Visual phototransduction is a complex of processes that is responsible for the change (phototransformation) of pigments and their subsequent regeneration. This is necessary to transfer information from the outside world to neurons. Due to biochemical processes, under the influence of light with different wavelengths, structural changes occur in the structure of pigments that are located in the bilayer lipid region of the membranes of the outer lobe of the photoreceptor.

Changes in photoreceptors

The photoreceptors of all vertebrates, including humans, can respond to light rays by changing the photopigments that are located in bilayer membranes in the region of the outer lobe of cones and rods.

The visual pigment itself is a protein (opsin), which is a derivative of vitamin A. Beta-carotene itself is found in foods, and is also synthesized in retinal cells (photoreceptor layer). These opsins or chromophores in a bound state are localized in the depths of the bipolar discs in the zone of the outer lobes of the photoreceptors.

About half of the opsins are in the bilayer lipid layer, which is connected externally by short protein loops. Each rhodopsin molecule has seven transmembrane regions that surround the chromophore in the bilayer. The chromophore is located horizontally in the photoreceptor membrane. The outer disk of the membrane region has a large number of visual pigment molecules. After a photon of light has been absorbed, the pigment substance passes from one isoform to another. As a result, the molecule undergoes conformational changes, and the receptor structure is restored. At the same time, metarhodopsin activates the G-protein, which triggers a cascade of biochemical reactions.

Photons of light act on the visual pigment, which leads to the activation of a cascade of reactions: photon - rhodopsin - metarhodopsin - transducin - an enzyme that hydrolyzes cGMP. As a result of this cascade, a closing membrane is formed on the external receptor, which is associated with cGMP and is responsible for the operation of the cation channel.

In the dark, cations (mainly sodium ions) penetrate through open channels, which lead to partial depolarization of the photoreceptor cell. At the same time, this photoreceptor releases a mediator (amino acid glutamate), which acts on the inaptic endings of second-order neurons. With slight light excitation, the rhodopsin molecule isomerizes into the active form. This leads to the closure of the ion transmembrane channel, and, accordingly, stops the cation flow. As a result, the photoreceptor cell hyperpolarizes, and mediators cease to be released in the zone of contact with second-order neurons.

In the dark, sodium (80%), calcium (15%), magnesium, and other cations flow through the transmembrane channels. To remove excess calcium and sodium during darkness, a cation exchanger operates in the photoreceptor cells. It was previously thought that calcium is involved in the photoisomeration of rhodopsin. However, there is now evidence that this ion plays other roles in phototransduction. Due to the presence of a sufficient concentration of calcium, rod photoreceptors become more receptive to light, and the recovery of these cells after illumination is also significantly increased.

Cone photoreceptors are able to adjust to the level of illumination, so the human eye is able to perceive objects in different lighting conditions (from shadows under a tree to objects located on brilliantly lit snow). Rod photoreceptors have less adaptability to light levels (7-9 units and 2 units for cones and rods, respectively).

Photopigments of exteroreceptors of cones and rods of the retina

The photopigments of the cone and rod apparatus of the eye include:

• Iodopsin;
• Rhodopsin;
• Cyanolab.

All these pigments differ from each other in the amino acids that make up the molecule. In this regard, pigments absorb a certain wavelength, more precisely a range of wavelengths.

Cone exteroreceptor photopigments

The cones of the retina contain iodopsin and a variety of iodopsin (cyanolab). Everyone distinguishes three types of iodopsin, which are tuned to a wavelength of 560 nm (red), 530 nm (green) and 420 nm (blue).

On the existence and identification of cyanolalab

Cyanolab is a type of iodopsin. In the retina, blue cones are located regularly in the peripheral zone, green and red cones are localized randomly over the entire surface of the retina. At the same time, the density of distribution of cones with green pigments is greater than that of red ones. Blue cones have the lowest density.

The following facts testify in favor of the theory of trichromacy:

• The spectral sensitivity of two cone pigments was determined using densitometry.
• Using microspectrometry, three pigments of the cone apparatus were determined.
• The genetic code responsible for the synthesis of red, blue and green cones has been identified.
• The scientists were able to isolate the cones and measure their physiological response to irradiation with light of a specific wavelength.

The theory of trochromasia was previously unable to explain the presence of four primary colors (blue, yellow, red, green). It was also difficult to explain why dichromatic people are able to distinguish between white and yellow. Currently, a new retinal photoreceptor has been discovered, in which melanopsin plays the role of the pigment. This discovery put everything in its place and helped answer many questions.

