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Confocal laser scanning microscopy and three-dimensional reconstruction of cell clusters in serous fluids.

Thick cell clusters are a common finding in reactive and malignant effusions. In order to arrive at a diagnosis, clusters are evaluated for certain cytomorphologic features including size, shape, smooth vs. scalloped borders, and three-dimensional (3-D) configuration. By conventional microscopy, the image of these clusters is often blurred due to limitations in resolution. Consequently, the exact internal structure and cellular arrangement within these clusters cannot be adequately determined. Utilizing confocal laser scanning microscopy (CLSM), we examined serous fluids from a variety of conditions. Cases included mesothelioma, adenocarcinoma, and papillary adenocarcinoma. Smears were stained with 0.01% ethidium bromide and 1% eosin Y, followed by analysis with an ACAS 570 image analyzer (Meridian Instruments, Inc. Okemos, MI). Serial confocal fluorescence images were acquired, which allowed 3-D reconstruction of the clusters. Mesothelioma clusters (excluding those with obvious central collagen cores by light microscopy) appeared to be formed of the following configurations: 1) randomly coiled cords of cells, 2) small papillae encompassing central cores, and 3) tissue fragments with pseudoacinar formation. In contrast, adenocarcinomas had a more orderly pattern, with tightly cohesive cells and true acinar formation.

Adenocarcinoma↗

Abnormal retinotopic organization of the dorsal lateral geniculate nucleus of the tyrosinase-negative albino cat.

Visual defects associated with hypopigmentation have been studied extensively in Siamese and albino cats. Previous research on tyrosinase-negative albino cats has shown that (1) approximately 95% of all nasal and temporal retinal fibers cross at the optic chiasm, and (2) ocular dominance columns normally found in cortex are replaced with hemiretinal domains. In this study, we compared the retinotopic organization of the dorsal lateral geniculate nucleus (LGNd) and visual cortex in albino cats. Extracellular recordings were conducted in the LGNd, area 17, and area 18 of six albino cats. Receptive fields (RFs) were plotted for all sites. We find that, as in albino visual cortex, the albino LGNd contains (1) normal cells with RFs in the visual hemifield contralateral to the recording site (RFc), (2) abnormal cells with RFs in the ipsilateral hemifield (RFi), (3) abnormal cells with dual, mirror-symmetric RFs, one in each hemifield (RFd), and (4) abnormal cells with broad RFs that span the vertical meridian (RFb). Our data indicate that lamina A and lamina A1 consist predominantly of normal RFc and abnormal RFi cells, respectively. The C laminae contain a mixture of RFc, RFi, RFd, and RFb cells. The interlaminar zones contained RFd cells, RFb cells, or both. Thus, the albino LGNd is arranged into hemiretinal and not ocular dominance laminae. Finally, the percentage of normal cells is significantly larger in area 17 (84%) and area 18 (70%) than in the LGNd (46%), suggesting a suppression of abnormal activity in albino cat cortex, which could underlie the existing competence of visual function in albinos.

Albinism, Oculocutaneous↗

Organization of callosal linkages in visual area V2 of macaque monkey.

In visual area V2 of the macaque monkey callosal cells accumulate in finger-like bands that extend 7-8 mm from the V1/V2 border, or approximately half the width of area V2. The present study investigated whether or not callosal connections in area V2 link loci that are located at the same distance from the V1/V2 border in both hemispheres. We analyzed the patterns of retrograde labeling in V2 resulting from restricted injections of fluorescent tracers placed at different distances from the V1/V2 border in contralateral area V2. The results show that varying the distance of V2 tracer injections from the V1/V2 border led to a corresponding variation in the location of labeled callosal cells in contralateral V2. Injections into V2 placed on or close to the V1 border produced labeled cells that accumulated on or close to the V1 border in contralateral V2, whereas injections into V2 placed away from the V1 border produced labeled cells that accumulated mainly away from the V1 border. These results provide evidence that callosal fibers in V2 preferentially link loci that are located at similar distances from the V1/V2 border in both hemispheres. Relating this connectivity pattern to the topographic map of V2 suggests that callosal fibers link topographically mirror-symmetrical regions of V2, i.e., callosal fibers near the V1/V2 border interconnect areas representing visual fields on, or close to, the vertical meridian, whereas callosal connections from regions away from the V1/V2 border interconnect visuotopically mismatched visual fields that extend onto opposite hemifields.

