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Constancy of DNA content in adrenal medulla nuclei of cold-treated rats.

Reports of changes in DNA content of certain types of cells following exposure to conditions of stress has led to the suggestion that two kinds of DNA may be present. One is genetic DNA, and the other is called "metabolic" DNA. In a further attempt to investigate the possibility of this phenomenon, determinations of DNA content were made on Feulgen-stained nuclei of adrenal glands and kidneys in cold-treated rats. Feulgen-stained nuclei were measured by two-wavelength microspectrophotometry. Particular attention was given to the handling of the smears in hydrolysis and staining. Mean values of Feulgen-DNA contents in a total of 720 nuclei demonstrated (a) a constancy of DNA content within 2% in individual nuclei both in adrenal medulla and kidney cortex, (b) no more than an average of 2% difference in DNA content between control and experimental nuclei, and (c) no more than an average of 1.5% difference in DNA content between normal kidney cortex nuclei and normal adrenal medulla nuclei. These results confirm the view that the more precise the measurement, the more accurately the constancy rule is obeyed. Moreover, there is no support for the concept of a metabolic DNA in the rat adrenal medulla.

Adrenal Medulla↗

DNA-protein binding in interphase chromosomes.

The metachromatic dye, azure B, was analyzed by microspectrophotometry when bound to DNA fibers and DNA in nuclei with condensed and dispersed chromatin. The interaction of DNA and protein was inferred from the amount of metachromasy (increased beta/alpha-peak) of azure B that resulted after specific removal of various protein fractions. Dye bound to DNA-histone fibers and frog liver nuclei fixed by freeze-methanol substitution shows orthochromatic, blue-green staining under specific staining conditions, while metachromasy (blue or purple color) results from staining DNA fibers without histone or tissue nuclei after protein removal. The dispersed chromatin of hepatocytes was compared to the condensed chromatin of erythrocytes to see whether there were differences in DNA-protein binding in "active" and "inactive" nuclei. Extraction of histones with 0.02 N HCl, acidified alcohol, perchloric acid, and trypsin digestion all resulted in increased dye binding. The amount of metachromasy varied, however; removal of "lysine-rich" histone (extractable with 0.02 N HCl) caused a blue color, and a purplish-red color (micro-peak absorption) resulted from prolonged trypsin digestion. In all cases, the condensed and the dispersed chromatin behaved in the same way, indicating the similarity of protein bound to DNA in condensed and dispersed chromatin. The results appear to indicate that "lysine-rich" histone is bound to adjacent anionic sites of a DNA molecule and that nonhistone protein is located between adjacent DNA molecules in both condensed and dispersed chromatin.

Animals↗

The nature, significance, and evaluation of the Schwarzschild-Villiger (SV) effect in photometric procedures.

The Schwarzschild-Villiger effect has been experimentally demonstrated with the optical system used in this laboratory. Using a photographic mosaic specimen as a model, it has been shown that the conclusions of Naora are substantiated and that the SV effect, in large or small magnitude, is always present in optical systems. The theoretical transmission error arising from the presence of the SV effect has been derived for various optical conditions of measurement. The results have been experimentally confirmed. The SV contribution of the substage optics of microspectrophotometers has also been considered. A simple method of evaluating a flare function f(A) is advanced which provides a measure of the SV error present in a system. It is demonstrated that measurements of specimens of optical density less than unity can be made with less than 1 per cent error, when using illuminating beam diameter/specimen diameter ratios of unity and uncoated optical surfaces. For denser specimens it is shown that care must be taken to reduce the illuminating beam/specimen diameter ratio to a value dictated by the magnitude of a flare function f(A), evaluated for a particular optical system, in order to avoid excessive transmission error. It is emphasized that observed densities (transmissions) are not necessarily true densities (transmissions) because of the possibility of SV error. The ambiguity associated with an estimation of stray-light error by means of an opaque object has also been demonstrated. The errors illustrated are not necessarily restricted to microspectrophotometry but may possibly be found in such fields as spectral analysis, the interpretation of x-ray diffraction patterns, the determination of ionizing particle tracks and particle densities in photographic emulsions, and in many other types of photometric analysis.

