Early evolution of multifocal optics for well-focused colour vision in vertebrates.

Jawless fishes (Agnatha; lampreys and hagfishes) most closely resemble the earliest stage in vertebrate evolution and lamprey-like animals already existed in the Lower Cambrian [about 540 million years ago (MYA)]. Agnathans are thought to have separated from the main vertebrate lineage at least 500 MYA. Hagfishes have primitive eyes, but the eyes of adult lampreys are well-developed. The southern hemisphere lamprey, Geotria australis, possesses five types of opsin genes, three of which are clearly orthologous to the opsin genes of jawed vertebrates. This suggests that the last common ancestor of all vertebrate lineages possessed a complex colour vision system. In the eyes of many bony fishes and tetrapods, well-focused colour images are created by multifocal crystalline lenses that compensate for longitudinal chromatic aberration. To trace the evolutionary origins of multifocal lenses, we studied the optical properties of the lenses in four species of lamprey (Geotria australis, Mordacia praecox, Lampetra fluviatilis and Petromyzon marinus), with representatives from all three of the extant lamprey families. Multifocal lenses are present in all lampreys studied. This suggests that the ability to create well-focused colour images with multifocal optical systems also evolved very early.

For whales and seals the ocean is not blue: a visual pigment loss in marine mammals.

Most terrestrial mammals have colour vision based on two spectrally different visual pigments located in two types of retinal cone photoreceptors, i.e. they are cone dichromats with long-to-middle-wave-sensitive (commonly green) L-cones and short-wave-sensitive (commonly blue) S-cones. With visual pigment-specific antibodies, we here demonstrate an absence of S-cones in the retinae of all whales and seals studied. The sample includes seven species of toothed whales (Odontoceti) and five species of marine carnivores (eared and earless seals). These marine mammals have only L-cones (cone monochromacy) and hence are essentially colour-blind. For comparison, the study also includes the wolf, ferret and European river otter (Carnivora) as well as the mouflon and pygmy hippopotamus (Artiodactyla), close terrestrial relatives of the seals and whales, respectively. These have a normal complement of S-cones and L-cones. The S-cone loss in marine species from two distant mammalian orders strongly argues for convergent evolution and an adaptive advantage of that trait in the marine visual environment. To us this suggests that the S-cones may have been lost in all whales and seals. However, as the spectral composition of light in clear ocean waters is increasingly blue-shifted with depth, an S-cone loss would seem particularly disadvantageous. We discuss some hypotheses to explain this paradox.

The development of the crystalline lens is sensitive to visual input in the African cichlid fish, Haplochromis burtoni.

We investigated whether the development of the vertebrate crystalline lens is sensitive to visual input. The optical properties of fish lenses were examined as a function of lens size and the optical rearing conditions. Fish (Haplochromis burtoni, Cichlidae) were reared in white light (control group), under spectral deprivation (monochromatic lights), deprivation of the cone system (scotopic illumination), and complete visual deprivation (darkness). Longitudinal spherical aberrations (LSAs) and refractive index profiles of the lenses were measured with thin laser beams. The performance of the lens was modeled by ray-tracing calculations from measured LSAs. In lenses from the control group, LSA and f/R (focal length relative to lens radius) decreased as a function of age. The optical properties of the lenses were modified after rearing in darkness, scotopic illumination, and in monochromatic lights due to changes in the refractive index profile. Rearing in darkness and scotopic illumination reduced the optical quality of the lens. In animals reared under spectral deprivation, the lens did not create well-focused images for all spectral cone types in the same plane, as it does in animals reared in white light. We conclude that visual input seems to play an important role in the development of the lens. The control mechanisms remain unknown.

Rearing in different photic and chromatic environments modifies spectral responses of cone horizontal cells in adult fish retina.