Animals↗

Apolipoproteins (apoproteins) and LPL variation in Mennonite populations of Kansas and Nebraska.

Apoproteins (also known as apolipoproteins) have been studied extensively because of their role in lipid transport, association between specific genotypes and elevated serum lipid levels, and increased risk of heart disease. There is considerable genetic variation in the geographic distributions of these markers, with a north-south cline of the APOE*4 allele observed in Europe by Lucotte et al. ([1997] Hum Biol 69:253-262). This study compares the frequencies of seven APO (APOA1 -75 bp, APOA1 +83 bp, APOB Ins/Del, APOB XbaI, APOC3 SstI, and APOE) and LPL loci in Mennonite populations from Kansas and Nebraska. In total, 277 individuals were sampled from Goessel, Meridian, Garden View, and Lone Tree in 2002-2004. In addition, DNA samples that were collected in 1981 from Henderson, Nebraska, were genotyped for the seven APO and LPL loci. Of the seven APO and LPL loci tested, only one locus, APOB XbaI, departed significantly from Hardy-Weinberg equilibrium, with an unexpected excess of observed heterozygotes. The frequencies of the several APO loci are unique among the Mennonites, separating them from other European populations. A bidimensional scaling representation of Reynold's co-ancestry distances based on allelic frequencies of the seven APO and LPL markers in five Mennonite congregations fails to represent schematically the known patterns of fission. It is unclear whether the observed patterns are due to selection operating on these loci or whether genetic drift, small populations sizes, or a lack of statistical power of these biallelic loci distort the observed genetic relationship among congregations.

Apolipoproteins↗

Genetic loci for pathological myopia are not associated with juvenile myopia.

The purpose of this study was to evaluate chromosomal regions previously linked to pathological myopia for linkage to juvenile myopia in a sample of myopic children and their families. Of 125 families with a myopic child participating in the Orinda longitudinal study of myopia, 53 submitted 221 buccal swab samples for genetic analysis. Myopia in proband children was defined as -0.75 D or more myopia in both meridians on cycloplegic autorefraction (1% tropicamide). Affected status in parents and siblings was obtained by survey. DNA was extracted from buccal mucosal cells, amplified by polymerase chain reaction (PCR), and then analyzed with seven markers for chromosome 12 and five markers for chromosome 18 in the regions previously associated with pathological myopia. LOD scores were not significant for any marker tested. The largest positive LOD score was 0.15 for GATA30F04. Model-free methods using a SimIBD approach suggested a possible linkage at one marker, GATA6H09 (P = 0.003), but these results were not supported by transmission disequilibrium test (TDT) analysis. The statistical power to detect LOD scores of > or =1.0, assuming homogeneity, was estimated at 93.2%. We found no confirmatory evidence of linkage between juvenile myopia and regions of chromosomes 12 and 18 previously associated with pathological myopia.

Child↗

Confrontation visual field techniques in the detection of anterior visual pathway lesions.

The accuracy of a variety of finger and color confrontation tests in identifying chiasmal and optic nerve visual field defects was assessed in patients whose field defects had been established beforehand by a conventional achromatic kinetic technique on the Goldmann perimeter. Kinetic and static finger confrontation methods identified an average of 42% of the 28 chiasmal hemianopic defects. False negatives included eyes with hemianopias complete to the largest (V4e) Goldmann isopter. False positives (average, 15%) occurred in eyes containing nerve fiber bundle defects with borders that fell near the vertical fixational meridian. Kinetic and static color confrontation techniques were 78.6% sensitive to hemianopias. Accuracy did not differ significantly whether the red target was presented kinetically or statically against the tangent screen, projected on the Autoplot screen, or held in the examiner's hand without attention to background. False positives (average, 23%) were slightly greater than with finger confrontation methods and occurred not only in eyes with nerve fiber bundle defects but also in eyes with no defects in reference visual fields. Finger confrontation identified 11% or fewer of optic nerve field defects, while some color techniques detected as many as 31 1/3%. There were no false positives.