Coloring Agents↗

Cytological and cytochemical studies of HeLa cells infected with adeno-viruses.

The sequential morphological and cytochemical alterations in HeLa cells infected with adenovirus types 1 to 4 are described. Each of the four viruses studied led to consistent and reproducible cytological changes not observed in uninfected control cultures. All four agents produced striking and characteristic changes in the nuclei of infected cells. Alterations in the cytoplasm, though present, were less marked, particularly in the early stages of infection. In cells infected with type 1 or 2 adenovirus, rounded intranuclear inclusions which progressed from eosinophilic and Feulgen-negative to basophilic and Feulgen-positive, together with a homogeneous glassy, Feulgen-positive nuclear background, were prominent features. Cells infected with type 3 or 4 adenovirus exhibited marked rearrangement of basophilic nuclear material and sharply defined crystal-like inclusions, predominantly intranuclear in location, which also varied from Feulgen-negative to positive. In terms of detailed cytological effects, therefore, the four agents could be divided into two subgroups, viz., types 1 and 2 on the one hand and types 3 and 4 on the other. Measurement of DNA in individual nuclei by means of Feulgen microspectrophotometry revealed the values in infected cells to be increased above the levels of the uninfected controls.

Adenoviridae↗

Rhodopsin of the larval mosquito.

Larvae of the mosquito Aedes aegypti have a cluster of four ocelli on each side of the head. The visual pigment of each ocellus of mosquitoes reared in darkness was characterized by microspectrophotometry, and found to be the same. Larval mosquito rhodopsin (lambda(max) = 515 nm) upon short irradiation bleaches to a stable photoequilibrium with metarhodopsin (lambda(max) = 480 nm). On long irradiation of glutaraldehyde-fixed tissues or in the presence of potassium borohydride, bleaching goes further, and potassium borohydride reduces the product, retinal, to retinol (vitamin A(1)). In the presence of hydroxylamine, the rhodopsin bleaches rapidly, with conversion of the chromophore to retinaldehyde oxime (lambda(max) about 365 nm).

Aedes↗

Characteristics of Drosophila rhodopsin in wild-type and norpA vision transduction mutants.

The properties of the major visual pigment of Drosophila melanogaster were evaluated. The visual pigment was isolated from other protein components using acrylamide gel electrophoresis and spectral identification. Sodium dodecyl sulfate (SDS) acrylamide gels of the isolated visual pigment gave a single protein subunit with a mol wt of 37,000 daltons. The rhodopsin480 molar extinction coefficient was 35,000 liter/mol-cm (+/- 2,700 SE). The metarhodopsin580 molar extinction coefficient was approximately 56,000 liter/mol-cm. Microspectrophotometry was used to compare the rhodopsin concentrations in wild-type flies and norpA vision transduction mutants. At 2 days of age (12 h dark-12 h light cycle, 19 degrees C) all of the norpA flies exhibited a similar rhodopsin concentration (75% of the wild-type strain). By 21 days of age some of the norpA alleles showed substantially reduced rhodopsin concentrations (16-43% of normal), whereas others showed no major age-dependent decreases (68-77%). Temperature and light-dark cycle affected the reduction. Alleles with no receptor potential exhibited the largest decreases in rhodopsin concentration. The data indicate that the norpA phototransduction mutant has a defect in the system responsible for maintaining the rhodopsin480 concentration. This defect in the rhodopsin maintenance system does not appear to be the cause of the reduced electroretinogram (ERG) amplitude observed in some of these mutants, but instead is a consequence of the decrease in ERG amplitude, or the flaw(s) responsible for the decrease in ERG amplitude.

Age Factors↗

Euphausiid visual pigments. The rhodopsins of Euphausia superba and Meganyctiphanes norvegica (Crustacea, Euphausiacea).

