Fig. 2: A mitochondrial electron surplus induces sleep.
a, Proton-pumping complexes I, III and IV convert the energy of electron transfers from NADH to O2—through intermediates CoQ and cytochrome c (cyt c)—into a proton electrochemical gradient, ∆p, across the IMM. Ucp4 discharges, whereas illumination (hν) of mito-dR charges, the IMM. The return of extruded protons to the matrix spins the blades of the ATP synthase and produces ATP, which leaves the matrix via sesB in exchange for cytoplasmic ADP. Neuronal ATP consumption is activity-dependent, in part because the plasma membrane Na+–K+ ATPase must restore ion gradients dissipated by action and excitatory synaptic currents. An oversupply (relative to ATP demand) of electrons to CoQ increases the risk of single-electron reductions of O2 to O2− at complexes I and III. AOX mitigates this risk. b,c, Summed-intensity projections of dFBN dendrites expressing iATPSnFR plus RFP (b) or ATeam (c), in rested and sleep-deprived (SD) flies. Emission ratios are intensity-coded according to the keys below and increase after sleep deprivation (P < 0.0001 (b) and P = 0.0003 (c); two-sided Mann–Whitney test). d, Arousing heat elevates ATP in dFBNs expressing iATPSnFR plus tdTomato. Mean fluorescence was quantified in 20-s windows immediately before and after stimulation (P = 0.0152, two-sided paired t-test) and is plotted as a change in fluorescence intensity ratio (∆R/R) with co-expressed tdTomato relative to pre-stimulation baseline. e, Optogenetic stimulation dissipates ATP in dFBNs expressing iATPSnFR and CsChrimson, but not in dFBNs lacking CsChrimson (P = 0.0076, two-sided t-test). ∆F/F is the change in fluorescence intensity relative to pre-stimulation baseline. f, Sleep in flies expressing R23E10 ∩ VGlut-GAL4-driven Ucp4A or Ucp4C and parental controls (P ≤ 0.0381, Holm–Šídák test after analysis of variance (ANOVA)). g,h, Sleep during the first 60 min after illumination (g; P ≤ 0.0432, Dunn’s test after Kruskal–Wallis ANOVA) and cumulative sleep percentages in flies expressing R23E10 ∩ VGlut-GAL4-driven mito-dR, with or without retinal, and parental controls (h; ∆p photogeneration effect: P < 0.0001, time × ∆p photogeneration interaction: P < 0.0001, mixed-effects model). Asterisks indicate significant differences (P < 0.05) from both parental controls or in planned pairwise comparisons. Data are means ± s.e.m.; n, number of dendritic regions (b,c) or flies (d–h). Scale bars, 5 μm (b,c). For statistical details see Supplementary Table 1.