Hemianopsia↗

The influence of preexisting oscillations on the binocular optokinetic response.

Full-field binocular optokinetic responses were examined in patients with congenital nystagmus. If the stimulus moved in the same plane as the congenital nystagmus, a gaze-modulated response was recorded for both idiopathic and albino observers. When the congenital nystagmus was almost totally suppressed, neither a gaze-evoked nor a sawtooth response was seen. For stimulus movements in a plane orthogonal to the congenital nystagmus, normal sawtooth optokinetic responses were exhibited by both groups. Experimental evidence supports the hypothesis that the optokinetic response in the meridian of the spontaneous oscillation has been adapted.

Adult↗

Architecture of the ciliary muscle of Gallus domesticus.

There are species-related anatomical differences in the ciliary muscle of the avian eye. The arrangement of muscular fibers in the avian eye is not well defined. To clarify this situation, we studied the architecture of ciliary muscle of Gallus domesticus by light and scanning electron microscopy (SEM). Our results showed the existence of three main muscular groups that we defined as anterior, posterior, and intermediary. These muscle divisions correspond to the description of the ciliary muscle as previously stated by Crampton (1813), Brucke, and Muller (1856). The striated fibers have a meridian orientation. The anterior and posterior muscular groups are inserted in the sclera, around the Schlemm's canal wall and ciliary process stroma. The vitreal intermediary muscle has fibers inserted in Schlemm's canal wall and ciliary process stroma. The framework of these muscular fibers may according to its insertions participate in the visual accommodation mechanism and outflow of the aqueous humor system.

Animals↗

Variations in the structure of the prelunate gyrus in Old World monkeys.

Anatomical and electrophysiological studies have revealed a complex organization in the macaque prelunate gyrus. We investigated the morphology and architecture of the prelunate gyrus in Old World monkeys. In Macaca nemestrina, we observed a sulcus crossing the prelunate gyrus within 2 mm of the vertical meridian representation. In other macaque species and other cercopithecines, we observed substantial variations in sulcal morphology across the prelunate gyrus. We did not find a sulcus in all species, and the location and depth of that indentation on the gyrus varied among species. A deep sulcus was observed in all species that emerged earlier in evolution than macaques, such as guenons, baboons, and colobines. We analyzed the regional and parcellation features of the prelunate gyrus in three macaque species, M. maura, M. mulatta, and M. radiata, and in Erythrocebus patas, with emphasis on the relation of structure to the distribution of prelunate visual areas. Nonphosphorylated neurofilament protein immunoreactivity permitted the delineation of a novel area in the prelunate gyrus of Old World monkeys, located around the prelunate sulcus. Species-specific patterns were also observed in the prelunate gyrus of the patas monkey compared to macaques. These observations, as well as a cladistic analysis of the data, suggest an expanded and diversified organization of the prelunate gyrus in some cercopithecoids that may reflect adaptation to specific ecological environments. It was, however, progressively lost in most macaques, being retained only in species that diverged early in the evolution of the genus Macaca, such as M. nemestrina and M. maura.

Animals↗

Topographic approach to the study of the human body.

Recent developments in medical imaging techniques such as ultrasound, computed tomography (CT), and magnetic resonance imaging (MRI) have been explosive. These modalities provide 3D information about the human body and assess tissue damage in various pathological conditions. To complement the diagnostic usefulness of these imaging techniques, we have designed a system of topographic coordinates based on the principles of global projection cartography in which lines of latitude and longitudes are assigned to the surface of the human body. We designated the median sagittal plane as corresponding to the Greenwich Meridian (zero longitude) in global cartography. From the median sagittal plane (M0), vertical lines of longitude or "great circles" divide the body into 12 zones that are 30 degrees apart. Parallel lines of latitude are assigned according to surface anatomy landmarks. Studying the 3D reconstruction of anatomical structures is important for: 1) devising a system of coordinates; 2) allowing biomedical measurements to be made; and 3) drawing maps that may be useful in some clinical procedures (e.g., biopsies).

Anatomy↗

Neuronal development in larval polychaete Phyllodoce maculata (Phyllodocidae).

