This dataset presents a ZT14 (two hours after lights OFF) three-dimensional Serial Block-Face Scanning Electron Microscopy (SBEM) volume of the dorsal terminals of small Lateral ventral Neurons (s-LNvs) in the adult Drosophila melanogaster brain. These neurons, which express the neuropeptide Pigment-Dispersing Factor (PDF), serve as key circadian pacemakers that synchronize the brain’s clock network.
SBEM was performed using a Zeiss GeminiSEM 300 equipped with a Gatan 3View system and a focal nitrogen gas injection setup to minimize charging and enhance image quality under high vacuum conditions. Samples were prepared from adult female Drosophila melanogaster brains expressing mito::APEX2 under the Pdf-GAL4 driver, labeling mitochondria within PDF-expressing neurons. Fixation was carried out with 2.5% glutaraldehyde and 2% paraformaldehyde in 0.15 M cacodylate buffer, followed by a DAB reaction, osmium tetroxide, potassium ferrocyanide, uranyl acetate, and lead aspartate staining. The samples were embedded in Durcupan resin at 60 °C for 72 hours. SBEM datasets were collected at an accelerating voltage of 2.5 kV, with a z-step size of 50 nm and 5 nm pixel resolution in x and y. Regions of interest were identified and registered in advance using Micro-Computed Tomography (Micro-CT) and confocal fluorescence imaging for correlative light and electron microscopy (CLEM).
