Threshold response properties of larval local interneurons. (a) LN2 cells in the antennal lobe stained to reveal G-CaMP (left; anti-GFP antibody, green) and gamma-aminobutyric acid (middle; anti-GABA, magenta). Arrows in the merged image (right) indicate GABA-positive LN2 neurons. (b) Imaging LN activation at the terminal of the Or42a neuron, as marked by an Or42a-nsyb:tdTomato reporter. The leftmost panel is a merged image of intrinsic fluorescence of G-CaMP (green) and nsyb::tdTomato (magenta). The boundary of the antennal lobe is marked with a white dashed line and the Or42a glomerulus with a black dashed line. The other three panels show antennal lobe calcium responses to paraffin oil (solvent) and three odorants (10-2 dilution) taken 400 ms after stimulus onset, represented as ΔF/F (%) (scale at right). (c) Top panel: schematic for measuring functional activation of LN2 neurons in the antennal lobe (blue boxes) of wild-type, Or42a-, Or42b-, and Or42a+Or42b-functional larvae. Bottom panel: responses of LN2 neurons in larvae of indicated genotype to eight odorants (10-2 dilution except as indicated) and paraffin oil (solvent) represented as false color-coded time traces (%ΔF/F; scale at bottom right). Traces from n = 6–9 animals per stimulus are stacked. (d) Responses in LNs in larvae of indicated genotype to an ethyl butyrate concentration series and paraffin oil (solvent) represented as ΔF/F (%) (scale at right). Traces from n = 6–8 animals per genotype and stimulus are stacked.