Cells In Upper Layers Research Articles

Features of polyingression and primitive streak ingression through the basal lamina in the chicken blastoderm

The de-epithelialization of cells of the upper layer during the phenomena of polyingression and primitive streak ingression was studied by analyzing, from the time of laying to the end of gastrulation, the ultrastructure of the basal lamina underlying the upper layer. The electron density of the basal lamina and associated extracellular materials was enhanced by addition of tannic acid to the fixative. Special attention was also paid to the spatial and temporal distribution of blebs at the basal surface of the upper layer, and to the contribution of the de-epithelialized cells to the formation of the deep layer. The results indicate that a nascent basal lamina is already present at the time of laying, especially beneath regions of the area pellucida where polyingression is not apparent. From the onset of incubation, the basal lamina rapidly develops, and it is interrupted by a large number of blebs. However, during the first 6-8 h of incubation, i.e., stages 1-2 of Vakaet (Arch. Biol. (Liège) 81:387-426, 1970), a downward movement of de-epithelialized cells that insert into the deep layer and form the endophyll persists cranially. This phenomenon of polyingression, which starts during the intrauterine period, probably extends from caudal to cranial and comes to an end by stage 3. During these first three stages, the number of blebs progressively decreases, especially in the cranial part of the area pellucida, and a thicker, continuous basal lamina associated with numerous interstitial bodies is laid down. The caudal part of the upper layer is still actively blebbing at that time. Due to the convergence of this area toward the axis of the blastoderm, which leads to ingression at and elongation of the primitive streak up to and including stage 6, the number of blebs at the basal surface of the upper layer progressively decreases. From stage 7 on, blebs are virtually absent; shortening of the primitive streak and formation of the head process begin. At the level of the head process, primitive streak ingression has ceased and a novel basal lamina is progressively deposited beneath the upper layer. By stage 9, a thick, smooth basal lamina physically separates the upper layer from the head mesenchyme. Summarizing, at the time of gastrulation, the presence of blebs that perforate the basal lamina is correlated with the de-epithelialization of cells. Before incubation, however, de-epithelialization of upper-layer cells occurs before the assembly of the basal lamina.(ABSTRACT TRUNCATED AT 400 WORDS)

Filaggrin expression in cutaneous and mucosal human papillomavirus induced lesions

A series of 32 human papillomavirus induced lesions derived from epidermis and mucosa was studied for the modulation of filaggrin-profilaggrin (F-PF) expression according to the degree of virus infection as compared to normal skin and mucosa biopsies. This investigation was carried out on frozen sections using indirect immunofluorescence for filaggrin detection and group specific viral antigen and by in situ hybridization with biotinylated probes for viral DNA detection and typing. The 9 cutaneous warts showed an increase of F-PF expression in upper layer cells as compared to normal epidermis, which could be related to the high production of virus (viral antigen and HPV types 1 or 2). The 5 condyloma acuminata displayed also an enhanced expression of these components which was located in several upper layers but virus infection was confirmed in 2 of them with HPV types 6, 11 or 16. The 6 laryngeal papillomas exhibited a granular reactivity pattern for F-PF in suprabasal cell layers with an increase in the upper layers; viral antigen was found in 4 cases and HPV DNA types 6, 11 or 16 were detected in 4 specimens. Conversely among 12 cervical intraepithelial neoplasia, F-PF was expressed only in very superficial layers in few cases, without any correlation with the DNA detection (6, 11 or 16, 18). Taken together these data are suggestive of an intense expression of F-PF in benign lesions which can replicate the virus and a discrete or an absent expression of these components in premalignant or malignant lesions.

Complementary global maps for orientation coding in upper and lower layers of the monkey's foveal striate cortex.