The rhabdoms of Euphausia superba contain one digitonin-extractable rhodopsin, lambda max 485 nm. The rhodopsin undergoes unusual pH-dependent spectral changes: above neutrality, the absorbance decreases progressively at 485 nm and rises near 370 nm. This change is reversible and appears to reflect an equilibrium between a protonated and an unprotonated form of the rhodopsin Schiff-base linkage. Near neutral pH and at 10 degrees C, the rhodopsin is partiaLly converted by 420-nm light to a stable 493-nm metarhodopsin. The metarhodopsin is partially photoconverted to rhodopsin by long-wavelength light in the absence of NH2OH; in the presence of NH2OH, it is slowly converted to retinal oxime and opsin. The rhodopsin of Meganyctiphanes norvegica measured in fresh rhabdoms by microspectrophotometry has properties very similar to those of the extracted rhodopsin of E. superba. Its lambda max is 488 nm and it is partially photoconverted by short wavelength irradiation to a stable photoconvertible metarhodopsin similar to that of E. superba. In the presence of light and NH2OH, the M. norvegica metarhodopsin is converted to retinal oxime and opsin. Our results indicate that previous determinations of euphausiid rhodopsin absorbance spectra were incorrect because of accessory pigment contamination.

Animals↗

Experimental validation of arthroscopic cartilage stiffness measurement using enzymatically degraded cartilage samples.

In order to evaluate the ability of the arthroscopic indentation instrument, originally developed for the measurement of cartilage stiffness during arthroscopy, to detect cartilage degeneration, we compared changes in the stiffness with the structural and constitutional alterations induced by enzymes on the tissue in vitro. The culturing of osteochondral plugs on Petri dishes was initiated in Minimum Essential Medium with Earle's salts and the baseline stiffness was measured. Then, the experimental specimens were digested using 50 microg ml(-1) trypsin for 24 h, 0.1 U ml(-1) chondroitinase ABC or 30 U ml(-1) purified collagenase (type VII) for 24 h or 48 h (n = 8-15 per group). The control specimens were incubated in the medium. After the enzyme digestion, the end-point stiffness was measured and the specimens for the microscopic analyses were processed. The proteoglycan (PG) distribution was analysed using quantitative microspectrophotometry and the quantitative evaluation of the collagen network was made using a computer-based polarized light microscopy analysis. Decrease (p < 0.05) of cartilage stiffness was found after 24 h trypsin (36%) and 48 h chondroitinase ABC (24%) digestion corresponding to a decrease (p < 0.01) of up to 80% and up to 30% in the PG content respectively. Decrease of the superficial zone collagen content or arrangement (78%, p < 0.001) after 48 h collagenase digestion also induced a decrease (30%, p < 0.001) in cartilage stiffness. We conclude that our instrument is capable of detecting early structural and compositional changes related to cartilage degeneration.

Animals↗

Effect of local heating and cooling on cambial activity and cell differentiation in the stem of Norway spruce (Picea abies).

UNLABELLED: BACKGROUND AND AIMS The effect of heating and cooling on cambial activity and cell differentiation in part of the stem of Norway spruce (Picea abies) was investigated. METHODS: A heating experiment (23-25 degrees C) was carried out in spring, before normal reactivation of the cambium, and cooling (9-11 degrees C) at the height of cambial activity in summer. The cambium, xylem and phloem were investigated by means of light- and transmission electron microscopy and UV-microspectrophotometry in tissues sampled from living trees. KEY RESULTS: Localized heating for 10 d initiated cambial divisions on the phloem side and after 20 d also on the xylem side. In a control tree, regular cambial activity started after 30 d. In the heat-treated sample, up to 15 earlywood cells undergoing differentiation were found to be present. The response of the cambium to stem cooling was less pronounced, and no anatomical differences were detected between the control and cool-treated samples after 10 or 20 d. After 30 d, latewood started to form in the sample exposed to cooling. In addition, almost no radially expanding tracheids were observed and the cambium consisted of only five layers of cells. Low temperatures reduced cambial activity, as indicated by the decreased proportion of latewood. On the phloem side, no alterations were observed among cool-treated and non-treated samples. CONCLUSIONS: Heating and cooling can influence cambial activity and cell differentiation in Norway spruce. However, at the ultrastructural and topochemical levels, no changes were observed in the pattern of secondary cell-wall formation and lignification or in lignin structure, respectively.