We investigated chromatic processing in the outer retina of the cichlid fish Aequidens pulcher. Intracellular recordings from cone-specific horizontal cells (CHCs) revealed that the two morphologically identified types (H1 and H2) also have different spectral responses. H1-L cells hyperpolarize to all wavelengths ("luminosity"). H2-Cb cells depolarize to long wavelengths and hyperpolarize to short wavelengths ("chromaticity", biphasic). Furthermore, we verified by immunocytochemistry that H2-Cb cells of A. pulcher predominantly contact the middle-wavelength-sensitive (MWS) members of double cones. Developmental plasticity in the cone-CHC networks was induced by rearing fish under conditions of spectral deprivation and different levels of white light. H1-L spectral responses were unaffected by the rearing conditions. Different intensity levels of white light and deprivation of long wavelengths during rearing both induced changes in the spectral responses of H2-Cb. Deprivation of short and middle wavelengths had no effect. Our results indicate that spectral processing in the outer retina of fishes can be modulated in response to different visual experiences and suggest that developmental fine tuning of the color-vision system occurs at early levels of visual processing.

Effects of long-term spectral deprivation on the morphological organization of the outer retina of the blue acara (Aequidens pulcher).

To investigate the developmental plasticity of colour vision, we reared fish with a trichromatic cone system (Aequidens pulcher) under three near-monochromatic lights, differentially stimulating each spectral cone type from the larval stage to the age of at least one year. Control conditions comprised white lights of two intensities. The treatments did not affect the visual pigments, hut led to significant changes in cone outer segment lengths. Furthermore, in the blue-reared group the density of single cones within the retina was reduced by two-thirds after 18 months of exposure, while no changes were observed in the other groups. The connectivity of cone horizontal cells with the single cones was influenced by the intensity and spectral composition of the rearing lights: H1 cells were more sensitive to the spectral component, whereas H2 cells responded to intensity cues. In the blue-light group the dynamics of horizontal cell synaptic spinule formation and degradation were severely compromised. These observations show that long-term spectral deprivation leads to significant morphological changes at the level of photoreceptors and horizontal cells. While the reactions of photoreceptors may be interpreted mostly in terms of compensation, the functional consequences of the changes observed on the horizontal cell level remain to be determined electrophysiologically.

Use of paper selectively absorbing long wavelengths to reduce the impact of educational near work on human refractive development.

BACKGROUND/AIMS: Educational near work has been identified as a major risk factor for the development of juvenile progressive myopia. A study was undertaken to determine whether differences in focal length resulting from longitudinal chromatic aberration of the eye can be exploited to reduce the impact of near work on refractive development. METHODS: Infrared photorefraction was used to determine refractive states in young adult volunteers performing a task similar to reading and writing under various spectral environments. The potential benefits of the observed differences in accommodation demand were studied with a computational model of emmetropisation and myopia progression. RESULTS: The refractive state was largely independent of the colour temperature of the illumination light (white paper) and the colour of commercially available papers (white illumination). Selective elimination of long wavelengths, however, significantly reduced the accommodation stimulus by about 0.5 dioptres. CONCLUSION: Results from model calculations suggest that the use of paper which selectively absorbs long wavelengths may significantly reduce the myopiagenic effects of educational near work.

Morphological changes in the retina of Aequidens pulcher (Cichlidae) after rearing in monochromatic light.

We investigate the processing of chromatic information in the outer retina of a cichlid fish, Aequidens pulcher. The colour opponent response characteristics of some classes of cone-specific horizontal cells in the fish retina are the result of feedforward-feedback loops with cone photoreceptors. To interfere with the reciprocal transmissions of signals, animals were reared in monochromatic lights which preferentially stimulated the spectrally different cone types. Here we report the effects on the cones. Their absorbance spectra were largely unaffected, indicating no change in photopigment gene expression. Significant changes were observed in the cone outer segment lengths and the frequencies of spectral cone types. Quantum catch efficiency and survival of cones appear to be controlled in a spectrally selective way. Our results suggest that the retina responds to spectral deprivation in a compensatory fashion aimed at balancing the input from the different cone types to second order neurons.

Multifocal lenses compensate for chromatic defocus in vertebrate eyes.

