Analysis of Glial Distribution in Drosophila Adult Brains

Abstract

Neurons and glia are the two major cell types in the nervous system and work closely with each other to program neuronal interplay. Traditionally, neurons are thought to be the major cells that actively regulate processes like synapse formation, plasticity, and behavioral output. Glia, on the other hand, serve a more supporting role. To date, accumulating evidence has suggested that glia are active participants in virtually every aspect of neuronal function. Despite this, fundamental features of how glia interact with neurons, and their spatial relationships, remain elusive. Here, we describe the glial cell population in Drosophila adult brains. Glial cells extend and tightly associate their processes with major structures such as the mushroom body (MB), ellipsoid body (EB), and antennal lobe (AL) in the brain. Glial cells are distributed in a more concentrated manner in the MB. Furthermore, subsets of glia exhibit distinctive association patterns around different neuronal structures. Whereas processes extended by astrocyte-like glia and ensheathing glia wrap around the MB and infiltrate into the EB and AL, cortex glia stay where cell bodies of neurons are and remain outside of the synaptic regions structured by EB or AL.

Keywords: Drosophila; Glia; Mushroom body.

Fig. 1

Glia extend processes around the MB, EB, and AL. Adult brains carrying Repo>mCD8GFP (green) were co-stained with FasII (magenta, A–D2) to reveal the structure of the MB and EB. E–F2 Adult brains carrying Repo>mCD8GFP (green) were co-stained with nc82 (magenta) to reveal the structure of the AL. Confocal projections of sections that exhibit most of each structure (~10–15 sections) were shown (A–A2, C–C2, E–E2) and corresponding enlarged views were shown below (B–B2, D–D2, F–F2). Note that GFP-positive glial processes were seen around the MB, particularly at the tip and the peduncle (arrows in B2), and the EB (arrow in D2). These processes also infiltrated into the EB (D2) and AL (arrow in F2).

Fig. 2

Analysis of glial cell numbers in the adult brain. Adult brains carrying mb247>mCD8GFP or C105>mCD8GFP (green) were co-stained with Repo (white, A1 and B1) to reveal glial nuclei. C–C2 Adult brains co-stained with Elav (green) and Repo (white, C1). Three different bilaterally symmetrical regions (total of 6) were randomly chosen with sizes of 0.14 mm² (1, 2, 5, and 6) and 0.4 mm² (3 and 4). D–F Statistical analysis of glial cell numbers in the vicinity of the MB and EB. Areas 5, 10, and 15 μm away from the α and β + γ lobes of the MB were drawn appropriately and quantified for glial cell number (D). Rings enclosing the EB of diameter 40 (inner), 60, and 80 μm (outer) were designated and the glial cell numbers counted (E). The sum of MB-associated glia was divided by the number of MB neurons that differentiated into the α, β, and γ lobes to obtain the glia/MB neuron ratio (F). Numbers of glia and neurons were counted in areas 1–6 to obtain the glia/neuron ratio (G). Data are shown as mean ± SEM. Eight to ten brains were dissected and the numbers of glia and neurons were counted.

Fig. 3

Ensheathing glia extend processes around the MB, EB, and AL. Adult brains carrying NP6520>mCD8GFP (green, A–C2) co-stained with FasII (white, A1 and B1) or nc82 (white, C1) to reveal the structure of the MB, EB, and AL. Confocal projections merging sections exhibiting most of each structure (~10–15 sections) were shown. Note that GFP-positive glial processes were seen around the MB (A) and EB (B). These processes also infiltrated into the EB (B) and AL (C).

Fig. 4

Astrocyte-like glia extend processes around the MB, EB, and AL. Adult brains carrying Alrm>mCD8GFP (green, A–C2) were co-stained with FasII (white, A1 and B1) or nc82 (white, C1) to reveal the structure of the MB, EB, and AL. Confocal projections merging sections exhibiting most of each structure (~10–15 sections) were shown. Note that the overall GFP expression for astrocyte-like glia was higher than that for ensheathing glia. GFP-positive glial processes were seen around the MB (A) and EB (B). These processes also infiltrated into the EB (B) and AL (C).

Fig. 5

Cortex glial processes extend around the cell bodies of neurons. Adult brains carrying NP577>mCD8GFP (green, A–C2) were co-stained with FasII (white, A1 and B1) or nc82 (white, C1) to reveal the structure of the MB, EB, and AL. Confocal projections merging sections exhibiting most of each structure (~10–15 sections) were shown. Note that GFP-labeled glial processes were mainly seen near the cell bodies of neurons. No specific processes were detected around the MB and they hardly infiltrated into the EB and AL.