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The medial part of the nucleus of Edinger-Westphal (EWM) in birds mediates light-regulated adaptive increases in choroidal blood flow (ChBF). We sought to characterize the effect of loss of EWM-mediated ChBF regulation on photoreceptor health in pigeons housed in either moderate intensity diurnal or constant light (CL). Photoreceptor abundance following complete EWM destruction was compared to that following a lesion in the pupil control circuit (as a control for spread of EWM lesions to the nearby pupil-controlling lateral EW) or following no EW damage. Birds were housed post-lesion in a 12 h 400 lux light/12 h dark light cycle for up to 16.5 months, or in constant 400 lux light for up to 3 weeks. Paraformaldehyde-glutaraldehyde fixed eyes were embedded in plastic, sectioned, slide-mounted, and stained with toluidine blue/azure II. Blinded analysis of photoreceptor outer segment abundance was performed, with outer segment types distinguished by oil droplet tint and laminar position. Brains were examined histologically to assess lesion accuracy. Disruption of pupil control had no adverse effect on photoreceptor outer segment abundance in either diurnal light or CL, but EWM destruction led to 50-60% loss of blue/violet cone outer segments in both light conditions, and a 42% loss of principal cone outer segments in CL. The findings indicate that adaptive regulation of ChBF by the EWM circuit plays a role in maintaining photoreceptor health and mitigates the harmful effect of light on photoreceptors, especially short wavelength-sensitive cone photoreceptors.

The zebrafish photopic electroretinogram (ERG) sums isolatable elements. In each element, red-, blue-, green-, and UV- (r, g, b, and u) cone signals combine in a way that reflects retinal organization. ERG responses to monochromatic stimuli of different wavelengths and irradiances were recorded on a white rod suppressing background using superfused eyecups. Onset elements were isolated with glutamatergic blockers and response subtractions. CNQX-blocked ionotropic (AMPA/kainate) glutamate receptors; l-AP4 or CPPG-blocked metabotropic (mGluR6) glutamate receptors; TBOA-blocked glutamate transporters; and l-aspartate inactivated all glutamatergic mechanisms. Seven elements emerged: photopic PIII, the l-aspartate-isolated cone response; b1, a CNQX-sensitive early b-wave element of inner retinal origin; PII, a photopic, CNQX-insensitive composite b-wave element from ON bipolar cells; PIIm, an l-AP4/CPPG-sensitive, CNQX-insensitive, metabotropic subelement of PII; PIInm, an l-AP4/CPPG/CNQX-insensitive nonmetabotropic subelement of PII; a1nm, a TBOA-sensitive, CNQX/l-AP4/CPPG-insensitive, nonmetabotropic, postphotoreceptor a-wave element; and a2, a CNQX-sensitive a-wave element linked to OFF bipolar cells. The first five elements were fit with a spectral model that demonstrates independence of cone-color pathways. From this, Vmax and half-saturation values (k) for the contributing r-, g-, b-, and u-cone signals were calculated. Two signal patterns emerged. For PIII or PIInm, the Vmax order was Vr > Vg >> Vb approximately Vu. For b1, PII, and PIIm, the Vmax order was Vr approximately Vb > Vg > Vu. In either pattern, u-cone amplitude (Vu) was smallest, but u-cone sensitivity (ku362) was greatest, some 10-30 times greater than r cone (kr570). The spectra of b1/PII/PIIm elements peaked near b- and u-cone absorbance maxima regardless of criteria, but the spectra of PIII/PIInm elements shifted from b- toward r-cone absorbance maxima as criterion levels increased. The greatest gains in Vmax relative to PIII occurred for the b- and u-cone signals in the b1/PII/PIIm b-wave elements. This suggests a high-gain prolific metabotropic circuitry for b- and u-cone bipolar cells.

