Reflective properties of iridophores and fluorescent eyespots in the loliginid squid Alloteuthis subulata and Loligo vulgaris
1 The Marine Biological Association of the UK, Citadel Hill, Plymouth PL1 2PB, UK and
2 Department of Animal and Plant Sciences, University of Sheffield, Sheffield S10 2TN, UK
*e-mail: l.mathger{at}mba.ac.uk
Accepted March 27, 2001
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Summary |
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In both species, the mantle muscle is almost transparent. Stripes of iridophores run along the length of each side of the mantle, some of which, when viewed at normal incidence in white light, reflect red, others green or blue. When viewed obliquely, the wavebands best reflected move towards the blue/ultraviolet end of the spectrum and their reflections are almost 100% polarised. These are properties of quarter-wavelength stacks of chitin and cytoplasm, predicted in theoretical analyses made by Sir A. F. Huxley and Professor M. F. Land. The reflecting surfaces of the individual iridophores are almost flat and, in a given stripe, these surfaces are within a few degrees of being parallel. Both species of squid have conspicuous, brightly coloured reflectors above their eyes. These eyespots have iridescent layers similar to those found on the mantle but are overlaid by a green fluorescent layer that does not change colour or become polarised as it is viewed more obliquely. In the sea, all reflections from the iridophore stripes will be largely confined to the blue-green parts of the spectrum and all reflections in other wavebands, such as those in the red and near ultraviolet, will be weak. The functions of the iridophores reflecting red at normal incidence must be sought in their reflections of blue-green at oblique angles of incidence. These squid rely for their camouflage mainly on their transparency, and the ventral iridophores and the red, green and blue reflective stripes must be used mainly for signalling. The reflectivities of some of these stripes are relatively low, allowing a large fraction of the incident light to be transmitted into the mantle cavity. Despite their low reflectivities, the stripes are very conspicuous when viewed from some limited directions because they reflect light from directions for which the radiances are much higher than those of the backgrounds against which they are viewed. The reflective patterns seen, for example, by neighbouring squid when schooling depend on the orientation of the squid in the external light field and the position of the squid relative to these neighbours.
Key words: squid, Alloteuthis subulata, Loligo vulgaris, iridophore, reflectivity, signalling, concealment, fluorescence, quarter-wavelength stack.
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Introduction |
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Internal to the layer of chromatophore organs in the skin squid possess light-reflecting cells (iridophores) (Williams, 1909). There has been very little work on these cells. Most of the studies have focused on morphological (Arnold, 1967; Mirow, 1972; Arnold et al., 1974) and physiological (Cooper and Hanlon, 1986; Cooper et al., 1990; Hanlon et al., 1990) aspects of squid iridophores. Here, we show that the squid A. subulata and L. vulgaris possess distinct stripes of iridophores that reflect different wavebands when viewed in white light. Some squid iridophores have plates which have been shown by interference microscopy to have refractive indices (n) close to 1.56, which is the refractive index of chitin, whilst the optical thicknesses were those expected for quarter-wavelength (/4) stacks of alternating chitin and cytoplasm plates (Denton and Land, 1971). As shown by Huxley (Huxley, 1968) and Land (Land, 1972), light reflected from an ideal
/4 multilayer stack, for which the optical thicknesses of the plates and spaces are a quarter of the wavelength best reflected at normal incidence, can be characterised as follows. (i) Effect of oblique incidence: as the angle of the light incident to a multilayer reflector is made more oblique, the waveband of the reflected light moves towards the short (blue/ultraviolet) end of the spectrum; (2) Polarisation: depending on the angle of incidence, the light reflected from a
/4 stack will be plane polarised to a greater or lesser extent. At small angles of incidence, the reflected light is hardly polarised whilst at Brewsters angle (49.5° for chitin and water) the reflected light is completely polarised. Stacks that are not ideal have very similar reflective properties as long as each pair of plates and spaces has an optical thickness of
/2 and the thickness of a plate is not less than
/8 (Denton and Land, 1971).
