Development of ramified microglia from early macrophages in the zebrafish optic tectum

Abstract

Microglia, the resident macrophage precursors of the brain, are necessary for the maintenance of tissue homeostasis and activated by a wide range of pathological stimuli. They have a key role in immune and inflammatory responses. Early microglia stem from primitive macrophages, however the transition from early motile forms to the ramified mature resident microglia has not been assayed in real time. In order to provide such an assay, we used zebrafish transgenic lines in which fluorescent reporter expression is driven by the promoter of macrophage expressed gene 1 (mpeg1; Ellet et al. [2011]: Blood 117(4): e49-e56,). This enabled the investigation of the development of these cells in live, intact larvae. We show that microglia develop from highly motile amoeboid cells that are engaged in phagocytosis of apoptotic cell bodies into a microglial cell type that rapidly morphs back and forth between amoeboid and ramified morphologies. These morphing microglia eventually settle into a typical mature ramified morphology. Developing microglia frequently come into contact with blood capillaries in the brain, and also frequently contact each other. Up to 10 days postfertilization, microglia were observed to undergo symmetric division. In the adult optic tectum, the microglia are highly branched, resembling mammalian microglia. In addition, the mpeg1 transgene also labeled highly branched cells in the skin overlying the optic tectum from 8-9 days postfertilization, which likely represent Langerhans cells. Thus, the development of zebrafish microglia and their cellular interactions was studied in the intact developing brain in real time and at cellular resolution.

Glial Cell Development and Function in Zebrafish

“…In zebrafish and mammals, mature microglia are present throughout the parenchyma of the brain and spinal cord (Sieger and Peri 2013;Svahn et al 2013). In the healthy CNS, the cell bodies of microglia are largely stationary, but they actively extend and retract processes as they inspect their environment for any sign of perturbation (Nimmerjahn 2005).…”

“…Unlike neurons, OLs, and radial glia/astrocytes, which originate from neuroepithelial progenitors, microglia derive from primitive macrophages that move into the CNS from the periphery and later differentiate as CNS-resident microglia (Herbomel et al 2001;Ginhoux et al 2010;Schulz et al 2012;Sieger and Peri 2013;Svahn et al 2013). The first macrophages in zebrafish arise from anterior mesodermal cells and begin to migrate throughout the embryo by 24 hours postfertilization (hpf ) (Herbomel et al 1999(Herbomel et al , 2001Le Guyader et al 2008).…”

“…Primitive macrophages in the CNS begin to differentiate as microglia as early as 60 hpf (Herbomel et al 2001;Peri and Nusslein-Volhard 2008). Primitive macrophages and microglia are both actively phagocytic, and microglia maintain expression of some early macrophage genes, such as mfap4 and mpeg1 (Zakrzewska et al 2010;van Ham et al 2012van Ham et al , 2014Svahn et al 2013). …”

Caspase-mediated apoptosis induction in zebrafish cerebellar Purkinje neurons

“…5D, see number of microglia in EtOH and 4OHT control specimens). Microglia are involved in developmental apoptosis in the larval zebrafish brain (Cole and Ross, 2001), which still occurs in the optic tectum at 3 dpf, whereas it has ceased in the cerebellum (Peri and Nüsslein-Volhard, 2008;Sieger and Peri, 2013;Svahn et al, 2013). To address the potential involvement of microglia in the clearance of apoptotic debris, we induced the ablation of PCs at 4 dpf in cerebellum (B,D), accompanied by microglial activation based on ramified to ameboid morphology changes (E).…”

Mitophagy and neurodegeneration

“…14 Furthermore, zebrafish neurons possess typical structural features, including the soma, dendrites, and the axon, which can be myelinated or demyelinated. Astrocytes, 27 oligodendrocytes, 28 and microglia 29,30 have all been positively identified in the zebrafish brain. The identification of these cell types is important because their interactions with neurons and their involvement in disease processes is becoming increasingly relevant, and these in vivo interactions are something that cannot be easily replicated in vitro.…”