Evidence for tactile 3D shape discrimination by octopus.

Octopuses integrate visual, chemical and tactile sensory information while foraging and feeding in complex marine habitats. The respective roles of these modes are of interest ecologically, neurobiologically, and for development of engineered soft robotic arms. While vision guides their foraging path, benthic octopuses primarily search "blindly" with their arms to find visually hidden prey amidst rocks, crevices and coral heads. Each octopus arm is lined with hundreds of suckers that possess a combination of chemo- and mechanoreceptors to distinguish prey. Contact chemoreception has been demonstrated in lab tests, but mechanotactile sensing is less well characterized. We designed a non-invasive live animal behavioral assay that isolated mechanosensory capabilities of Octopus bimaculoides arms and suckers to discriminate among five resin 3D-printed prey and non-prey shapes (all with identical chemical signatures). Each shape was introduced inside a rock dome and was only accessible to the octopus' arms. Octopuses' responses were variable. Young octopuses discriminated the crab prey shape from the control, whereas older octopuses did not. These experiments suggest that mechanotactile sensing of 3D shapes may aid in prey discrimination; however, (i) chemo-tactile information may be prioritized over mechanotactile information in prey discrimination, and (ii) mechanosensory capability may decline with age.

An experimental method for evoking and characterizing dynamic color patterning of cuttlefish during prey capture.

Cuttlefish are active carnivores that possess a wide repertoire of body patterns that can be changed within milliseconds for many types of camouflage and communication. The forms and functions of many body patterns are well known from ethological studies in the field and laboratory. Yet one aspect has not been reported in detail: the category of rapid, brief and high-contrast changes in body coloration ("Tentacle Shot Patterns" or TSPs) that always occur with the ejection of two ballistic tentacles to strike live moving prey ("Tentacles Go Ballistic" or TGB moment). We designed and tested a mechanical device that presented prey in a controlled manner, taking advantage of a key stimulus for feeding: motion of the prey. High-speed video recordings show a rapid transition into TSPs starting 114 ms before TGB (N = 114). TSPs are then suppressed as early as 470-500 ms after TGB (P < 0.05) in unsuccessful hunts, while persisting for at least 3 s after TGB in successful hunts. A granularity analysis revealed significant differences in the large-scale high-contrast body patterning present in TSPs compared to the camouflage body pattern deployed beforehand. TSPs best fit the category of secondary defense called deimatic displaying, meant to briefly startle predators and interrupt their attack sequence while cuttlefish are distracted by striking prey. We characterize TSPs as a pattern category for which the main distinguishing feature is a high-contrast signaling pattern with aspects of Acute Conflict Mottle or Acute Disruptive Pattern. The data and methodology presented here open opportunities for quantifying the rapid neural responses in this visual sensorimotor set of behaviors.

Octopus arms exhibit exceptional flexibility.

The octopus arm is often referred to as one of the most flexible limbs in nature, yet this assumption requires detailed inspection given that this has not been measured comprehensively for all portions of each arm. We investigated the diversity of arm deformations in Octopus bimaculoides with a frame-by-frame observational analysis of laboratory video footage in which animals were challenged with different tasks. Diverse movements in these hydrostatic arms are produced by some combination of four basic deformations: bending (orally, aborally; inward, outward), torsion (clockwise, counter-clockwise), elongation, and shortening. More than 16,500 arm deformations were observed in 120 min of video. Results showed that all eight arms were capable of all four types of deformation along their lengths and in all directions. Arms function primarily to bring the sucker-lined oral surface in contact with target surfaces. Bending was the most common deformation observed, although the proximal third of the arms performed relatively less bending and more shortening and elongation as compared with other arm regions. These findings demonstrate the exceptional flexibility of the octopus arm and provide a basis for investigating motor control of the entire arm, which may aid the future development of soft robotics.

Dynamic masquerade with morphing three-dimensional skin in cuttlefish.