To reduce visibility and send out visual signals in particular directions, squid exploit the special properties of light in the sea. These properties differ in a number of aspects from those found on land.
The intensity of terrestrial daylight changes little over the waveband 400700nm (i.e. near-ultraviolet to deep red) (Le Grand, 1952). Sea water, however, absorbs and scatters light selectively and, with increasing depth, the wavebands of light transmitted are increasingly confined to wavelengths between 400 and 580nm (Atkins, 1945; Tyler and Smith, 1970; Jerlov, 1976). Measurements of the spectral irradiance of waters close to Plymouth were made by Atkins (Atkins, 1945). More detailed measurements on very similar water masses are given by Tyler and Smith (Tyler and Smith, 1970). Both sets of measurements are shown in Fig.1A. It can be seen that even at depths as shallow as 19m the wavebands are largely confined to the blue-green parts of the spectrum, the cut-off being much sharper on the red side of the spectrum than on the blue side. Like many marine animals, the three species of squid used in our experiments have visual pigments that absorb best in the blue-green. For A. subulata, maximal absorption (max) is at 499nm, and for Loligo forbesi
max=494nm (Morris et al., 1993) (see Fig.1A).
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Being completely transparent is evidently the ideal solution for camouflage, and very many midwater animals are highly transparent (McFall-Ngai, 1990; Johnson and Widder, 1998). There are, however, some parts of the body, such as the eyes and the inksac, that could not function if they were transparent, so other camouflage methods have evolved in addition to transparency. When swimming very close to a dark sea bottom, chromatophores most probably play an important role in matching the background. In the midwaters of the sea, where the light intensity changes dramatically depending on the angle of view, pigments such as those contained by chromatophores will make chromatophores very conspicuous when, for example, the squid is seen from below. Light reflectors may be very efficient for camouflage in midwaters and it is important to recall that they may also enable the possessor to give strong visual signals that may be useful, for example, in schooling.
The squid used in our experiments all appeared very transparent, with the chromatophores mostly in a retracted state. When we observed squid in tanks illuminated by white light, the reflectors could easily be seen. The iridophores appeared in stripes above the eyes and on the mantle, and the colours they reflected covered the full extent of our visible spectrum, i.e. from red to deep blue. When the squid were viewed in far blue/ultraviolet light, the stripes above the eyes were seen to fluoresce brightly in the green parts of the spectrum and the emitted green colour was found to be independent of the angle of view. Here, we only describe the reflective patterns produced by the reflective stripes. The complex interactions between chromatophores and iridophores are still under investigation.
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Materials and methods |
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Spectral reflectivity, iridophore orientation, transparency and fluorescence
Spectral reflectivities of squid iridophores were measured using the apparatus shown in Fig.2A. The reflecting material was placed on a small tilting table (Denton and Nicol, 1965). Illumination was by a narrow beam of white light, which was set to give known angles of incidence to the reflecting structures. Perspex wedges could be attached to the side of the tilting table to obtain an angle of 55° incidence. Using a dissecting microscope, images of the reflections of a group of iridophores were formed on a small aperture leading to a photomultiplier tube (PMT) (Thorn EMI 9924B). A diaphragm was placed on the objective lens of the dissecting microscope, so that only light over a range of angles of 5° could enter the microscope. Interference filters (Balzer), which transmitted only narrow wavebands of light, were placed in the light beam leading to the PMT. Polaroid filters attached to the light sources could be rotated by 90° so as to polarise the incident light with its electric vector in either the perpendicular or parallel planes of incidence. A polaroid attached to the base of the dissecting microscope was positioned in the same plane of polarisation as the polaroid of the light source and so acted as an analyser to the reflections from the specimens. Relative reflectivities of the preparations were found by comparing, for each filter, the output of the PMT to light reflected from a specimen with that of a freshly cleaned block of magnesium carbonate, a surface that is known to reflect nearly 100% of light of all visible wavelengths (Benford, 1947).
