Neurobiology of polarization vision in the locust Schistocerca gregaria

The polarization pattern of the blue sky serves as an important reference for spatial orientation in insects. To understand the neural mechanisms involved in sky compass orientation we have analyzed the polarization vision system in the locust Schistocerca gregaria. As in other insects, photoreceptors adapted for the detection of sky polarization are concentrated in a dorsal rim area (DRA) of the compound eye. Stationary flying locusts show polarotactic yaw-torque responses when illuminated through a rotating polarizer from above. This response is abolished after painting the DRAs. Central stages of the polarization vision system, revealed through tracing studies, include dorsal areas in the lamina and medulla, the anterior lobe of the lobula, the anterior optic tubercle, the lateral accessory lobe and the central complex. Physiological analysis of polarization-sensitive (POL) neurons has focussed on the optic tubercle and on the central complex. Each POL neuron was maximally excited at a certain e-vector (phimax) and was maximally inhibited at an e-vector perpendicular to phimax. The neurons had large visual fields, and many neurons received input from both eyes. The neuronal organization of the central complex suggests a role as a spatial compass within the locust brain.     

Integration of polarization and chromatic cues in the insect sky compass

Animals relying on a celestial compass for spatial orientation may use the position of the sun, the chromatic or intensity gradient of the sky, the polarization pattern of the sky, or a combination of these cues as compass signals. Behavioral experiments in bees and ants, indeed, showed that direct sunlight and sky polarization play a role in sky compass orientation, but the relative importance of these cues are species-specific. Intracellular recordings from polarization-sensitive interneurons in the desert locust and monarch butterfly suggest that inputs from different eye regions, including polarized-light input through the dorsal rim area of the eye and chromatic/intensity gradient input from the main eye, are combined at the level of the medulla to create a robust compass signal. Conflicting input from the polarization and chromatic/intensity channel, resulting from eccentric receptive fields, is eliminated at the level of the anterior optic tubercle and central complex through internal compensation for changing solar elevations, which requires input from a circadian clock. Across several species, the central complex likely serves as an internal sky compass, combining E-vector information with other celestial cues. Descending neurons, likewise, respond both to zenithal polarization and to unpolarized cues in an azimuth-dependent way.     

Transmedulla Neurons in the Sky Compass Network of the Honeybee (Apis mellifera) Are a Possible Site of Circadian Input

Honeybees are known for their ability to use the sun’s azimuth and the sky’s polarization pattern for spatial orientation. Sky compass orientation in bees has been extensively studied at the behavioral level but our knowledge about the underlying neuronal systems and mechanisms is very limited. Electrophysiological studies in other insect species suggest that neurons of the sky compass system integrate information about the polarization pattern of the sky, its chromatic gradient, and the azimuth of the sun. In order to obtain a stable directional signal throughout the day, circadian changes between the sky polarization pattern and the solar azimuth must be compensated. Likewise, the system must be modulated in a context specific way to compensate for changes in intensity, polarization and chromatic properties of light caused by clouds, vegetation and landscape. The goal of this study was to identify neurons of the sky compass pathway in the honeybee brain and to find potential sites of circadian and neuromodulatory input into this pathway. To this end we first traced the sky compass pathway from the polarization-sensitive dorsal rim area of the compound eye via the medulla and the anterior optic tubercle to the lateral complex using dye injections. Neurons forming this pathway strongly resembled neurons of the sky compass pathway in other insect species. Next we combined tracer injections with immunocytochemistry against the circadian neuropeptide pigment dispersing factor and the neuromodulators serotonin, and γ-aminobutyric acid. We identified neurons, connecting the dorsal rim area of the medulla to the anterior optic tubercle, as a possible site of neuromodulation and interaction with the circadian system. These neurons have conspicuous spines in close proximity to pigment dispersing factor-, serotonin-, and GABA-immunoreactive neurons. Our data therefore show for the first time a potential interaction site between the sky compass pathway and the circadian clock.     

