Visual pigments, cone oil droplets, ocular media and predicted spectral sensitivity in the domestic turkey (Meleagris gallopavo)

A microspectrophotometric survey conducted on the retinal photoreceptors of the domestic turkey (Meleagris gallopavo) revealed the presence of five different types of vitamin A1-based visual pigment (rhodopsin) in seven different types of photoreceptor. A single class of rod contained a medium wavelength-sensitive visual pigment (wavelength of maximum absorbance, lambda max, 504 nm). Four different types of single cone contained visual pigment maximally sensitive to wavelengths in either the long (LWS, lambda max 564 nm), medium (MWS, lambda max 505 nm), short (SWS, lambda max 460 nm) or violet (VS, lambda max 420 nm) spectral ranges. The LWS, MWS and SWS single cones contained pigmented oil droplets with cut-off wavelengths (lambda cut) at 514, 490 and 437 nm, respectively. The VS single cone contained a transparent oil droplet which displayed no significant absorbance above 330 nm. A single class of double cone was also identified, both the principal and accessory members of which contained the LWS cone visual pigment. The principal member contained an oil droplet with a lambda cut at 436 nm. No oil droplet was observed in the accessory member. The use of a glycerol-based cell mountant, which reduced wavelength dependent measurement artefacts in the microspectrophotometric measurements, is described. Predictions of cone effective spectral sensitivity, incorporating measurements of the spectral transmission of the ocular media, suggest that turkeys have considerable sensitivity to wavelengths in the ultraviolet-A (UV-A, 315-400 nm) spectral range. This has implications for both the visual ecology of wild birds and the welfare of intensively farmed individuals.     

Visual pigments and oil droplets in the retina of a passerine bird, the canary Serinus canaria: microspectrophotometry and opsin sequences.

The visual receptors of the passeriform bird Serinus canaria, the canary, have been examined microspectrophotometrically and the sequences of the opsins determined. Rods have a maximum absorbance (lambda max) at 506 nm. Four spectral classes of single cone are present: long-wave-sensitive (LWS) containing a photopigment with lambda max at 569 nm, middle-wave-sensitive (MWS) with lambda max at 505 nm, short-wave-sensitive (SWS) with lambda max at 442 nm, and ultraviolet-sensitive (UVS) with lambda max at about 366 nm. Double cones possess the 569-nm pigment in both members. Typical combinations of photopigment and oil droplet occur in most cone classes. An ambiguity exists in the oil droplet of the single LWS cones. In some birds, LWS cones are paired with an R-type droplet, whereas in the majority of canaries the LWS pigment is paired with a droplet similar to the P-type of double cones. Mechanisms of spectral tuning within each opsin class are discussed.     

Visual pigment and oil droplet characteristics of the bobolink (Dolichonyx oryzivorus), a new world migratory bird

The retinal photoreceptors of a New World migratory bird, the bobolink (Dolichonyx oryzivorus) were examined using microspectrophotometry. Based on the absorbance spectra of their visual pigments and oil droplets, retinal photoreceptors include five classes of single cones, one double cone, and one rod. The single cones contain a long-wavelength pigment (lambda(max)=564nm) paired with a red or clear droplet, a medium-wavelength pigment (lambda(max)=505nm) with an orange or yellow droplet, and an ultraviolet pigment (lambda(max)=372nm) with a transparent droplet. No violet- or blue-sensitive single cone was found. The Bobolink double cone is unique: the Accessory member possesses the violet sensitive pigment (lambda(max)=403nm), in contrast to the long-wavelength sensitive pigment present in the double cones of other avian species.     

Ionochromic properties of long-wave-sensitive cones in the goldfish retina: an electrophysiological and microspectrophotometric study

Long-wave-sensitive (LWS) cone visual pigments are sensitive to the concentration of chloride ions and show a spectral shift to shorter wavelengths when exposed to low chloride levels. We have used the aspartate-isolated mass receptor potential of the electroretinogram (ERG) to establish whether the spectrally shifted cone pigment is functional. In the goldfish, Carassius auratus, the lambda(max) of the LWS porphyropsin is displaced from about 622 nm to around 606 nm when chloride is replaced by gluconate. The electrical response of the LWS cones (but not MWS cones and rods) is selectively and reversibly abolished by the removal of chloride ions. The effect is concentration dependent and a decrease to half the normal chloride ion concentration is sufficient to cause a decrease in the response.     