Cell Differentiation↗

Regional O2 consumption in canine left ventricular myocardium in experimental acute aortic valvular insufficiency.

Moderate and severe acute aortic valvular insufficiency (AVI) were produced for 30 min in anesthetised, open-chest dogs, in order to measure O2 consumption in subepicardial (EPI) and subendocardial (ENDO) regions of the left ventricular free wall. Regional O2 consumption (MVO2) was determined from regional O2 extraction and blood flow data determined by absorbence microspectrophotometry and radioactive microspheres. In all groups, venous saturations were lower and O2 extractions higher in the ENDO than in the EPI. Oxygen extractions increased in both regions in both severities of AVI, but more so with severe AVI. The ENDO became perfused relative to the EPI with increasing severity of AVI. Left ventricular MVO2 increased with AVI; however, ENDO MVO2 increased less than EPI MVO2. ENDO/EPI blood flow ratios decreased from 1.21 to 0.93 in moderate AVI and to 0.76 in severe AVI. ENDO/EPI MVO2 ratio decreased from 1.44 in controls to 0.94 in moderate and 0.54 in severe AVI. The ENDO/EPI MVO2 ratios began to decrease when DPTI/SPTI was less than 0.4 to 0.5. We conclude that AVI increases MVO2, but the ENDO cannot maintain its MVO2 relative to EPI because of proportionately less ENDO flow, and maximal O2 extraction.

Acute Disease↗

Effect of aortic constriction on the oxygen supply-consumption balance in partially ischaemic myocardium.

A study was carried out to determine the effect of distal aortic constriction on the regional oxygen supply-consumption balance in ischaemic and non-ischaemic myocardium. Left anterior descending coronary artery flow was reduced by 50% in anaesthetised mongrel dogs, and half these animals were subjected to distal ascending aortic constriction (ventricular systolic-aortic systolic pressure gradient 50 mm Hg). Blood flows were determined using radioactive microspheres, and arterial and venous oxygen saturations were determined using microspectrophotometry. When left anterior descending artery flow was decreased by 50% tissue flow, as measured by microspheres in the ischaemic region in both control and aortic constricted animals, decreased. Aortic constriction increased flow in the non-ischaemic zone compared with the same region in control animals. The arteriovenous oxygen content difference increased similarly in the ischaemic region in control and aortic constricted animals compared with their respective non-ischaemic zones. Aortic constriction decreased this difference in the non-ischaemic region compared with these same regions in control animals. Oxygen consumption in the non-ischaemic and ischaemic regions of aortic constricted animals was higher than in control animals. The oxygen supply to consumption ratio was lower in the ischaemic zone in both control and aortic constricted animals than in their respective non-ischaemic regions. Aortic constriction does not appear to alter the oxygen supply-consumption balance in the ischaemic myocardium compared with control animals. Thus the increased work performed by the heart in response to the aortic constriction is adequately matched by an increased flow even in the flow restricted region.

Animals↗

Relationship between local oxygen consumption and local and external cardiac work: effect of tachycardia.

