Development of 14 microsatellite markers in the Queensland koala (Phascolarctos cinereus adustus) using next generation sequencing technology

We report the development of 14 new microsatellite markers in the Queensland koala (Phascolarctos cinereus adustus). Ten unrelated Queensland koala individuals from the San Diego Zoo, USA, were genotyped. The number of alleles per locus ranged from 2 to 7, with an average of 5.14 alleles per locus. Across all loci, the average observed and expected heterozygosity values were both 0.69. These polymorphic microsatellite loci will be useful for genetic studies relevant to the conservation of the koala, a species listed as vulnerable.     

Retinal ganglion cell topography in elasmobranchs

Retinal wholemounts are used to examine the topographic distribution of retinal cells within the ganglion cell layer in a range of elasmobranchs from different depths. The retina is examined for regional specializations for acute vision in six species of selachians, Galeocerdo cuvieri, Hemiscyllium ocellatum, Scyliorhinus canicula, Galeus melastomus, Etmopterus spinax, Isistius brasiliensis, one species of batoid, Raja bigelowi and one species of chimaera, Hydrolagus mirabilis. These species represent a range of lifestyles including pelagic, mesopelagic and benthic habitats, living from shallow water to the sea bottom at a depth of more than 3000 m. The topography of cells within the ganglion cell layer is non-uniform and changes markedly across the retina. Most species possess an increased density of cells across the horizontal (dorsal) meridian or visual streak, with a density range of 500 to 2,500 cells per mm(2) with one or more regional increases in density lying within this specialized horizontal area. It is proposed that the higher spatial resolving power provided by the horizontal streak in these species mediates panoramic vision in the lower frontal visual field. Only I. brasiliensis possesses a concentric arrangement of retinal iso-density contours in temporal retina or an area centralis, thereby increasing spatial resolving power in a more specialized part of the visual field, an adaptation for its unusual feeding behavior. In Nissl-stained material, amacrine and ganglion cell populations could be distinguished on the criteria of soma size, soma shape and nuclear staining. Quantitative analyses show that the proportion of amacrine cells lying within the ganglion cell layer is non-uniform and ranges between 0.4 and 12.3% in specialized retinal areas and between 8.2 and 48.1% in the peripheral non-specialized regions. Analyses of soma area of the total population of cells in the ganglion cell layer also show that the pelagic species possess significantly smaller soma (9-186 micrometer(2)) than benthic and/or deep-sea species (16-338 micrometer(2)), and that a number of different morphological classes of cells are present including a small population of giant ganglion cells.     

Retinal topography of the harp seal Pagophilus groenlandicus

The total number, size, topographic distribution, and cell density of ganglion cells were studied in retinal wholemounts of the harp seal Pagophilus groenlandicus. Ganglion cell size varied from 10 to 60 mum. A distinct group were large ganglion cells more than 30-35 mum in diameter which were similar to alpha-cells known in terrestrial mammals. The number of alpha-cells constituted 5.3-5.9% of the total ganglion cell population. The cell size distribution was bimodal, with the second mode composed of alpha-cells. The topographic distribution of ganglion cells showed a definite area of high cell density similar to area centralis of terrestrial carnivores. This area was located in the temporal retinal quadrant, 7-8 mm (16-18 degrees ) from the optic disk. In this area, the peak cell densities in two wholemounts were 2,500 and 1,650 (mean 2,075) cells/mm(2). With a posterior nodal distance of 25.5 mm (underwater), this density corresponded to 495 and 327 (mean 411) cells/deg(2). These values predict a retinal resolution of 2.7-3.3' (11.1-9.0 cycles/deg) in water and 3.6-4.4' (8.3-6.8 cycles/deg) in air. Topographic distributions of alpha-cells was qualitatively similar to that of the total ganglion cell population, but the density of alpha-cells constituted only a few percent (mostly 3-7.5%) of the total ganglion cell density.     