Four photoreceptor types in the ground squirrel retina as evidenced by immunocytochemistry

Previous morphological and electrophysiological data show that the ground squirrel retina consists of one rod and two different cone systems. In the present study, immunological evidence is presented that four different photoreceptor cell types are located in the retina of the European ground squirrel (Citellus citellus). In addition to the green and blue cones two rod-like photoreceptor cell types (types 1 and 2) can be distinguished using color specific anti-visual pigment antibodies.     

Visual pigment composition in zebrafish: Evidence for a rhodopsin-porphyropsin interchange system

Numerous reports have concluded that zebrafish (Danio rerio) possesses A1-based visual pigments in their rod and cone photoreceptors. In the present study, we investigated the possibility that zebrafish have a paired visual pigment system. We measured the spectral absorption characteristics of photoreceptors from zebrafish maintained in different temperature regimes and those treated with exogenous thyroid hormone using CCD-based microspectrophotometry. Rods from fish housed at 15 degrees C and 28 degrees C were not significantly different, having lambda max values of 503 +/- 5 nm (n = 106) and 504 +/- 6 nm (n = 88), respectively. Thyroid hormone treatment (held at 28 degrees C), however, significantly shifted the lambda max of rods from 503 +/- 5 nm (n = 194) to 527 +/- 8 nm (n = 212). Cone photoreceptors in fish housed at 28 degrees C (without thyroid hormone treatment) had lambda max values of 361 +/- 3 nm (n = 2) for ultraviolet-, 411 +/- 5 nm (n = 18) for short-, 482 +/- 6 nm (n = 9) for medium-, and 565 +/- 10 nm (n = 14) for long-wavelength sensitive cones. Thyroid hormone treatment of fish held at 28 degrees C significantly shifted the lambda max of long-wavelength sensitive cones to 613 +/- 11 nm (n = 20), substantially beyond that of the lambda max of the longest possible A1-based visual pigment (approximately 580 nm). Thyroid hormone treatment produced smaller shifts of lambda max in other cone types and increased the half-band width. All shifts in photoreceptor lambda max values resulting from thyroid hormone treatment matched predictions for an A1- to A2-based visual pigment system. We therefore conclude that zebrafish possess a rhodopsin-porphyropsin interchange system that functions to spectrally tune rod and cone photoreceptors. We believe that these observations should be carefully considered during analysis of zebrafish spectral sensitivity.     

Rod and cone inputs to bipolar and horizontal cells of the Xenopus retina

This report summarizes some recent studies of the Xenopus retina in which intracellular recordings were made from photoreceptors, horizontal and bipolar cells. The studied cells were identified by injection of Lucifer yellow. Rod spectral sensitivity functions conformed to the density spectrum of a 524 nm pigment, those of cones to that of a 612 nm pigment. Horizontal cell responses reflected both these classes of photoreceptor input. Rod input evoked a slow waveform, with Vmax less than or equal to 18 mV, cone input a faster waveform with Vmax = 30-40 mV. In the mesopic state the horizontal response reflected both waveforms. Rod and cone inputs to the horizontal cells appeared not to act independently, in that a steady weak green background greatly enhanced the response to a superimposed red flash, but not the reverse. A third photoreceptor type (blue-sensitive rod, Y lambda max = 445 nm) provided input to a chromatic bipolar cell which was hyperpolarized by blue light and depolarized by red light. Such chromatic bipolars had broad areas of spatial integration and lacked center-surround organization.     

