Orco regulates the circadian activity of pheromone-sensitive olfactory receptor neurons in hawkmoths

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The mating behavior of nocturnal Manduca sexta hawkmoths is under strict temporal control. It is orchestrated via circadian- and ultradian oscillations in sex-pheromone stimuli as social zeitgeber. The extremely sensitive pheromone-detecting olfactory receptor neurons (ORNs) that innervate the long trichoid sensilla on the male’s antennae are peripheral circadian clocks. They express the transcriptional-translational feedback loop (TTFL) circadian clockwork, best characterized in Drosophila melanogaster . In hawkmoths, it is still unknown whether or how the ORN’s TTFL clockwork regulates the daily rhythms in pheromone-sensitivity and in temporal resolution of ultradian pheromone pulses as prerequisites to the temporal regulation of hawkmoth mating behavior.

We hypothesize that, rather than the slow TTFL clock, a more rapidly adaptive post-translational feedback loop (PTFL) clockwork, associated with the ORN’s plasma membrane, allows for temporal control of pheromone detection via generation of multiscale endogenous membrane potential oscillations. The potential oscillations of the PTFL clock could rapidly synchronize to oscillations of pheromone stimuli at different timescales, thus enable the prediction of stimulus patterns as a mechanism for active sensing. With in vivo long-term tip recordings of long trichoid sensilla of male hawkmoths, we analyzed the spontaneous spiking activity indicative of the ORNs’ endogenous membrane potential oscillations. Consistent with our hypothesis of a multiscale PTFL clock in hawkmoth ORNs, spontaneous spiking was modulated on ultradian and circadian timescales, with maximum activity at night. When we blocked the evolutionarily conserved olfactory receptor coreceptor (Orco), the circadian modulation was abolished but the ultradian frequency modulation of the spontaneous activity remained. Consistent with PTFL control, Orco was not under the circadian control of the TTFL clock. We could replicate the experimental data in a conductance-based computational model of Orco. In this model, Orco conductivity changed as a function of fluctuating 2 ^nd messenger levels. This study demonstrates that a PTFL is sufficient to impose a circadian pattern on ORN sensitivity.

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Significance statement

It is generally assumed that all circadian rhythms in an organism are driven by a transcriptional-translational feedback loop (TTFL) clock. In this study, we demonstrate with in vivo recordings of hawkmoth pheromone-sensitive olfactory receptor neurons (ORNs) that the olfactory receptor coreceptor (Orco) is the key pacemaker channel for controlling circadian but not ultradian rhythms in spontaneous spiking activity. Since Orco expression is not driven by the TTFL clock, its conductance is rather controlled by a post-translational feedback loop (PTFL) membrane clock. Accordingly, our computational model suggests that periodic changes in the conductivity of an Orco ion channel, which is mediated by cycling levels of cyclic nucleotides, tune ORN sensitivity. This highlights the role of the contribution of posttranslational modifications to the generation of circadian rhythmicity.