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Transparency was measured with a similar arrangement to that shown in Fig.2A. The light source was placed below the Perspex box containing the tilting table, so that the light beam passed through the specimen on the tilting table into the PMT. The ratio of light entering the microscope with and without the specimen in the light beam was taken as the measure of transparency. The animals used had not fed prior to the measurements.
To measure fluorescence from the fluorescent layers of the eyespots, the fluorescent tissue was homogenised and extracted in ethanol. Excitation and emission spectra for these preparations were determined using a Perkin-Elmer 3000 fluorescence spectrometer.
Photographs of iridophores were taken with a Nikon (UFX-II) and a Pentax camera using 400 and 1600 ASA slide and print films (Fujicolor).
Theoretical calculations of spectral reflectivity
The theoretical calculations of spectral reflectivity were made by D. M. Rowe after equations given by Huxley (Huxley, 1968) and Land (Land, 1972). They include spectral reflectivity for both planes of polarisation of an ideal /4 stack consisting of 10 alternating high (n=1.56) and low (n=1.33) refractive index platelets.
Results
Theoretical calculations for spectral reflectivity
We give in Fig.3, as an example of the properties of ideal /4 stacks, the results of theoretical calculations of reflectivity and polarisation for obliquities to 80° incidence of a stack with 10 plates of chitin and spaces of cytoplasm, where
max=660nm. Fig.3A shows the spectral reflectivity for the incident light polarised in the plane perpendicular to the plane of incidence. It can be seen that the reflected wavebands move towards the blue/ultraviolet end of the spectrum with increasing obliquity of the incident light. In Fig.3B, the incident light is polarised in the plane parallel to the plane of incidence. At Brewsters angle (49.5°), all reflected light will be polarised in the plane perpendicular to the plane of incidence.
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Except at the most anterior end, where they are very closely packed, the red stripe iridophores are loosely spaced and their combined area covers approximately half the surface of the stripe. The reflectivities at the anterior end of the mantle are approximately 23 times higher than those found at a position halfway along the length of the mantle. When facing the animals from the front, the reflections from the collar iridophores are very conspicuous. The collar iridophores and the remainder of the red stripe are not always in a reflective state at the same time: they have been observed with the red stripe being non-reflective.
The green stripes
There are two very bright reflecting stripes between the fins on the dorsal side of the mantle of L. vulgaris (Fig.5). These have been described as dorsal iridophore sheen (e.g. Hanlon, 1982; Hanlon et al. 1999). They are not present in A. subulata. These stripes are densely packed with iridophores, which are 100200µm long. The iridophores of this stripe have been observed to extend some way towards the anterior end of the mantle, and small patches of green iridescence can sometimes be observed above the red stripes on the dorsal side. These patches, described as iridophore splotches (e.g. Hanlon, 1982), have spectral reflectivity and polarisation characteristics identical to those of the green stripes. In Fig.5B, it can be seen that the iridophores of this stripe lie approximately parallel to the skin surface, so that their normals make angles of approximately 85° with the horizontal. In white light and at low angles of incidence, these stripes reflect green. At angles between 45 and 55° incidence, the reflections become blue and polarised. Photographs of the reflections are shown in Fig.7D,E. Fig.9 shows their spectral reflectivity at three angles of incidence.
The ventral iridophores
The iridophores making up the ventral side are densely packed, approximately 200300µm long and 50µm wide. Their long axes lie parallel to the antero-posterior axis of the squid mantle. Their flat surfaces make angles of between 55 and 60° with the skin surface within which they lie. They consequently lie within a range of angles with the horizontal from approximately 25° at the side of the mantle to 10° near the most ventral parts of the mantle (Fig.5B).
In white light and when viewed at normal incidence, the ventral iridophores appear red (Fig.7F). The spectral shifts and polarisation patterns of the ventral iridophores are similar to those described for the mantle red stripe (see Fig.8). Because of the orientation of the iridophores, however, the light seen when the ventral iridophores are viewed obliquely comes from the inside of the mantle (Fig.10). In all squid examined, these iridophores were in a reflective state.