Also in recent studies, sections of the retina of birds were studied using a fluorescent microscope. This revealed four types of cones (purple, green, red and blue). Due to the opponent's color vision, photoreceptors and neurons complement each other.

Rod photopigment rhodopsin

Rhodopsin belongs to the family of G-linked proteins, which is so named because of the mechanism of transmembrane signaling. At the same time, G-proteins located in the near-membrane space are involved in the process. In the study of rhodopsin, the structure of this pigment was established. This discovery is very important for biology and medicine, because rhodopsin is the ancestor of the GPCR family of receptors. In this regard, its structure is used in the study of all other receptors, and also determines the functionality. Rhodopsin is named so because it has a bright red color (from Greek it literally translates as pink vision).

Day and night vision

By studying the absorption spectra of rhodopsin, it can be seen that reduced rhodopsin is responsible for the perception of light in low light conditions. In daylight, this pigment decomposes, and the maximum sensitivity of rhodopsin shifts to the blue spectral region. This phenomenon is called the Purkinje effect.

In bright light, the rod ceases to perceive daylight rays, and the cone takes over this role. In this case, the excitation of photoreceptors occurs in three regions of the spectrum (blue, green, red). These signals are then converted and sent to central structures brain. As a result, a color optical image is formed. It takes about half an hour to completely restore rhodopsin in low light conditions. During all this time, there is an improvement in twilight vision, which reaches a maximum at the end of the pigment recovery period.

Biochemist M.A. Ostrovsky held a series fundamental research and showed that rods containing the pigment rhodopsin are involved in the perception of objects in low light conditions and are responsible for night vision, which is black and white.

Light absorption is required to stimulate photoreceptor cells. Absorption of a photon of light should cause structural changes in the light-absorbing grouping

Rice. 37.21. Micrograph of retinal rod cells obtained in a scanning electron microscope. (Reprinted with the kind permission of Dr. Deric Bownds.)

(chromophore). The photosensitive substance of the rods is rhodopsin, which consists of an opsin protein and a prosthetic group represented by 11-cis-retinal (Fig. 37.24). Rhodopsin is a transmembrane protein with a mass

Rice. 37.22. Schematic representation of the retinal rod.

Its N-terminus is located in the aqueous phase inside the disk, and its C-terminus is located on the other side of the disk membrane, in the cytosol. The N-terminal region of rhodopsin contains two oligosaccharide units covalently attached to an aspartic side chain.

Rice. 37.23. The outer segment of the retinal rod under an electron microscope. Stacked disks are visible. (Allen J. M., ed., Molecular organization and biological function, Harper and Row, 1967.)

Rice. 37.24. Structure of 11-cis-retinal, all-trans-retinal and all-trans-retinol (vitamin A).

These sugars seem to belong important role in the directed movement of rhodopsin from the inner segment to the discs.

Rice. 37.25. Disc formation by invagination of the plasma membrane. The arrows show the polarity of the rhodopsin molecules.

The fact is that rhodopsin, like other eukaryotic membrane proteins, is synthesized on ribosomes attached to the endoplasmic reticulum. The newly synthesized rhodopsin enters the Golgi apparatus and only then reaches the plasma membrane. New discs are formed at the base of the inner segment by invagination of the plasma membrane, which is why the carbohydrate units of rhodopsin are localized inside the disc, although initially, being part of the plasma membrane, they face the extracellular space (Fig. 37.25).

Opsin, like other proteins lacking prosthetic groups, does not absorb visible light. The color of rhodopsin and its sensitivity to light are determined by the presence of 11-cis-retinal, which is a highly effective chromophore. Due to 11-cis-retinal, rhodopsin has a wide absorption band in the visible region of the spectrum with a maximum at which perfectly corresponds to solar radiation. Also noteworthy is the intensity of absorption of visible light by rhodopsin. The extinction coefficient of rhodopsin at is very high, namely (Fig. 37.26). The total power of absorption of visible light by rhodopsin approaches the maximum values ​​for organic compounds. The high chromophore qualities of 11-cis-retinal are due to the fact that it is a polyene. The alternation of six single and double (unsaturated) bonds in it creates a long unsaturated system for electron transfer.

11-cis-retinal is attached to rhodopsin through a Schiff base, which is formed by binding the aldehyde group

Rice. 37.26. Absorption spectrum of rhodopsin.