Surgical lesion of the anterior optic tract abolishes polarotaxis in tethered flying locusts, <Emphasis Type="Italic">Schistocerca gregaria</Emphasis>

Many insects can detect the polarization pattern of the blue sky and rely on polarization vision for sky compass orientation. In laboratory experiments, tethered flying locusts perform periodic changes in flight behavior under a slowly rotating polarizer even if one eye is painted black. Anatomical tracing studies and intracellular recordings have suggested that the polarization vision pathway in the locust brain involves the anterior optic tract and tubercle, the lateral accessory lobe, and the central complex of the brain. To investigate whether visual pathways through the anterior optic tract mediate polarotaxis in the desert locust, we transected the tract on one side and tested polarotaxis (1) with both eyes unoccluded and (2) with the eye of the intact hemisphere painted black. In the second group of animals, but not in the first group, polarotaxis was abolished. Sham operations did not impair polarotaxis. The experiments show that the anterior optic tract is an indispensable part of visual pathways mediating polarotaxis in the desert locust.     

A distinct layer of the medulla integrates sky compass signals in the brain of an insect

Mass migration of desert locusts is a common phenomenon in North Africa and the Middle East but how these insects navigate is still poorly understood. Laboratory studies suggest that locusts are able to exploit the sky polarization pattern as a navigational cue. Like other insects locusts detect polarized light through a specialized dorsal rim area (DRA) of the eye. Polarization signals are transmitted through the optic lobe to the anterior optic tubercle (AOTu) and, finally, to the central complex in the brain. Whereas neurons of the AOTu integrate sky polarization and chromatic cues in a daytime dependent manner, the central complex holds a topographic representation of azimuthal directions suggesting a role as an internal sky compass. To understand further the integration of sky compass cues we studied polarization-sensitive (POL) neurons in the medulla that may be intercalated between DRA photoreceptors and AOTu neurons. Five types of POL-neuron were characterized and four of these in multiple recordings. All neurons had wide arborizations in medulla layer 4 and most, additionally, in the dorsal rim area of the medulla and in the accessory medulla, the presumed circadian clock. The neurons showed type-specific orientational tuning to zenithal polarized light and azimuth tuning to unpolarized green and UV light spots. In contrast to neurons of the AOTu, we found no evidence for color opponency and daytime dependent adjustment of sky compass signals. Therefore, medulla layer 4 is a distinct stage in the integration of sky compass signals that precedes the time-compensated integration of celestial cues in the AOTu.     

Evidence for the possible existence of a second polarization-vision pathway in the locust brain

For spatial orientation and navigation, many insects derive compass information from the polarization pattern of the blue sky. The desert locust Schistocerca gregaria detects polarized light with a specialized dorsal rim area of its compound eye. In the locust brain, polarized-light signals are passed through the anterior optic tract and tubercle to the central complex which most likely serves as an internal sky compass. Here, we suggest that neurons of a second visual pathway, via the accessory medulla and posterior optic tubercle, also provide polarization information to the central complex. Intracellular recordings show that two types of neuron in this posterior pathway are sensitive to polarized light. One cell type connects the dorsal rim area of the medulla with the medulla and accessory medulla, and a second type connects the bilaterally paired posterior optic tubercles. Given the evidence for a role of the accessory medulla as the master clock controlling circadian changes in behavioral activity in flies and cockroaches, our data open the possibility that time-compensated polarized-light signals may reach the central complex via this pathway for time-compensated sky-compass navigation.     

Polarized skylight orientation in the desert antCataglyphis

Summary The desert antCataglyphis bicolor is able to use the pattern of polarized light in the sky as compass. By confronting the ant to single spots of artificially and naturally polarized light it is shown howCataglyphis uses the polarization pattern.When exposed to a horizontal e-vector,Cataglyphis was always oriented correctly. Orientation errors occurred, however, when other e-vector directions were presented. This indicates that the e-vector positions assumed by the ant do not coincide with the e-vector positions actually realized in the sky. From this it is concluded thatCataglyphis has no detailed knowledge of the actual azimuthal positions of the e-vectors. Instead, it is relying on a simplified celestial map of the polarization patterns in the sky (Fig. 7).Usually, the ant did not confuse celestial spots with identical e-vector directions. Even at sunset when the polarization pattern is completely ambiguous, correct orientation occurred. This suggests that the ant uses additional celestial cues such as the degree of polarization, the color or the intensity to find its way home when the sun is obscured.     