The focal length of the vertebrate eye is a function of wavelength, i.e. the eye suffers from longitudinal chromatic aberration. Chromatic defocus is a particularly severe problem in eyes with high light-gathering ability, since depth of field is small due to a pupillary opening that is large in relation to the focal length of the eye. Calculations show that in such eyes only a narrow spectral band of light can be in focus on the retina. For the major part of the visual spectrum, spatial resolution should be limited by the optics of the eye and far lower than the resolving power achievable by the retinal cone photoreceptor mosaic. To solve this problem, fishes with irises unresponsive to light have developed lenses with multiple focal lengths. Well-focused images are created at the wavelengths of maximum absorbance of all spectral cone types. Multifocal lenses also appear to be present in some terrestrial species. In eyes with mobile irises, multifocal lenses are correlated with pupil shapes that allow all zones of the lens, with different refractive powers, to participate in the imaging process, irrespective of the state of pupil constriction.

Effects of retinal dopamine depletion on the growth of the fish eye.

We investigated the suitability of fishes as animal models to study the involvement of the retinal dopaminergic system in the visually guided control of eye growth (emmetropization). Advantages of such a model system are (i) that all dopaminergic cells in the retina can be destroyed without apparent damage to other neurons, (ii) simple optical design and short depth of field of the eye, and (iii) continuous growth throughout life. Depleting the retina of dopamine in Aequidens pulcher (Cichlidae) had no apparent effect on refractive state, since size and focal length of the eye were reduced by the same amount. Furthermore, imposed defocus was compensated at a normal rate in spite of the absence of retinal dopamine. In A. pulcher, the dopaminergic system of the retina trus appears not to have an essential role in emmetropization. Our results furthermore suggest that in eyes of more complicated optical design, manipulation of the retinal dopaminergic system may lead to unrelated effects indistinguishable from direct interference with emmetropization. A major disadvantage of the fish model was that refractive state of the eye could not be measured accurately in vivo with standard methods.

A fluorescent double stain for visualization of neural tissue by confocal laser scanning microscopy.

We present a fast and simple method for a general, fluorescent double stain that differentially labels various cellular components and visualizes all cells in confocal laser scanning microscopy. The technique is useful for two- and three-dimensional visualization of neural tissue and facilitates quantification of a variety of neuroanatomical parameters. Examples from cerebellum and retina are shown to demonstrate the broad applicability.

Connectivity patterns of cone horizontal cells in blue acara (Aequidens pulcher, Cichlidae) reared in different light regimes.

Two types of cone horizontal cells were identified morphologically in the retina of a trichromatic fish by fluorescent labelling with Lucifer Yellow and confocal laser scanning microscopy. H1 cells are located adjacent to the outer plexiform layer, have large somata, small dendritic fields, and contact all cone types. H2 cells are positioned vitread to the H1 cells, have small somata, and large dendritic fields. Their dendrites invaginate the synaptic pedicles of short wavelength sensitive single cones and show a significant preference for one of the spectrally different members of the double cones, presumably the middle wavelength sensitive member. We tested the impacts of different visual environments on the development of these connectivity patterns and found minor changes induced by rearing in white light of different intensities or monochromatic blue light.

Horizontal cell spinule dynamics in fish are affected by rearing in monochromatic light.

Blue acaras (Aequidens pulcher, Cichlidae) were reared for 1 yr in white or monochromatic "red", "green" and "blue" lights to study the function and control mechanisms of horizontal cell (HC) spinules in the synaptic pedicles of cones. Ratios of spinules per synaptic ribbon (S/R) were determined in tangential sections in both single and double cones. We found that the S/R ratios in light adapted retinae decreased with decreasing wavelength of the rearing light in all cone types. Conversely, there was an increasing number of incompletely formed spinules with the highest frequency in the blue light group. Dark adaptation resulted in the complete degradation of mature spinules. However, significant numbers of incompletely degraded spinules were observed in the group reared in blue light. Fish reared in blue light which were transferred to white light formed mature spinules when light adapted and still had vestigial spinules when dark adapted. The mechanisms of spinule formation and degradation and the control of spinule dynamics appear to be fully developed in fish reared in monochromatic light. However, long-term chromatic deprivation seems to induce a compensatory modulation of spinule dynamics. A working hypothesis is formulated that interprets the observed effects as manifestations of differences in the activition of dopaminergic interplexiform cells (light adapted) and the sensitivity to glutamate of HCs (dark adapted). Our findings are consistent with the hypothesis that spinules are involved in sign-inverting feedback transmission from HCs to cones.

The eye of the blue acara (Aequidens pulcher, Cichlidae) grows to compensate for defocus due to chromatic aberration.