Retinal cones are depolarized in darkness, keeping voltage-gated Ca2+ channels open and sustaining exocytosis of synaptic vesicles. Light hyperpolarizes the membrane potential, closing Ca2+ channels and suppressing exocytosis. Here, we quantify the Ca2+ concentration in cone terminals, with Ca2+ indicator dyes. Two-photon ratiometric imaging of fura-2 shows that global Ca2+ averages approximately 360 nM in darkness and falls to approximately 190 nM in bright light. Depolarizing cones from their light to their dark membrane potential reveals hot spots of Ca2+ that co-label with a fluorescent probe for the synaptic ribbon protein ribeye, consistent with tight localization of Ca2+ channels near ribbons. Measurements with a low-affinity Ca2+ indicator show that the local Ca2+ concentration near the ribbon exceeds 4 M in darkness. The high level of Ca2+ near the ribbon combined with previous estimates of the Ca2+ sensitivity of release leads to a predicted dark release rate that is much faster than observed, suggesting that the cone synapse operates in a maintained state of synaptic depression in darkness.

We used suction-pipette recording and fluo-4 fluorescence to study light-induced Ca2+ release from the visible double cones of zebrafish. In Ringer, light produces a slow decrease in fluorescence which can be fitted by the sum of two decaying exponentials with time constants of 0.5 and 3.8 s. In 0Ca2+-0Na+ solution, for which fluxes of Ca2+ across the outer segment plasma membrane are greatly reduced, light produces a slow increase in fluorescence. Both the decrease and increase are delayed after incorporation of the Ca2+ chelator BAPTA, indicating that both are produced by a change in Ca2+. If the Ca2+ pool is first released by bright light in 0Ca2+-0Na+ solution and the cone returned to Ringer, the time course of Ca2+ decline is much faster than in Ringer without previous light exposure. This indicates that the time constants of 0.5 and 3.8 s actually reflect a sum of Na+/Ca2+-K+ exchange and light-induced release of Ca2+. The Ca2+ released by light appears to come from at least two sites, the first comprising 66% of the total pool and half-released by bleaching 4.8% of the pigment. Release of the remaining Ca2+ from the second site requires the bleaching of nearly all of the pigment. If, after release, the cone is maintained in darkness, a substantial fraction of the Ca2+ returns to the release pool even in the absence of pigment regeneration. The light-induced release of Ca2+ can produce a modulation of the dark current as large as 0.75 pA independently of the normal transduction cascade, though the rise time of the current is considerably slower than the normal light response. These experiments show that Ca2+ can be released within the cone outer segment by light intensities within the physiological range of photopic vision. The role this Ca2+ release plays remains unresolved.

Chromaticity-type (C-type) horizontal cells of the turtle retina receive antagonistic inputs from cones of different spectral types, and therefore their response to background illumination is expected to reflect light adaptation of the cones and the interactions between their antagonistic inputs. Our goal was to study the behavior of C-type horizontal cells during background illumination and to evaluate the role of wavelength in background adaptation. The photoresponses of C-type horizontal cells were recorded intracellularly in the everted eyecup preparation of the turtle Mauremys caspica during chromatic background illuminations. The voltage range of operation was either reduced or augmented, depending upon the wavelengths of the background and of the light stimuli, while the sensitivity to light was decreased by any background. The response-intensity curves were shifted to brighter intensities and became steeper as the background lights were made brighter regardless of wavelength. Comparing the effects of cone iso-luminant backgrounds on the Red/Green C-type horizontal cells indicated that background desensitization in these cells could not solely reflect background adaptation of cones but also depend upon response compression/expansion and changes in synaptic transmission. This leads to wavelength dependency of background adaptation in C-type horizontal cells, that is expressed as increased light sensitivity (smaller threshold elevation) and improved suprathreshold contrast detection when the wavelengths of the background and light stimuli were chosen to exert opponent effects on membrane potential.

