a7, and ctsba (Figure 3C), the latter a lysosomal gene found to be enriched in tectal populations of microglia 39 (See Table S5 for all glial markers). Thus, at 7dpf the larval optic tectum contains at least three non-neuronal cell types and 22 neuronal populations.…”

Section: The Larval Ot Contains Both Mature and Developing Neurons As Well As Several Glial Populationsmentioning

confidence: 99%

“…Two appear to represent oligodendrocytes, which have been previously identified in the deeper portions of the tectal PVL 50 . A small population of microglia with an amoeboid morphology has been recently described to localize near neurogenic regions and express the myeloid reporter gene mpeg1.1, as well as lysosomal genes ctsla and ctsba 39 . We found a single small population to express mpeg1.1, and ctsba; as well as slc7a7, which functions in microglia development 51 .…”

Section: Non-neuronal Cells Of the Optic Tectummentioning

confidence: 99%

Single-Cell RNA Sequencing Characterizes the Molecular Heterogeneity of the Larval Zebrafish Optic Tectum

Martin

Babbitt

Pickens

et al. 2021

Preprint

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SUMMARYThe optic tectum (OT) is a multilaminated midbrain structure that acts as the primary retinorecipient in the zebrafish brain. Homologous to the mammalian superior colliculus, the OT is responsible for the reception and integration of stimuli, followed by elicitation of salient behavioral responses. While the OT has been the focus of functional experiments for decades, less is known concerning specific cell types, microcircuitry, and their individual functions within the OT. Recent efforts have contributed substantially to the knowledge of tectal cell types; however, a comprehensive cell catalog is incomplete. Here we contribute to this growing effort by applying single-cell RNA-sequencing (scRNA-seq) to characterize the transcriptomic profiles of tectal cells labeled by the transgenic enhancer trap line y304Et(cfos:Gal4;UAS:Kaede). We sequenced 13,320 cells, a 4X cellular coverage, and identified 25 putative OT cell populations. Within those cells, we identified several mature and developing neuronal populations, as well as non-neuronal cell types including oligodendrocytes, microglia, and radial glia. Although most mature neurons demonstrate GABAergic activity, several glutamatergic populations are present, as well as one glycinergic population. We also conducted Gene Ontology analysis to identify enriched biological processes, and computed RNA velocity to infer current and future transcriptional cell states. Finally, we conducted in situ hybridization to validate our bioinformatic analyses and spatially map select clusters. In conclusion, the larval zebrafish OT is a complex structure containing at least 25 transcriptionally distinct cell populations. To our knowledge, this is the first time scRNA-seq has been applied to explore the OT alone and in depth.

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“…Finally, microglia, the resident innate immune cell of the CNS, have several roles in CNS development, including phagocytosing cell debris as well as pruning developing synapses and myelin (Hughes and Appel, 2020;Mazaheri et al, 2014;Silva et al, 2021;Stevens et al, 2007;Villani et al, 2019). To our knowledge, our study is the first examination of the role of inflammation and complement signaling in PNS development beyond the NCC stage (Carmona-Fontaine et al, 2011).…”

Section: Discussionmentioning

confidence: 91%

Cd59 and inflammation orchestrate Schwann cell development

Wiltbank

Steisnon

Criswell

et al. 2022

Preprint

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Efficient neurotransmission is essential for organism survival and is enhanced by myelination. However, the genes that regulate myelin and myelinating glial cell development have not been fully characterized. Data from our lab and others demonstrates that cd59, which encodes for a small GPI-anchored glycoprotein, is highly expressed in developing zebrafish, rodent, and human oligodendrocytes (OLs) and Schwann cells (SCs), and that patients with CD59 dysfunction develop neurological dysfunction during early childhood. Yet, the function of CD59 in the developing nervous system is currently undefined. In this study, we demonstrate that cd59 is expressed in a subset of developing SCs. Using cd59 mutant zebrafish, we show that developing SCs proliferate excessively, which leads to reduced myelin volume, altered myelin ultrastructure, and perturbed node of Ranvier assembly. Finally, we demonstrate that complement activity is elevated in cd59 mutants and that inhibiting inflammation restores SC proliferation, myelin volume, and nodes of Ranvier to wildtype levels. Together, this work identifies Cd59 and developmental inflammation as key players in myelinating glial cell development, highlighting the collaboration between glia and the innate immune system to ensure normal neural development.

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In situ and transcriptomic identification of microglia in synapse-rich regions of the developing zebrafish brain

Cited by 44 publications

References 74 publications

Single-Cell RNA Sequencing Characterizes the Molecular Heterogeneity of the Larval Zebrafish Optic Tectum

Single-Cell RNA Sequencing Characterizes the Molecular Heterogeneity of the Larval Zebrafish Optic Tectum

Single-Cell RNA Sequencing Characterizes the Molecular Heterogeneity of the Larval Zebrafish Optic Tectum

Cd59 and inflammation orchestrate Schwann cell development

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