Source: UNIVERSITY OF CALIFORNIA, DAVIS submitted to
COMPARATIVE STUDIES OF THE VERTEBRATE EYE
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
TERMINATED
Funding Source
Reporting Frequency
Annual
Accession No.
0133523
Grant No.
(N/A)
Project No.
CA-D*-NPB-4910-H
Proposal No.
(N/A)
Multistate No.
(N/A)
Program Code
(N/A)
Project Start Date
Oct 1, 2002
Project End Date
Jun 28, 2007
Grant Year
(N/A)
Project Director
Sillman, A. J.
Recipient Organization
UNIVERSITY OF CALIFORNIA, DAVIS
410 MRAK HALL
DAVIS,CA 95616-8671
Performing Department
NEUROBIOLOGY, PHYSIOLOGY AND BEHAVIOR
Non Technical Summary
(1) The visual abilities of animals that are endangered (sturgeon), essential to a healthy ecosystem (snake, alligator), or commercially important (sturgeon) are poorly understood. (2) Heavy metal poisoning affects the vision of humans as well as domestic and wild animals. We will study the photoreceptors of sturgeon, snake and alligator to determine distribution and light absorption, the ability to process color, and the effect of heavy metals on light absorption.
Animal Health Component
(N/A)
Research Effort Categories
Basic
100%
Applied
(N/A)
Developmental
(N/A)
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
3050899102080%
3140899115020%
Goals / Objectives
: 1) Determine whether or not the feeding behavior of larval white sturgeon is influenced by manipulation of the spectral environment. 2) Characterize the absorption spectra and chromophore types of the visual pigments in the photoreceptors of the green sturgeon, pallid sturgeon, rattlesnake and alligator. 3) Determine whether or not heavy metals inhibit visual pigment regeneration in the retina. 4) Determine hotoreceptor distribution in the retinas of sturgeons and paddlefish. 5) Sequence the DNA and characterize the visual pigment opsins in sturgeons, white shark, rattlesnake and night snake. 6) Determine the extractability and chloride sensitivity of night snake visual pigment.
Project Methods
In situ microspectrophotometry will be employed to measure absorbance in single photoreceptors. Classical digitonin extraction techniques will be employed to place visual pigments into solution for ordinary spectrophotometric analysis. Brightfield and Nomarski light microscopy will be used to map photoreceptor distribution in retinal whole-mounts. Fundus reflectometry will be used to study the kinetics of visual pigment regeneration in isolated eyecup preparations. DNA isolation, amplification and cloning will permit determination of amino acid sequences in visual pigment opsins.

Progress 10/01/02 to 06/28/07

Outputs
For both paddlefish (Polyodon spathula) and green sturgeon (Acipenser medirostrus retinal photoreceptor density was uniform throughout. There was no evidence for either a typical fovea or visual streak. Mean cone density for paddlefish was 6,401 +/- 1,216 cones/mm2; mean rod density 10,271 +/- 1,205 rods/mm2. Mean cone density for the green sturgeon was 4,689 +/- 891 cones/mm2; mean rod density 16,005 +/- 1667 rods/mm2. Cones comprised 38% of the photoreceptor population in paddlefish; 23% in green sturgeon. Mean diameter of paddlefish oil globules in the ventral portion of the retina was 4.47 +/- 0.84 micrometers; dorsal portion 5.04 +/- 0.84 micrometers. Mean diameter of green sturgeon oil globules in the ventral portion of the retina was 4.65 +/- 1.07 micrometers; dorsal portion 5.15 +/- 1.04 micrometers. Ventral portions of the retinas of both species are richer in the smaller oil globules. In a hybrid sturgeon (white sturgeon, Acipenser transmontanus, crossed with lake sturgeon, Acipenser fulvescens) the rod pigment absorbs maximally near 542 nm, as does the middle wavelength sensitive cone pigment. The long wavelength sensitive cone pigment absorbs maximally near 616 nm; the short wavelength sensitive cone pigment near 471 nm. Lake sturgeon rods contain a visual pigment absorbing maximally at 541 +/- 2 nm. The short wavelength sensitive cone photoreceptor contains a visual pigment absorbing maximally at 448 +/- 1 nm; middle wavelength sensitive cone at 538 +/- 1 nm; long wavelength sensitive cone at 619 +/- 3 nm. All visual pigments were best fit by nomograms for visual pigments based on vitamin A2. Lake sturgeon rods are typical of vertebrates in general and are similar to rods found in other species of sturgeon and in the North American paddlefish. Mean length of the outer segment is 39.7 +/- 6.2 micrometers; mean diameter is 6.2 +/- 0.5 micrometers. Cones are also similar to those found in other Acipenserids. They are single, robust cones with a globular inner segment from which extends a tapering outer segment. Mean diameter of the inner segment at its widest point is 7.7 +/- 1.4 micrometers. Mean diameter at the base of the outer segment is 5.0 +/- 0.8 micrometers. Two cones were seen with relatively undamaged outer segments of 15.5 miocrometers and 14.8 micrometers, respectively. Mean packing density of rod photoreceptors in the lake sturgeon, as determined with SEM, is 22,624 +/- 3,509 rods/mm2. Cones comprise 34.7 +/- 4.5 % of the lake sturgeon's photoreceptor population.

