Pál Röhlich

Two different visual pigments in one retinal cone cell

The retina of the mouse, rabbit, and guinea pig is divided into a superior area dominated by green-sensitive (M) cones and an inferior area in which cones possess practically only short wavelength-sensitive (S) photopigments. The present study shows that the transitional zone between these retinal areas is populated by cones labeled by both the M and S cone photopigment-specific antibodies COS-1 and OS-2. It is concluded that the overwhelming majority of the transitional cones express both visual pigments. A small population of the transitional cones was strongly labeled exclusively by OS-2 (genuine S cones). The results indicate that, in contrast to the generally accepted idea of one visual pigment per one cone cell, cones of certain mammalian species can express different opsins simultaneously under natural conditions. We speculate that the coexpression may be due to the overlapping of regulatory factors determining the M and S fields.

Two cone types of rat retina detected by anti-visual pigment antibodies

The presence of two distinct cone types was demonstrated in the retina of the rat using two cone-specific monoclonal anti-visual pigment antibodies. Cones labelled by antibody COS-1 constituted the large majority (about 93%) of cones, and are most probably responsible for the green photopic sensitivity of the rat. About 7% of the cones were recognized by antibody OS-2, and are thought to be blue-sensitive elements. While OS-2 positive cones were evenly distributed throughout the retina, there were slight differences in the distribution of COS-1 positive cones. The cones made up about 0.85% of all photoreceptor cells. Although the OS-2 positive cones occur in a very low number (0.05% of all photoreceptors) and probably do not appreciably contribute to the photopic system of the rat, their presence in the rat strengthens the presumption that most mammalian species exhibit a dual cone system with a shortwave and a middle-to-longwave sensitivity.

Distribution of cone photoreceptors in the mammalian retina

The retina of mammals contains various amounts of cone photoreceptors that are relatively evenly distributed and display a radially or horizontally oriented area of peak density. In most mammalian species two spectrally different classes of cone can be distinguished with various histochemical and physiological methods. These cone classes occur in a relatively constant ratio, middle‐to‐longwave sensitive cones being predominant over short‐wave cones. Recent observations do not support the idea that each cone subpopulation is uniformly distributed across the retina. With appropriate type‐specific markers, unexpected patterns of colour cone topography have been revealed in certain species. In the mouse and the rabbit, the “standard” uniform pattern was found to be confined exclusively to the dorsal retina. In a ventral zone of variable width all cones express short‐wave pigment, a phenomenon whose biological significance is not known yet. Dorso‐ventral asymmetries have been described in lower vertebrates, matching the spectral distribution of light reaching the retina from various sectors of the visual field. It is not clear, however, whether the retinal cone fields in mammals carry out a function similar to that of their counterparts in fish and amphibians. Since in a number of mammalian species short‐wave cones are the first to differentiate, and the expression of the short‐wave pigment seems to be the default pathway of cone differentiation, we suggest that the short‐wave sensitive cone fields are rudimentary areas conserving an ancestral stage of the photopigment evolution.

The sensory cilium of retinal rods is analogous to the transitional zone of motile cilia

SummaryThe connecting cilium of rat retinal rods was studied by freeze-fracture and thin-sectioning techniques. Transverse strands of intramembranous particles could be observed on fracture face B of the ciliary plasma membrane. The strands were essentially similar to those found at the transitional zone of motile cilia (“ciliary necklace”). The large number of intramembranous particles obscured the pattern on fracture face A of the membrane.On longitudinal sections of the cilia, beads showing a periodicity similar to the necklace strands were observed. Each bead consisted of two structures apposed to both sides of the plasma membrane. Transverse sections of the cilia revealed radial Y-shaped structures that connected each ciliary doublet with the plasma membrane. Axial tubules, central sheath, radial spokes and dynein arms were missing in the connecting cilium.Comparing the fine structure of the retinal cilia with that of motile cilia it becomes evident that the connecting cilium is analogous in structure with the transitional zone of motile cilia. The present observations suggest that periodic membrane beads along the plasma membrane on thin sections correspond to strands of necklace particles as observed on freeze-fractured membranes. The arrangement of the particles in transverse strands is probably ensured by the radial connecting structures.

Monoclonal antibody-recognizing cone visual pigment

Monoclonal antibodies were raised to a crude photoreceptor-membrane suspension from chicken retinas. Clones producing antibodies against cone outer segments were selected by screening with immunocytochemistry on semithin sections of the retina. One monoclonal antibody, called COS-1, specifically labelled outer segments of double cones and one type of single cones; outer segments of rods and several single cones were not stained. On immunoblots of retinal photoreceptor membranes, this antibody recognized a protein with an apparent molecular mass of 33,000. The visual pigment character of the 33,000 protein was indirectly established by another monoclonal antibody, OS-2, which labelled all outer segments on semithin sections and four bands (33,000-, 36,000-, 38,000-40,000- and a composite band between 66,000-72,000 MW) on immunoblots. Of these, the 36,000- and the 72,000 MW protein bands were identified with an anti-rhodopsin polyclonal antibody as rhodopsin monomer and dimer. Monoclonal antibody OS-2 is assumed therefore to represent an antibody against a common epitope of all visual pigments of the chicken. The monoclonal antibody COS-1 was found to bind to certain cone outer segments of many other vertebrate species as well.