Retinal topography in two species of flamingo (Phoenicopteriformes: Phoenicopteridae)

In this study, we assessed eye morphology and retinal topography in two flamingo species, the Caribbean flamingo (Phoenicopterus ruber) and the Chilean flamingo (P. chilensis). Eye morphology is similar in both species and cornea size relative to eye size (C:A ratio) is intermediate between those previously reported for diurnal and nocturnal birds. Using stereology and retinal whole mounts, we estimate that the total number of Nissl-stained neurons in the retinal ganglion cell (RGC) layer in the Caribbean and Chilean flamingo is ~1.70 and 1.38 million, respectively. Both species have a well-defined visual streak with a peak neuron density of between 13,000 and 16,000 cells mm⁻² located in a small central area. Neurons in the high-density regions are smaller and more homogeneous compared to those in medium- and low-density regions. Peak anatomical spatial resolving power in both species is approximately 10–11 cycles/deg. En-face images of the fundus in live Caribbean flamingos acquired using spectral domain optical coherence tomography (SD-OCT) revealed a thin, dark band running nasotemporally just dorsal to the pecten, which aligned with the visual streak in the retinal topography maps. Cross-sectional images (B-scans) obtained with SD-OCT showed that this dark band corresponds with an area of retinal thickening compared to adjacent areas. Neither the retinal whole mounts, nor the SD-OCT imaging revealed any evidence of a central fovea in either species. Overall, we suggest that eye morphology and retinal topography in flamingos reflects their cathemeral activity pattern and the physical nature of the habitats in which they live.     

Retinal topography and spectral sensitivity of the Port Jackson shark (Heterodontus portusjacksoni)

In this study, we investigated the visual system of the Port Jackson shark Heterodontus portusjacksoni, a shallow-dwelling benthic species and generalist predator endemic to the temperate coastal waters around southern Australia. Measurements of retinal spectral sensitivity in juvenile sharks, made using single flash and heterochromatic flicker photometry under conditions of dark- or light-adaptation, indicated a peak sensitivity at around 500 nm, with no evidence of a spectral shift with increasing levels of light adaptation. Histological sections of the retina revealed a heavily rod dominated retina containing only a few small cell profiles in the photoreceptor layer that might represent a sparse cone population or may be immature rods. Assessment of retinal topography in juvenile sharks indicated the presence of a distinct specialisation for increased visual spatial acuity in the form of a horizontal streak of higher rod photoreceptor (~80,000 rods mm⁻²) and ganglion cell (~1,800 cells mm⁻²) densities across the horizontal meridian of the eye. This specialization would be adaptive for panoramic sampling of the part of the visual field corresponding to the substrate-water interface and remove the need for H. portusjacksoni to move its eyes extensively when resting on the sea floor. The estimated upper limit of spatial resolving power in juvenile H. portusjacksoni was 3.14 cycles deg⁻¹, which is at the lower end of values measured in elasmobranchs. Taken together, these results suggest that the retina of H. portusjacksoni is well adapted for nocturnal vision.     

Retinal ganglion cell topography and spatial resolving power in the river hippopotamus (Hippopotamus amphibius)

The river hippopotamus (Hippopotamus amphibius), one of the closest extant relatives to cetaceans, is a large African even‐toed ungulate (Artiodactyla) that grazes and has a semiaquatic lifestyle. Given its unusual phenotype, ecology, and evolutionary history, we sought to measure the topographic distribution of retinal ganglion cell density using stereology and retinal wholemounts. We estimated a total of 243,000 ganglion cells of which 3.4% (8,300) comprise alpha cells. The topographic distribution of both total and alpha cells reveal a dual topographic organization of a temporal and nasal area embedded within a well‐defined horizontal streak. Using maximum density of total ganglion cells and eye size (35 mm, axial length), we estimated upper limits of spatial resolving power of 8 cycles/deg (temporal area, 1,800 cells/mm²), 7.7 cycles/deg (nasal area, 1,700 cells/mm²), and 4.2 cycles/deg (horizontal streak, 250 cells/mm²). Enhanced resolution of the temporal area toward the frontal visual field may facilitate grazing, while resolution of the horizontal streak and nasal area may help the discrimination of objects (predators, conspecifics) in the lateral and posterior visual fields, respectively. Given the presumed role of alpha cells to detect brisk transient stimuli, their similar distribution to the total ganglion cell population may facilitate the detection of approaching objects in equivalent portions of the visual field. Our finding of a nasal area in the river hippopotamus retina supports the notion that this specialization may enhance visual sampling in the posterior visual field to compensate for limited neck mobility as suggested for rhinoceroses and cetaceans.     