Impacts
All species of sturgeon are considered threatened to one degree or another. Knowledge gained through study of one of their primary sensory modalities should foster better decisions with respect to environmental policy and management. Farmed sturgeon are becoming increasingly more valuable as a food source. The data may allow for more effective conservation and aquaculture.

Publications

  • No publications reported this period


Progress 01/01/06 to 12/31/06

Outputs
Electron micrographs of the lake sturgeon (Acipenser fulvescens) retina were analyzed. The rods are typical of vertebrates in general and are similar to rods found in other species of sturgeon and in the North American paddlefish (Polyodon spathula). Mean length of the rod outer segment is 39.7 plus or minus 6.2 micro meters (n = 36; range: 28.1 - 54.0 micro meters); mean diameter is 6.2 plus or minus 0.5 micro meters (n = 35; range: 4.8 - 7.2 micro meters). Cones are also similar to those found in other Acipenserids. They are single cones, well described as robust with a somewhat globular inner segment from which extends a tapering outer segment. Mean diameter of the cone inner segment at its widest point is 7.7 plus or minus 1.4 micro meters (n = 30; range: 5.1 - 12.1 micro meters). Mean diameter at the base of the outer segment is 5.0 plus or minus 0.8 micro meters (n= 29; range: 3.8 - 7.2 micro meters). Two cones were seen with relatively undamaged outer segments of 15.5 micro meters and 14.8 micro meters, respectively. By counting the tips of rod photoreceptors seen from the distal aspect we found that the mean packing density of rod photoreceptors in the lake sturgeon, as determined with the scanning electron microscope (SEM), is 22,624 plus or minus 3,509 rods/mm2. Although this value is in reasonable agreement with packing densities found in five other Acipenseriformes using the same SEM technique, it undoubtedly is higher than it should be. Other studies with the retinal whole mount method, which was not used here, yielded values much lower than those arrived at using the SEM. Tissue shrinkage and preferential sampling probably account for the higher values obtained with the SEM. By counting rods and cones in micrographs of edges such as that shown in Fig. 1A, we calculated that cones comprise 34.7 plus or minus 4.5 % of the lake sturgeon's photoreceptor population (n = 8; range: 28.6 - 40.4%). This value agrees well with those obtained in previous SEM studies.

Impacts
All species of sturgeon are considered threatened to one degree or another. Knowledge gained through study of one of their primary sensory modalities should foster better decisions with respect to environmental policy and management. Farmed sturgeon are becoming increasingly more valuable as a food source. The data may allow for more effective conservation and aquaculture.

Publications

  • Sillman, A. J., Ong, E. K. and Loew, E. R. (2007). Spectral absorbance, structure and population density of photoreceptors in the retina of the lake sturgeon (Acipenser fulvescens). Can. J. Zool. Submitted.


Progress 01/01/05 to 12/31/05

Outputs
Absorbance spectra acquired by means of single cell, in situ microspectroscopy were analyzed to obtain accurate wavelengths of maximum absorbance for the photoreceptors of the Lake Sturgeon, Acipenser fulvescens. The rod photoreceptor contained a visual pigment absorbing maximally at 541 +/- 2 nm (n = 18, range: 537 - 543 nm). The short wavelength sensitive cone photoreceptor contained a visual pigment absorbing maximally at 448 +/- 1 nm (n = 7, range: 446 - 450 nm); middle wavelength sensitive cone at 538 +/- 1 nm (n = 11, range: 536 - 540 nm); long wavelength sensitive cone at 619 +/- 3 nm (n = 8, range: 615 - 623 nm). No ultraviolet sensitive visual pigment was identified. All visual pigments were best fit by nomograms for visual pigments based on vitamin A2. The data are consistent with those obtained previously from other sturgeon species.

Impacts
All species of sturgeon are considered threatened to one degree or another. Knowledge gained through study of one of their primary sensory modalities should foster better decisions with respect to environmental policy and management. Farmed sturgeon are becoming increasingly more valuable as a food source. The data may allow for more effective conservation and aquaculture.

Publications

  • Sillman, A. J., Beach, A. K., Dahlin, D. A. and Loew, E. R. (2005). Photoreceptors and visual pigments in the retina of the fully anadromous green sturgeon (Acipenser medirostrus) and the potamodromous pallid sturgeon (Scaphirhynchus albus). J. Comp. Physiol. A, 191:799-811.


