Snapping shrimp have helmets that protect their brains by dampening shock waves.

Shock waves are supersonic high-amplitude pressure waves that cause barotrauma when they transfer kinetic energy to the tissues of animals.1-4 Snapping shrimp (Alpheidae) produce shock waves and are exposed to them frequently, so we asked if these animals have evolved mechanisms of physical protection against them. Snapping shrimp generate shock waves by closing their snapping claws rapidly enough to form cavitation bubbles that release energy as an audible "snap" and a shock wave when they collapse.5-8 We tested if snapping shrimp are protected from shock waves by a helmet-like extension of their exoskeleton termed the orbital hood. Using behavioral trials, we found shock wave exposure slowed shelter-seeking and caused a loss of motor control in Alpheus heterochaelis from which we had removed orbital hoods but did not significantly affect behavior in shrimp with unaltered orbital hoods. Shock waves thus have the potential to harm snapping shrimp but may not do so under natural conditions because of protection provided to shrimp by their orbital hoods. Using pressure recordings, we discovered the orbital hoods of A. heterochaelis dampen shock waves. Sealing the anterior openings of orbital hoods diminished how much they altered the magnitudes of shock waves, which suggests these helmet-like structures dampen shock waves by trapping and expelling water so that kinetic energy is redirected and released away from the heads of shrimp. Our results indicate orbital hoods mitigate blast-induced neurotrauma in snapping shrimp by dampening shock waves, making them the first biological armor system known to have such a function. VIDEO ABSTRACT.

Automated methods for efficient and accurate electroretinography.

Electroretinography (ERG) is a foundational method for assessing visual system physiology, but accurate ERG can be time- and labor-intensive, often involving manual adjustment of the wavelength and intensity of light stimuli and real-time comparison of physiological responses to inform those adjustments. Furthermore, current approaches to ERG often require expertise beyond that necessary for the electrophysiological preparation itself. To improve both the efficiency and accessibility of ERG, we designed an automated system for stimulus presentation and data acquisition. Here, we test this novel system's ability to accurately assess spectral sensitivity in the well-characterized visual system of the crayfish Procambarus clarkii using three approaches: the first, based on response magnitude, maximizes efficiency; the second is a well-established method we use to further validate our efficient approach's accuracy. Third, we explore the potential benefits of extensible automation using a method assessing the interplay between temporal acuity and spectral sensitivity. Using our system, we are able to acquire accurate results in ERG experiments quickly (testing the entire visible spectrum in 8 min, 30 s using our response magnitude approach). Moreover, data collected via all three methods yielded results consistent with each other and previous work on P. clarkii.

The orbital hoods of snapping shrimp have surface features that may represent tradeoffs between vision and protection.

Snapping shrimp (Alpheidae) are decapod crustaceans named for the snapping claws with which they produce cavitation bubbles. Snapping shrimp use the shock waves released by collapsing cavitation bubbles as weapons. Along with their distinctive claws, snapping shrimp have orbital hoods, extensions of their carapace that cover their heads and eyes. Snapping shrimp view the world through their orbital hoods, so we asked if the surfaces of the orbital hoods of the snapping shrimp Alpheus heterochaelis have features that minimize the scattering of light. Using SEM, we found that surface features, primarily microbial epibionts, covered less space on the surfaces of the orbital hoods of A. heterochaelis (∼18%) than they do elsewhere on the carapace (∼50%). Next, we asked if these surface features influence aerophobicity. By measuring the contact angles of air bubbles, we found the orbital hoods of A. heterochaelis are less aerophobic than other regions of the carapace. Surfaces that are less aerophobic are more likely to have cavitation bubbles adhere to them and are more likely to have shock waves cause new cavitation bubbles to nucleate upon them. Computational modeling indicates the orbital hoods of A. heterochaelis face a functional trade-off: fewer surface features, such as less extensive communities of microbial epibionts, may minimize the scattering of light at the cost of making the adhesion and nucleation of cavitation bubbles more likely.

A snapping shrimp has the fastest vision of any aquatic animal.

Animals use their sensory systems to sample information from their environments. The physiological properties of sensory systems differ, leading animals to perceive their environments in different ways. For example, eyes have different temporal sampling rates, with faster-sampling eyes able to resolve faster-moving scenes. Eyes can also have different dynamic ranges. For every eye, there is a light level below which vision is unreliable because of an insufficient signal-to-noise ratio and a light level above which the photoreceptors are saturated. Here, we report that the eyes of the snapping shrimp Alpheus heterochaelis have a temporal sampling rate of at least 160 Hz, making them the fastest-sampling eyes ever described in an aquatic animal. Fast-sampling eyes help flying animals detect objects moving across their retinas at high angular velocities. A. heterochaelis are fast-moving animals that live in turbid, structurally complex oyster reefs and their fast-sampling eyes, like those of flying animals, may help them detect objects moving rapidly across their retinas. We also report that the eyes of A. heterochaelis have a broad dynamic range that spans conditions from late twilight (approx. 1 lux) to direct sunlight (approx. 100 000 lux), a finding consistent with the circatidal activity patterns of this shallow-dwelling species.