The morphology and spectral absorption characteristics of the retinal photoreceptors in the southern hemisphere lamprey Geotria australis (Agnatha) were studied using light and electron microscopy and microspectrophotometry. The retinae of both downstream and upstream migrants of Geotria contained two types of cone photoreceptor and one type of rod photoreceptor. Visual pigments contained in the outer segments of these three photoreceptor types had absorbance spectra typical of porphyropsins and with wavelengths of maximum absorbance (downstream/upstream) at 610/616 nm (long-wavelength-sensitive cone, LWS), 515/515 nm (medium-wavelength-sensitive cone, MWS), and 506/500 nm (medium-wavelength-sensitive rod). A "yellow" photostable pigment was present in the myoid region of all three types of photoreceptor in the downstream migrant. The same short-wavelength-absorbing pigment, which prevents photostimulation of the beta band of the visual pigment in the outer segment, was present in the rods and LWS cones of the upstream migrant, but was replaced by a large transparent ellipsosome in the MWS cones. Using microspectrophotometric and anatomical data, the quantal spectral sensitivity of each photoreceptor type was calculated. Our results provide the first evidence of a jawless vertebrate, represented today solely by the lampreys and hagfishes, with two morphologically and physiologically distinct types of cone photoreceptors, in addition to a rod-like photoreceptor containing a colored filter (a cone-like characteristic). In contrast, all other lampreys studied thus far have either (1) one type of cone and one type of rod, or (2) a single type of rod-like photoreceptor. The evolution or retention of a second type of cone in adult Geotria is presumably an adaptation to life in the brightly lit surface waters of the Southern Ocean, where this species lives during the marine phase of its life cycle. The functional significance of the unique visual system of Geotria is discussed in relation to its life cycle and the potential for color vision.

Murine cones contain two opsins in the same cone, one ultraviolet (UV) and the other middle-wavelength sensitive (M). A long-wavelength flash only affecting M-opsin suppresses the cone electroretinogram (ERG) produced by light absorption of UV-cone opsin raising the hypothesis that activation of M-cone opsin suppresses UV-cone opsin responses in the same cone. Here we show that pharmacologic blockade of synaptic transmission in the superfused murine retina, which eliminates interaction from second-order neurons, fails to prevent suppression of the UV-opsin driven pathway by long-wavelength stimuli. This proves that the antagonism must be occurring in the same cone, co-expressing both opsins. Our results show that UV-opsin suppression successively ceases in presence of the M-opsin activating background light, which implies that cone light adaptation is controlled at the opsin stage, before activation of transducin. It also reveals the time course of a transient desensitization of cones due to post-opsin factors in the transduction cascade.

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.

The present study has identified a population of cone photoreceptors in the murine retina that are uniquely immunoreactive for protein kinase C (PKC). Wavelength-sensitive cone subtypes are segregated along the dorso-ventral axis in the mouse retina with ventral retina occupied exclusively by ultraviolet wavelength-sensitive (UVWS) cones, and dorsal retina dominated by middle wavelength-sensitive cones. PKC-positive cones are found primarily in the ventral retina, and double-label immunocytochemistry using a short wavelength-sensitive opsin antibody confirms that they specifically correspond to the UVWS cone subtype. The PKC antibody, as documented in other mammals, also identifies rod bipolar cells in the mouse retina. UVWS cones and bipolar cells have previously been shown to share transcriptional regulatory elements, as observed in transgenic mice encoding a portion of the human SWS-opsin promoter controlling the lacZ reporter gene. In such mice, the transgene product, beta-galactosidase, is expressed in populations of both cones and bipolar cells. The present study confirms that lacZ-expressing photoreceptors are indeed PKC-positive photoreceptors, but that the lacZ-expressing bipolar cells are not the PKC-positive rod bipolar cells. These cells must correspond to a type of cone bipolar cell.