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Fluorescent and iridescent layers above the eyes
The fluorescent layers
In both species examined, the fluorescent layers can cover all or part of the other reflecting layers of the eyespots. When viewed in white light, these layers emit yellow-green light (Fig.7H), and the colour of the emitted light does not change with angle of view and it is not polarised. They look equally bright from all angles of view and approximate to being perfect diffusers. In blue light, the fluorescent layers fluoresce in the green parts of the spectrum, at around 500nm (Fig.7H, inset). The wavelengths that excite the fluorescence best were found to be between 400 and 420nm. The measurements using extracted tissue showed no appreciable change in spectral characteristics after extraction. It was found that the emission peak (max) was at 485nm, with maximum excitation at 415nm (Fig.12). These measurements revealed a further excitation peak at around 300nm, but since spectral irradiance in the sea drops off sharply at wavelengths around 350nm (Le Grand et al., 1954), this excitation peak cannot play an important role in the fluorescence from this tissue in the sea.
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Discussion
Transparency, /4 multilayer reflectors and fluorescence
When a squid was observed with chromatophores retracted, it could be seen that most parts of the squid are highly transparent. In the midwater environment, largely transparent animals may be quite effectively camouflaged against predation (for a review, see McFall-Ngai, 1990). It may, however, be more difficult for a transparent squid to send visual signals to neighbouring squid without a compromise to the animals transparency. The reflective stripes of the two species of squid studied have properties that represent a compromise between transparency and signalling. Since the squid disturb the general patterns of light around them very little, even small reflections can give clear signals.
The reflective properties of all the reflective stripes, except those of the fluorescent layers, resemble those of /4 stacks. The bandwidth of the reflections, their spectral shifts with changes in angle of incidence and the polarisation patterns measured for the various iridophore stripes are in good agreement with the theoretical predictions of Huxley (Huxley, 1968) and Land (Land, 1972) (see Fig.4). This agrees with the findings of Denton and Land (Denton and Land, 1971), who showed that the chitin plates from the reflectors above the eyes of L. forbesi have optical thicknesses approximating to a quarter of the wavelength of the light that is best reflected at normal incidence. It has been shown by transmission electron microscopy that the iridophore platelets of some squid are arranged in stacks (Arnold, 1967; Mirow, 1972; Arnold et al., 1974) and that they have optical thicknesses approximating those expected for
/4 reflectors (Cooper et al., 1990). Hanlon et al. (Hanlon et al., 1983) performed experiments in which they shone a beam of white light onto the anterior end of the red stripe and found that, when viewed from various angles, the preparation became coloured. They argued that thin-film devices do not produce a spectrum of colours with a given angle of incident light and that the edges of the iridophores act as diffraction gratings (see also Cloney and Brocco, 1983). We found that the changes in colour with changes in angle of incidence and the polarisation properties agree well with the theory of
/4 stacks. From the measurements using aluminium foil, it is expected that the iridophores of the blue stripes and the ventral side are orientated with their flat surfaces at an angle to the skin surface so that the platelets lie edge on to the skin surface. The results we report here suggest that the colours are produced by constructive interference of light reflected from the platelets rather than by diffraction from the edges of the platelets.
On the eyespots, we found two types of reflector, one with the properties of /4 stacks, the other fluorescent and with the properties of a diffuser. The iridophore stripes underneath the fluorescent layer will act as a tapetum, reflecting green and blue light, whose energy is transferred to the fluorescent layer and emitted as green light. The result of the fluorescence is that a certain number of quanta corresponding with light in a waveband around 415nm are transformed to light in a waveband around 485nm. Although the quanta produced are smaller than those exciting the fluorescent layers, they will be approximately 10 times more effective at bleaching the photosensitive pigment of the squid.