Progress 01/01/04 to 12/31/04

Outputs
Hybrid sturgeon, created by crossing white sturgeon (Acipenser transmontanus) with lake sturgeon (Acipenser fulvescens) were maintained in fresh water, flow-through tanks for seven years. The retinas from two fish, each about 1 meter total body length, were excised, isolated from the pigment epithelium, and macerated. The outer segments of the rod and cone photoreceptors were examined by means of in situ, single cell microspectrophotometry. Four different visual pigments were identified. The rod pigment absorbs maximally near 542 nm, as does the middle wavelength sensitive cone pigment. The long wavelength sensitive cone pigment absorbs maximally near 616 nm; the short wavelength sensitive cone pigment near 471 nm. We found no photoreceptor containing a visual pigment sensitive to ultraviolet light. All visual pigments were best fit by nomograms for visual pigments based on vitamin A2. The data are consistent with those obtained previously from other sturgeon species.

Impacts
All species of sturgeon are considered threatened to one degree or another. Knowledge gained through study of one of their primary sensory modalities should foster better decisions with respect to environmental policy and management. Farmed sturgeon are becoming increasingly more valuable as a food source. The data may allow for more effective conservation and aquaculture.

Publications

  • Sillman, A. J. and Dahlin, D. A. (2004). The photoreceptors and visual pigments of sharks and sturgeons. In: The Senses of Fishes: Adaptations for the Reception of Natural Stimuli, G. von der Emde, J. Mogdans and B. G. Kapoor (eds.), Narosa Publishing House, New Delhi, pp31-54.
  • Sillman, A. J. and Dahlin, D. A. (2004). Photoreceptor topography in the duplex retina of the paddlefish (Polyodon spathula). J. Exp. Zool., 301A:674-681.


Progress 01/01/03 to 12/31/03

Outputs
Whole mounts were prepared from the isolated retinas of two Acipenseriform fishes, the paddlefish (Polyodon spathula) and the green sturgeon (Acipenser medirostrus). Using a Nikon Eclipse Model E600 microscope and Scion Image video-enhancement software, we measured the diameter of the oil globules located in the inner segment of each cone photoreceptor. Mean diameter of paddlefish oil globules in the ventral portion of the retina was 4.47 +/- 0.84 micrometers (n = 272; range: 2.0 to 6.4 micrometers); in the dorsal portion 5.04 +/- 0.84 micrometers (n = 324; range: 2.8 to 7.2 micrometers). Mean diameter of green sturgeon oil globules in the ventral portion of the retina was 4.65 +/- 1.07 micrometers (n = 83; range: 1.8 to 6.8 micrometers); in the dorsal portion 5.15 +/- 1.04 micrometers (n = 90; range: 2.8 to 7.2 micrometers). Visual inspection of the histograms shows a definite skewing of the data such that the ventral portions of the retinas of both species are richer in the smaller oil globules. The oil globules in Acipenseriform species are colorless, having no wavelength-specific absorbance between 350 and 750 nm, and therefore do not affect the light sensitivity of the cones. However, different size oil globules may be associated with cones having visual pigments absorbing in different regions of the spectrum. The correlation of oil globule size with microspectrophotometric data remains to be done.

Impacts
All species of sturgeon and paddlefish are considered threatened to one degree or another. Knowledge gained through study of one of their primary sensory modalities should foster better decisions with respect to environmental policy and management. Farmed sturgeon are becoming increasingly more valuable as a food source. The data may allow for more effective conservation and aquaculture.

Publications

  • No publications reported this period


Progress 01/01/02 to 12/31/02

Outputs
Data were analyzed from previously prepared whole mounts of retinal tissue from the freshwater paddlefish (Polyodon spathula) and the fully anadromous green sturgeon (Acipenser medirostrus). Mapping of the photoreceptor population of each species was done using a Nikon Eclipse Model E600 microscope and Scion Image video-enhancement software. Data from several retinas were superimposed and, using the optic disk as the central point, quadrants and radii were delineated. Similarly, the superimposed retinas were divided up into horizontal rows so as to determine whether or not a visual streak is present. The data were compared by analysis of variance followed by T-tests to determine significance of differences between the specific regions. For both species, photoreceptor density was uniform throughout the retina. There was no sign of areas of especially high cone density and, therefore, no evidence for the presence of either a typical fovea or a visual streak. Mean cone density for the paddlefish was 6,401 +/- 1,216 cones/mm2; mean rod density 10,271 +/- 1,205 rods/mm2. Mean cone density for the green sturgeon was 4,689 +/- 891 cones/mm2; mean rod density 16,005 +/- 1667 rods/mm2. Thus, cones comprised 38% of the photoreceptor population in paddlefish; 23% in green sturgeon. The 23% value for the green sturgeon retina was virtually identical to that obtained previously for the white sturgeon (A. transmontanus).