The effect of background luminance on cone sensitivity functions

Implementations of the Wald-Marré technique have employed fixed luminance backgrounds to isolate cone sensitivity mechanisms. We evaluated the effect of varying the adaptation level on the relative isolation of the different cone types. For MWS and LWS cone isolation we used a 15 Hz flickering test light to isolate the achromatic channel, and we modelled the resulting spectral sensitivity functions as a linear sum of LWS and MWS input. We found only mild improvement in relative cone isolation with increasing adaptation level. The LWS and MWS cone mechanisms showed decreasing sensitivity with adaptation level and reached limiting Weber behavior above 1000 Td. SWS cones were isolated with a 2.4 Hz flickering light. SWS cone isolation improved with adapting level, reaching a plateau above 1000 Td. The SWS cone mechanism showed decreasing sensitivity with adaptation level but did not reach a limiting Weber region. Our data indicate that the use of fixed high adaptation levels has different effects on the cone mechanisms. Absolute sensitivity loss for LWS or MWS mechanism will not be revealed. LWS and MWS thresholds will appear normal unless there is an adaptation abnormality. On the other hand, the SWS cone thresholds would be sensitive to both absolute and increment sensitivity loss. More than one adaptation condition is needed to separate different types of sensitivity loss characteristic of eye disease.     

Contribution of the retinal ON channels to scotopic and photopic spectral sensitivity

Visual information encoded by the middle-wavelength-sensitive (MWS) and long-wavelength-sensitive (LWS) cones in the primate retina are processed by both depolarizing (ON) and hyperpolarizing (OFF) bipolar cells. In contrast, signals from the short-wavelength-sensitive (SWS) cones and dark-adapted rod photoreceptors are thought to be carried almost exclusively by ON bipolar cells (Gouras & Evers, 1985). Consequently, it would be expected that functional inactivation of the retinal ON channels at the bipolar cell level would produce selective deficits in visual functions mediated by rods and SWS cones. We have examined this hypothesis by injecting rhesus monkeys with 2-amino-4-phosphonobutyric acid (APB), a pharmacological agent that reduces the responsiveness of retinal ON neurons, and psychophysically measuring the changes in spectral sensitivities. Under adaptation conditions that isolated rod function, APB caused, as expected, a substantial loss in rod-mediated spectral sensitivity. However, under photopic conditions, cone-mediated spectral sensitivity, including that associated with the SWS cones, was relatively unaffected. These results demonstrate distinct organizational differences between the rod and cone systems; specifically, they indicate that the rod system is more dependent upon retinal ON channels than the cone system. Our failure to find a selective visual deficit related to SWS cone function under photopic viewing conditions suggests that the OFF system can mediate stimulus detection throughout the visible spectrum and that the ability of the OFF system to process signals from the SWS cones has been underestimated.     

Cone visual pigments of the Australian marsupials, the stripe-faced and fat-tailed dunnarts: sequence and inferred spectral properties

Studies of color vision in marsupial mammals have been very limited. Two photoreceptor genes have been characterized from the tammar wallaby, but a third cone pigment was suggested by microspectrophotometric measurements on cone photoreceptors in two other species, including the fat-tailed dunnart, Sminthopsis crassicaudata. To determine the sequence and infer absorption maxima of the cone photoreceptor pigments of S. crassicaudata and the related stripe-faced dunnart (Sminthopsis macroura), we have used evolutionarily conserved sequences of the cone pigments of other species, including the tammar wallaby, to design primers to amplify the S. macroura and S. crassicaudata pigment sequences by the polymerase chain reaction (PCR) using genomic DNA or retinal cDNA as a template. These primers will be useful for amplifying cone opsin coding sequences from a variety of vertebrates. Amplified products were directly sequenced to determine gene structure and coding sequences. The inferred amino acid sequences of the cone visual pigments indicated that both species have middle-wave-sensitive (MWS) pigments with a predicted absorption maximum (lambda(max)) at 530 nm, and ultraviolet-sensitive (UVS) pigments with a predicted lambda(max) at 360 nm. The MWS pigments of the two species differ by two, and UVS by three amino acid positions. No evidence was obtained for a third cone pigment in either species.     