Masquerade is a defence tactic in which a prey resembles an inedible or inanimate object thus causing predators to misclassify it. Most masquerade colour patterns are static although some species adopt postures or behaviours to enhance the effect. Dynamic masquerade in which the colour pattern can be changed is rare. Here we report a two-step sensory process that enables an additional novel capability known only in cuttlefish and octopus: morphing three-dimensional physical skin texture that further enhances the optical illusions created by coloured skin patterns. Our experimental design incorporated sequential sensory processes: addition of a three-dimensional rock to the testing arena, which attracted the cuttlefish to settle next to it; then visual processing by the cuttlefish of physical textures on the rock to guide expression of the skin papillae, which can range from fully relaxed (smooth skin) to fully expressed (bumpy skin). When a uniformly white smooth rock was presented, cuttlefish moved to the rock and deployed a uniform body pattern with mostly smooth skin. When a rock with small-scale fragments of contrasting shells was presented, the cuttlefish deployed mottled body patterns with strong papillae expression. These robust and reversible responses indicate a sophisticated visual sensorimotor system for dynamic masquerade.

Tactical decisions for changeable cuttlefish camouflage: visual cues for choosing masquerade are relevant from a greater distance than visual cues used for background matching.

Cuttlefish use multiple camouflage tactics to evade their predators. Two common tactics are background matching (resembling the background to hinder detection) and masquerade (resembling an uninteresting or inanimate object to impede detection or recognition). We investigated how the distance and orientation of visual stimuli affected the choice of these two camouflage tactics. In the current experiments, cuttlefish were presented with three visual cues: 2D horizontal floor, 2D vertical wall, and 3D object. Each was placed at several distances: directly beneath (in a circle whose diameter was one body length (BL); at zero BL [(0BL); i.e., directly beside, but not beneath the cuttlefish]; at 1BL; and at 2BL. Cuttlefish continued to respond to 3D visual cues from a greater distance than to a horizontal or vertical stimulus. It appears that background matching is chosen when visual cues are relevant only in the immediate benthic surroundings. However, for masquerade, objects located multiple body lengths away remained relevant for choice of camouflage.

How visual edge features influence cuttlefish camouflage patterning.

Rapid adaptive camouflage is the primary defense of soft-bodied cuttlefish. Previous studies have shown that cuttlefish body patterns are strongly influenced by visual edges in the substrate. The aim of the present study was to examine how cuttlefish body patterning is differentially controlled by various aspects of edges, including contrast polarity, contrast strength, and the presence or absence of "line terminators" introduced into a pattern when continuous edges are fragmented. Spatially high- and low-pass filtered white or black disks, as well as isolated, continuous and fragmented edges varying in contrast, were used to assess activation of cuttlefish skin components. Although disks of both contrast polarities evoked relatively weak disruptive body patterns, black disks activated different skin components than white disks, and high-frequency information alone sufficed to drive the responses to white disks whereas high- and low-frequency information were both required to drive responses to black disks. Strikingly, high-contrast edge fragments evoked substantially stronger body pattern responses than low-contrast edge fragments, whereas the body pattern responses evoked by high-contrast continuous edges were no stronger than those produced by low-contrast edges. This suggests that line terminators vs. continuous edges influence expression of disruptive body pattern components via different mechanisms that are controlled by contrast in different ways.

Defensive responses of cuttlefish to different teleost predators.

We evaluated cuttlefish (Sepia officinalis) responses to three teleost predators: bluefish (Pomatomus saltatrix), summer flounder (Paralichthys dentatus), and black seabass (Centropristis striata). We hypothesized that the distinct body shapes, swimming behaviors, and predation tactics exhibited by the three fishes would elicit markedly different antipredator responses by cuttlefish. Over the course of 25 predator-prey behavioral trials, 3 primary and 15 secondary defense behaviors of cuttlefish were shown to predators. In contrast, secondary defenses were not shown during control trials in which predators were absent. With seabass-a benthic, sit-and-pursue predator-cuttlefish used flight and spent more time swimming in the water column than with other predators. With bluefish-an active, pelagic searching predator-cuttlefish remained closely associated with the substrate and relied more on cryptic behaviors. Startle (deimatic) displays were the most frequent secondary defense shown to seabass and bluefish, particularly the Dark eye ring and Deimatic spot displays. We were unable to evaluate secondary defenses by cuttlefish to flounder-a lie-and-wait predator-because flounder did not pursue cuttlefish or make attacks. Nonetheless, cuttlefish used primary defense during flounder trials, alternating between cryptic still and moving behaviors. Overall, our results suggest that cuttlefish may vary their behavior in the presence of different teleost predators: cryptic behaviors may be more important in the presence of active searching predators (e.g., bluefish), while conspicuous movements such as swimming in the water column and startle displays may be more prevalent with relatively sedentary, bottom-associated predators (e.g., seabass).