Impacts
All species of sturgeon and paddlefish are considered threatened to one degree or another. Knowledge gained through study of one of their primary sensory modalities should foster better decisions with respect to environmental policy and management. Farmed sturgeon are becoming increasingly more valuable as a food source. The data may allow for more effective conservation and aquaculture.

Publications

  • No publications reported this period


Progress 01/01/01 to 12/31/01

Outputs
Whole mount preparations were made from either mildly fixed or fresh retinas isolated from the fully anadromous green sturgeon (Acipenser medirostris), the semi-anadromous white sturgeon (Acipenser transmontanus), and the freshwater paddlefish (Polyodon spathula), with the intent of (1) determining whether or not acipenseriform retinas contained areas of specialization and (2) obtaining accurate photoreceptor packing density values. The retinas, isolated from the pigmented epithelium were mounted on glass slides and the photoreceptor population (both rods and cones) was mapped using a Nikon Eclipse Model E600 microscope and Scion Image video-enhancement software. At a magnification of 200X, images 0.69 x 0.53 mm were captured and archived so that the entire retina could be reconstructed after the cell counts were made. The data from several retinas were then superimposed and, using the optic disk as the central point, quadrants and radii (at 1.0 mm intervals) were delineated. Composite data were compared first by analysis of variance (ANOVA), after which T-tests were used to determine the significance of differences between specific regions. Data analysis is not yet completed for the green sturgeon and paddlefish. For the white sturgeon, other than a tendency for the photoreceptors to be less densely packed at the far periphery, no region of the retina appeared to be significantly different from any other region. Mean rod density for the entire white sturgeon retina was 15,329 +/- 2,679 rods/mm2, whereas mean cone density for the entire retina was 3,855 +/- 832 cones/mm2. Thus, cones comprise 20% of the total photoreceptor population in the retina of the white sturgeon. These values are substantially lower than previous estimates made from studies with the scanning electron microscope (SEM). However, for several reasons, there is little doubt that the values obtained with the retinal whole mount preparation described here are the more accurate.

Impacts
All species of sturgeon and paddlefish are considered threatened to one degree or another. Knowledge gained through study of one of their primary sensory modalities should foster better decisions with respect to environmental policy and management. Farmed sturgeons are becoming increasingly more valuable as a food source. The data may allow for more effective conservation and aquaculture.

Publications

  • Sillman, A. J., Johnson, J. L. and Loew, E. R. (2001). Retinal photoreceptors and visual pigments in Boa constrictor imperator. J. Exp. Zool., 290:359-365.


Progress 01/01/00 to 12/31/00

Outputs
Rod and cone photoreceptors of two species of Acipenseriform fishes, the fully anadromous green sturgeon (Acipenser medirostris) and the freshwater pallid sturgeon (Scaphirhynchus albus), were examined with the microspectrophotometer (MSP) and the scanning electron microscope. The photoreceptor population is similar in the two species, in terms of both structure and distribution. Rods dominate but cones comprise approximately 40% of the population. Rods may be of two types. The common rod is a robust cell with a long, cylindrical outer segment of large diameter. A rare rod has a much narrower outer segment. Cones are of one morphological type, single cones with large diameter inner segments connected to large, tapered outer segments. A colorless oil globule was present in the inner segment of each cone. Oil globules of one diameter are associated with cones with one type of spectral absorbance. Wavelengths of peak absorbance have not yet been computed, but MSP measurements showed the presence, in both species, of cones that are long wavelength-, middle wavelength- and short wavelength-sensitive, respectively. Rods absorb maximally near 540 nm. Preliminary analysis indicates that all the visual pigments are based on vitamin A2. Paddlefish (Polyodon spathula) were raised to useful size (c. 45 cm) and wholemounts of their retinas were prepared. The entire photoreceptor population of each retina was scanned and captured digitally, with the intent of mapping rod and cone distribution and determining whether or not the paddlefish retina has regional specializations.

Impacts
All species of sturgeon and paddlefish are considered threatened to one degree or another. Knowledge gained through study of one of their primary sensory modalities should foster better decisions with respect to environmental policy and management. Farmed sturgeon are becoming increasingly more valuable as a food source. The data may allow for more effective conservation and aquaculture.

Publications

  • WEIDNER, W. J., WADDELL, D. S. and SILLMAN, A. J. 2000. Low levels of cadmium chloride alter the immunoprecipitation of corneal cadherin-complex proteins. Archives of Toxicology, in press.