Experience‐related reorganization of giant synapses in the lateral complex: Potential role in plasticity of the sky‐compass pathway in the desert ant Cataglyphis fortis

Cataglyphis desert ants undergo an age‐related polyethism from interior workers to relatively short‐lived foragers with remarkable visual navigation capabilities, predominantly achieved by path integration using a polarized skylight‐based sun compass and a stride‐integrating odometer. Behavioral and physiological experiments revealed that the polarization (POL) pattern is processed via specialized UV‐photoreceptors in the dorsal rim area of the compound eye and POL sensitive optic lobe neurons. Further information about the neuronal substrate for processing of POL information in the ant brain has remained elusive. This work focuses on the lateral complex (LX), known as an important relay station in the insect sky‐compass pathway. Neuroanatomical results in Cataglyphis fortis show that LX giant synapses (GS) connect large presynaptic terminals from anterior optic tubercle neurons with postsynaptic GABAergic profiles of tangential neurons innervating the ellipsoid body of the central complex. At the ultrastructural level, the cup‐shaped presynaptic structures comprise many active zones contacting numerous small postsynaptic profiles. Three‐dimensional quantification demonstrated a significantly higher number of GS (～13%) in foragers compared with interior workers. Light exposure, as opposed to age, was necessary and sufficient to trigger a similar increase in GS numbers. Furthermore, the increase in GS numbers was sensitive to the exclusion of UV light. As previous experiments have demonstrated the importance of the UV spectrum for sky‐compass navigation in Cataglyphis, we conclude that plasticity in LX GS may reflect processes involved in the initial calibration of sky‐compass neuronal circuits during orientation walks preceding active foraging. © 2015 Wiley Periodicals, Inc. Develop Neurobiol 76: 390–404, 2016     

Evidence for instantaneous e-vector detection in the honeybee using an associative learning paradigm

Many insects use the polarization pattern of the sky for obtaining compass information during orientation or navigation. E-vector information is collected by a specialized area in the dorsal-most part of the compound eye, the dorsal rim area (DRA). We tested honeybees' capability of learning certain e-vector orientations by using a classical conditioning paradigm with the proboscis extension reflex. When one e-vector orientation (CS+) was associated with sugar water, while another orientation (CS-) was not rewarded, the honeybees could discriminate CS+ from CS-. Bees whose DRA was inactivated by painting did not learn CS+. When ultraviolet (UV) polarized light (350 nm) was used for CS, the bees discriminated CS+ from CS-, but no discrimination was observed in blue (442 nm) or green light (546 nm). Our data indicate that honeybees can learn and discriminate between different e-vector orientations, sensed by the UV receptors of the DRA, suggesting that bees can determine their flight direction from polarized UV skylight during foraging. Fixing the bees' heads during the experiments did not prevent learning, indicating that they use an 'instantaneous' algorithm of e-vector detection; that is, the bees do not need to actively scan the sky with their DRAs ('sequential' method) to determine e-vector orientation.     