Core-shell nanospheres behind the blue eyes of the bay scallop Argopecten irradians.

The bay scallop Argopecten irradians (Mollusca: Bivalvia) has dozens of iridescent blue eyes that focus light using mirror-based optics. Here, we test the hypothesis that these eyes appear blue because of photonic nanostructures that preferentially scatter short-wavelength light. Using transmission electron microscopy, we found that the epithelial cells covering the eyes of A. irradians have three distinct layers: an outer layer of microvilli, a middle layer of random close-packed nanospheres and an inner layer of pigment granules. The nanospheres are approximately 180 nm in diameter and consist of electron-dense cores approximately 140 nm in diameter surrounded by less electron-dense shells 20 nm thick. They are packed at a volume density of approximately 60% and energy-dispersive X-ray spectroscopy indicates that they are not mineralized. Optical modelling revealed that the nanospheres are an ideal size for producing angle-weighted scattering that is bright and blue. A comparative perspective supports our hypothesis: epithelial cells from the black eyes of the sea scallop Placopecten magellanicus have an outer layer of microvilli and an inner layer of pigment granules but lack a layer of nanospheres between them. We speculate that light-scattering nanospheres help to prevent UV wavelengths from damaging the internal structures of the eyes of A. irradians and other blue-eyed scallops.

Vision in the snapping shrimp Alpheus heterochaelis.

Snapping shrimp engage in heterospecific behavioral associations in which their partners, such as goby fish, help them avoid predators. It has been argued that snapping shrimp engage in these partnerships because their vision is impaired by their orbital hood, an extension of their carapace that covers their eyes. To examine this idea, we assessed the visual abilities of snapping shrimp. We found the big claw snapping shrimp, Alpheus heterochaelis, has spatial vision provided by compound eyes with reflecting superposition optics. These eyes view the world through an orbital hood that is 80-90% as transparent as seawater across visible wavelengths (400-700 nm). Through electroretinography and microspectrophotometry, we found the eyes of A. heterochaelis have a temporal sampling rate of >40 Hz and have at least two spectral classes of photoreceptors (λmax=500 and 519 nm). From the results of optomotor behavioral experiments, we estimate the eyes of A. heterochaelis provide spatial vision with an angular resolution of ∼8 deg. We conclude that snapping shrimp have competent visual systems, suggesting the function and evolution of their behavioral associations should be re-assessed and that these animals may communicate visually with conspecifics and heterospecific partners.

The mirror-based eyes of scallops demonstrate a light-evoked pupillary response.

Light levels in terrestrial and shallow-water environments can vary by ten orders of magnitude between clear days and overcast nights. Light-evoked pupillary responses help the eyes of animals perform optimally under these variable light conditions by balancing trade-offs between sensitivity and resolution [1]. Here, we document that the mirror-based eyes of the bay scallop Argopecten irradians and the sea scallop Placopecten magellanicus have pupils that constrict to ∼60% of their fully dilated areas within several minutes of light exposure. The eyes of scallops contain two separate retinas and our ray-tracing model indicates that, compared to eyes with fully constricted pupils, eyes from A. irradians with fully dilated pupils provide approximately three times the sensitivity and half the spatial resolution at the distal retina and five times the sensitivity and one third the spatial resolution at the proximal retina. We also identify radial and circular actin fibers associated with the corneas of A. irradians that may represent muscles whose contractions dilate and constrict the pupil, respectively.

Evidence for spatial vision in Chiton tuberculatus, a chiton with eyespots.

To better understand relationships between the structures and functions of the distributed visual systems of chitons, we compare how morphological differences between the light-sensing structures of these animals relate to their visually guided behaviors. All chitons have sensory organs - termed aesthetes - embedded within their protective shell plates. In some species, the aesthetes are interspersed with small, image-forming eyes. In other species, the aesthetes are paired with pigmented eyespots. Previously, we compared the visually influenced behaviors of chitons with aesthetes to those of chitons with both aesthetes and eyes. Here, we characterize the visually influenced behaviors of chitons with aesthetes and eyespots. We find that chitons with eyespots engage in behaviors consistent with spatial vision, but appear to use spatial vision for different tasks than chitons with eyes. Unlike chitons with eyes, Chiton tuberculatus and C. marmoratus fail to distinguish between sudden appearances of overhead objects and equivalent, uniform changes in light levels. We also find that C. tuberculatus orients to static objects with angular sizes as small as 10 deg. Thus, C. tuberculatus demonstrates spatial resolution that is at least as fine as that demonstrated by chitons with eyes. The eyespots of Chiton are smaller and more numerous than the eyes found in other chitons and they are separated by angles of <0.5 deg, suggesting that the light-influenced behaviors of Chiton may be more accurately predicted by the network properties of their distributed visual system than by the structural properties of their individual light-detecting organs.