Progress 01/01/99 to 12/31/99

Outputs
Rod and cone photoreceptors of the boa constrictor (Boa constrictor imperator) were analyzed with the microspectrophotometer. The absorbance spectra for the visual pigments within these cells proved to be quite similar to that of ball python photoreceptors. Peak absorbance for the rod pigment is near 494nm. Peak absorbance for the visual pigment in the most common boa constrictor cone is near 551nm. A much rarer, and smaller, cone contained a visual pigment absorbing in the ultraviolet, with peak absorbance near 360nm. Comparison of the pigment curves with a nomogram revealed that the visual pigments of boa constrictor contain chromophores based on vitamin A. Scanning electron microscopy confirmed the presence of three distinct photoreceptor types in boa constrictor. Rods are typical in that their outer segments are long and cylindrical. The most common cone is relatively large with a robust, tapered outer segment. The rare cone is significantly smaller, with a more delicate tapered outer segment. Spectral transmission of the optical media of both the boa constrictor and the ball python were measured with a spectrophotometer. The lens, spectacle and cornea in both species functioned as cut-off filters, transmitting light very effectively down to about 320 nm. Green sturgeon were acquired and are being maintained for the purpose of characterizing their photoreceptors and visual pigments. Paddlefish larvae were obtained and these fish are being raised to a size appropriate for analysis of photoreceptor distribution throughout the entire retina using retinal wholemounts. Work continued using western immunoblots to study the effects of heavy metals on corneal endothelial integrity. Both 1.0 micromolar and 75.0 micromolar concentrations of cadmium chloride decreased the amount of immunoprecipitation with pan cadherin antibody. However, whereas the 1.0 micromolar dose virtually eliminated immunoprecipitation, the 75.0 dose only reduced it, albeit significantly. This suggests two modes of action of cadmium on adhesion molecules.

Impacts
The snake and (endangered) sturgeon data foster better decisions on environmental policy and management. Sturgeon flesh and caviar are becoming increasingly more valuable as a food source. The sturgeon data allow for more effective conservation and aquaculture. The heavy metal data are important because those metals are hazards to human and animal health.

Publications

  • Sillman, A. J., Carver, J. K. and Loew, E. R. 1999. The photoreceptors and visual pigments in the retina of a boid snake, the ball python (Python regius). J. Exp. Biol., 202:1931-1938.


Progress 01/01/98 to 12/01/98

Outputs
Rod and cone photoreceptors of the ball python (Python regius) retina were analyzed with the microspectrophotometer. Peak absorbance of the rod visual pigment is 494 nm. Two cone pigments were identified. The common cone, with a stubby outer segment, contains a pigment with peak absorbance at 551 nm. The rare cone, with a long, slender outer segment, contains an ultraviolet sensitive pigment with peak absorbance at 360 nm. All pigments are based on vitamin A1. The photoreceptors of three different strains of Lake sturgeon (Acipenser fulvescens) larvae were analyzed with the microspectrophotometer at two week intervals from day of hatch through three months of age. Results were similar to those from white sturgeon (A. transmontanus) and shovelnose sturgeon (Scaphirhynchus platorynchus) in that there is one rod pigment with peak absorbance near 500 nm, as well as long-, middle- and short-wavelength sensitive cone pigments. Exact values for peak absorbances have not yet been calculated. All pigments are based on vitamin A2. Tissue from lake sturgeon was preserved for future DNA analysis. Western blot analysis, enhanced chemoluminescence and immuno-electron microscopy were employed to examine the effects of cadmium (CdC12) on corneal endothelial cadherin activity. Cd appears to decrease immunoprecipitation of cadherins and the associated regulatory proteins, alpha and beta catenin. This effect may be associated with altered protein phosphorylation. Altered cadherin complex activity may, in part, explain the damaging effects of heavy metals on corneal endothelial permeability.

Impacts
(N/A)

Publications

  • SILLMAN, A. J., O'LEARY, C. J., TARANTINO, C. D. and LOEW, E. R. The photoreceptors and visual pigments of two species of Acipenseriformes, the shovelnose sturgeon (Scaphirhynchus platorynchus) and the paddlefish (Polyodon spathula).