The use of background matching vs. masquerade for camouflage in cuttlefish Sepia officinalis.

Cuttlefish, Sepia officinalis, commonly use their visually-guided, rapid adaptive camouflage for multiple tactics to avoid detection or recognition by predators. Two common tactics are background matching and resembling an object (masquerade) in the immediate area. This laboratory study investigated whether cuttlefish preferentially camouflage themselves to resemble a three-dimensional (3D) object in the immediate visual field (via the mechanism of masquerade/deceptive resemblance) rather than the 2D benthic substrate surrounding them (via the mechanisms of background matching or disruptive coloration). Cuttlefish were presented with a combination of benthic substrates (natural rocks or artificial checkerboard and grey printouts) and 3D objects (natural rocks or cylinders with artificial checkerboards and grey printouts glued to the outside) with visual features known to elicit each of three camouflage body pattern types (Uniform, Mottle and Disruptive). Animals were tested for a preference to show a body pattern appropriate for the 3D object or the benthic substrate. Cuttlefish responded by masquerading as the 3D object, rather than resembling the benthic substrate, only when presented with a high-contrast object on a substrate of lower contrast. Contrast is, therefore, one important cue in the cuttlefish's preference to resemble 3D objects rather than the benthic substrate.

Extreme aggression in male squid induced by a ß-MSP-like pheromone.

Male-male aggression is widespread in the animal kingdom and subserves many functions related to the acquisition or retention of resources such as shelter, food, and mates. These functions have been studied widely in the context of sexual selection, yet the proximate mechanisms that trigger or strengthen aggression are not well known for many taxa. Various external sensory cues (visual, audio, chemical) acting alone or in combination stimulate the complex behavioral interactions of fighting behaviors. Here we report the discovery of a 10 kDa protein, termed Loligo ß-microseminoprotein (Loligo ß-MSP), that immediately and dramatically changes the behavior of male squid from calm swimming and schooling to extreme fighting, even in the absence of females. Females synthesize Loligo ß-MSP in their reproductive exocrine glands and embed the protein in the outer tunic of egg capsules, which are deposited on the open sea floor. Males are attracted to the eggs visually, but upon touching them and contacting Loligo ß-MSP, they immediately escalate into intense physical fighting with any nearby males. Loligo ß-MSP is a distant member of the chordate ß-microseminoprotein family found in mammalian reproductive secretions, suggesting that this gene family may have taxonomically widespread roles in sexual competition.

Night vision by cuttlefish enables changeable camouflage.

Because visual predation occurs day and night, many predators must have good night vision. Prey therefore exhibit antipredator behaviours in very dim light. In the field, the giant Australian cuttlefish (Sepia apama) assumes camouflaged body patterns at night, each tailored to its immediate environment. However, the question of whether cuttlefish have the perceptual capability to change their camouflage at night (as they do in day) has not been addressed. In this study, we: (1) monitored the camouflage patterns of Sepia officinalis during the transition from daytime to night-time using a natural daylight cycle and (2) tested whether cuttlefish on a particular artificial substrate change their camouflage body patterns when the substrate is changed under dim light (down to starlight, 0.003 lux) in a controlled light field in a dark room setting. We found that cuttlefish camouflage patterns are indeed adaptable at night: animals responded to a change in their visual environment with the appropriate body pattern change. Whether to deceive their prey or predators, cuttlefish use their excellent night vision to perform adaptive camouflage in dim light.

Cuttlefish dynamic camouflage: responses to substrate choice and integration of multiple visual cues.