The existing view on neuronal development in polychaetes, as undergoing neurogenesis beginning in the rudiments of central ganglia and then extending peripherally, has been contrasted with the latest findings in molluscs, their sister trochozoan group, which show a peripheral to central mode of neurogenesis. The current study addresses this issue by examining early neuronal development in the polychaete Phyllodoce maculata using immunolabeling against acetylated alpha-tubulin, serotonin, and the FMRFamide. The first nervous cell was detected 20 hours before hatching, at the early trochophore stage. A solitary serotonergic neuron was located at the posterior-dorsal extreme of the larva and issued anterior projecting fibers, which outline the future ventral nerve cords and prototroch nerve. Two more serotonergic dorsal peripheral cells and three peripheral FMRFamidergic cells appeared soon thereafter. The fibers of these early cells formed a scaffolding, which prefigured the future adult nervous system (cerebral ganglion, ventral cords, prototroch and esophageal nerve rings) in prehatched trochophores. Shortly before hatching, the larval nervous system developed, including the apical organ, meridianal nerves in the episphere, and posttrochal nerves that innervate the feeding apparatus. After hatching, the rudiments of the adult nervous system started to develop along the paths already established by the earliest peripheral neurons. Thus, the general strategy of neurogenesis in a representative polychaete trochophore appears to resemble that of molluscs. The first neuronal cells to appear are peripheral in origin, located near the posterior margins of the embryo. Their similar anatomical appearance suggests that they share a similar functional role in trochophore development and behavior.

Animals↗

Resolving the organization of the New World monkey third visual complex: the dorsal extrastriate cortex of the marmoset (Callithrix jacchus).

We tested current hypotheses on the functional organization of the third visual complex, a particularly controversial region of the primate extrastriate cortex. In anatomical experiments, injections of retrograde tracers were placed in the dorsal cortex immediately rostral to the second visual area (V2) of New World monkeys (Callithrix jacchus), revealing the topography of interconnections between the "third tier" cortex and the primary visual area (V1). The data indicate the presence of a dorsomedial area (DM), which represents the entire upper and lower quadrants of the visual field, and which receives strong, topographically organized projections from the superficial layers of V1. The visuotopic organization and boundaries of DM were confirmed by electrophysiological recordings in the same animals and by architectural characteristics which were distinct from those found in ventral extrastriate cortex rostral to V2. There was no electrophysiological or histological evidence for a transitional area between V2 and DM. In particular, the central representation of the upper quadrant in DM was directly adjacent to the representation of the horizontal meridian that marks the rostral border of V2. The present results argue in favor of the hypothesis that the third visual complex in New World monkeys contains different areas in its dorsal and ventral components: area DM, near the dorsal midline, and a homolog of area 19 of other mammals, located more lateral and ventrally. The characteristics of DM suggest that it may correspond to visual area 6 (V6) of Old World monkeys.

Action Potentials↗

Receptive fields of single cells and topography in mouse visual cortex.

The visual cortex was studied in the mouse (C57 Black/6J strain) be recording from single units, and a topographic map of the visual field was constructed. Forty-five percent of the neurons in striate cortex responded best to oriented line stimuli moving over their receptive fields; they were classified as simple (17%), complex (25%) and hypercomplex (3%). Of all preferred orientations horizontal was most common. Fifty-five percent of recpetive fields were circularly symmetric: these were on-center (25%), off-center (7%) and homogeneous on-off in type (23%). Optimal stimulus velocities were much higher than those reported in the cat, mostly varying between 20 degrees and 300 degrees/sec. The field of vision common to the two eyes projected to more than one-third of the striate cortex. Although the contralateral eye provided the dominating influence on cells in this binocular area, more than two-thirds of cells could also be driven through the ipsilateral eye. The topography of area 17 was similar to that found in other mammals: the upper visual field projected posteriorly, the most nasal part mapped onto the lateral border. Here the projection did not end at the vertical meridian passing through the animal's long axis, but proceeded for at least 10 degrees into the ipsilateral hemifield of vision, so that at least 20 degrees of visual field were represented in both hemispheres. The magnification in area 17 was rather uniform throughout the visual field. In an area lateral to area 17 (18a) the fields were projected in condensed mirror image fashion with respect to the arrangement of area 17. Medial to area 17 a third visual area (area 18) was again related to 17 as a condensed mirror image.