Coding of azimuthal directions via time-compensated combination of celestial compass cues

Many animals use the sun as a reference for spatial orientation [1-3]. In addition to sun position, the sky provides two other sources of directional information, a color gradient [4] and a polarization pattern [5]. Work on insects has predominantly focused on celestial polarization as an orientation cue [6, 7]. Relying on sky polarization alone, however, poses the following two problems: E vector orientations in the sky are not suited to distinguish between the solar and antisolar hemisphere of the sky, and the polarization pattern changes with changing solar elevation during the day [8, 9]. Here, we present neurons that overcome both problems in a locust's brain. The spiking activity of these neurons depends (1) on the E vector orientation of dorsally presented polarized light, (2) on the azimuthal, i.e., horizontal, direction, and (3) on the wavelength of an unpolarized light source. Their tuning to these stimuli matches the distribution of a UV/green chromatic contrast as well as the polarization of natural skylight and compensates for changes in solar elevation during the day. The neurons are, therefore, suited to code for solar azimuth by concurrent combination of signals from the spectral gradient, intensity gradient, and polarization pattern of the sky.     

Behavioral analysis of polarization vision in tethered flying locusts

For spatial navigation many insects rely on compass information derived from the polarization pattern of the sky. We demonstrate that tethered flying desert locusts (Schistocerca gregaria) show e-vector-dependent yaw-torque responses to polarized light presented from above. A slowly rotating polarizer (5.3° s^–1) induced periodic changes in yaw torque corresponding to the 180° periodicity of the stimulus. Control experiments with a rotating diffuser, a weak intensity pattern, and a stationary polarizer showed that the response is not induced by intensity gradients in the stimulus. Polarotaxis was abolished after painting the dorsal rim areas of the compound eyes black, but remained unchanged after painting the eyes except the dorsal rim areas. During rotation of the polarizer, two e-vectors (preferred and avoided e-vector) induced no turning responses: they were broadly distributed from 0 to 180° but, for a given animal, were perpendicular to each other. The data demonstrate polarization vision in the desert locust, as shown previously for bees, flies, crickets, and ants. Polarized light is perceived through the dorsal rim area of the compound eye, suggesting that polarization vision plays a role in compass navigation of the locust.     

Navigation by light polarization in clear and turbid waters

Certain terrestrial animals use sky polarization for navigation. Certain aquatic species have also been shown to orient according to a polarization stimulus, but the correlation between underwater polarization and Sun position and hence the ability to use underwater polarization as a compass for navigation is still under debate. To examine this issue, we use theoretical equations for per cent polarization and electric vector (e-vector) orientation that account for the position of the Sun, refraction at the air-water interface and Rayleigh single scattering. The polarization patterns predicted by these theoretical equations are compared with measurements conducted in clear and semi-turbid coastal sea waters at 2 m and 5 m depth over sea floors of 6 m and 28 m depth. We find that the per cent polarization is correlated with the Sun's elevation only in clear waters. We furthermore find that the maximum value of the e-vector orientation angle equals the angle of refraction only in clear waters, in the horizontal viewing direction, over the deeper sea floor. We conclude that navigation by use of underwater polarization is possible under restricted conditions, i.e. in clear waters, primarily near the horizontal viewing direction, and in locations where the sea floor has limited effects on the light's polarization.     

Neural mechanisms in insect navigation: polarization compass and odometer

Insect navigation relies on path integration, a procedure by which information about compass bearings pursued and distances travelled are combined to calculate position. Three neural levels of the polarization compass, which uses the polarization of skylight as a reference, have been analyzed in orthopteran insects. A group of dorsally directed, highly specialized ommatidia serve as polarization sensors. Polarization-opponent neurons in the optic lobe condition the polarization signal by removing unreliable and irrelevant components of the celestial stimulus. Neurons found in the central complex of the brain possibly represent elements of the compass output. The odometer for measuring travelling distances in honeybees relies on optic flow experienced during flight, whereas desert ants most probably use proprioreceptive cues.     

