Living under intense predation pressure has driven cephalopods (octopus, squid, and cuttlefish) to develop an impressive and rapid dynamic camouflaging capability, taking avail of their complex skin structure to defend themselves against predators, approach prey, and communicate (Moynihan, 1985; Packard, 1988; Hanlon and Messenger, 1998; Borrelli et al., 2006; Hanlon et al., 2009; Mäthger et al., 2009).

Cephalopods multilayered and sophisticated skin structure consists of a dense network of pigmented muscle-driven chromatophore cells, which are neurally stimulated to actuate and affect local skin coloring (reviewed in Hanlon and Messenger, 1998). When camouflaging, these visually-oriented mollusks alter their appearance to match their surroundings which, providing researchers a glimpse into the animal's apparent visual perspective (Messenger, 1977; Muntz and Gwyther, 1988; Hanlon and Messenger, 1998; Chiao and Hanlon, 2001b; Barbosa et al., 2007; Hanlon et al., 2009; Josef et al., 2012, 2017; Schwarz, 2015). Theoretically, by manipulating the visual environment it might be possible to characterize and test the background effect and eliciting these various displays.

Initially researchers used static printed backgrounds to elicit and study camouflaging behaviors (Chiao and Hanlon, 2001a,b; Grable et al., 2002; Barbosa et al., 2004; Chiao et al., 2005; Shohet et al., 2006). Background patterns were statically or permanently mounted to the bottom of the experimental tank, offering limited alternatives in any given scenario. Although much was learned about cephalopod's behavior using these important static backgrounds, the rapid and dynamic camouflage capability of these animals called for an agile and more responsive experimental design in which the animal might even passively or interactively affect its own surroundings.

More recently, computational technologies have been utilized in hopes of objectively quantifying and categorizing cephalopods skin patterns (Barbosa et al., 2008; Hanlon et al., 2009; Chiao et al., 2010; Zylinski et al., 2010; Josef et al., 2012, 2017; Orenstein et al., 2016) and other marine organisms behavior under gradient sensory cues (Berdahl et al., 2013). Modulating the visual surroundings while concurrently studying an organism's behavior provides a subjective and interactive modality which opened new avenues for behavioral, physiological and psychological research. This approach also provided a platform for addressing old research questions and asking new ones.

In a previous study, a first evidence that video playback is a valuable tool for studying cephalopod behaviors was demonstrated by Pronk et al. (2010), showing that gloomy octopus (Octopus tetricus) responds to video playback of prey items. Nonetheless, the technique was developed to specifically induce preying behavior and was not design to create an immersive experience, nor elicited a camouflaging response. In a more recent work, Jaffe et al. (2011) identified the need for a dynamic testing system. They designed a similar system, named the Sub-Sea Holodeck (SSH), which provided an immersive environment for studying camouflaging cephalopods. The system described in their important work provided a crucial baseline and preliminary results, confirming the system's applicability to cephalopod research. While the holodeck array is a most impressive piece of engineering, it mandated multiscale design and fabrication.

As a cephalopod behavior scientist I believed a simpler, more cost-effective design would be of great value. Building upon Jaffe et al. (2011) inspiring work, I developed the Cephalopod Experimental Projected Habitat (CEPH) system, which is comprehensively detailed below. While this responsive artificial lighting and projecting apparatus does not simulate ocean light conditions, it does create dynamic visual stimuli of either artificial or natural scenes. Although the system described herein was originally designed to study cephalopods camouflage behavior, its flexibility and non-specificity offers new opportunities to expand such research to other marine organisms. I believe that this apparatus will allow new multidisciplinary and novel questions to be answered both within and outside of the marine behavioral study field.

In the following work, I bring CEPH's novel experimental design as well as some of the preliminary data demonstrating its reliability.

In this section, I describe the system setup and configuration of the experimental lab, also illustrated in a diagram (Figure 1). The lab has been specifically designed to hold a large experimental tank in which I conduct all experiments. The room's walls and ceiling are painted matte black to minimize any possible reflections and eliminate any obvious visual landmarks. A Viewsonic DLP short-throw projector (model PJD5553LWS) is suspended from the ceiling, facing straight down and projects directly onto the bottom. A high-resolution Basler acA1920-50gc GigE camera (capable of 50 fps) is also suspended from the ceiling and is used to capture all activity within the tank. The projector and camera are both connected to a high performance Windows-based desktop machine which is located in the lab. This device runs the customized Matlab-based software described below. Experimental room temperature was maintained at 22 Celsius with relative humidity under 45% at all times, providing adequate conditions and protecting all electronic components from corrosion and failure. Although cephalopods does not require ethical authorization under the Institutional Animal Care and Use Committee (IACUC), the experiments carried out in this study complied with the European legislation for animal experiments and with EU directive 2010/63 on the protection of animals used for scientific purposes (Smith et al., 2013; Fiorito et al., 2014).

In the following section, I describe some of the preliminary results of our system. I believe these early data provide insight into the potential offered by this system.

The CEPH offers versatile, variable, and reliable and dynamically changing underwater visual surroundings. Building and experimenting with the CEPH system, some limitations, and advantages were noticeable. Pros: The light regime, intensity, and the visual input are easily displayed, modified and potentially allow addressing a versatile set of question. Another point is that a light coming from the water surface and reflected from the tank back to the animal's eye resembles the light field in real scenario vs. the Plasma/LCD display alternatives. Moreover, the simple and user-friendly set-up allowed non-qualified staff, students and interns to assist, design, and execute their idea with ease. Furthermore, the entire system may be remotely controlled which allows researchers to design and conduct their experiments from anywhere in the world (with the exception of animal maintenance and handling). Cons: As mentioned, the CEPH room had to be painted matt black, which could be of issue in different laboratories and locations. Also, the GigE camera we used requires a significant amount of light to obtain a clear and high-resolution image, which might be problematic when studying light sensitive organisms. In our case, octopuses had to be continuously and carefully monitored due to the infamous octopus escaping behavior and the open-lid arrangement of the tank.

Given the octopus's response to prey and predators, I believe that the CEPH's validity as an experimental testing environment has been verified. Obviously, responses to projected visual stimuli can only be intuitive; nonetheless, the animals' responses to the presence of prey (Stone crab) and predator (Shark) strongly suggest that the animals truly perceive what is expect them to see. Further, the fact that the octopuses matched the background intensity (Unilateral display) further supports the adequacy of using these visual stimuli in a variety of behavioral experiments including camouflage and communication. By recreating and isolating the effect of visual cues on marine organisms, the CEPH offers unprecedented opportunity to study the effect of artificial and natural light fields at various spatio-temporal resolutions on the behavior, navigation and ecology of visually oriented organisms (e.g., the natural flickering light on the seabed, Video S5). Creatively, by adding measured and known sensory stimuli researchers may even use this system to study sensory cue integration in a controlled environment.

I believe that the CEPH system's simplicity, affordability and the ease of assembly makes it an important tool available for use in multiple future experimental arenas, hopefully expanding our understanding of animals' perception and behavior.

NJ contributed in conceiving the idea, planed and built it. Also responsible of data collection, manuscript writing and media preparation.