Spatial orientation based on multiple visual cues in non-migratory monarch butterflies.

Monarch butterflies (Danaus plexippus) are prominent for their annual long-distance migration from North America to their overwintering area in Central Mexico. To find their way on this long journey, they use a sun compass as their main orientation reference but will also adjust their migratory direction with respect to mountain ranges. This indicates that the migratory butterflies also attend to the panorama to guide their travels. Although the compass has been studied in detail in migrating butterflies, little is known about the orientation abilities of non-migrating butterflies. Here, we investigated whether non-migrating butterflies - which stay in a more restricted area to feed and breed - also use a similar compass system to guide their flights. Performing behavioral experiments on tethered flying butterflies in an indoor LED flight simulator, we found that the monarchs fly along straight tracks with respect to a simulated sun. When a panoramic skyline was presented as the only orientation cue, the butterflies maintained their flight direction only during short sequences, suggesting that they potentially use it for flight stabilization. We further found that when we presented the two cues together, the butterflies incorporate both cues in their compass. Taken together, we show here that non-migrating monarch butterflies can combine multiple visual cues for robust orientation, an ability that may also aid them during their migration.

The roles of vision and antennal mechanoreception in hawkmoth flight control.

Flying animals need continual sensory feedback about their body position and orientation for flight control. The visual system provides essential but slow feedback. In contrast, mechanosensory channels can provide feedback at much shorter timescales. How the contributions from these two senses are integrated remains an open question in most insect groups. In Diptera, fast mechanosensory feedback is provided by organs called halteres and is crucial for the control of rapid flight manoeuvres, while vision controls manoeuvres in lower temporal frequency bands. Here, we have investigated the visual-mechanosensory integration in the hawkmoth Macroglossum stellatarum. They represent a large group of insects that use Johnston's organs in their antennae to provide mechanosensory feedback on perturbations in body position. Our experiments show that antennal mechanosensory feedback specifically mediates fast flight manoeuvres, but not slow ones. Moreover, we did not observe compensatory interactions between antennal and visual feedback.

Comparative system identification of flower tracking performance in three hawkmoth species reveals adaptations for dim light vision.

Flight control in insects is heavily dependent on vision. Thus, in dim light, the decreased reliability of visual signal detection also prompts consequences for insect flight. We have an emerging understanding of the neural mechanisms that different species employ to adapt the visual system to low light. However, much less explored are comparative analyses of how low light affects the flight behaviour of insect species, and the corresponding links between physiological adaptations and behaviour. We investigated whether the flower tracking behaviour of three hawkmoth species with different diel activity patterns revealed luminance-dependent adaptations, using a system identification approach. We found clear luminance-dependent differences in flower tracking in all three species, which were explained by a simple luminance-dependent delay model, which generalized across species. We discuss physiological and anatomical explanations for the variance in tracking responses, which could not be explained by such simple models. Differences between species could not be explained by the simple delay model. However, in several cases, they could be explained through the addition on a second model parameter, a simple scaling term, that captures the responsiveness of each species to flower movements. Thus, we demonstrate here that much of the variance in the luminance-dependent flower tracking responses of hawkmoths with different diel activity patterns can be captured by simple models of neural processing.This article is part of the themed issue 'Vision in dim light'.

Adaptations for nocturnal and diurnal vision in the hawkmoth lamina.

Animals use vision over a wide range of light intensities, from dim starlight to bright sunshine. For animals active in very dim light the visual system is challenged by several sources of visual noise. Adaptations in the eyes, as well as in the neural circuitry, have evolved to suppress the noise and enhance the visual signal, thereby improving vision in dim light. Among neural adaptations, spatial summation of visual signals from neighboring processing units is suggested to increase the reliability of signal detection and thus visual sensitivity. In insects, the likely neural candidates for carrying out spatial summation are the lamina monopolar cells (LMCs) of the first visual processing area of the insect brain (the lamina). We have classified LMCs in three species of hawkmoths with considerably different activity periods but very similar ecology-the diurnal Macroglossum stellatarum, the nocturnal Deilephila elpenor and the crepuscular-nocturnal Manduca sexta. Using this classification, we investigated the anatomical adaptations of hawkmoth LMCs suited for spatial summation. We found that specific types of LMCs have dendrites extending to significantly more neighboring cartridges in the two nocturnal and crepuscular species than in the diurnal species, making these LMC types strong candidates for spatial summation. Moreover, while the absolute number of cartridges visited by the LMCs differed between the two dim-light species, their dendritic extents were very similar in terms of visual angle, possibly indicating a limiting spatial acuity. The overall size of the lamina neuropil did not correlate with the size of its LMCs.

A clearer view of the insect brain-combining bleaching with standard whole-mount immunocytochemistry allows confocal imaging of pigment-covered brain areas for 3D reconstruction.

In the study of insect neuroanatomy, three-dimensional (3D) reconstructions of neurons and neuropils have become a standard technique. As images have to be obtained from whole-mount brain preparations, pigmentation on the brain surface poses a serious challenge to imaging. In insects, this is a major problematic in the first visual neuropil of the optic lobe, the lamina, which is obstructed by the pigment of the retina as well as by the pigmented fenestration layer. This has prevented inclusion of this major processing center of the insect visual system into most neuroanatomical brain atlases and hinders imaging of neurons within the lamina by confocal microscopy. It has recently been shown that hydrogen peroxide bleaching is compatible with immunohistochemical labeling in insect brains, and we therefore developed a simple technique for removal of pigments on the surface of insect brains by chemical bleaching. We show that our technique enables imaging of the pigment-obstructed regions of insect brains when combined with standard protocols for both anti-synapsin-labeled as well as neurobiotin-injected samples. This method can be combined with different fixation procedures, as well as different fluorophore excitation wavelengths without negative effects on staining quality. It can therefore serve as an effective addition to most standard histology protocols used in insect neuroanatomy.