Orientation in a desert lizard (Uma notata): time-compensated compass movement and polarotaxis

Summary The diurnal escape response of fringetoed lizards (Uma notata) startled by predators demonstrates clear directional orientation not likely to depend on local landmarks in the shifting sands of their desert environment. Evidence that celestial orientation is involved in this behavior has been sought in the present experiments by testing the effects of (1) phase shifting the animal's internal clock by 6 h and (2) by training the lizards to seek shelter while exposed to natural polarization patterns. In the first case, 90° shifts in escape direction were demonstrated in outdoor tests, as expected if a time-compensated sun or sky polarized light compass is involved. In the second instance, significant bimodale-vector dependent orientation was found under an overhead polarizing light filter but this was only evident when the response data were transposed to match the zenithe-vector rotation dependent on the sun's apparent movement through the sky. This extends to reptiles the capacity to utilize overheade-vector directions as a time-compensated sky compass. The sensory site of this discrimination and the relative roles of sun and sky polarization in nature remain to be discovered.     

Ultrastructure and orientation of ommatidia in the dorsal rim area of the locust compound eye

In many insect species, a dorsal rim area (DRA) in the compound eye is adapted to analyze the sky polarization pattern for compass orientation. In the desert locust Schistocerca gregaria, these specializations are particularly striking. The DRA of the locust consists of about 400 ommatidia. The facets have an irregular shape, and pore canals are often present in the corneae. Screening pigment is missing in the region of the dioptric apparatus suggesting large receptive fields. The rhabdoms are shorter, but about four times larger in cross-section than the rhabdoms of ordinary ommatida. Eight retinula cells contribute to the rhabdom. The microvilli of retinula cell 7 and of cells 1, 2, 5, 6, 8 are highly aligned throughout the rhabdom and form two blocks of orthogonal orientation. The microvilli in the minute rhabdomeres of retinula cells 3 and 4, in contrast, show no particular alignment. As in other insect species, microvillar orientations are arranged in a fan-like pattern across the DRA. Photoreceptor axons project to distinct areas in the dorsal lamina and medulla. The morphological specializations in the DRA of the locust eye most likely maximize the polarization sensitivity and suggest that the locust uses this eye region for analysis of the sky polarization pattern.     

Celestial orientation in bees: the use of spectral cues

Summary The spectral cues used in the bee's celestial compass are investigated by presenting bees dancing on a horizontal comb with unpolarized (or polarized) spectral stimuli. Where appropriate, the use of e-vector information is prevented by painting out the specialized dorsal margin of the bee's eye (POL area, Fig. 1). This area has been shown to mediate e-vector information (Fig. 3; Wehner 1982), whereas the remainder of the dorsal retina is sufficient for mediating spectral information (Fig. 4).Spectral cues are used by the bees to discriminate between sun and sky (Fig. 4). According to physical reality (Fig. 2), a long-wavelength stimulus is taken as the sun, whereas a short-wavelength stimulus is expected by the bee to lie anywhere within the antisolar half of the sky (Figs. 5 and 6). This is in accord with the bee's e-vector compass in which e-vectors are confined to the antisolar half of the sky (Fig. 9).In general, spectral cues do not provide precise compass information except when a full celestial colour gradient is available including the solar and the antisolar meridian (Figs. 7 and 8).     

Optic lobe projections of marginal ommatidia in Calliphora erythrocephala specialized for detecting polarized light

Summary The structure of ommatidia at the dorsal eye margin of the fly, Calliphora erythrocephala is specialized for the detection of the e-vector of polarized light. Marginal zone ommatidia are distinguished by R7/R8 receptor cells with large-diameter, short, untwisted rhabdomeres and long axons to the medulla. The arrangement of the R7 microvillar directions along the marginal zone is fan-shaped. Ommatidia lining the dorsal and frontal edge of the eye lack primary screening pigments and have foreshortened crystalline cones. The marginal ommatidia from each eye view a strip that is 5 °–20 ° contralateral to the fly's longitudinal axis and that coincides with the outer boundaries of the binocular overlap.Cobalt injection into the retina demonstrates that photoreceptor axons arising from marginal ommatidia define a special area of marginal neuropil in the second visual neuropil, the medulla. Small-field neurons arising from the marginal medulla area define, in turn, a special area of marginal neuropil in the two deepest visual neuropils, the lobula and the lobula plate. From these arise local assemblies of columnar neurons that relay the marginal zones of one optic lobe to equivalent areas of the opposite lobe and to midbrain regions from which arise descending neurons destined for the the thoracic ganglia.Optically, the marginal zone of the retina represents the lateral edge of a larger area of ommatidia involved in dorsofrontal binocular overlap. This binocularity area is also represented by special arrangements of columnar neurons, which map the binocularity area of one eye into the lobula beneath the opposite eye. Another type of binocularity neuron terminates in the midbrain.These neuronal arrangements suggest two novel features of the insect optic lobes and brain: (1) Marginal neurons that directly connect the left and right optic lobes imply that each lobe receives a common input from areas of the left and right eye, specialized for detecting the pattern of polarized light. (2) Information about the e-vector pattern of sky-light polarization may be integrated with binocular and monocular pathways at the level of descending neurons leading to thoracic motor neuropil.     