Animals↗

The pulvinar nucleus of Galago senegalensis.

The present study was undertaken to analyze the connections of the pulvinar nucleus in a prosimian. The experiments, which rely on the Fink-Heimer ('67) method for staining degenerating axons and their terminals, fall into two parts: first, the tracing of ascending tectal projections to the caudal thalamus and second, the tracing of projections from this thalamic target to the cortex. Large lesions of the superior colliculus resulted in dense degeneration in the caudal half of the inferior subdivision of the pulvinar complex. This pathway could be identified when the lesion was restricted to the superficial layers of the superior colliculus, signifying that it is a visual pathway. In general, the projections of the deep and superficial layers of the superior colliculus were distinct and in this respect Galago resembles Tupaia. The inferior pulvinar nucleus in turn projects to area MT, a conspicuous subdivision of the temporal cortex. The superior division of the pulvinar, in contrast to the inferior division, is not a major target of ascending projections from the superior colliculus and projects to the areas of the occipital and temporal lobe intercalated between areas MT and 17. When these results are compared with similar studies in nonprimates, notably studies of Tupaia, a striking difference in organization emerges. In Tupaia, and in distantly related mammals such as the squirrel, the target of the tecto-pulvinar system includes area 18 adjacent to area 17. This feature is important since the two parallel projection systems seem to be related to each other in terms of the way in which the zero vertical meridian is spatially represented. However, in Galago the subdivision of the pulvinar receiving projections arising from the superior colliculus does not project to area 18. Area 18 is indeed the target of pulvinar projections, but these projections arise from that portion of the pulvinar which is not a recipient of ascending tectal projections. It is not easy to see how this primate organization, if indeed the Galago is representative of primates, evolved from the organization reflected in Tupais.

Animals↗

Interhemispheric neocortical connections of the corpus callosum in the normal mouse: a study based on anterograde and retrograde methods.

Interhemispheric neocortical connections are widely distributed through the corpus callosum in the mouse. Callosal connections are present in all cytoarchitectonic fields except field 25. The distal extemity representations of SmI, and MsI the representation of the mystacial vibrissae in SmI, and the more peripheral field representation of VI are relatively acallosal. Dense projections lie in the midline or truncal representations of SmI, MsI, SmII, at the vertical meridian representations bordering field 17, and medial to the AI representation. The radial distribution of terminals is bimodal in most cytoarchitectonic fields. It is unimodal in the supracallosal segment of field 29b and fields 49 and 27, trimodal in fields 13 and 35. The cells of origin of callosal fibers appear to have the same topographic pattern of distribution as the callosal terminals, observing the same steep and gradual density gradients. No cells giving rise to callosal axons are identified in the acallosal regions of fields 2 and 17. Further, superficial focal lesions in cortical areas which receive callosal connections give rise only to homotopic contralateral degeneration. Acallosal areas of 17 and 2 give rise to no callosal connections. The cells of origin of callosal connections are located at all laminar levels of the cortex and include pyramidal and polymorphic cells but not the granule cells of layer IV.

Animals↗

Demonstration of bilateral projection of the central retina of the monkey with horseradish peroxidase neuronography.