The interphotoreceptor matrix: Electron microscopic and histochemical observations on the vertebrate retina

Comparative fine structural investigations made on frog, chicken, rat, monkey and human retinas confirmed the presence of an amorphous intercellular substance, interphotoreceptor matrix, around the outer and inner segments of the visual cells. A higher concentration of the interphotoreceptor matrix was frequently found on the surface of the outer segments. Tubules in the apical cytoplasm of the pigment epithelial cells were often filled with substance of morphological similar appearance which may indicate the site of formation of the matrix. The acid mucopolysaccharide character of the matrix was demonstrated by light-microscopic histochemical (metachromasia, Alcian blue and Hale reaction) and electron histochemical observations (binding of colloidal iron, PTA staining at acid pH).

Photoreceptor distribution in the retinas of subprimate mammals.

Relevant data on the distribution of color cones are summarized, with special emphasis on the marked dorsoventral asymmetries observed in a number of mammalian species. In addition, an overview is given of studies that demonstrate the coexistence of two visual pigments within the same cone cell. The biological significance of these phenomena is discussed in conjunction with comparative immunocytochemical analyses of subprimate retinas. Based on various cone distribution patterns and temporal and spatial visual pigment coexpression, two models of cone photoreceptor differentiation are suggested.

The photoreceptors and visual pigments of the garter snake (Thamnophis sirtalis): A microspectrophotometric, scanning electron microscopic and immunocytochemical study

Abstract Scanning electron microscopy, immunocytochemistry, and single cell microspectrophotometry were employed to characterize the photoreceptors and visual pigments in the retina of the garter snake, Thamnophis sirtalis. The photoreceptor population was found to be comprised entirely of cones, of which four distinct types were identified. About 45.5% of the photoreceptors are double cones consisting of a large principal member joined near the outer segment with a much smaller accessory member. About 40% of the photoreceptors are large single cones, and about 14.5% are small single cones forming two subtypes. The outer segments of the large single cones and both the principal and accessory members of the doubles contain the same visual pigment, one with peak absorbance near 554 nm. The small single cones contain either a visual pigment with peak absorbance near 482 nm or one with peak absorbance near 360 nm. Two classes of small single cones could be distinguished also by immunocytochemistry and scanning electron microscopy. The small single cones with the 360-nm pigment provide the garter snake with selective sensitivity to light in the near ultraviolet region of the spectrum. This ultraviolet sensitivity might be important in localization of pheromone trails.

Binding sites of photoreceptor-specific antibodies COS-1, OS-2 and AO

The chicken red-sensitive cone visual pigment (iodopsin) and several synthetic peptides of cone and rod visual pigments were used to find the binding sites of our photoreceptor-specific antibodies with immunocytochemistry. The ability of iodopsin to block immunolabeling with monoclonal antibodies COS-1 and OS-2 furnished direct evidence that both antibodies are specific to visual pigments. Immunocytochemistry on whole-mount retinas with and without detergent, as well as electron microscopic labeling of cone photoreceptor membranes revealed the binding sites of COS-1 and OS-2 to be on the cytoplasmic side of the membrane. By testing several synthetic peptides, mainly from the C-terminal region of the cone visual pigments, we found that the domain consisting of the last 6 amino acids of the human red/green-, and the chicken red-sensitive cone pigments completely blocked immunolabeling with COS-1, while the sequence consisting of the last 12 amino acids of the human blue cone pigment was effective to block the binding of OS-2. Both monoclonals can be regarded therefore C-terminal specific antibodies. OS-2 was found to bind to the dark-adapted photopigment more strongly than to the light-adapted one. The binding of the polyclonal rhodopsin antibody AO was almost entirely inhibited by the N-terminal synthetic peptide of bovine rhodopsin indicating that this antibody binds primarily to the N-terminal domain of rhodopsin in a tissue environment.

Cones in the retina of the Mongolian gerbil,Meriones unguiculatus: an immunocytochemical and electrophysiological study

Immunocytochemistry revealed in the retina of the Mongolian gerbil three immunologically distinct photoreceptor cell types. Rods comprising about 87% of the total receptor population were selectively recognized by an antirhodopsin serum (AO). The most abundant cone type (11-13% of photoreceptors) was labeled by the monoclonal antibody COS-1, specific in mammals to the middle-to-long-wave sensitive cone visual pigments. A minor cone population (2.5-5% of the cones) reacted with the monoclonal antibody OS-2, shown earlier to bind to the blue cones in mammalian species. Color substitution experiments revealed on the ERG level a color discrimination capability which must be attributed to the cooperative activity of green-sensitive (COS-1 positive) and blue-sensitive (OS-2 positive) cones. We conclude that the Mongolian gerbil has a well developed cone system, and that it may possess dichromatic green-blue color vision.