If we value the pursuit of knowledge, we must be free to follow wherever that search may lead us. The free mind is not a barking
dog, to be tethered on a ten-foot chain.
--Adlai E. Stevenson Jr.
You can know the name of a bird in all the languages of the world, but when you're finished, you'll know absolutely nothing
whatever about the bird... So let's look at the bird and see what it's doing -- that's what counts. I learned very early the
difference between knowing the name of something and knowing something.
--Richard Feynman
The first active chemical components of color vision were discovered in 1932, when George Wald , studying in the laboratory of Otto Warburg
in Berlin-Dahlen, found the light sensitive component of frogs' retinas. He found that the protein yielded both the protein opsin and a
compound, "retinene" (retinaldehyde) when exposed to light. Wald then moved to Karrer's laboratory in Zurich, Switzerland and extracted
enough material for Karrer to confirm that it was an analog of vitamin A (retinol). Despite being related to night blindness, it was a promising
discovery that the vitamin's transform was involved directly in the physiological process of vision. 1
Thus, the light sensitive visual pigment consist of two pieces, a chromophore, rhodopsin, imbedded in a larger protein. It took years of
work by many laboratories to develop a working model of rhodopsin (Fig. 1) as a bundle of transmembrane helices that encompass a binding
pocket for the light-sensitive 11-cis-retinal. When light strikes a molecule of rhodopsin
(protein containing 348 amino acids organized as seven helices across the disk membrane with it amino terminus outside the cell2)
in the rod cell, a photon is absorbed, causing isomerization of rhodopsin’s 11-cis-retinal to the all-trans isomer.
This causes a change in the conformation of rhodopsin that stimulates transducin , setting off a biochemical amplification cascade that results
in a drop in cGMP concentration. That in turn leads to hyperpolarization of the plasma membrane and the signaling of second-order neurons.
Since the time of Wald’s demonstration that rhodopsin was the chromophore of vision and Hubbard’s contemporary proposal that
photodetection involved an isomerism of the chromophore, conventional wisdom has adopted that position. To support these proposals,
Collins proposed that a Schiff-base was the mechanism connecting the protein opsin to the retinoid, retinol , in forming rhodopsin. This
structure is defined as retinylidene-opsin. But since this proposal was found to have serious problems on energy grounds, an additional
conceptual proposal was made by Bownds suggesting protonation of the Schiff base , to form N-retinyl-opsin. No confirmation of these
reactions under biological conditions has occurred.
Hubbard showed the two isomers in solution differed in free energy by 1.1 kcal/mole at 253
Debate still rages on the precise description of the photoisomerization process at the molecular level. Shapiro recently reported molecular
mechanical calculations to model Warshel's "bicycle pedal" model concerted rotation about pivotal carbon-carbon double bonds as compared
to Liu's hula-twist model involving more than one double bond in rhodopsin as viable.4
These models apply to the chemical mechanisms for low-light level sensing (scotopic vision) rod cells and sunlight sensing color
distinguishing (photopic vision) cone cells.
COLOR SENSING
Our ability to differentiate colors requires that different visual cells respond characteristically to different parts of the spectrum. Theories
concerning the physiological basis of color vision originated with Isaac Newton , Thomas Young and Hermann von Helmholtz. These theories
were based on perception experiments.
An anecdote, documented in the Thomas Young Centenary celebration at Cornell, gave rise to the proposal of three types of cones in the
human eye. The proposal linked the “trichromatic theory” of light from Young, Helmholtz, and Maxwell with observations of many sources
which he incorporated into his mechanism of vision. Young is quoted as saying, “As it is almost impossible to conceive each sensitive point
of the retina to contain infinite number of particles, each capable of vibrating in perfect unison with every possible undulation,” Young
wrote, “It becomes necessary to suppose the number limited, for instance to three principle colors, red, yellow and blue.”
Young’s brilliant idea, thrown off casually in the course of a lecture, was forgotten, or lay dormant for fifty years, until Hermann von Helmholtz,
in the course of his own investigation of vision, resurrected it and gave it a new precision. …For Helmholtz, as for Young, color was a direct
expression of the wavelengths of light absorbed by each receptor, the nervous system just translating one into the other.5
See interesting discussion6.