Localization of iodopsin in the chick retina during in vivo and in vitro cone differentiation

Using highly specific antibodies against a chick red-sensitive cone pigment, iodopsin, we investigated the localization of iodopsin in the developing and mature chick retina. The chick retina contains several different photoreceptor types, including a rod, a double cone with a principal and accessory cone, and four different types of single cones. Immunocytochemical observations revealed that outer segments (OS) of one of the single cones (type 1) and both cells of the double cone were strongly immunoreactive to anti-iodopsin antibodies. The Golgi regions and small vesicular structures in the inner segments (IS) of these cells also were intensely stained, indicating a continuous synthesis of iodopsin and its addition to the newly formed cone OS. In the differentiating cones of the developing but immature chick retina, iodopsin immunoreactivity was found at the plasma membranes of both the IS and the terminals (pedicles). This suggests that unidirectional transport of iodopsin to the outer segment may be established during cone differentiation. Immunostaining in the outer plexiform layer (OPL) produced two bands, suggesting that the pedicles of the double cones and type 1 single cones terminate at different positions in this layer. Application of the antibodies to a cell culture system of the chick retina revealed that cells immunoreactive to anti-iodopsin differ slightly in morphology from those reactive with anti-rhodopsin. Since antibodies to iodopsin and rhodopsin stained different types of photoreceptors in the intact chick retina, it will be possible to analyze cell lineage of rods and cones in vitro by use of these antibodies.     

Mapping absorbance spectra,cone fractions,and neuronal mechanisms to photopic spectral sensitivity in the zebrafish

The four spectral cone types in the zebrafish retina each contribute to photopic visual sensitivity as measured by the b-wave of the electroretinogram (ERG). The goal of the current study was to evaluate a model of photopic b-wave spectral sensitivity in the zebrafish that mapped first-order cellular and biophysical aspects of cone photoreceptors (visual pigment absorbance spectra and cone fractions) onto a second-order physiological aspect of cone-derived neural activity in the retina. Good correspondence between the model and photopic ERG data was attained using new visual pigment absorbance data for zebrafish cones (lambda(max) of the L, M, and S cones were 564, 473, and 407 nm, respectively), visual pigment templates, and linearly gained cone fractions. The model inferred four distinct cone processing channels that contribute to the photopic b-wave, two of which are antagonistic combinations of cone-derived signals (L-M and M-S), and two of which are noncombinatorial signals from S and U cones. The nature of the gains and the processing channels suggested general rules of cone-specific inputs to second-order neurons. The model further suggested that the zebrafish retina utilizes neuronal mechanisms for enhancing sensitivity to luminance contrast at short wavelengths and chromatic contrast at middle and long wavelengths. The results indicated that first-order cellular and biophysical aspects of cone photoreceptors can successfully explain physiological aspects of cone-derived neuronal activity in the zebrafish retina.     

Idiopathic retinal degeneration in the dog: differential patterns of [3H]uridine incorporation and HIOMT-like immunoreactivity in surviving photoreceptors

Photoreceptor cell pathology was investigated in an 8-yr-old mixed-breed dog which had displayed visual symptoms of 1 month duration. An electroretinogram detected no light-evoked responses. Light and electron microscopic features showed marked thinning and atrophy of the outer both the tapetal and non-tapetal retina appeared to be involved. In the non-tapetal region, a majority of the rod inner segments were missing, while scattered mitochondria-filled stubby inner segments of cones were readily identified. Inner segments of both rods and cones were observed in the tapetal region. Photoreceptor outer segments were completely absent from the affected retina, and no outer segment debris was observed between the photoreceptor layer and the retinal pigmented epithelium (RPE). Autoradiographic analysis of 3-mm retinal disks from the degenerate retina following incubation with [3H]uridine indicated that only 61% +/- 13 S.D. of the remaining nuclei of rod photoreceptors were undergoing RNA synthesis, whereas more than 99% of cone nuclei incorporated the label. Normal and degenerate retina were also analysed for localization of hydroxyindole-O-methyltransferase (HIOMT)-like immunoreactivity. While the normal retina showed immunoreactivity in both rod and cone photoreceptors with more intense immunoreactivity present in cones, the degenerate retina showed HIOMT-like immunoreactivity only in the remaining cone photoreceptors. The results of this study of idiopathic photoreceptor degeneration of the canine retina suggest that although both photoreceptor types are involved, rods are more severely affected than cones.     