Retinal ganglion cell topography and spatial resolving power in the white rhinoceros (Ceratotherium simum)

This study sought to determine whether the retinal organization of the white rhinoceros (Ceratotherium simum), a large African herbivore with lips specialized for grazing in open savannahs, relates to its foraging ecology and habitat. Using stereology and retinal wholemounts, we estimated a total of 353,000 retinal ganglion cells. Their density distribution reveals an unusual topographic organization of a temporal (2,000 cells/mm²) and a nasal (1,800 cells/mm²) area embedded within a well‐defined horizontal visual streak (800 cells/mm²), which is remarkably similar to the retinal organization in the black rhinoceros. Alpha ganglion cells comprise 3.5% (12,300) of the total population of ganglion cells and show a similar distribution pattern with maximum densities also occurring in the temporal (44 cells/mm²) and nasal (40 cells/mm²) areas. We found higher proportions of alpha cells in the dorsal and ventral retinas. Given their role in the detection of brisk transient stimuli, these higher proportions may facilitate the detection of approaching objects from the front and behind while grazing with the head at 45 °. Using ganglion cell peak density and eye size (29 mm, axial length), we estimated upper limits of spatial resolving power of 7 cycles/deg (temporal area), 6.6 cycles/deg (nasal area), and 4.4 cycles/deg (horizontal streak). The resolution of the temporal area potentially assists with grazing, while the resolution of the streak may be used for panoramic surveillance of the horizon. The nasal area may assist with detection of approaching objects from behind, potentially representing an adaptation compensating for limited neck and head mobility. J. Comp. Neurol., 525:2484–2498, 2017. © 2017 Wiley Periodicals, Inc.     

Retinal topography and microhabitat diversity in a group of dragon lizards

The well-studied phylogeny and ecology of dragon lizards and their range of visually mediated behaviors provide an opportunity to examine the factors that shape retinal organization. Dragon lizards consist of three evolutionarily stable groups based on their shelter type, including burrows, shrubs, and rocks. This allows us to test whether microhabitat changes are reflected in their retinal organization. We examined the retinae of three burrowing species (Ctenophorus pictus, C. gibba, and C. nuchalis), and three species that shelter in rock crevices (C. ornatus, C. decresii, and C. vadnappa). We used design-based stereology to sample both the photoreceptor array and neurons within the retinal ganglion cell layer to estimate areas specialized for acute vision. All species had two retinal specializations mediating enhanced spatial acuity: a fovea in the retinal center and a visual streak across the retinal equator. Furthermore, all species featured a dorsoventrally asymmetric photoreceptor distribution with higher photoreceptor densities in the ventral retina. This dorsoventral asymmetry may provide greater spatial summation of visual information in the dorsal visual field. Burrow-dwelling species had significantly larger eyes, higher total numbers of retinal cells, higher photoreceptor densities in the ventral retina, and higher spatial resolving power than rock-dwelling species. C. pictus, a secondary burrow-dwelling species, was the only species that changed burrow usage over evolutionary time, and its retinal organization revealed features more similar to rock-dwelling species than other burrow-dwelling species. This suggests that phylogeny may play a substantial role in shaping retinal organization in Ctenophorus species compared to microhabitat occupation.     