How did we arrive at confirming Young's notion? It was from a clinical perspective of John Dalton . John Dalton described his own color
blindness in 1794, as confusing scarlet with green and pink with blue. Dalton supposed that his vitreous humor was tinted blue, selectively
absorbing longer wavelengths. He willed that after his death his eyes should be dissected and the color of his vitreous humor determined.
In July 1844 Joseph Ransome dissected his eyes and found his assumption false. He saved his cells in a preservative. In the 1990s British
biologists studied samples of his eyes and found that Dalton had the red photopigment gene, but lacked the green photopigment gene. He
was a green- dichromat.
Dalton was a deuteranope, lacking the middlewave photopigment of the retina.7
Biologists have examined the eyes of all species and found many similarities and some distinct differences that lead to differences in visual
acuity. One third do not detect light at all. A second third sense diffused illumination. The remainder, which include humans, focus an
image carried by light on to a array of organized light sensitive cells, the retina. The retina resides in the rear of the human eye covering
about 2500 mm^2 and ranging from 100 to 230 Um thick. Detailed mapping of human retina reveals that for the most part rods, low
light intensity sensing cells responsible for scotopic vision, are located over all of the retina at about 100,000 per mm^2. The density of rods
decreases, however, in the fovea centralis region and near the edges of the retina.
This foveal region has a diameter of 1.0 -- 1.5 mm and contains mostly cone photoreceptors more densely packed, near 200,000 per mm^2.
The fovea is located about 4-8 degrees from the optical axis and views a small fraction of the visual field. Estimates are 4-6 million
cones and 90-140 million rods in each human retina.8
CONE CELLS COLOR VISION
Color vision in cone cells involves a path of sensory transduction essentially identical to rod cells. Three types of cone cells are
specialized to detect different spectral regions using three related photoreceptor proteins—opsins. Each cone cell expresses only
one type of opsin. The opsins are closely related in size, amino acid sequence and presumably three dimensional structure. The
differences are great enough to place the chromophore in different environments inducing different absorption spectra. Protein
interactions cause spectral tuning of the chromophore absorption yielding variations in the maximum absorption wavelength.
This “opsin shift” from ‘bare’ 11-cis-retinal to rhodopsins in rods estimated as 500 MU and rhodopsins in cones within the region
between 400 and 600 MU. Candidate spectral tuning sites have been identified by mapping the amino acid sequence of fish opsin
onto a three dimensional model containing tuning sites and specific substitutions in amino acid secondary order in the opsin protein.
Potential tuning sites are identified as those that either point into the chromophore binding pocket or face other helices. This set of sites was
then extended to include all sites shown by Palczewski et al 2000 to be in close proximity to the chromophore or Schiff base. Based on
specific substitutions in specific protein locations, shifts in lambda max are observed.9
Fundamentally the lambda max of a visual pigment depends on at least two factors. First, the strength of the electrostatic interaction
between the Glutamine at 113 counterion and the protonated Schiff base is critical; substituents that increase the strength of this interaction
and stabilize the ground state will result in a shorter wavelength shift, those that reduce the interaction result in a longer wavelength shift.
Second, photoexcitation of the chromophore induces a significant increase in electron delocalization and a corresponding change in dipole
moment, with a shift of net positive charge towards the beta-ionene ring upon excitation (Kropf and Hubbard 1958 Mathies and Stryer
1976.) Interactions with charged, polar, or polarizable residues that alter delocalization will lead to a change in the energy difference ground
and excited states.
COLOR SENSING IS BUT A PART OF COLOR VISION
For every complex problem there is a simple solution. And it is always wrong. H. L. Mencken, 1949
Nearly a century after Maxwell provided a demonstration of color and linked it to Young's trichromatic color sensing in the human eye,
experiments could not be explained by the trichromatic theory were reported. Initially phenomenological terms described the effects.
D. H. Tanzer described nicely color constancy (same color under differing lighting conditions) and reflection from different surfaces were explained as
anomalies resulting from physical influences to a standard condition10.