Iridophore reflections in the light environment of the sea
Although some squid iridophores reflect light in the red part of the spectrum, the intensities of these reflections in the sea will be relatively small because of the low intensity of the red light available (see Fig.1A). It is thought that the squid used in our study have only one visual pigment absorbing maximally at around 500nm (Morris et al., 1993). A pigment of this kind will have very low absorption in the red, and the eyes of the squid will be very insensitive to such light. Although, at near-normal incidence, the red stripe iridophores reflect red light, at higher angles of incidence the reflections become those of shorter (green and blue) wavelengths. These are the wavebands that penetrate best into the sea and to which the eyes of these squid are most probably sensitive. Fig.13 shows how large these effects can be. Fig.13A summarises the results on spectral reflectivity (R) obtained for the red stripe. Fig.13B shows the effects of the spectral intensity of daylight in the sea (E
) (in the example given here at a depth of 19m) and the spectral sensitivity of the photosensitive pigment of A. subulata on the reflections seen by these squid (see Ratio 1 below). It may be seen from this figure that, even at depths of a few metres, the reflections from the iridophores will be confined largely to the blue-green parts of the spectrum, and these will be much more effective than other wavebands in stimulating the squids visual pigment. The blue and green stripes reflect best in the blue-green at low angles of incidence, so that the reflections are most effective at these angles. The fluorescent layers emit blue-green light at both normal and oblique angles of incidence.
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At angles around Brewsters angle, the reflections from the iridophore stripes are highly polarised. Both the plane and the degree of polarisation change dramatically depending on the movement of the squid. For schooling squid, this could be a powerful information source, because cephalopods have the ability to discriminate light polarised in different planes (Moody and Parriss, 1960; Saidel et al., 1983; Shashar et al., 1996; Shashar et al., 2000). The polarisation patterns are complicated. Near the surface of the sea, they are influenced by the changing patterns of polarisation in the sky. These are mainly ascribed to scattering of directional light, as a result of which the orientation of the e-vector will depend on the direction of view relative to the bearing of the sun, the suns altitude and the depth of the water (Waterman, 1954; Waterman, 1955; Waterman and Westell, 1956). Generally, the degree of polarisation is between 5 and 35% (Waterman and Westell, 1956; Ivanoff and Waterman, 1958; Ivanoff, 1956). In one exceptional case, Ivanoff (Ivanoff, 1959) found values of polarisation as high as 85%.
Visibility and invisibility in the sea
In Fig.14 we show, as an example, a squid mantle in cross section and an observer looking at a reflective area on the surface of the squid. The visibility of the reflecting area to the observer will depend on a number of factors: (i) the radiance of the light from the direction from which the reflections arise (L1), (ii) the radiance of the background light field against which the area is viewed (L
2), (iii) the spectral reflectivity of the reflector (R
) at an angle of incidence
, (iv) the spectral irradiance of submarine daylight (E
) and (v) the spectral sensitivity of the eye of the observer (S
). The intensities of the reflections with respect to the backgrounds against which they are viewed are given by the ratio
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We assume that, at a given depth, the spectral distribution of energy in the sea is the same in all directions of view (see upwelling and downwelling irradiance measurements of Tyler and Smith, 1970). We use (i) the values for the angular distribution of light given by Denton et al. (Denton et al., 1972), after the equation of Tyler (Tyler, 1963) (Fig.1B) and (ii) the spectral irradiance data from Tyler and Smiths (Tyler and Smith, 1970) measurements in the Gulf of California (19m depth) (Fig.1A). (iii) The absorption curve S of the squids visual pigment (taken as the measure of spectral sensitivity) was calculated after Knowles and Dartnall (Knowles and Dartnall, 1977) with
max=499nm, which is the wavelength at which the rhodopsin of A. subulata absorbs best (Morris et al., 1993) (Fig.1A).