Scene from above: Retinal ganglion cell topography and spatial resolving power in the giraffe (Giraffa camelopardalis)

The giraffe (Giraffa camelopardalis) is a browser that uses its extensible tongue to selectively collect leaves during foraging. As the tallest extant terrestrial mammal, its elevated head height provides panoramic surveillance of the environment. These aspects of the giraffe's ecology and phenotype suggest that vision is of prime importance. Using Nissl‐stained retinal wholemounts and stereological methods, we quantitatively assessed the retinal specializations in the ganglion cell layer of the giraffe. The mean total number of retinal ganglion cells was 1,393,779 and their topographic distribution revealed the presence of a horizontal visual streak and a temporal area. With a mean peak of 14,271 cells/mm², upper limits of spatial resolving power in the temporal area ranged from 25 to 27 cycles/degree. We also observed a dorsotemporal extension (anakatabatic area) that tapers toward the nasal retina giving rise to a complete dorsal arch. Using neurofilament‐200 immunohistochemistry, we also detected a dorsal arch formed by alpha ganglion cells with density peaks in the temporal (14–15 cells/mm²) and dorsonasal (10 cells/mm²) regions. As with other artiodactyls, the giraffe shares the presence of a horizontal streak and a temporal area which, respectively, improve resolution along the horizon and in the frontal visual field. The dorsal arch is related to the giraffe's head height and affords enhanced resolution in the inferior visual field. The alpha ganglion cell distribution pattern is unique to the giraffe and enhances acquisition of motion information for the control of tongue movement during foraging and the detection of predators. J. Comp. Neurol. 521:2042–2057, 2013. © 2012 Wiley Periodicals, Inc.     

Two visual pathways to the telencephalon in the nurse shark (Ginglymostoma cirratum). I. Retinal projections

The central projections of the retina in the nurse shark were studied by anterograde transport of horseradish peroxidase and tritiated proline. With regard to efferent retinal fibres, both techniques gave completely identical results. Projections were found to pretectal area, dorsal thalamus, basal optic nucleus, and optic tectum, all at the contralateral side. The retinal target cells in the dorsal thalamus are restricted to the ventrolateral optic nucleus and the posterior optic nucleus. No evidence was found for an earlier-reported projection to the lateral geniculate nucleus. The present findings show that the ventrolateral optic nucleus exhibits homological features of the dorsal lateral geniculate nucleus in other vertebrate groups, whereas the lateral geniculate nucleus of the nurse shark is much more comparable to the nucleus rotundus of teleosts and birds and would be more appropriately so named. The application of the HRP technique also allowed us to study afferents to the retina by retrograde transport of tracer. Retrogradely labeled cells were observed in the contralateral optic tectum and are apparently similar to those reported for teleosts and birds.     

Retinal cone topography of artiodactyl mammals: influence of body height and habitat

This study addresses the correlation of retinal topography with factors such as the visual environment, life style, and behavior for a major mammalian group, the artiodactyls. To provide a broader basis for semiquantitative comparison, short-wavelength-sensitive (S)- and middle-to-long-wavelength-sensitive (M)-opsin cone receptor populations from 25 species from five artiodactyl families and of the African elephant were labeled and sampled. The resulting topographic maps were analyzed with respect to the position and extension of high-density regions. For better parameter differentiation, systematic relationships were statistically normalized. In all species examined, two classes of cones have been detected. In most species, the S-cone maxima were located in the temporodorsal retina, but there are exceptions such as the roe deer with accumulation in the ventral retina. For M-cones, as a consequence of their role in terrain/food assessment and predator detection, the standard topography is L-shaped: a horizontal visual streak including a temporal area centralis is extended by a temporal rim. Its extension is correlated with the animal's body height (P = 0.0017): small species (pudu, mouse deer) tend to have a visual streak only, whereas the giraffe shows a complete dorsal arch of elevated densities. Furthermore, a size-independent habitat correlation was revealed for a similar M-cone pattern (P < 0.0001): mountainous species show a striking specialization around the dorsal retina, pointing to the importance of the inferior visual field in precipitous terrain.     