Progress 01/01/97 to 12/01/97

Outputs
Scanning electron micrographs of ball python (Python regius) retina were analyzed and photoreceptor structure and distribution were quantified. About 8% of the photoreceptors throughout the retina are cones. There appears to be some regional variability in that the cone population ranged from 5% to 15%. Although all cones are of the single variety, at least two different morphotypes could be identified on the basis of relative size of outer and inner segments. Rods, which comprise 92% of the photoreceptor population, are uniformly distributed throughout the retina with mean density of 457,740 /+/-/ 11,517 cells/mm(superscript 2). The rods are fairly uniform in size with a mean outer segment diameter of 1.6 /+\-/ 0.2 (mu)m and a mean outer segment length of 27.4 /+\-/ 2.2 (mu)m. Data from bullfrog rhodopsin extraction experiments examining the effects of a heavy metal (PbCI[2]) on visual pigment regeneration were suggestive but equivocal. Thus, a fundus reflectometer was designed and constructed to allow dynamic measurements of visual pigment regeneration in situ. Research continued on the effects of heavy metals on corneal endothelial permeability. Immunoblotting techniques revealed the presence of cadherin and both alpha and beta catenins in the bullfrog cornea. Experiments were performed to determine whether or not cadherins and/or catenins are up or down regulated by either of two concentrations of cadmium chloride, but more data need to be collected before a conclusion can be drawn.

Impacts
(N/A)

Publications

  • SILLMAN, A. J., GOVARDOVSKII, V. I., ROHLICH, P., SOUTHARD, J. A. and LOEW, E. R. 1997. The photoreceptors and visual pigments of the garter snake (Thamnophis sirtalis): a microspectrophotometric,
  • WEIDNER, W. J. and SILLMAN, A. J. 1997. Low levels of cadmium chloride damage the corneal endothelium. Arch. Toxicol., 71:455-460.
  • LOEW, E. R. and SILLMAN, A. J. 1998. Spectral sensitivity of a phototactic behavior of larval white sturgeon, Acipenser transmontanus. Vision Res., in press.


Progress 01/01/96 to 12/30/96

Outputs
Hybrid sturgeon produced from parent lines of white sturgeon and lake sturgeon were raised, and their retinas were then examined with the scanning electron microscope. Their photoreceptors are very similar to those of the white sturgeon. That is, about 60% of the photoreceptors are rods with very large, cylindrical outer segments. The remaining 40% are cones with relatively large, tapered outer segments. Only single cones could be identified and there appears to be only one morphotype. The cone outer segment was characterized by the presence of an accessory outer segment virtually identical to that seen in white sturgeon cones. The scanning electron microscope was also used to examine the photoreceptors of a boid snake, the ball python. In contrast to the all cone retina of the garter snake, the python has a rod dominated retina with only about 20% of the photoreceptors identified as cones. The cones are all single cones; no double or twin cones were observed. There may be more than one cone morphotype as some of the cones appeared to be larger than others. The rods, with their long, slender, cylindrical outer segments, are typical of vertebrate rod photoreceptors. A very rich visual pigment extract was prepared from the retinas of shovelnose sturgeon and analyzed spectrophotometrically with the partial bleaching technique.

Impacts
(N/A)

Publications

  • SILLMAN, A. J., LETSINGER, G. A., PATEL, S., LOEW, E. R. and KLIMLEY, A. P. 1996. Visual pigments and photoreceptors in two species of shark, "Triakis semifasciata" and "Mustelus henlei". J. Exp. Zool. 276:1-10.


Progress 01/01/95 to 12/30/95

Outputs
Microspectrophotometric examination of the retina of the garter snake, THAMNOPHIS SIRTALIS, revealed the presence of a previously undetected cone photoreceptor with peak light absorbance in the ultraviolet. This UV sensitive photoreceptor is probably the very small single cone identified previously both immunocytochemically and with the scanning electron microscope. Analysis of scanning electron micrographs show that the garter snake retina has only cones, with a packing density of from 12,000 to 18,000 cells per square millimeter, depending on the subspecies. Double cones and large single cones dominate, comprising 43% and 42% of the photoreceptor population, respectively. Small single cones comprise 11% of the photoreceptors; only about 4% of the cones are very small singles. Scanning electron microscopy of the retinas of the shovelnose sturgeon (SCAPHIRHYNCHUS PLATORYNCHUS) and the paddlefish (POLYODON SPATHULA) revealed duplex retinas with photoreceptors very similar in structure and distribution to those of the white STURGEON (ACIPENSER TRANSMONTANUS). Scanning electron microscopy of bullfrog corneas exposed to micromolar concentrations of cadmium chloride showed structural damage to the corneal endothelium, thus confirming previous vital staining data showing that cadmium chloride interferes with the corneal endothelium's barrier function. Similar structural damage was seen after exposure to another heavy metal, lead chloride.

Impacts
(N/A)

Publications

  • No publications reported this period.


