An Integrated Micro- and Macroarchitectural Analysis of the Drosophila Brain by Computer-Assisted Serial Section Electron Microscopy
(A, B) Montage of electron micrographs of representative brain section (A; co, cortex; ne, neuronal cell body; np, neuropile). Note that this and all other electron microscopic images of the figures presented in this article are a composite of multiple tiled digital photographs stitched together with the TrakEM2 software, as explained in Materials and Methods. Occasional slight discontinuities in image brightness (as for example at lower right corner of panel A) correspond to borders of tiles. Bundles of long axons can be recognized and identified with specific, lineage-related fascicles known from confocal microscopy. Upper boxed area, shown at higher magnification in panel B, contains branch of antenno-cerebral tract (APT; blue) and adjacent axon tract formed by one of the DPM lineages (PAT). Note glial processes (gl) accompanying axon fascicles. Green box in (A) demarcates size and position of a microvolume that was cropped out of stack for further analysis (see panels F–H). Dotted purple line demarcates calyx (CX) compartment. (C) Schematic drawing showing L1 brain hemisphere with circuit consisting of antennal lobe-calyx-spur/lobes of mushroom body. Fascicles forming this circuit (e.g., APT, mushroom body) can be identified in TEM stack and help to interpret short segments of neurites contained within microvolumes (e.g., microvolume from within the calyx). (D, E) Confocal cross-sections of first instar larval brain hemisphere (D) and adult brain hemisphere (E) labeled with anti-Brp to visualize neuropile. Set of lineage-related axon fascicles that can be followed from first instar to adult are rendered in different colors. These fascicles (among them the APT shown in panels A, B) form a system of invariant landmarks that can be recognized in confocal stacks and TEM stacks. Abbreviations: APT, antenno-protocerebral tract; DPC, dorso-posterior commissure; loSM, longitudinal superior-medial fascicle; MEF, medial equatorial fascicle; obP, oblique posterior fascicle; SLP, superior lateral protocerebrum; VLP, ventro-lateral protocerebrum. (F–H) 3D views of selected objects within the calyx microvolume. Microvolume contained 42 large diameter (>0.2 µm) neurite segments. Of these, 14 were randomly picked and are shown individually in panels F1–F14 (volume in side view; y and z indicate direction of axes of volume). (G, H) Top view of large diameter neurite segments combined (G: all 42 segments; H: 14 segments shown individually in panel F). Red dots indicate position of presynaptic sites. Scale bars: 4 µm (A); 0.5 µm (B); 10 µm (C, D); 1 µm (F, G, H).