Extended Data Fig. 5: Thermosensory Lateral Horn Output Neuron II (TLHON-II) is the primary output of TPN-V and is critical for heat avoidance behavior in D. melanogaster.
From: Evolution of temperature preference in flies of the genus Drosophila
(a) Connectomic diagram illustrating the pathways that relay temperature information from the hot thermosensory receptor neurons (TRNs) to the Thermosensory Lateral Horn Output Neuron II (TLHON-II). TPN-V is the major output of the aristal hot TRNs (comprising 52.67% of hot TRN output synapses to TPNs) and the major direct path of connectivity from hot TRNs to Thermosensory Lateral Horn Output Neuron II (TLHON-II), which is in turn the major target of TPN-V (black arrows represent alternative direct pathways of connectivity between hot TRNs and TLHON-II, Ns=number of synapses). (b) D. melanogaster EM reconstruction of TPN-V (red) and TLHON-II (light blue). (c) TPN-V::TLHON-I synapses (yellow dots) and TPN-V::TLHON-II synapses within the LH (blue dots) tile the TPN-V axonal arbor within the LH (synapse location was reconstructed from EM data, see methods for details). (d) A split-Gal4 driver exclusively expressed in TLHON-II (maximum projections of 2-photon z-stacks of the brain and VNC of TLHON-II-Gal4>GFP animal). Scale bars = 25 μm. (e) Genetic silencing using TLHON-II:Gal4>Kir2.1 impacts heat avoidance (*p < 0.05, 2-way ANOVA, p = [0.748, 1.95e-3, 4.15e-5, 4.91e-7] for test temperatures 25, 30, 35, and 40, respectively) (f) Optogenetic activation via TLHON-II:Gal4>CsChrimson is sufficient to induce avoidance of red light (*p < 0.05 indicates significant difference from zero in two-way one sample t-test, p = [0.440, 0.983, 2.54e-5] for Gal4/+, UAS/+, and Gal4>UAS, respectively). (in e-f red lines=mean, grey boxes=one standard deviation, empty boxes=95% confidence interval for the mean, circles=groups of 15 and 25 flies each for e and f, respectively; N = 8 and 9 groups/condition for e and f, respectively). Abbreviations: Lateral Horn, LH; Calyx, CA; Posterior Antennal Lobe, PAL; Mushroom Body, MB; Superior Medial Protocerebrum, SMP.