Progress 01/01/94 to 12/30/94

Outputs
The photoreceptors of paddlefish and shovelnose sturgeon larvae were examined with the microspectrophotometer (MSP) from day of hatch to 10 months of age. Both species exhibit green sensitive rods and green sensitive cones first. Red sensitive cones develop later. Still later the shovelnose sturgeon develops blue sensitive cones, but the paddlefish retina remains devoid of anything but red and green sensitive cells. All visual pigments in both species are based on the vitamin A2 chromophore. Immunohistochemistry of garter snake retina revealed the presence of a very small, single, ultraviolet (UV) sensitive cone in addition to green sensitive and blue sensitive cones found previously with the MSP. Scanning electron microscopy (SEM) revealed the presence of large single cones, double cones, small single cones and very small single cones. SEM thus supports the histochemical data, however, MSP examination has failed to demonstrate the presence of a UV sensitive photoreceptor. Micromolar concentrations of cadmium chloride damage the integrity of the corneal endothelium in a dose dependent manner. The effect of cadmium is enhanced by the calcium ionophore A23187, but is not affected by altering external calcium concentration.

Impacts
(N/A)

Publications

  • SILLMAN, A. J., SORSKY, M. E. and LOEW, E. R. In Press. The visual pigments of wild white sturgeon (Acipenser transmontanus). Can. J. Zool.


Progress 01/01/93 to 12/30/93

Outputs
Scanning electron microscopy of the retinas of leopard and brown smoothhound sharks allowed positive identification of cone photoreceptors. Cones comprise but a small percentage of the photoreceptor population. This was confirmed by microspectrophotometric analysis which yielded records from only two possible cones, one with peak absorbance in the blue; one in the green. Microspectrophotometric analysis of photoreceptors and partial bleaching analysis of visual pigment extracts from wild sturgeon taken from estuarine waters showed that the spectral absorbance characteristics of wild sturgeon photoreceptors and cultured sturgeon photoreceptors are the same. The visual pigment chromophore of estuarine sturgeon is always based on vitamin A2, as is true of the cultured sturgeon. Microspectrophotometric analysis of garter snake photoreceptors revealed a blue sensitive photoreceptor in addition to the accepted green sensitive photoreceptor.

Impacts
(N/A)

Publications

  • LOEW, E. R. and SILLMAN, A. J. 1993. Age-related changes in the visual pigments of the white sturgeon (Acipenser transmontanus). Can. J. Zool., 71: 1552-1557.
  • SILLMAN, A. J., RONAN, S. J. and LOEW, E. R. In Press. Scanning electron microscopy and microspectrophotometry of the photoreceptors of ictalurid catfishes. J. Comp. Physiol. A.
  • SILLMAN, A. J. and WEIDNER, W. J. 1993. Low levels of inorganic mercury damage the corneal endothelium. Exp. Eye Res., 57: 549-555.


Progress 01/01/92 to 12/30/92

Outputs
Analysis of visual pigment extracts prepared from the retinas of leopard and brown smoothhound sharks showed that both juveniles and adults possess only one rod photopigment, based on vitamin A1. The spectral absorbance is somewhat different for the two species, but peak absorbance is always near 500nm. Single cell, in situ, microspectrophotometry, which confirmed these findings on rod photoreceptors, yielded no evidence for the presence of cone photoreceptors in either species. Scanning electron microscopy of the retinas of pacific hagfish revealed the presence of well developed rod outer segments with relatively undifferentiated inner segments. Structures resembling cone outer segments were seen, but their exact identity remains uncertain. Micromolar doses of inorganic mercury disturb the barrier function of bullfrog corneal endothelium. The effect is not altered by changes in external calcium, nor is it influenced by the calcium ionophore A23187, indicating that inorganic mercury disturbs the barrier function of the corneal endothelium through a mechanism which does not involve competition with external calcium or interaction with calcium channels. Transmission and scanning electron microscopy demonstrate significant ultrastructural damage to the endothelium exposed to inorganic mercury, including cellular swelling, increased vacuolization, focal denuding of Descemet's membrane, and diminished integrity at the intercellular junctions.

Impacts
(N/A)

Publications

  • No publications reported this period.


Progress 01/01/91 to 12/30/91

Outputs
Rod photoreceptors of the American alligator (ALLIGATOR MISSISSIPPIENSIS) all contain a visual pigment which absorbs light maximally near 500nm. Four different pigments are present in the cone photoreceptors. The most common cone is asingle cone containing a pigment with absorbance maximum near 535nm. Doubles cones are also common. The principal memeber contains a pigment with absorbance maximum near 566nm; the accessory member near 503nm. Blue cones, with maximum absorbance near 443nm are rare. All pigments are based on Vitamin A1. Channel catfish (ICTALURUS PUNCTATUS) cones develop before rods. Two different cone pigments are present. One absorbs maximally near 608nm; the other near 535nm. Rods absorb maximally near 540nm. Pigment content does not change with development. All catfish pigments are based on Vitamin A1. White sturgeon (ACIPENSER TRANSMONTANUS) rods, containing a pigment with maximum absorbance near 540nm, devlop before cones. A cone absorbing maximally near 536nm is the first to appear. Somewhere between three and six months of age a blue (c. 430nm) absorbing cone and a red (605nm) sensitive cone develop. Sturgeon pigments are all Vitamin A2 based. There is no change with changes in light environment or season. Horse photoreceptors were examined with the scanning electron microscope. Rod photoreceptors dominate. Ten to 20% of the photoreceptors are cones with relatively large inner segments and fine (c.2.75 microns) outer segments.