The visual system of the channel catfish (Ictalurus punctatus). I. Retinal ganglion cell morphology

Horseradish peroxidase was applied to lesions in the optic nerve of catfish (Ictalurus punctatus). The retinae were processed to reveal HRP-labelled ganglion cells. The histochemical techniques employed allowed fine details of the dendritic arbor to be resolved. Flat-mounted retinae were examined and the following characteristics were noted in individual ganglion cells: Soma area, shape, and depth; number and diameter of major dendrites; shape, area, and depth(s) within the inner plexiform layer (ipl) of the dendritic arbor; origin of the axon (from the soma or a dendrite). On the basis of these characteristics, eleven classes of ganglion cells were delineated: four classes of giant cells (G1-G4) and seven classes of smaller cells (S1-S7). G1 cells had dendrites arborizing in the most distal sublamina of the ipl. G1 cells in the dorsal retina had nasotemporally elongated dendritic arbors. G2 cells had dendrites in the proximal portion of the ipl. G3 cells were almost completely confined to a band running between the nasal and temporal retinal poles, through the center of the retina. In this location, the cells had dorsoventrally elongated dendritic arbors, which were bistratified in the ipl. G4 cells were displaced into the inner nuclear layer. S1 and S4 cells had axons arising from their somata, and dendrites arborizing in the distal and the proximal ipl, respectively. S2 cells were typified by their unstratified dendritic arbors. Similarly, S3 cells were characterised by their bistratified arbors. S5 cells arborized in the most proximal ipl sublamina. S6 cells were small ganglion cells with their somata lying in the inner nuclear layer. S7 cells tended to have complex dendritic arbors, and their axons arose from dendrites.     

Retinal projections to the superior colliculus and dorsal lateral geniculate nucleus in the tammar wallaby (Macropus eugenii): I. Normal topography

The topography of retinal projections to the superior colliculus and dorsal lateral geniculate nucleus of a wallaby, the tammar (Macropus eugenii), was investigated by an anatomical method. Small laser lesions were made in the retinas of experimental animals, and the remaining retinal projections were visualized by means of horseradish-peroxidase histochemistry. The position of each lesion was correlated with the position of the filling defects in the terminal label. The whole of the retina projects to the contralateral superior colliculus. The nasal retina is represented caudally, and the temporal retina rostrally. The ventral retina is represented medially, and the dorsal retina laterally. There is a projection to the ipsilateral superior colliculus, but it is patchy and its topography could not be determined by this method. The retinotopic map in the contralateral dorsal lateral geniculate nucleus has the nasal retina represented rostrally and the temporal retina caudally in the nucleus. The dorsal retina is represented ventrally, and the ventral retina is represented dorsally. It appears that the whole of the retina projects contralaterally, and in addition the temporal retina projects ipsilaterally. The maps of visual space through the two eyes were shown to be in topographic register in the binocular region by making a deposit of HRP in the visual cortex. This resulted in a column of retrogradely labeled cells in the nucleus. This column crossed the laminae, which are innervated by the ipsilateral and contralateral eye at right angles.     

Retinal topography in reef teleosts. I. Some species with well-developed areae but poorly-developed streaks

The retinal ganglion cell layer of five species of teleosts has been studied from Nissl-stained whole-mounts and the distribution of neuronal elements determined quantitatively. Isodensity contour maps of neurons in the ganglion cell layer revealed areas of high density (areae centrales) predominantly in the temporal retina, but other areae were also found in the nasal and dorso-nasal retina. Neuronal densities within the ganglion cell layer at the areae centrales ranged from 0.4 x 10(4) to 4.7 x 10(4) cells/mm2. Species that were found to lack a horizontal streak of high ganglion cell density appear to be those whose behaviour suggests they possess an interrupted view of the sand-water horizon and are 'enclosed' species. Concentric density contours around an area centralis seem to be associated with enclosed environments. The relationship between retinal topography and niche is also discussed.     

Retinal ganglion cell layer of the Caspian seal Pusa caspica: topography and localization of the high-resolution area

Retinal topography, cell density and sizes of ganglion cells in the Caspian seal (Pusa caspica) were analyzed in retinal whole mounts stained with cresyl-violet. The topographic distribution of ganglion cells displayed an area of high cell density located in the temporal quadrant of the retina and was similar to the area centralis of terrestrial carnivores. It extended nasally, above the optic disk, as a streak of increased cell density. In different whole mounts, the peak cell density in the high-density area ranged from 1,684 to 1,844 cells/mm2 (mean 1,773 cells/mm2). The cell density data predict a retinal resolution of around 8.5 cycles/degree in water. A distinctive feature of the Caspian seal's retina is the large size of ganglion cells and the low cell density compared to terrestrial mammals. The ganglion cell diameter ranged from 10 to 58 μm. Cell size histograms featured bimodal patterns with groups of small and large ganglion cells. The large cells appeared similar to α-cells of terrestrial mammals and constituted 7% of the total ganglion cell population.     