This study was designed to determine the extent of coupling between regional myocardial segment work and corresponding regional oxygen consumption, and to examine whether tachycardia induced changes in regional work are translated into corresponding changes in external cardiac work. In the open chest anaesthetised dog, the heart was paced at frequencies of 120-270 beats.min-1. Global and regional myocardial O2 supply, consumption, and balance were evaluated at each heart rate, and correlated with corresponding functional changes. Global cardiac function was evaluated from aortic flow, blood pressure, and left ventricular pressure. Coronary sinus flow and O2 saturation were used to calculate O2 consumption. The integrated multiple of myocardial shortening (ultrasonic dimension crystals) by corresponding force (strain gauge arch) during an averaged beat was used to express regional segment work. Regional coronary blood flow was measured with radioactive microspheres, and microspectrophotometry was used to evaluate O2 saturation in small arteries and veins. These indices were used to calculate regional myocardial oxygen consumption. NADH redox levels were recorded by surface fluorometry, and were found to increase with heart rate by up to 67%. Increasing heart rate from 120 to 180 beats.min-1 increased regional work from 3040(SEM 220) to a peak of 4290(280) mm.g-1.min-1, whereas external cardiac work did not increase [67.0(2.6) to 65.3(4.4) mm Hg.litre-1.min-1] and fell further at the highest rates. Regional oxygen consumption increased from 6.16(0.47) to 8.29(0.53) ml O2.min-1.100 g-1 and was linearly related to regional work at all heart rates (r = 0.971, p less than 0.05). External cardiac work fell by about 26% whereas global myocardial oxygen consumption increased by 49% during tachycardia. It is concluded that myocardial oxygen consumption is more closely related to regional segment work than to external work, and that tachycardia significantly raises the oxygen cost of external work of the heart.

Animals↗

Augmented efficiency of regional myocardial work by ouabain.

STUDY OBJECTIVE: The effect of ouabain on regional myocardial mechanics and corresponding regional myocardial oxygen consumption was investigated. DESIGN: Regional myocardial segment length (ultrasonic dimension crystals) and force development (miniature strain gauge) were measured. The integrated multiples of myocardial shortening by corresponding force during an averaged beat was used to express regional segment work (= area under the systolic portion of the length-force loop). External cardiac work was calculated from aortic blood pressure and cardiac output. Global and regional myocardial O2 consumption (MVO2) were evaluated at baseline and during intravenous infusion of ouabain (7 micrograms.kg-1.min-1). Regional coronary blood flow was measured with radioactive microspheres, and microspectrophotometry of frozen myocardial biopsies was used to evaluate O2 saturation in small arteries and veins. These variables were used to calculate regional myocardial MVO2. Arterial and coronary sinus O2 saturation was used to calculate global left ventricular O2 consumption. Myocardial efficiency was expressed as regional work/regional myocardial MVO2. SUBJECTS: Ten open chest anaesthetised dogs, weight about 20 kg, were used. MEASUREMENTS AND MAIN RESULTS: Ouabain increased regional myocardial segment work [from 5291(SEM 1067) to 8916(2338) mm.g-1.min-1] to a greater extent than regional myocardial MVO2 [from 6.33(0.75) to 8.54(0.75) ml O2.min-1.100 g-1]. Regional efficiency was increased from 734(78) to 1036(160) g.mm.ml O2(-1).100 g-1. External efficiency was not raised. Percent shortening was increased to a greater extent than peak force development (+76% v + 38%), possibly indicating increased myocardial compliance. The length-force loop was shifted upward and to the right. CONCLUSIONS: Ouabain increases regional efficiency of myocardial contraction.

Animals↗

Local adenosine infusion and myocardial oxygen supply and consumption in reperfused canine myocardium.

STUDY OBJECTIVE: The aim was to determine whether a local infusion of adenosine could increase coronary blood flow to restore fully the O2 supply to consumption balance in reperfused myocardium. DESIGN: The left anterior descending coronary artery was occluded for 2 h followed by a 4 h reperfusion with and without simultaneous local infusion of adenosine (50 micrograms.kg-1.min-1). In another group, adenosine was infused for 4 h. Small artery and vein O2 saturations (microspectrophotometry) and regional flow (radioactive microspheres) were determined. SUBJECTS: 25 anaesthetised open chest mongrel dogs, divided in three groups, were used in this study. MEASUREMENT AND MAIN RESULTS: In the central group, 2 h occlusion decreased regional blood flow below normal. After 4 h of reperfusion, blood flow was less than normal in the affected area, at 57(SD39) affected v 85(31) ml.min-1.100 g-1 control. O2 extraction was higher in the reperfused area, with 61% of veins having O2 saturations less than 35%. For the experimental group, adenosine was infused into the left anterior descending artery at a dose of 50 micrograms.kg-1.min-1. Adenosine coupled with reperfusion increased regional blood flow above control in the affected area, to 267(238) ml.min-1.100 g-1, but this was less than was found with adenosine alone. O2 extraction in this area was lower than with reperfusion alone, with only 1% of veins having O2 saturations less than 35%. CONCLUSIONS: A large flow reserve is present in reperfused tissue and the low O2 saturations found in reperfused myocardium cannot be due to vessel blockage (greater than 20 microns in diameter). Thus local adenosine can be used to mobilise the reserve to improve the O2 supply to consumption balance.