Expression of G Proteins in the Eyes and Parietovisceral Ganglion of the Bay Scallop Argopecten irradians.

A multitude of image-forming eyes are spread across the bodies of certain invertebrates. Recent efforts have characterized how these eyes function, but less progress has been made toward describing the neural structures associated with them. Scallops, for example, have a distributed visual system that includes dozens of eyes whose optic nerves project to the lateral lobes of the parietovisceral ganglion (PVG). To identify sensory receptors and chemical synapses associated with the scallop visual system, we studied the expression of four G protein α subunits (Gαi, Gαo, Gαq, and Gαs) in the eyes and PVG of the bay scallop Argopecten irradians (Lamarck, 1819). In the eyes of A. irradians, we noted expression of Gαo by the ciliary photoreceptors of the distal retina, expression of Gαq by the rhabdomeric photoreceptors of the proximal retina, and the expression of Gαo and Gαq by the cells of the cornea; we did not, however, detect expression of Gαi or Gαs in the eyes. In the PVG of A. irradians, we noted widespread expression of Gαi, Gαo, and Gαq. The expression of Gαs was limited to fine neurites in the lateral and ventral central lobes, as well as large unipolar neurons in the dorsal central lobes. Our findings suggest that light detection by the eyes of A. irradians is conferred primarily by photoreceptors that express Gαo or Gαq, that the corneal cells of scallops may contain sensory receptors and/or receive neural input, and that G protein labeling is useful for visualizing substructures and identifying specific populations of cells within the nervous systems of invertebrates.

Diverse Distributions of Extraocular Opsins in Crustaceans, Cephalopods, and Fish.

Non-visual and extraocular photoreceptors are common among animals, but current understanding linking molecular pathways to physiological function of these receptors is lacking. Opsin diversity in extraocular tissues suggests that many putative extraocular photoreceptors utilize the "visual" phototransduction pathway-the same phototransduction pathway as photoreceptors within the retina dedicated to light detection for image sensing. Here, we provide a brief overview of the current understanding of non-visual and extraocular photoreceptors, and contribute a synopsis of several novel putative extraocular photoreceptors that use both visual and non-visual phototransduction pathways. Crayfish, cephalopods, and flat fish express opsins in diverse tissues, suggesting the presence of extraocular photoreceptors. In most cases, we find that these animals use the same phototransduction pathway that is utilized in the retinas for image-formation. However, we also find the presence of non-visual phototransduction components in the skin of flounders. Our evidence suggests that extraocular photoreceptors may employ a number of phototransduction pathways that do not appear to correlate with purpose or location of the photoreceptor.

Short- and long-wavelength-sensitive opsins are involved in photoreception both in the retina and throughout the central nervous system of crayfish.

Crayfish have two classes of photoreceptors in the retinas of their reflecting superposition eyes. Long-wavelength-sensitive photoreceptors, comprised of microvilli from R1-7 cells, make up the main rhabdoms. Eighth retinular cells, located distal to the main rhabdoms, house short-wavelength-sensitive photoreceptors. While the opsin involved in long-wavelength sensitivity has long been known, we present the first description of the short-wavelength-sensitive opsin in the retina of the red swamp crayfish, Procambarus clarkii. The expression patterns of these SWS and LWS opsin proteins in the retina are consistent with the previously described locations of SWS and LWS receptors. Crayfish also have a well-characterized extraocular photoreceptor, called the caudal photoreceptor, located in the sixth abdominal ganglion. To search for retinal opsins in the caudal photoreceptor (and elsewhere in the CNS), we used RT-PCR and immunohistochemical labeling. We found both SWS and LWS opsin transcripts not only in the sixth abdominal ganglion, but also in all ganglia of the nerve cord. Immunolabeling shows that both opsins are expressed in nerve fibers that extend from the brain through the entire length of the CNS. Thus, the same two photopigments are used both for vision in the retina and for extraocular functions throughout the CNS of crayfish.

An Unexpected Diversity of Photoreceptor Classes in the Longfin Squid, Doryteuthis pealeii.