The mantle red stripe
Fig.15A shows diagrammatically how the orientations and the reflective properties of the red stripe iridophores will affect the visibility of the red stripe to three observers (X, Y and Z) at different positions in a plane perpendicular to the long axis of the squid. It may be seen from this figure that a red stripe iridophore reflects light that always arises from a direction in which the radiance is much higher than that of the background. Observer X, for example, located in a plane normal to the surface of the iridophore (=0°), will compare the unpolarised reflections arising from an angle of
1=40° to the downward vertical with the approximately 30 times dimmer background at an angle of
2=140°. The fraction of the light reflected at near-normal incidence (15° incidence) in the blue-green, calculated from the experimental results shown in Fig.8A, is approximately 4%. Assuming that the angular distribution of light in the sea is of the type shown in Fig.1B, we find that these reflections are sufficient to approximately equal the background light. Observer Y (Fig.15A) in a position
=45° to the normal of the iridophore, will compare the heavily polarised reflections arising from an angle of
1=5° to the vertical with the 40 times dimmer background light at an angle of
2=95° with the vertical. At 45° incidence, approximately 20% of the light striking the iridophore in the perpendicular plane of polarisation is reflected. Assuming that the background light is largely unpolarised and that the squid sums both planes of polarisation, we calculate that the reflections are approximately four times brighter than the background. If the squid eye treats the planes of polarisation separately, the reflections in the perpendicular plane of polarisation will have an intensity eight times that of the background, whilst those in the parallel plane of polarisation have an intensity lower than that of the background. Observer Z (Fig.15A) will compare the heavily polarised reflections arising from an angle of
1=85° to the vertical with the background from an angle of
2=175°. This observer is also positioned at
=45° to the normal of the iridophore, and we find that these reflections are enough to approximately equal the background light.
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On the basis of trigonometric relationships, we can calculate that, for observer Y in Fig.15B, the reflections of the polarised light seen within an angle of view of ±30° will appear at least four times brighter than the background. Both reflectivity and degree of polarisation will decrease with an angle of view greater than that, since the angle of incidence to the iridophores becomes larger than 60°, at which the reflections will be maximal in the near ultraviolet. The intensity of ultraviolet relative to blue-green light diminishes with increasing depth (Le Grand et al., 1954), so that the reflections in this part of the spectrum become limited by the weakness of the ultraviolet light present. The same trigonometric calculations show that the reflections seen by observer Z (Fig.15B) will approximately match the background radiance over the entire visual field, thus making the squid hard to detect.
The mantle blue and green stripes and the ventral iridophores
The iridophores of the mantle blue stripe can be seen only when viewed from the side and, for the examples given above, only observers X and Y will see this stripe. This is because these iridophores only reflect light visible to us for a small range of angles around normal incidence. At high angles of incidence, we expect that the reflections will be strongest in the ultraviolet. Again, the iridophores of the blue stripe reflect light arising from a direction in which the light is much brighter than the background and, despite their low reflectivity in the blue-green (15%), the stripes will appear approximately four times brighter than the background.
The green stripes found between the fins of L. vulgaris and the green patches found anterior to the green stripes give very bright blue-green reflections both upwards and sideways. These reflections may be useful signals to other squid. We have observed on a number of occasions how L. vulgaris males show the accentuated testis component (described by, for example, Hanlon et al., 1999), in which the green stripes appear as conspicuous flashes in an area in which the chromatophores are retracted.
Midwater animals are most visible when seen from below against the background of the strongest radiance in the sea. If the light travelling downwards is absorbed by the animal, the loss cannot be replaced by reflections of light from any other direction. Being transparent makes a midwater animal less likely to be detected by predators and in comparison with other muscular animals, such as herring and mackerel, the squid that were the subjects of the present study are very transparent. The loose spacing of the red stripe iridophores, their low reflectivity and high blue-green transmission allow approximately 90% of the incident light to be transmitted through the skin. Despite opacity changes that occur during mantle muscle activity (Abbott and Lowy, 1956), we found that the mantle muscle itself transmits approximately 95% of blue-green light and that at most 30% of the transmitted blue-green light is lost in the internal organs. The iridophores of the ventral side are orientated so that the blue-green light that enters the mantle cavity falls on them obliquely (Fig.16) and so the light is channelled downwards by the ventral iridophores, which reflect light strongly at oblique angles of incidence. At normal incidence, at which reflectivity in the blue-green is low, the iridophores transmit the incident blue-green light very efficiently, so that the total light directed downwards minimises the shadow cast to an observer below the animal.