Rice. 37.27. The primary act upon excitation by light is the isomerization of the 11-cis isomer of the Schiff base formed by retinal. into full-trans form. The double bond between is shown in green.

11-cis-retinal with the e-amino group of a specific lysine residue in opsin. The spectral properties of rhodopsin indicate that the Schiff base is in the protonated form.

The precursor of 11-cis-retinal is all-trans-retinol (vitamin A), which cannot be synthesized de novo in mammals. All-trans retinol (Figure 37.24) is converted to 11-cis-retinal in two steps. First, the alcohol group is oxidized to an aldehyde group in the presence of retinol dehydrogenase and NADP+ as an electron acceptor. Then, under the action of retinal isomerase, the double bond between isomerizes from the trans form to the cis form. Vitamin A deficiency leads to night ("night") blindness and ultimately to damage to the outer segments of the rods.

visual pigments

visual pigments

The structure of rhodopsin

Cones and color vision

color blindness

Properties of photoreceptor channels

Molecular structure of cGMP-gated channels

Signal transduction in photoreceptors

Metabolic cascade of cyclic GMP

Vertebrate receptors that depolarize on exposure to light

Signal amplification in the cGMP cascade

Signals in response to single light quanta

Literature

visual pigments

Visual pigments are concentrated in the membranes of the outer segments. Each stick contains about 10 8 pigment molecules. They are organized into several hundred discrete discs (about 750 in monkey wand) that are not connected to the outer membrane. In cones, the pigment is located in special pigment folds that are a continuation of the outer cell membrane of the photoreceptor. Pigment molecules make up about 80% of all disk proteins. Visual pigments are so densely packed in the membranes of the outer segment that the distance between two visual pigment molecules in a rod does not exceed 10 nm. Such dense packing increases the probability that a photon of light passing through a layer of photoreceptor cells will be captured. The following question arises: how do signals arise when light is absorbed by visual pigments?

Light absorption by visual pigments

The events that occur when light is absorbed by the rod pigment, rhodopsin, were studied using psychophysiological, biochemical, and molecular techniques. The visual pigment molecule consists of two components: a protein, called opsin, and a chromophore, 11-cis-vitamin A-aldehyde, called retinal (Fig. 1). It should be clarified that the chromophore contains a chemical group that gives color to the compound. The quantitative characteristics of the absorption capacity of the pigments were studied using spectrophotometry. When rhodopsin, the visual pigment of the rods, was illuminated with light of different wavelengths, blue-green light with a wavelength of about 500 nm was best absorbed. A similar result was also obtained by illuminating a single rod under a microscope with beams of light with different wavelengths. An interesting relationship has been found between the absorption spectrum of rhodopsin and our perception of twilight light. Quantitative psychophysical studies performed on humans have shown that bluish-green daylight with a wavelength of about 500 nm is optimal for the perception of twilight light in the dark. During the day, when rods are inactive and only cones are used, we are most sensitive to the red color corresponding to the absorption spectrum of cones (we will talk about this later).

When one photon is absorbed by rhodopsin, retinal undergoes photoisomerization and passes from the 11-cis to the trans configuration. This transition occurs very quickly: in about 10-12 seconds. After that, the protein part of the pigment also undergoes a series of transformational changes, with the formation of a number of intermediate products. One of the conformations of the protein moiety, metarhodopsin II, is most important for signal transduction (we will discuss this later in this chapter). Figure 2 shows the sequence of events during decolorization and regeneration of active rhodopsin. Metarhodopsin II is formed after 1 ms. Pigment regeneration after its disintegration occurs slowly, within a few minutes; this requires the transport of retinal from photoreceptors to the pigment epithelium.

The structure of rhodopsin

On the molecular level opsin protein consists of 348 amino acid residues, forming 7 hydrophobic zones, each of which consists of 20-25 amino acids, making up 7 transmembrane helices. The N-terminus of the molecule is located in the extracellular space (i.e., inside the rod disk), and the C-terminus is located in the cytoplasm.

Fig.1. Structure of vertebrate rhodopsin embedded in the photoreceptor membrane. The spiral is somewhat unrolled to show the location of the retinal (indicated in black). C - C-terminus, N - N-terminus.