Prey camouflage is an evolutionary response to predation pressure. Cephalopods have extensive camouflage capabilities and studying them can offer insight into effective camouflage design. Here, we examine whether cuttlefish, Sepia officinalis, show substrate or camouflage pattern preferences. In the first two experiments, cuttlefish were presented with a choice between different artificial substrates or between different natural substrates. First, the ability of cuttlefish to show substrate preference on artificial and natural substrates was established. Next, cuttlefish were offered substrates known to evoke three main camouflage body pattern types these animals show: Uniform or Mottle (function by background matching); or Disruptive. In a third experiment, cuttlefish were presented with conflicting visual cues on their left and right sides to assess their camouflage response. Given a choice between substrates they might encounter in nature, we found no strong substrate preference except when cuttlefish could bury themselves. Additionally, cuttlefish responded to conflicting visual cues with mixed body patterns in both the substrate preference and split substrate experiments. These results suggest that differences in energy costs for different camouflage body patterns may be minor and that pattern mixing and symmetry may play important roles in camouflage.

Mottle camouflage patterns in cuttlefish: quantitative characterization and visual background stimuli that evoke them.

Cuttlefish and other cephalopods achieve dynamic background matching with two general classes of body patterns: uniform (or uniformly stippled) patterns and mottle patterns. Both pattern types have been described chiefly by the size scale and contrast of their skin components. Mottle body patterns in cephalopods have been characterized previously as small-to-moderate-scale light and dark skin patches (i.e. mottles) distributed somewhat evenly across the body surface. Here we move beyond this commonly accepted qualitative description by quantitatively measuring the scale and contrast of mottled skin components and relating these statistics to specific visual background stimuli (psychophysics approach) that evoke this type of background-matching pattern. Cuttlefish were tested on artificial and natural substrates to experimentally determine some primary visual background cues that evoke mottle patterns. Randomly distributed small-scale light and dark objects (or with some repetition of small-scale shapes/sizes) on a lighter substrate with moderate contrast are essential visual cues to elicit mottle camouflage patterns in cuttlefish. Lowering the mean luminance of the substrate without changing its spatial properties can modulate the mottle pattern toward disruptive patterns, which are of larger scale, different shape and higher contrast. Backgrounds throughout nature consist of a continuous range of spatial scales; backgrounds with medium-sized light/dark patches of moderate contrast are those in which cuttlefish Mottle patterns appear to be the most frequently observed.

The scaling effects of substrate texture on camouflage patterning in cuttlefish.

Camouflage is the primary defense in cuttlefish. The rich repertoire of their body patterns can be categorized into three types: uniform, mottle, and disruptive. Several recent studies have characterized spatial features of substrates responsible for eliciting these body patterns on natural and artificial backgrounds. In the present study, we address the role of spatial scales of substrate texture in modulating the expression of camouflage body patterns in cuttlefish, Sepia officinalis. Substrate textures were white noise patterns first filtered into various octave-wide spatial frequency bands and then thresholded to generate binary (black/white) images. Substrate textures differed in spatial frequency but were identical in all other respects; this allowed us to examine the effects of spatial scale on body patterning. We found that as the spatial scale of substrate texture increased, cuttlefish body patterns changed from uniform, to mottle, to disruptive, as predicted from the camouflage mechanism of background matching. For substrates with spatial scales larger than skin patterning components, cuttlefish showed reduced disruptive patterning. These results are consistent with the idea that the body pattern deployed by a cuttlefish attempts to match the energy spectrum of the substrate, and underscore recent reports suggesting that substrate spatial scale is a key determinant of body patterning responses in cuttlefish.

Cuttlefish camouflage: the effects of substrate contrast and size in evoking uniform, mottle or disruptive body patterns.