Green-sensitive cone photoreceptors are selectively labeled by Procion yellow dye in goldfish retina

Selective labeling of intravitreal Procion yellow dye by presumed blue-sensitive cone photoreceptors has been demonstrated in primate retina. To determine whether Procion yellow is selective for this cone type in an unrelated vertebrate species, labeling by this dye was studied in goldfish retina, where cone pigment type can be directly inferred from photoreceptor morphology. At low vitreal concentrations of the dye (less than 0.4%), only cone outer segments were labeled. At vitreal concentrations of 0.4-0.5%, the inner segments of short-double cones and a subset of long single cones (presumed green-sensitive cones) were selectively stained. At still higher vitreal concentrations (0.6-0.7%), the inner segments of short-single cones and miniature short-single cones (presumed blue-sensitive cones) showed evidence of Procion label, but were not as heavily labeled. The inner segments of long-double cones and a subset of long-single cones (presumed red-sensitive cones) did not label at any of these concentrations. These results show that Procion yellow is not a selective marker for blue-sensitive cones in the goldfish retina. In addition, stained rod and cone nuclei were observed at each dye concentration, including those concentrations at which no inner segments were labeled.     

Molecular cloning and characterization of five opsin genes from the marine flatfish Atlantic halibut (Hippoglossus hippoglossus)

Most molecular studies on the visual system in fish have been performed on freshwater teleosts such as goldfish and zebrafish where cones and rods appear simultaneously. Many marine fishes have long larval phase in the upper pelagic zone before transformation into a juvenile and a benthic life style. The retina at the larval stages consists of only single cone cells; later during metamorphosis double cones and rods develop. The flatfish Atlantic halibut (Hippoglossus hippoglossus) is a typical example of a marine species with such a two-step retina development. In this study, we have cloned five different opsins from Atlantic halibut larvae and juvenile retinas. Sequence comparisons with other opsins and phylogenetic analysis show that the five genes belong to the opsins of long-wavelength sensitive (L); middle-wavelength sensitive, M(Cone) and M(Rod); and short-wavelength sensitive, S(Blue) and S(Ultraviolet), respectively. In situ hybridization analysis reveals expression in double cone (L and M(Cone)), single cone (S(Blue) and S(Ultraviolet)), and rod (M(Rod)) types of photoreceptor cells in juvenile halibut retina. The visual system in Atlantic halibut seems therefore to have all four types of cone photoreceptors in addition to rod photoreceptors. This work shows for the first time molecular isolation of a complete set of retinal visual pigment genes from a marine teleost and describes the first cloning of an ultraviolet-sensitive opsin type from a marine teleost.     

LWS cone effects on rod threshold and saturation in achromats with residual cone function

Rod saturation on flashed and steady red backgrounds was investigated in normals and three achromats, two of whom were found to have some residual cone function. LWS cones selectively reduce the background level at which rod saturation occurs and elevate rod thresholds at flashed background levels well below saturation. Both of these LWS cone actions are also present in eyes with greatly reduced LWS cone function. In normal eyes LWS cones also elevate rod thresholds on steady backgrounds. We thus conclude that LWS cones influence rods through different mechanisms under transient (flashed) and steady-state background stimulation and that the increase in rod visual sensitivity observed during prolonged presentation of a background is due to a time-dependent reduction of LWS cone influence on rods. Finally, the finding that rod-cone interactions of the same magnitude found in normals can be seen in individuals where the cones' ability to mediate vision is severely reduced suggests the rod saturation paradigm as a sensitive technique for revealing residual LWS cone function.     