In the primate, ganglion cells of the temporal retina project ipsilaterally and those of the nasal retina, contralaterally into the optic tract. The vertical meridian passing through the fovea defines the border between these two populations of ganglion cells and has been demonstrated in four Macaque monkeys after unilateral injection of horseradish peroxidase into the dorsal lateral geniculate nucleus and examination of the pattern of retrograde labeling of those ganglion cells projecting to the injected side. A median 1 degree vertical strip in which ipsi- and contralaterally projecting ganglion cells intermingle was found, confirming the report by Stone et al. ('73). In addition, occasional extrafoveal labeled ganglion cells were found as far as 2 degrees from the vertical midline in the otherwise unlabeled hemiretinae. These ganglion cells were not numerous and had somata of all sizes, suggesting that they do not constitute a separate class of ganglion cells as found in the temporal retina of the cat. In contrast to the description by Stone et al. ('73), the strip of vertical overlap did not show a constant width through the fovea, since mixing of labeled and unlabeled ganglion cells was found in a band approximately 1/2 degree wide along both the nasal and temporal rims of the foveal pit which is 500 mum (2 degrees) in diameter. Beyond these 1/2 degree arcs, the appropriate hemiretina was either completely unlabeled, or contained virtually every ganglion cell labeled on the side projecting to the injected dorsal lateral geniculate nucleus. The scattered labeled ganglion cells rimming an otherwise unlabeled hemifovea represent a possible anatomical basis for the phenomenon of "macular" or "foveal sparing" in which unilateral damage to the occipital cortex produces homonymous hemianopsia with sparing of a small island of centralmost vision extending about 1 degree from the foveal center. From this study, it is not possible to define the receptive fields or specific photoreceptor connections of the ganglion cells labeled with horseradish peroxidase, so that at the present time quantitative correlations cannot be made between the numbers of ganglion cells remaining on the affected side of the fovea and the extent of preservation of visual function in the spared zone. The presence of labeled ganglion cells rimming the fovea in its entirety is compatible with the sequence of foveal development in late prenatal life. After lateral displacement both nasally and temporally of ganglion cells which initially lay in the median vertical overlap strip of 1 degree, in the adult retina a strip approximately 1/2 degree wide around the perimeter of the foveola should contain a mixture of ipsi- and contralaterally projecting ganglion cells. The total population of ganglion cells beyond this 1/2 degree band should be completely ipsi- or contralateral in their projection patterns, as is observed...

Animals↗

Postnatal neurogenesis in the kitten retina.

Postnatal neurogenesis in the kitten retina was studied using 3H-thymidine radioautography. Kittens were injected with 3H-thymidine at 1 day, 10 days, 3 weeks or 4 weeks after birth and allowed to survive until 14 weeks of age. Labeled neuronal nuclei were not found in the ganglion cell layer in any of the retinas, but they were seen in the other nuclear layers of the same retinas. In retinas from kittens injected at one day after birth, the peripheral 80% of the length of the retina (in sections cut parallel to the dorsoventral meridian) contained labeled nuclei; the central 20%, around the optic disc, contained no labeled nuclei. Near the ora serrata most nuclei in both inner and outer nuclear layers were labeled. Away from the ora serrata the proportion of labeled to unlabeled nuclei gradually decreased. Labeled nuclei extended farther centrally in the the inner than the outer nuclear layer. The same pattern of labeling was repeated in retinas from kittens injected at ten days after birth, but fewer nuclei were labeled, and the central, unlabeled region around the optic disc was longer--55% of the length of the retina. Only a few nuclei near the ora serrata were labeled in retinas from kittens injected at three weeks after birth, and no labeled neurons were found in kittens injected at four weeks. From these results we conclude that all of the ganglion cells in the kitten retina are present by one day after birth, as are all of the other neurons in the central retina. In peripheral regions of the inner and outer nuclear layers, proliferation of cells destined to become neurons continues up to three weeks after birth.

Animals↗

Gradients between nasal and temporal areas of the cat retina in the properties of retinal ganglion cells.

Evidence is presented of gradients between nasal and temporal areas of the cat's retina in the properties of their ganglion cell populations. Mean ganglion cell size is greater in temporal retina than in nasal retina, partly because the alpha- and beta-cells of temporal retina are distinctly bigger than their counterparts in nasal retina, and partly because more medium-sized cells, and fewer small cells, are to be found in temporal retina. This high proportion of medium-sized ganglion cells may reflect a high proportion of beta-cells or of the medium sized gamma-cells described by Stone and Clarke ('80). Several of these differences can be related to prior morphological, electrophysiological, and behavioural observations in the cat, and similar differences have been reported in several other mammalian species. Evidence is presented that, in the cat, at least some of these differences are less marked near the vertical meridian of the retina than more temporally or nasally. The present results may therefore, be evidence of a nasal-temporal gradient in retinal structure and funtion common to many mammals, and distinct from previously recognised gradients in ganglion cell properties related to the area centralis and visual streak specialisations.

Animals↗