Evidence for calibration of magnetic migratory orientation in Savannah sparrows reared in the field

The orientation system of migratory birds consists of a magnetic compass and compasses based upon celestial cues. In many places, magnetic compass directions and true or geographic compass directions differ (referred to as magnetic declination). It has been demonstrated experimentally in several species that the innate preferred direction of magnetic orientation can be calibrated by celestial rotation, an indicator of geographic directions. This calibration process brings the two types of compass into conformity and provides the birds with a mechanism that compensates for the spatial variation in magnetic declination. Calibration of magnetic orientation has heretofore been demonstrated only with hand-raised birds exposed to very large declination (90° or more). Here we show that the magnetic orientation of wild birds from near Albany, New York, USA (declination = 14° W) was N–S, a clockwise shift of 26° from the NNW–SSE direction of birds raised entirely indoors. Hand-raised birds having visual experience with either the daytime sky or both day and night sky orientated N–S, similar to wild-caught birds. These data provide the first confirmation that calibration of magnetic orientation occurs under natural conditions and in response to modest declination values.     

How dim is dim? Precision of the celestial compass in moonlight and sunlight

Prominent in the sky, but not visible to humans, is a pattern of polarized skylight formed around both the Sun and the Moon. Dung beetles are, at present, the only animal group known to use the much dimmer polarization pattern formed around the Moon as a compass cue for maintaining travel direction. However, the Moon is not visible every night and the intensity of the celestial polarization pattern gradually declines as the Moon wanes. Therefore, for nocturnal orientation on all moonlit nights, the absolute sensitivity of the dung beetle's polarization detector may limit the precision of this behaviour. To test this, we studied the straight-line foraging behaviour of the nocturnal ball-rolling dung beetle Scarabaeus satyrus to establish when the Moon is too dim--and the polarization pattern too weak--to provide a reliable cue for orientation. Our results show that celestial orientation is as accurate during crescent Moon as it is during full Moon. Moreover, this orientation accuracy is equal to that measured for diurnal species that orient under the 100 million times brighter polarization pattern formed around the Sun. This indicates that, in nocturnal species, the sensitivity of the optical polarization compass can be greatly increased without any loss of precision.     

Polarized light helps monarch butterflies navigate

During their spectacular migratory journey in the fall, North American monarch butterflies (Danaus plexippus) use a time-compensated sun compass to help them navigate to their overwintering sites in central Mexico. One feature of the sun compass mechanism not fully explored in monarchs is the sunlight-dependent parameters used to navigate. We now provide data suggesting that the angle of polarized skylight (the e-vector) is a relevant orientation parameter. By placing butterflies in a flight simulator outdoors and using a linear polarizing filter, we show that manipulating the e-vector alters predictably the direction of oriented flight. Butterflies studied in either the morning or afternoon showed similar responses to filter rotation. Monarch butterflies possess the anatomical structure needed for polarized skylight detection, as rhabdoms in the dorsalmost row of photoreceptor cells in monarch eye show the organization characteristic of polarized-light receptors. The existence of polarized-light detection could allow migrants to accurately navigate under a variety of atmospheric conditions and reveals a critical input pathway into the sun compass mechanism.