By rearing fish in various monochromatic illuminations we investigated (1) the potential for compensation of refractive error due to chromatic aberration, (2) the contributions of the chromatic channels to emmetropization, and (3) the role of color cues in the control of eye growth. Cichlid fish (Aequidens pulcher) were reared for 6 months (12 h light/12 h dark) in monochromatic lights (623.5, 534.1, 485.0 nm; spectral purity 5-10 nm). Light levels were isoirradiant at 1.1.10(12) quanta/s/cm2. Two control groups were reared in white light with down-welling illuminances of 0.2 and 33 lx. Nasotemporal diameters (NTDs) of the eyes were measured in relation to lens size. Due to the oblique axis of highest acuity vision in cichlids, NTD is considered to be a more important dimension than axial length. Variances in NTD were equally small in all rearing groups. NTDs were enlarged with increasing wavelengths of the rearing lights with highly significant values over controls in the red-light group. The wavelength-dependent size of the eyes matched the changes in focal length due to longitudinal chromatic aberration. Complete recovery from eye enlargement was observed after fish reared in red light were exposed to a white light regime for 5 weeks. Small variances in NTD in all groups indicated stringent control of eye growth in the absence of color cues. The reversibility of the increase in NTD in fish reared in red light suggests that the eyes were emmetropized by visually guided mechanisms. Eye size in fish reared in white light was intermediate between the values expected if only blue-sensitive single or the red- and green-sensitive double cones contributed to the control of eye growth. This suggests that all chromatic channels participate in emmetropizing the fish eye.

Refractive index distribution and spherical aberration in the crystalline lens of the African cichlid fish Haplochromis burtoni.

Refractive index distribution in the teleost crystalline lens was measured with a nondestructive method in freshly excised lenses of the African teleost fish Haplochromis burtoni. Independently, spherical aberration was measured in a parallel set of lenses. The measured refractive index profiles show a continual decrease of refractive index from the center to the surface of the lens. The H. burtoni lens is of high optical quality and slightly overcorrected for spherical aberration. Details of the small residual spherical aberration were accurately predicted by ray-tracing model calculations based on the measured refractive index profile. The refractive index profile and the spherical aberration both show more complex characteristics than suggested by earlier measurements and lens models.

Regulation of eye growth in the African cichlid fish Haplochromis burtoni.

Fish were reared in 6 conditions: broad spectrum white light, total darkness, scotopic illumination, and 3 monochromatic colors matched to the absorption spectra of the three cone types to study the influence of the light regime on the regulation of eye growth in the African cichlid fish Haplochromis burtoni. Fish reared in total darkness showed high variability in naso-temporal diameter and axial length of the eye. Animals reared in darkness and in scotopic illumination had significantly larger eyes relative to lens size in comparison to fish reared in white light. Eye size and shape was nearly identical in fish reared in monochromatic and in white light. Because of overlap in the absorption spectra of the three cone types of H. burtoni it could not be resolved whether the regulatory mechanism receives input from all three cone types or only from the green (523 nm) sensitive cones. It is clear from our results, however, that neither the blue (455 nm) nor the yellow sensitive (562 nm) cones alone are responsible for eye size regulation. It seems equally unlikely that all three cone types have to act in concert for normal growth of the eye.

Optics of the harbor porpoise eye in water.

A two-dimensional ray-tracing model for the harbor porpoise eye is constructed from new measurements, mainly on two enucleated eyes, and from data found in the literature. Model calculations show that the crystalline lens has too much refractive power to focus light on the retina. The cornea has a high refractive index and acts as a diverging lens of considerable refractive power. The cornea corrects the eye to near emmetropia for axial and temporal (caudal) directions of view. The eye is approximately 5-D myopic for nasal (frontal) directions of view. The iris serves a dual role as a stop: the iris determines the shapes of bundles of light that enter the lens and the iris blocks light that leaves the lens anterior to its equator.

Methods to estimate dispersion in vertebrate ocular media.

By utilizing existing data on refractive indices of the ocular media I developed methods to convert refractive indices within the visible spectrum. The calculated values have approximately the same accuracy as measurements with standard methods. An independent test confirmed the validity of the formulas.