Color appearance judgments revealed fundamental differences in visual processing of incremental and decremental lights. First, the balance of cone activation required for a light to appear achromatic was different for increments and decrements (Judd, 1940; Helson & Michels, 1948). Second, adaptation--the visual system's adjustment to background light--affected achromatic decrements more than increments. Third, the regulation of adaptation for incremental and decremental stimuli depended differently on background signals from the three cone types. We interpret these asymmetries as differences in mechanisms of adaptation in the ON and OFF pathways, and suggest that they evolved to accommodate the range and physical sources of color signals in the two pathways.

Photocurrents of cones in the retinas of a small fish, Danio aequipinnatus (Cyprinidae) were recorded with suction pipette electrodes. Spectral sensitivity was measured between 277 and 697 nm. Four spectral classes of cone were found, with lambdamax at 560, 480, 408, and 358 nm. For the latter, we provide the first complete characterization of spectral sensitivity of a vertebrate ultraviolet (UV) photoreceptor. All cones responded with similar kinetics, except for a subset of the 560-nm cones, which were distinctly faster. The alpha-bands of the three cones absorbing maximally in the visible have the same bandwidth when log sensitivity is plotted versus normalized frequency, and in this respect they are indistinguishable from primate cones ("Mansfield's rule"). An eighth-degree polynomial in lambdamax/lambda based on this combined data set (fish, primate) is presented as a template that is likely to have predictive value in describing cone spectra from other vertebrates. The alpha-band of the UV cone, however, is somewhat narrower than predicted by this function, is similar to other UV visual pigments, and an eighth-degree polynomial that describes its shape is also presented. These measurements also provide information on the beta-band (i.e. cis peak region), difficult to obtain by microspectrophotometry. The beta-band of cone pigments is found at longer wavelengths as the alpha-band shifts toward the red. A secondary rise in cone sensitivity around 280 nm indicates that photons absorbed by aromatic amino acids in the opsin (gamma-band) excite the transduction cascade, but the quantum efficiency is not as high as when absorption occurs in the retinal-protein chromophore.

We have developed a new model of phototransduction that accounts for the dynamics of primate and human cone flash responses in both their linear and saturating range. The model incorporates many of the known elements of the phototransduction cascade in vertebrate photoreceptors. The input stage is a new analytic expression for the activation and inactivation of cGMP-phosphodiesterase (PDE). Although the Lamb and Pugh (1992) model (of a delayed ramp for the rising phase of the PDE response in amphibian rods) provided a good fit for the first 2 log units of stimulus intensity without parameter adjustments, the remaining 4 log units of the data required nonlinear modifications of both delay and gain (slope). We show that this nonlinear behavior is a consequence of the delay approximation and develop a completely linear model to account for the rising phase of amphibian rod photocurrent responses over the full intensity range (approximately 6 log units). We use the same dynamic model to account for primate cone responses by decreasing the time constants of PDE activation and introducing an enhanced inactivation process. This PDE response activates a nonlinear calcium feedback stage that modulates guanylate cyclase synthesis of cyclic GMP. By adjustment of the throughput and feedback parameters, the full model successfully captures most of the features of the primate and human cone flash responses throughout their dynamic range. Our analysis suggests that initial processes in the transduction cascade may be qualitatively different from comparable processes in rods.

About 3% of all mouse photoreceptors are cones. An earlier electrophysiological study indicated that there were two classes of cone in the mouse retina having peak sensitivities (lambda max) of about 360 nm and 511 nm. Recent immunocytochemical results show there are two types of cones that have distinctive regional segregation patterns. We used regional stimulation of the retina in conjunction with electroretinogram (ERG) flicker photometry to see if the two cone types identified electrophysiologically are regionalized in a fashion suggested by the anatomical results. We find they are. Relative sensitivity to ultraviolet and visible light stimulation qualitatively parallels that predicted by immunocytochemical labelling. One result of this remarkable regionalization of cone types is that the mouse retina is relatively more sensitive to ultraviolet light stimulation when that light is directed toward the ventral half of the retina.