Fig.2. The efflorescence of rhodopsin in the light. In the dark, 11-cis-retinal is tightly bound to the protein opsin. Photon capture leads to the isomerization of all cis retinal in the retinal throne. In this case, the opsin all-tron-retinal complex quickly turns into metarhodopsin II, which dissociates into opsin and all-tron retinal. The regeneration of rhodopsin depends on the interaction of photoreceptors and pigment cells. Metarhodopsin II activates and maintains the second messenger system.

Retinal is connected to opsin through a lysine residue located in the seventh transmembrane segment. Opsin belongs to a family of proteins with 7 transmembrane domains, which also includes metabotropic mediator receptors, such as adrenergic and muscarinic receptors. Like rhodopsin, these receptors signal to second messengers via G-protein activation. Rhodopsin is remarkably stable in the dark. Bayor calculated that spontaneous thermal isomerization of the rhodopsin molecule takes about 3000 years, or 10 23 more than for photoisomerization.

Cones and color vision

The amazing research and experiments performed by Young and Helmholtz in the 19th century drew attention to the very important issue of color vision, and the scientists themselves gave a clear and accurate explanation of this phenomenon. Their conclusion about the existence of three various types color photoreceptors has stood the test of time and has been subsequently validated at the molecular level. Again, we can quote Helmholtz, who compared the perception of light and sound, color and sound tone. One can envy the clarity, power and beauty of his thought, especially when compared to the confusing vitalistic concepts that were widespread in the 19th century:

All differences in color tones depend on the combination in various proportions of the three primary colors ... red, green and violet ... Just as the perception of sunlight and its warmth depends ... on whether the rays of the sun hit the nerves, coming from the receptors of vision or from the receptors of thermal sensitivity. As Young suggested in his hypothesis, the difference in the perception of different colors simply depends on which of the 3 types of photoreceptors is more activated by this light. When all three types are equally excited, the result is white...

Rice. 3. Sensitivity spectra of human photoreceptors and various visual pigments. (A) Sensitivity spectra of three color visual pigments showing absorption peaks at wavelengths corresponding to cyan, green and red. (B) Sensitivity spectra of cones to blue, green, and red, and rods (shown in black) in macaques. The responses were recorded using suction electrodes, averaged and normalized. The curves of the rod spectrum were obtained in the study of visual pigments in humans. (C) Comparison of monkey and human cone spectra using a color sensitivity test. The continuous curve shows an experiment to determine the sensitivity to color in humans, when presented with light of different wavelengths. The dotted line shows the results predicted based on the registration of currents in individual cones, after correcting for the absorption of light in the lens and pigments on the way to the outer segment. The agreement between the results of both experiments is surprisingly high.

If we project onto White screen two beams of light of different colors at the same time ... we see only one color, more or less different from both of these colors. We can better understand the outstanding fact that we are able to perceive all the shades in the composition of external light by a mixture of three primary colors, if we compare the eye with a dry ... In the case of sound ... we hear longer waves as low tones, and short waves - both high and piercing, besides this, the ear is able to catch a lot at the same time sound waves, i.e. many notes. However, they β in this case do not merge into one complex chord, just as different colors ... merge into one complex color. The eye cannot tell the difference if we change orange to red or yellow; but if we hear the notes do and mi sounding at the same time, such a sound does not seem to us like a note re. If the ear perceived musical tones as the eye perceives colors, each chord could be represented by a combination of three constant notes, one very low, one very high, and one intermediate, producing all possible musical effects only by changing the relative loudness of these three notes. .. However, we are able to see the smooth transition of colors from one to another through an infinite number of shades and gradations ... The way we perceive each of the colors ... depends mainly on the structure of our nervous system. It must be admitted that at present, neither in humans nor in tetrapods has an anatomical basis been described to confirm the theory of color perception.

These accurate and far-sighted predictions have been confirmed by a series of different observations. Using spectrophotometry, Wald, Brouck, McNicol, and Dartnell et al showed the presence of three types of cones with different pigments in the human retina. Baylor and colleagues also managed to divert currents from the cones of monkeys and humans. The three cone populations were found to have different but overlapping ranges of sensitivity to the blue, green, and red portions of the spectrum. The optimal wavelengths for the excitation of electrical signals exactly coincided with the peaks of light absorption by visual pigments, established using spectrophotometry and psychophysical experiments to measure the sensitivity of the eye to the color spectrum. Ultimately, Natais cloned and sequenced the genes encoding the opsin pigment in three types of cones that are sensitive to red, green, and blue.