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The effects of position on the visibility of the reflective stripes
In Fig.17A we illustrate the reflective patterns of all the iridophore stripes observed from the positions of observers X, Y and Z. Observer X will see the fluorescent eyespots, the bright and polarised reflections at the anterior and posterior ends of the red stripe, the unpolarised reflections from the blue stripe and the polarised reflections from the green stripes and green patches. While observer Y will only see the bright and polarised reflections of the red stripe and the unpolarised reflections from the blue stripe, observer Z will see the fluorescent eyespots and the unpolarised reflections from the green stripes and green patches.
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Changes in the orientation of a squid with respect to the light field around it will change the brightness, position and polarisation of the reflective stripes. If, for example, the squid rolls, one of the red stripes will become brighter than the other; if the squid yaws, the stripes will appear longer, or shorter, depending on the position of the observer.
It would be advantageous for the squid to be able to detect the differences in brightness of individual iridophores. The iridophores are generally approximately 200µm in length. In a squid with a length of 30cm, the visual acuity needed to resolve iridophores of 200µm at a distance of 15cm would be 0.2 (i.e. approximately 5'). This is 10 times less than the highest acuity for man (Pirenne, 1948). Even allowing for the fact that the eye of L. vulgaris is approximately half the size of the human eye, we might expect that its acuity would be sufficient to enable the squid to resolve these iridophores.
Activity of iridophores and the effect of chromatophore activity
Some of the iridophore stripes found in L. vulgaris and A. subulata are not in a reflective state at all times. They vary from being non-reflective to weakly or strongly reflective. This has also been described by Hanlon (Hanlon, 1982), who observed that some iridophores of the squid Loligo plei are not visible at all times. It has been reported that the spectral reflectivities of some iridophores of the squid Lolliguncula brevis change in response to topical application of acetylcholine, with reflections becoming those of shorter wavelengths with increasing concentrations of acetylcholine (Cooper and Hanlon, 1986; Cooper et al., 1990; Hanlon et al., 1990). This wavelength shift has not been observed in L. vulgaris and A. subulata. It has, however, been possible to retrieve iridophore reflections, several hours after they had disappeared, by perfusing specimens with a solution of 50µmoll-1 carbachol (Sigma). One of us (L.M.M.) has observed that under stress (e.g. during capture of a squid) the mantle red stripes become very prominent, suggesting that iridophores could be controlled by the animal. The effect that iridophore activity would have on the reflectivities described in this paper would be to increase or decrease the intensity of the reflections with respect to the background radiance.
The iridophore stripes are overlain by hundreds of chromatophores, which can be observed in a variety of retraction states, producing a wide range of body patterns. The interaction between chromatophores and iridophores and their importance for camouflage and visual signalling is outside the scope of the present work and will be the subject for future study. It appears likely, however, that chromatophores can cover the reflections from the iridophore stripes. Their pigments are commonly of longer wavelengths (red, yellow and brown) and will absorb light in the blue-green parts of the spectrum. This will have the effect of making the squid darker if the chromatophores expand, reducing iridophore reflectivity; conversely, their retraction will increase reflectivity from the iridophore stripes.
Concluding remarks
In this paper, we report the existence of distinct iridophore stripes in squid. The stripes are described in terms of their spectral reflectivity, polarisation and orientation. Our measurements show that the transparency of the squid mantle and the ventral iridophores may minimise the visibility of a squid to observers below the animal. The iridophores of the ventral side have high reflectivity in the blue-green at oblique angles of incidence. This will channel the light, which passes through the mantle muscle, downwards, so that the squid minimises the shadow cast below the animal.
The fluorescent layers and the iridophores of the red, green and blue stripes are orientated in such a way that they reflect light, which always arises from directions in which the radiances are higher than those of the background. They consequently disrupt, rather than aid, camouflage and it seems that their function lies in communication between members of the same species, e.g. signalling between neighbours in schools.
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Acknowledgments |
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References |
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