Hippocampal neurons 4 jigsaw puzzle

Distribution of hippocampal neurons expressing EGFP from the Nr4a1/Nur77 promoter (Tg(Nr4a1-EGFP)GY139Gsat, www.gensat.org) in hippocampal CA1 colabelled for the vesicular glutamate transporter (VGluT1, red, Invitrogen polyclonal, 1:1000) and stained with DAPI (blue) to show cell layers. EGFP expression was amplified with chicken anti-GFP (Abcam, 1:2000). VGluT1 labels excitatory terminals in the dendritic fields of the pyramidal cells in stratum orien and stratum radiatum. Nr4a1-EGFP is heavily expressed by CA1 pyramidal cells and to a lesser extent in CA3 pyramidal cells with sparse cells in the dentate gyrus (see associated images). Section (40 um, sagittal) was mounted with Fluoromount G and imaged with a 32X, 0.4 NA LD Achroplan objective on a Zeiss Axiovert microscope. AF568 was excited at 562/40 nm and emission collected at 624/40 nm with Semrock filters and a single band dichroic (530-585 nm reflected, 601-800 nm transmitted). AF488 was excited at 482/35 nm and emission collected at 536/40 nm with Semrock filters and a single band dichroic (446-499 nm reflected, 513-735 nm transmitted). DAPI was excited at 350/50 nm and emission collected at 460/50 nm with a CLP400 dichroic (Chroma). Single images were captured for each channel using a Zeiss Axiocam MR CCD and pseudo-colored using Axiovision. Linear background subtraction was used on the presented image. See Davis and Puhl (2011), Pubmed ID 21305052, for a detailed staining protocol and images of EGFP expression in the striatum and amygdala in this strain. This image is part of a series characterizing EGFP expression from the Nr4a1 promoter throughout the brain.