Adenosine↗

Milrinone, a cyclic AMP-phosphodiesterase inhibitor, has differential effects on regional myocardial work and oxygen consumption in experimental left ventricular hypertrophy.

OBJECTIVE: The aim was to test the hypothesis that local myocardial work and O2 consumption would respond differentially to milrinone, a selective cyclic AMP-phosphodiesterase inhibitor, in left ventricular hypertrophy due to differences in myocardial cyclic AMP-phosphodiesterase activity. METHODS: The effect of milrinone on regional segment work and regional O2 consumption was measured in 12 open chest anaesthetised dogs with left ventricular hypertrophy induced by valvular aortic stenosis and in 10 age matched control dogs. Regional myocardial work was calculated as the integrated product of instantaneous force development (miniature transducer) and segment shortening (sonomicrometer). Regional O2 consumption was calculated from coronary blood flow (radiolabelled microspheres) and O2 saturations in small regional vessels (microspectrophotometry). Low Km phosphodiesterase activity was assayed by measuring the hydrolysis of radiolabelled cyclic AMP. RESULTS: Milrinone increased left ventricular dP/dtmax by approximately 60-70% in both control [2808(SEM 314) to 4584(660) mm Hg.s-1] and left ventricular hypertrophy [3279(258) to 5589(470) mm-Hg.s-1]. Regional work increased significantly in control [612(88) to 955(101) g.mm.min-1], while the increase was not significant in left ventricular hypertrophy [859(139) to 974(172) g.mm.min-1]. Regional O2 consumption increased significantly with milrinone in left ventricular hypertrophy [8.1(1.2) to 13.1(2.4) ml O2.min-1.100 g-1], but the increase was not significant in control [6.9(1.2) to 7.4(1.0) ml O2.min-1.100 g-1]. Myocardial stiffness during ejection was increased by milrinone to a significantly greater extent in animals with left ventricular hypertrophy. These effects were not related to differences in cyclic AMP-phosphodiesterase activity between control hearts and hearts with left ventricular hypertrophy [393(45) v 402(36) pmol.mg protein-1.1]. CONCLUSIONS: Differences between the hypertrophied and normal canine myocardium in response to milrinone are either due to altered levels of cyclic AMP production in left ventricular hypertrophy, to effects of milrinone that are unrelated to cyclic AMP-phosphodiesterase inhibition, or to other differences in hypertrophied hearts. The greater stiffness of the myocardium in left ventricular hypertrophy may require a greater energy expenditure to increase the amount of work it performs.

3',5'-Cyclic-AMP Phosphodiesterases↗

Karyotype, C- and G-band patterns and DNA content of Callimenus (=Bradyporus) macrogaster macrogaster.

Chromosomes and detailed karyotype information (the number, shape, relative length, arm ratio, centromeric index) of Callimenus (=Bradyporus) macrogaster macrogaster Lef. (Orthoptera: Tettigonioidea, Bradyporini) of Turkey belonging to the subfamily Bradyporinae are described. The diploid number of chromosomes was found to be 2n male symbol = 23, with 2 metacentric pairs, 2 submetacentric pairs, 6 acrocentric pairs. The X chromosome is metacentric. This species has a XX female symbol / X0 male symbol sex determining mechanism. The basic karyotype is complemented by a description of C- and G-banding patterns. The C-banding pattern in spermatogonial metaphase was characterized by the presence of paracentromeric C-bands in all chromosomes, and a distal C-band in chromosome 5. The G-banding pattern was complex. The 2C nuclear DNA content was found to be 10.26 +/- 0.16 picograms by microspectrophotometry.