Cephalopods are famous for their ability to change color and pattern rapidly for signaling and camouflage. They have keen eyes and remarkable vision, made possible by photoreceptors in their retinas. External to the eyes, photoreceptors also exist in parolfactory vesicles and some light organs, where they function using a rhodopsin protein that is identical to that expressed in the retina. Furthermore, dermal chromatophore organs contain rhodopsin and other components of phototransduction (including retinochrome, a photoisomerase first found in the retina), suggesting that they are photoreceptive. In this study, we used a modified whole-mount immunohistochemical technique to explore rhodopsin and retinochrome expression in a number of tissues and organs in the longfin squid, Doryteuthis pealeii. We found that fin central muscles, hair cells (epithelial primary sensory neurons), arm axial ganglia, and sucker peduncle nerves all express rhodopsin and retinochrome proteins. Our findings indicate that these animals possess an unexpected diversity of extraocular photoreceptors and suggest that extraocular photoreception using visual opsins and visual phototransduction machinery is far more widespread throughout cephalopod tissues than previously recognized.

Visual phototransduction components in cephalopod chromatophores suggest dermal photoreception.

Cephalopod mollusks are renowned for their colorful and dynamic body patterns, produced by an assemblage of skin components that interact with light. These may include iridophores, leucophores, chromatophores and (in some species) photophores. Here, we present molecular evidence suggesting that cephalopod chromatophores - small dermal pigmentary organs that reflect various colors of light - are photosensitive. RT-PCR revealed the presence of transcripts encoding rhodopsin and retinochrome within the retinas and skin of the squid Doryteuthis pealeii, and the cuttlefish Sepia officinalis and Sepia latimanus. In D. pealeii, Gqα and squid TRP channel transcripts were present in the retina and in all dermal samples. Rhodopsin, retinochrome and Gqα transcripts were also found in RNA extracts from dissociated chromatophores isolated from D. pealeii dermal tissues. Immunohistochemical staining labeled rhodopsin, retinochrome and Gqα proteins in several chromatophore components, including pigment cell membranes, radial muscle fibers, and sheath cells. This is the first evidence that cephalopod dermal tissues, and specifically chromatophores, may possess the requisite combination of molecules required to respond to light.

Characterization of visual pigments, oil droplets, lens and cornea in the whooping crane Grus americana.

Vision has been investigated in many species of birds, but few studies have considered the visual systems of large birds and the particular implications of large eyes and long-life spans on visual system capabilities. To address these issues we investigated the visual system of the whooping crane Grus americana (Gruiformes, Gruidae), which is one of only two North American crane species. It is a large, long-lived bird in which UV sensitivity might be reduced by chromatic aberration and entrance of UV radiation into the eye could be detrimental to retinal tissues. To investigate the whooping crane visual system we used microspectrophotometry to determine the absorbance spectra of retinal oil droplets and to investigate whether the ocular media (i.e. the lens and cornea) absorb UV radiation. In vitro expression and reconstitution was used to determine the absorbance spectra of rod and cone visual pigments. The rod visual pigments had wavelengths of peak absorbance (λmax) at 500 nm, whereas the cone visual pigment λmax values were determined to be 404 nm (SWS1), 450 nm (SWS2), 499 nm (RH2) and 561 nm (LWS), similar to other characterized bird visual pigment absorbance values. The oil droplet cut-off wavelength (λcut) values similarly fell within ranges recorded in other avian species: 576 nm (R-type), 522 nm (Y-type), 506 nm (P-type) and 448 nm (C-type). We confirm that G. americana has a violet-sensitive visual system; however, as a consequence of the λmax of the SWS1 visual pigment (404 nm), it might also have some UV sensitivity.

Male wing color properties predict the size of nuptial gifts given during mating in the Pipevine Swallowtail butterfly (Battus philenor).

In many animals, males bear bright ornamental color patches that may signal both the direct and indirect benefits that a female might accrue from mating with him. Here we test whether male coloration in the Pipevine Swallowtail butterfly, Battus philenor, predicts two potential direct benefits for females: brief copulation duration and the quantity of materials the male passes to the female during mating. In this species, males have a bright iridescent blue field on the dorsal hindwing surface, while females have little or no dorsal iridescence. Females preferentially mate with males who display a bright and highly chromatic blue field on their dorsal hindwing. In this study, we show that the chroma of the blue field on the male dorsal hindwing and male body size (forewing length) significantly predict the mass of material or spermatophore that a male forms within the female's copulatory sac during mating. We also found that spermatophore mass correlated negatively with copulation duration, but that color variables did not significantly predict this potential direct benefit. These results suggest that females may enhance the material benefits they receive during mating by mating with males based on the coloration of their dorsal hindwing.