Retinal ganglion cell topography in teleosts: a comparison between Nissl-stained material and retrograde labelling from the optic nerve

The retinal topography of cells within the ganglion cell layer of three teleost species is examined in Nissl-stained material in which all neuronal elements containing Nissl substance in the cytoplasm are counted. A topographic comparison is made with retrogradely labelled ganglion cells to differentiate the proportion of nonganglion cells not possessing an axon joining the optic nerve. In the three species studied 92%, 80%, and 66% were found to be the maximum proportion of true ganglion cells in the area centralis, horizontal streak, and periphery, respectively. The proportion of nonganglion cells in the total population of cells counted was 24%. The major contribution to this discrepancy is from peripheral nonspecialized regions of the retina. There is little difference in both topography and peak densities of retinal ganglion cells between the two techniques. The soma areas of both populations are analysed, with the homogeneous nonganglion cell population possessing cells between 5 and 15 micron2 and the heterogeneous ganglion cell soma between 5 and 68 micron2, increasing in size with eccentricity.     

Retinal topography in reef teleosts. II. Some species with prominent horizontal streaks and high-density areae

The retinal ganglion cell layer of five species of reef teleosts was studied from Nissl-stained whole-mounts and the distribution of neural elements determined quantitatively. Iso-density contour maps of neurons in the ganglion cell layer revealed a temporal area centralis (ranging from 3.5 to 8.3 x 10(4) cells/mm2) which often extended into a horizontal streak (ranging from 1.4 to 5.0 x 10(4) cells/mm2) across the retinal meridian. Species possessing a marked horizontal streak were found to inhabit open water and perceive their environment with an uninterrupted view of sand-water horizon. The behavioural significance of these horizontal areas of acute vision is also discussed.     

Retinal projections in the freshwater butterfly fish, Pantodon buchholzi (Osteoglossoidei). I. Cytoarchitectonic analysis and primary visual pathways.

The freshwater butterfly fish, Pantodon buchholzi, is a member of the most primitive radiation of teleosts. The retinofugal projections were studied in this fish with autoradiographic and horseradish peroxidase (HRP) methods, and the cytoarchitecture of the retinorecipient regions in the diencephalon and pretectum was analyzed with Bodian-, cresylecht-violet- and acetylcholinesterase-reacted sections. The rostral diencephalon of Pantodon contains a large retinorecipient nucleus, not previously identified in any other fish, i.e. nucleus rostrolateralis. Other nuclei that are described correspond to those previously recognized in other species. The majority of retinorecipient nuclei are positive for acetylcholinesterase, particularly those in the pretectum, as has been found in other species of teleosts. Most of the retinofugal fibers decussate in the optic chiasm. Some fibers project via the axial optic tract to preoptic nuclei and a region in the rostral hypothalamus. Fibers leave the medial optic tract to terminate in nucleus rostrolateralis and in dorsal and ventral thalamic nuclei, accessory optic and tubercular nuclei, periventricular and central pretectal nuclei, and sparsely in the deep tectal fascicle and terminal field. Dorsal optic tract fibers project to the dorsal accessory optic nucleus, superficial and central pretectal nuclei, and superficial and deep tectal layers. Ventral optic tract fibers project to the superficial pretectum, accessory optic nuclei, posterior tuberculum, nucleus corticalis in the central pretectum, and superficial tectal layer. Fibers that remain in the ipsilateral optic tract project to most of the targets reached by contralaterally projecting fibers. A few fibers in the contralateral medial optic tract redecussate via the posterior commissure to reach the ipsilateral periventricular pretectum. No labeled retinopetal cells caudal to the olfactory bulb were identified in any of the HRP cases.