Cuttlefish are cephalopod molluscs that achieve dynamic camouflage by rapidly extracting visual information from the background and neurally implementing an appropriate skin (or body) pattern. We investigated how cuttlefish body patterning responses are influenced by contrast and spatial scale by varying the contrast and the size of checkerboard backgrounds. We found that: (1) at high contrast levels, cuttlefish body patterning depended on check size; (2) for low contrast levels, body patterning was independent of "check" size; and (3) on the same check size, cuttlefish fine-tuned the contrast and fine structure of their body patterns, in response to small contrast changes in the background. Furthermore, we developed an objective, automated method of assessing cuttlefish camouflage patterns that quantitatively differentiated the three body patterns of uniform/stipple, mottle and disruptive. This study draws attention to the key roles played by background contrast and particle size in determining an effective camouflage pattern.

Disruptive coloration elicited on controlled natural substrates in cuttlefish, Sepia officinalis.

Cephalopods are known for their ability to change camouflage body patterns in response to changes in the visual background. Recent research has used artificial substrates such as checkerboards to investigate some specific visual cues that elicit the various camouflaged patterns in cuttlefish. In this study, we took information from experiments on artificial substrates and assembled a natural rock substrate (fixed with glue) with those features that are thought to elicit disruptive coloration in cuttlefish. The central hypothesis is that light rocks of appropriate size, substrate contrast and edge characteristics will elicit disruptive camouflage patterns in cuttlefish. By adding graded light sand in successively greater quantities to this glued rock substrate, we predicted that disruptive camouflage patterns would be replaced by progressively more uniform patterns as the visual features of rock size, contrast and edges were altered by the addition of sand. By grading the degree of disruptiveness in the animals' body patterns, we found that the results support this prediction, and that there is a strong correlation between fine details of the visual background properties and the resultant body pattern shown by the cuttlefish. Specifically, disruptive coloration was elicited (1) when one or a few light rocks of approximately the size of the animal's White square skin component were in the surrounding substrate (dark rocks alone did not elicit disruptive coloration), (2) there was moderate-to-high contrast between the light rocks and their immediate surrounds, and (3) the rock edges were well defined. Taken together, the present study provides direct evidence of several key visual features that evoke disruptive skin coloration on natural backgrounds.

Experimental evidence that ovary and oviducal gland extracts influence male agonistic behavior in squids.

Recent investigations of sensory and behavioral cues that initiate sexual selection processes in the squid Loligo pealeii have determined that egg capsules deposited on the substrate provide a strong visual and chemotactile stimulus to males, even in the absence of females (1, 2, 3). The visual stimulus of egg capsules attracts males to the eggs, and when the males touch the eggs, they encounter a chemical stimulus that leads to highly aggressive fighting behavior. We have recently demonstrated that egg capsule extracts implanted in artificial egg capsules elicit this aggressive behavior (4). In this communication, we present evidence that the salient chemical factor originates in the ovary and perhaps the oviducal gland of the female reproductive tract.

Contact chemosensory cues in egg bundles elicit male-male agonistic conflicts in the squid Loligo pealeii.

Male Loligo pealeii engage in frequent agonistic bouts to gain access to female mates while aggregated at communal egg beds. Male squids are attracted to eggs in the field and in the laboratory. It was recently demonstrated that visual detection followed by physical contact with egg capsules elicited male-male aggression. We tested specific physical and chemical features of the egg capsules that may cause this strong behavioral reaction. Male squids were presented with either natural or artificial egg stimuli and scored for four selected behaviors (egg touch, egg blowing, forward-lunge grab, and fin-beating), the last two of which are highly aggressive behaviors. First, squids were presented with natural eggs versus eggs sealed in agarose-coated tubes (ESACT), which eliminated both tactile and chemical stimuli. Second, males were presented with natural eggs versus eggs sealed in agarose coated tubes containing C18 Sep-Pak-purified extracts (TCPE) from squid egg capsules, which provided chemical cues from natural eggs without the physical stimulus of the egg capsules. Third, natural eggs versus heat-denatured eggs were tested to determine whether the active factor in natural eggs is heat-labile. Squids responded aggressively when contacting natural eggs and TCPE, whereas squids did not respond after touching ESACT or denatured eggs. These results suggest that aggressive behavior is elicited by a heat-labile factor that is embedded within squid egg capsules. This chemosensory cue appears to be a contact pheromone that stimulates the agonistic interactions that characterize the mating behavior of migratory squids on inshore spawning grounds.