Structure-function analysis of rods and cones in juvenile,adult, and aged C57bl/6 and Balb/c mice

To determine whether the photoreceptors change structurally and functionally during aging, and to analyze whether pigmentation in the retinal pigment epithelium might be a contributing factor. Young, adult, and aged C57BL/6 and Balb/c mice (1, 4, and 17 months of age) were housed under a 12-h light/12-h dark cycle, with an ambient light intensity at the eye level of the mice of 85 +/- 18 lux. Scotopic single-flash and photopic-flicker electroretinograms (ERGs) after complete dark adaptation were used to assess rod and cone function, respectively. Numbers of rod photoreceptors were counted in plastic sections, and rhodopsin levels were measured using absorption difference spectrophotometry. Numbers and types of cones were determined using lectin staining in retinal flatmounts and cone-specific antibodies in radial frozen sections. Young pigmented C57BL/6 and nonpigmented Balb/c mice had similar numbers of rods. In both mouse strains, there was an overall decline in rod photoreceptor number during aging, which was more pronounced in albino mice. Rod cell numbers correlated with a drop in the overall amount of rhodopsin and a reduction in the maximum a-wave of the rod ERG. The number of short-wavelength cones was unaffected by age and pigmentation, whereas an age-related decline was observed in mid-wavelength (MWL) cones in albino, but not in pigmented mice. In contrast, MWL cone function was reduced during aging in both strains. Flicker-fusion frequency was determined to be approximately 10 Hz lower in albino animals, which is due to prolonged b-waves in these ERGs. Age-related changes were found in both photoreceptor systems, rods and cones, and in both pigmented and nonpigmented mice. However, rod photoreceptors appear to be more susceptible to both aging and the lack of pigmentation, when compared to cones. These results may help as we begin to understand certain age-related retinal diseases.     

Spectral sensitivity of single cones in rainbow trout (Oncorhynchus mykiss): A whole-cell voltage clamp study

The UVS cone mechanism is known to light adapt at low intensities in comparison to other cones. We were interested in whether this property was related to higher sensitivity in UVS cones or to network adjustments in sensitivity. We investigated spectral sensitivity of 107 individual cone photoreceptors in rainbow trout (Oncorhynchus mykiss) using a whole-cell voltage clamp technique. Mean time-to-peak response was 339 ± 90 ms and flash sensitivity for a 100 ms flash was 4.37 × 10⁻³ ± 2.50 × 10⁻³ pA photons⁻¹ μm², with no significant differences between the UVS, SWS, MWS and LWS cone classes. The spectral sensitivity of the UVS, SWS and LWS cones conformed to the expression of SWS1, SWS2 and LWS opsin genes. The spectral sensitivity of MWS cones, however, showed clear evidence of co-expression of RH2a and RH2b opsin pigments. The fish used in this study bridged the ontogenetic stage where the MWS cones shift their expression from RH2b to RH2a.     

Cone photoreceptors in laminated retinal transplants.

PURPOSE: To investigate the contents of green- and blue-sensitive cone photoreceptors in laminated rabbit retinal transplants. METHODS: Eleven rabbits each received a sheet of embryonic neuroretina into the subretinal space in one eye. Vitrectomy was used in the procedure and properly polarized flat transplants were placed on the host pigment epithelium. After 17-309 days the transplants were examined immunohistochemically with specific antibodies against COS-1 (green-sensitive cones) and OS-2 (blue-sensitive cones). RESULTS: All grafts displayed normal lamination with well developed photoreceptor outer segments apposed to the host retinal pigment epithelium. Occasionally, rosettes were found at the transplant edges. Both COS-1 positive and OS-2 positive cones were detected. In the laminated part of the grafts, COS-1 positive cones were more numerous than OS-2 positive ones. In the rosetted parts of the transplants the relationship between the cones was reversed. CONCLUSION: Full-thickness embryonic rabbit retinal transplants develop into laminated retinas with well-developed photoreceptor outer segment. Both green- and blue-sensitive cone photoreceptors are present and the ratio between the two cone types is the same as in the normal adult rabbit retina.