Animals↗

Complex distribution of avian color vision systems revealed by sequencing the SWS1 opsin from total DNA.

To gain insights into the evolution and ecology of visually acute animals such as birds, biologists often need to understand how these animals perceive colors. This poses a problem, since the human eye is of a different design than that of most other animals. The standard solution is to examine the spectral sensitivity properties of animal retinas through microspectophotometry-a procedure that is rather complicated and therefore only has allowed examinations of a limited number of species to date. We have developed a faster and simpler molecular method, which can be used to estimate the color sensitivities of a bird by sequencing a part of the gene coding for the ultraviolet or violet absorbing opsin in the avian retina. With our method, there is no need to sacrifice the animal, and it thereby facilitates large screenings, including rare and endangered species beyond the reach of microspectrophotometry. Color vision in birds may be categorized into two classes: one with a short-wavelength sensitivity biased toward violet (VS) and the other biased toward ultraviolet (UVS). Using our method on 45 species from 35 families, we demonstrate that the distribution of avian color vision is more complex than has previously been shown. Our data support VS as the ancestral state in birds and show that UVS has evolved independently at least four times. We found species with the UVS type of color vision in the orders Psittaciformes and Passeriformes, in agreement with previous findings. However, species within the families Corvidae and Tyrannidae did not share this character with other passeriforms. We also found UVS type species within the Laridae and Struthionidae families. Raptors (Accipitridae and Falconidae) are of the violet type, giving them a vision system different from their passeriform prey. Intriguing effects on the evolution of color signals can be expected from interactions between predators and prey. Such interactions may explain the presence of UVS in Laridae and Passeriformes.

Amino Acid Sequence↗

The cone visual pigments of an Australian marsupial, the tammar wallaby (Macropus eugenii): sequence, spectral tuning, and evolution.

Studies on marsupial color vision have been limited to very few species. There is evidence from behavioral, electroretinographic (ERG), and microspectrophotometric (MSP) measurements for the existence of both dichromatic and trichromatic color vision. No studies have yet investigated the molecular mechanisms of spectral tuning in the visual pigments of marsupials. Our study is the first to determine the mRNA sequence, infer the amino acid sequence, and determine, by in vitro expression, the spectra of the cone opsins of a marsupial, the tammar wallaby (Macropus eugenii). This yielded some information on mechanisms and evolution of spectral tuning of these pigments. The tammar wallaby retina contains only short-wavelength sensitive (SWS) and middle-wavelength sensitive (MWS) pigment mRNAs. This predicts dichromatic color vision, which is consistent with conclusions from previous behavioral studies ( Hemmi 1999). We found that the wallaby has a SWS1 class pigment of 346 amino acids. Sequence comparison with eutherian SWS pigments predicts that this SWS1 pigment absorbs maximally (lambdamax) at 424 nm and, therefore, is a blue rather than a UV pigment. This (lambdamax) is close to that of the in vitro-expressed wallaby SWS pigment (lambdamax of 420 +/- 2 nm) and to that determined behaviorally (420 nm). The difference from the mouse UV pigment (lambdamax of 359 nm) is largely accounted for by the F86Y substitution, in agreement with in vitro results comparing a variety of other SWS pigments. This suggests that spectral tuning employing F86Y substitution most likely arose independently in the marsupials and ungulates as a result of convergent evolution. An apparently different mechanism of spectral tuning of the SWS1 pigments, involving five amino acid positions, evolved in primates. The wallaby MWS pigment has 363 amino acids. Species comparisons at positions critical to spectral tuning predict a lambdamax near 530 nm, which is close to that of the in vitro-expressed pigment (529 +/- 1 nm), but quite different from the value of 539 nm determined by microspectrophotometry. Introns interrupt the coding sequences of the wallaby, mouse, and human MWS pigment sequences at the same corresponding nucleotide positions. However, the length of introns varies widely among these species.

Amino Acid Sequence↗