Over time a more uniformly held view is that a second mechanism proposed by Ewald Herring in 1875 employing receptors that are
sensed and linked to reveal information. So, it is not that the trichromatic theory is invalid, but that color vision in humans is more
complex. This theory describes the contents of information obtained by the cones in terms of three types of "opponent activity":
red-green, blue-yellow, and white-black.
Edwin Land explored color vision that forced a departure from the physical view. He showed color was dependent on surveying a whole
view and comparing the wavelength composition from each point with the light reflected from the surround. Land's retinex model
proposed that the visual system made two comparisons first of all the wavelengths within the view coming up with a lightness record.
The second compared the three separate colors, red, green and blue. Again, we turn to Clinical evidence for support. Oliver Sacks
wrote about unique head injuries that could be temporarily influenced to reveal or lose color recognition.11
The visual process is edited and integrated in an amazingly rapid and coherent fashion. The information processing is not limited to
processes in the eye, which traces back to Galen who believed the retina and optic nerve were displaced portions of the brain.
(Some argue the concept originates with Aristotle.12)
To put it into perspective, Herring's theory treats some anomalies, but not all. Peter Gouras's more recent summary describes the brain's
role is more complicated with at least three mechanisms integrated by several sites in the brain.13
Investigations into this seem to begin with electro-retinography and anatomy leading to experimental results which have yet to
be explained by chemical phenomena.
We are seeing chemical work in terms of DNA characterization of gene coding giving rise to color recognition differences. Genes have been
identified for color blindness and other color sensing differences. It is likely the next series of advances will result from biochemical
experiments performing effect-cause-effect investigations on complicated systems in model forms.
Other provocative theories have been proposed about humans as blocked tetrachromats, simplifying the biological system with semiconductor
models and proposing likely rhodopsin binding models.14
It is well to remember these theories as we have learned from the tortuous path of our current understanding using Young's and Herring's
ideas as examples.
notes
1 http://math.ucr.edu/home/baez/physics/Quantum/see_photon.html
2 G. F. X. Schertler, C. Vial and R. Henderson, Nature 362, 770-2(1993).
3 R. Hubbard, j. Biol.Chem.241, 1814 (1966)
4 ja805586z%20JACS%20shapiro%2009.htm'
5 Oliver Sacks, "An Anthropologist on Mars," Alfred A. Knopf, New York, 1995, Ch. 1: The Case of the Colorblind Painter, pg. 3-41.
6 L. H. Anderson, I. B. Nielsen, M. B. Kristensen, M. O. A. El Ghazaly, S. Haacke, M. B. Nielsen, M. A. Petersen, J. Amer. Chem. Soc, 127, 12347 - 12350 (2005)
7 D. M. Hunt, K. S. Dulai, J. K. Bowmaker, J. D. Mollon, Science 267, 984 - 988 (1995)
8 Interestingly, the number of rods have been observed to decrease as humans age. David Bainbridge, "Beyond the Zonules of Zinn,"Harvvard University, Cambridge, 2008, p. 150.
webvision.med.utah.edu/FACTS.html
9 K. Palczewski, T. Kumasaka, T. Hoi, C. A. Behnke, H. Motoshima, B. A. Fox, I. LeTrong, D. C. Teller, T. Osakda, R. E. Stenkampf, M. Yamamoto, M. Miyano, Science 289, 739 - 745 (2000) ; R. E. Stenkampf, Acta Crystallographica D Biol. Crystallogr. 64 (Pt 8) 902 - 904 (2008).
10 D. H. Tanzer, "Physiology and Psychology of Human Color Vision," AI Memo 369- Massachusetts Institute of Technology, August, 1976.
11 Oliver Sacks, "An Anthropologist on Mars, Alfred A. Knopf, New York, 1995.
12 S. Finger, "Origins of Neuroscience: A History of the Explorations into Brain Function." Oxford University Press, Oxford, 2000, p. 40.
13 http://webvision.med.utah.edu/Color.html
14 J. T. Fulton, "Biologicla Vision: A 21st Century Tutorial," Vision Concepts, Trafford Publishing, 2004. http://manta.com/coms2/dnbcompany_0h4yr