The mechanism responsible for the dramatic color changes in underwater creatures such as squid and octopuses has been revealed. Understanding how cephalopods change color could lead to new approaches to making tunable filters and switchable photonic materials that more efficiently encode, transmit and decode information with light. Color in living organisms can be formed in two ways: pigmentation or anatomical structure. Structural colors arise from the physical interaction of light with biological nanostructures. A variety of organisms possess this ability, but the biological mechanisms underlying the process have been poorly understood. Two years ago, an interdisciplinary team from the University of California, Santa Barbara discovered the mechanism by which a neurotransmitter dramatically changes color in the common market squid. That neurotransmitter, acetylcholine, sets in motion a cascade of events that culminate in the addition of phosphate groups to a family of unique proteins called reflectins. This process allows the proteins to condense, driving the animal’s color-changing process. Now, the researchers are delving deeper to uncover the mechanism responsible for the dramatic color changes in squid and octopuses. Their latest research, which appears in PNAS (doi: 10.1073/pnas.1217260110), shows that specialized cells in the skin of squid, called iridocytes, contains pleats or invaginations of the cell membrane extending deep into the body of the cell. This creates layers or lamellae that operate as a tunable Bragg reflector. “We know cephalopods use their tunable iridescence for camouflage so that they can control their transparency or in some cases match the background,” said Daniel E. Morse, Wilcox Professor of Biotechnology in the Department of Molecular, Cellular and Developmental Biology, and director of the Marine Biotechnology Center/Marine Science Institute at UCSB. This shows the diffusion of the neurotransmitter applied to squid skin at upper right, which induces a wave of iridescence traveling to the lower left and progressing from red to blue. Each object in the image is a living cell, 10 µm long; the dark object in the center of each cell is the cell nucleus. Courtesy of UCSB. “They also use it to create confusing patterns that disrupt visual recognition by a predator and to coordinate interactions, especially mating, where they change from one appearance to another,” Morse said. “Some of the cuttlefish, for example, can go from bright red, which means stay away, to zebra-striped, which is an invitation for mating.” Antibodies were created to bind specifically to the reflectin proteins, which revealed that the reflectins are located exclusively inside the lamellae formed by the folds in the cell membrane. The events culminating in the condensation of the reflectins caused the osmotic pressure inside the lamellae to change drastically based on the expulsion of water, which shrinks and dehydrates the lamellae and reduces their thickness and spacing. The movement of water was demonstrated directly using deuterium-labeled heavy water. When the acetylcholine neurotransmitter is washed away and the cell can recover, the lamellae imbibe water, rehydrating and allowing them to swell to their original thickness. This reversible dehydration and rehydration, shrinking and swelling, changes the thickness and spacing, which, in turn, changes the wavelength of the light that’s reflected, thus “tuning” the color change over the entire visible spectrum. “This effect of the condensation on the reflectins simultaneously increases the refractive index inside the lamellae,” Morse said. “Initially, before the proteins are consolidated, the refractive index — you can think of it as the density — inside the lamellae and outside, which is really the outside water environment, is the same. There’s no optical difference, so there’s no reflection. “But when the proteins consolidate, this increases the refractive index so the contrast between the inside and outside suddenly increases, causing the stack of lamellae to become reflective, while at the same time they dehydrate and shrink, which causes color changes. The animal can control the extent to which this happens — it can pick the color — and it’s also reversible. The precision of this tuning by regulating the nanoscale dimensions of the lamellae is amazing.” The investigators also conducted mathematical analysis of the color change, confirming that the changes in refractive index perfectly correspond to the measurements made with live cells. This work was published in the Journal of the Royal Society Interface (doi: 10.1098/rsif.2013.0386). From left, Mary Baum, Amitabh Ghoshal, Daniel G. DeMartini and Daniel E. Morse, University of California, Santa Barbara. Courtesy of George Foulsham. A third paper, in the Journal of Experimental Biology, revealed that female market squid show a set of stripes that can be brightly activated to mimic the appearance of the male, which may function during mating to reduce the number of encounters and aggressive contacts from males. The most significant finding from the research is the discovery of a pair of stripes that switch from being completely transparent to bright white. “This is the first time that switchable white cells based on the reflectin proteins have been discovered,” Morse said. “The facts that these cells are switchable by the neurotransmitter acetylcholine, that they contain some of the same reflectin proteins, and that the reflectins are induced to condense to increase the refractive index and trigger the change in reflectance all suggest that they operate by a molecular mechanism fundamentally related to that controlling the tunable color.” These findings could have several practical applications, Morse said. “In telecommunications we’re moving to more rapid communication carried by light,” he said. “We already use optical cables and photonic switches in some of our telecommunications devices. The question is — and it’s a question at this point — can we learn from these novel biophotonic mechanisms that have evolved over millions of years of natural selection new approaches to making tunable and switchable photonic materials to more efficiently encode, transmit and decode information via light?” The scientists are now working with Raytheon Vision Systems in Goleta to investigate the development of tunable filters and switchable shutters for IR cameras. In the future, they may also investigate possible applications for synthetic camouflage. For more information, visit: www.ucsb.edu