MB304B>UAS-mCD8::GFP; aGFP (rabbit), NC82 (aBrp, mouse), GAR AF 488, GAM AF 555; Leica SP8, 20x immersion oil

Janelia Fly Light Split-GAL4 reference

3rd instar larvae were tested for their avoidance behavior on 1.5% Agarose/1.5% Agarose+1.5M Salt-Plate. MB Kenyon cells were optogenetically activated via Channelrhodopsin2-XXL during the experiment. TH-Gal4 positive dopaminergic cells were permanently ablated.

MB299B>UAS-mCD8::GFP; aGFP (rabbit), NC82 (aBrp, mouse), GAR AF 488, GAM AF 555; Leica SP8, 20x immersion oil

Janelia Fly Light Split-GAL4 reference

MB304B>UAS-mCD8::GFP; aGFP (rabbit), NC82 (aBrp, mouse), GAR AF 488, GAM AF 555; Leica SP8, 20x immersion oil

MB299B>UAS-mCD8::GFP; aGFP (rabbit), NC82 (aBrp, mouse), GAR AF 488, GAM AF 555; Leica SP8, 20x immersion oil

TH-D’-Gal4>UAS-mCD8::GFP; aGFP (rabbit), NC82 (aBrp, mouse), GAR AF 488, GAM AF 555; Leica SP8, 20x immersion oil

NP1568-Gal4>UAS-mCD8::GFP (adult brains); aGFP (rabbit), NC82 (aBrp, mouse), GAR AF 488, GAM AF 555; Leica SP8, 20x immersion oil

NP1568-Gal4/CyO>UAS-mCD8::GFP; aGFP (rabbit), NC82 (aBrp, mouse), GAR AF 488, GAM AF 555; Leica SP8, 20x immersion oil

NP0047-Gal4/CyO>UAS-mCD8::GFP; aGFP (rabbit), NC82 (aBrp, mouse), GAR AF 488, GAM AF 555; Leica SP8, 20x immersion oil

TH-D’-Gal4>UAS>>mCD8::GFP,UAS>>mCD8::GFP; aGFP (rabbit), NC82 (aBrp, mouse), GAR AF 488, GAM AF 555; Leica SP8, 20x immersion oil

TH-D1-Gal4>UAS>>mCD8::GFP,UAS>>mCD8::GFP; aGFP (rabbit), NC82 (aBrp, mouse), GAR AF 488, GAM AF 555; Leica SP8, 20x immersion oil

The mushroom bodies (MBs) are known to be the most prominent olfactory memory center in Drosophila. The MB Kenyon cells receive olfactory (via projection neurons, PNs) and gustatory (via dopaminergic neurons, DANs) input. The coincident detection of these stimuli elicits then the formation of associative memory. However, little is known whether and how this high order brain center affects naïve behaviors such as olfaction and gustation. We use fruit fly larvae to test the innate salt avoidance in a simple choice assay. Drosophila larvae are tested for 3 minutes for their preference for 1.5M NaCl. We show that salt avoidance is enhanced upon optogenetic activation of the Kenyon cells. This effect was also observed after ablation of pPAM DANs with simultaneous optogenetic activation of the KCs. We were able to phenocopy this effect by optogenetic activation of TH-Gal4 positive dopaminergic neurons, however not after artificial activation of single DANs of the DL1 cluster. In contrast, larvae showed no salt avoidance after optogenetic activation of sNPF-Gal4 positive neurons. Neuropeptidergic and dopaminergic signaling are tightly involved in modulation of the MBs. However, it is not clear whether our observations depend on coherent neuronal pathways. Therefore, we use simultaneous manipulations of different components of the MB, dopaminergic and sNPF circuitries to unravel the neural mechanisms of modulation of naïve gustatory behaviors in the Drosophila larvae.

5 days old larvae (3rd instar) were tested for their salt (1.5 M) avoidance behavior. Dopaminergic neurons from the TH-D’ cluster were activated upon ChR2-XXL expression and blue light exposure, respectively.