Impacts
(N/A)

Publications

  • NO PUBLICATIONS REPORTED THIS PERIOD.


Progress 01/01/90 to 12/30/90

Outputs
Using histological techniques and IN SITU, single cell microspectrophotometry, the development of photoreceptors and their visual pigments were studied in two species of benthic fish--the white sturgeon (ACIPENSER TRANSMONTANUS) which is a chondrostean, and the channel catfish (ICTALURUS PUNCTATUS) which is a modern teleost. In the catfish, cone photoreceptors develop first and no rod photoreceptors can be found in the yolk-fry. Both red sensitive and green sensitive cones are present with the visual pigment of the former absorbing maximally at 608nm and that of the latter at 535nm. The visual pigment of the later developing rod absorbs maximally at 540nm. The pigment content of the catfish retina did not change with adulthood. Development in the sturgeon retina is quite different. Rods which, like the catfish, contain a visual pigment which absorbs maximally at 540nm, develop before cones. The cone which initially does develop in the sturgeon retina is a green sensitive cone with maximal absorbance at 536nm. The red sensitive cone (maximal absorbance at 605 nm), which is the only cone found in the adult sturgeon, is not seen even at five months of age. In contrast to the catfish, the retina of the sturgeon undergoes substantial change during the course of development. At some point there is a switch from a green sensitive to a red sensitive system. This probably involves changes in the nature of the visual pigment protein which we hope to correlate with the life history of this animal.

Impacts
(N/A)

Publications

  • SILLMAN, A. J., RONAN S. J. and LOEW, E. R. 1991. Histology and microspectrophotometry of the photoreceptors of a crocodilian, ALLIGATOR MISSISSIPPIENSIS. Proc. Roy. Soc. Lond. B. 243:93-98.


Progress 01/01/89 to 12/30/89

Outputs
The visual pigments in the photoreceptors of the American alligator, Alligator mississippiensi, were studied by means of in situ, single cell microspectrophotometry. The rod photoreceptors all contained the same pigment, characterized by a light absorbance maximum near 500 nm. Four different pigments were found in the cone photoreceptors. The most frequently encountered cone was a single cone containing a visal pigment with absorbance maximum near 535nm. Double cones were also common. The principal member of the double contained a visual pigment with maximum absorbance near 566nm, whereas the pigment of the accessory member absorbed maximally near 503nm. Blue absorbing cones were present but were rarely encountered. The visual pigment of the blue cones absorbed maximally near 443nm. Comparison with appropriate theoretical nomogram curves indicated that all visual pigments were based on the vitamin A(1) chromophore and, therefore, were rhodopsins. The pigment data provide some insight into the evolution of color vision. They also allow more reasoned speculation about the visual capabilities of the extinct ruling reptiles to which the alligator is closely related. Examination of the retina with the scanning electron microscope reveals a retina which might best be described as in transition.

Impacts
(N/A)

Publications

  • SILLMAN, A.J., SPANFELNER, M.D. and LEOW, E.R. 1990. The photoreceptors and visual pigments in the retina of the white sturgeon, Acipenser transmontanus. Can. J. Zool. In press.


Progress 01/01/88 to 12/30/88

Outputs
Construction of a single cell microspectrophotometer (MSP) was completed and theinstrument was made fully operational, with the exception of the computer interface to be added later. Using a Nicolet digital oscilloscope to collect and store data it was possible to record absorbance spectra from the single photoreceptors of the sturgeon. The sturgeon retina was found to contain one type of cone photoreceptor and one type of rod photoreceptor with respect to visual pigments. The cone contains a vitamin A(subscript 2) based pigment absorbing maximally near 608nm. The rod contains a vitamin A(subscript 2) based pigment absorbing maximally at 545nm. The rod MSP data confirmed the finding made with digitonin extracts of the visual pigment. The extract analyses showed that the sturgeon retina has only a single rod pigment (maximum absorbance 545nm) which is a porphyropsin based on vitamin A(subscript 2). There was no sign of vitamin A(subscript 1) based rhodopsin. The sturgeon porphyropsin exhibited no chloride sensitivity and was found to be stable in the presence of hydroxylamine.

Impacts
(N/A)

Publications

  • SILLMAN, A.J. (1987). Extraction and characterization of the blue-sensitive visual pigment of the toad Bufo marinus, Can. J. Zool. 65:884-887.