A population of 269 cells recorded from the foveal representation of striate cortex in 2 rhesus macaque monkeys was examined for orientation preference as a function of receptive field position relative to the center of gaze. Cells recorded in supragranular and infragranular layers were segregated and compared. Within the foveolar region (0.0-0.5 degrees) supragranular cells showed a vertical bias which was not evident in the infragranular layers. At larger eccentricities (0.5-2.5 degrees) supragranular cells showed a radial bias (preferred orientation points toward the center of gaze), whereas infragranular cells showed a concentric bias (preferred orientation is tangent to a circle around the center of gaze). These results are consistent with our earlier report of an orientation shift between the supragranular and infragranular layers (Bauer et al. 1980, 1983; Dow and Bauer 1984). The diagonal orientation bias which we noted earlier (Bauer et al. 1980; Dow and Bauer 1984) in supragranular cells at eccentricities between 0.5 and 2.5 degrees can be explained by the radial bias, combined with a tendency for recording sites to favor receptive field locations closer to the diagonal meridia than to either the horizontal or vertical meridia. Given other evidence that upper layer cells in macaque striate cortex tend to show either orientation or color selectivity, while lower layer cells tend to show movement sensitivity (Dow 1974; Livingstone and Hubel 1984), the present data suggest a functional dichotomy between a supragranular system involved in fixational eye movements and pattern vision and an infragranular system activated primarily by optical flow fields during ambulation.

The same cell lineage is involved in scale formation and regeneration in the teleost fish Hemichromis bimaculatus

The elasmoid scales of the cichlid fish, Hemichromis bimaculatus, are localized within dermal pockets, the floors of which are separated from the stratum compactum by uninterrupted cellular sheets, the scale-pocket linings (SPL). TEM study of the fry skin shows that the SPL cells originate from the cell population constituting the dermal papilla of the scale. The upper-layer cells of the papilla, close to the epidermal-dermal junction, differentiate into scleroblasts that, subsequently, form the scale-bag, while the inner-layer cells, close to the stratum compactum, constitute a bi-layered sheet, the SPL. The SPL cells are joined one to another by numerous desmosomes and their cytoplasm is filled principally by microfilaments and free ribosomes. The SPL is also characterized by the presence of a basement membrane on its two faces. When a scale is experimentally pulled off, the scale-forming cells are removed with the dermis and the epidermis covering the free region of the scale, but the SPL is not damaged and epidermal fragments remain at the posterior edge of each scale-pocket. The epithelial cells migrate, from the epidermal fragments, on an extracellular matrix situated on the surface of the SPL, and the wound is closed from 3 to 6 h after scale removal. The scale-regenerating cells differentiate from the upper-layer cells of the SPL, initially in the central region of the scale-pocket where epithelial cells first contacted the SPL surface. Consequently, it is shown that scale-forming cells and scale-regenerating cells are derived from the same ontogenetic population, the dermal papilla.

Immunohistochemical distribution of monoclonal antibodies against keratin in papillomas and carcinomas from oral and nasopharyngeal regions

Papillomas (40) and squamous cell carcinomas (75) were examined for the presence of three keratin proteins with the use of an immunohistochemical technique. Polyclonal keratin antibody (TK, detecting 41 to 65 kDa keratin) and monoclonal antibodies KL 1 and PKK 1 (55 to 57 kDa and 41 to 56 kDa, respectively) were used. Squamous epithelium in normal oral mucosa showed marked TK staining in cells of upper strata and relatively slight staining in basal layer cells, moderate KL 1 staining in spinous and granular layers and was negative in basal cells. Positive PKK 1 staining was noted in cells of the basal layer. Columnar epithelium in the nasal mucosa showed TK staining in all layers, KL 1 staining on the apical side of epithelial cells and trace or negative staining in basal layer cells. There was moderate PKK 1 staining along the apical side of cells and variable staining in basal cells. Keratin distribution in oral papillomas was similar to that in normal oral epithelium, whereas in nasal and nasopharyngeal papillomas, keratin distribution was restricted to the upper layers. Tonsillar papillomas showed a strong TK reaction, negative KL 1 in upper layer cells, and marked PKK 1 staining in basal cells. Well-keratinized squamous carcinomas indicated an irregular TK distribution and decreased KL 1 and negative PKK 1 stainings. Intermediate and poorly differentiated keratinizing squamous carcinoma showed irregular staining patterns for the three classes of keratins studied. Immunohistochemically detectable keratins utilizing monoclonal antibodies were described as useful markers of epithelial tumors of squamous origin. Keratin expression within benign tumors was related to normal regional distribution, whereas in malignant tumors, keratin distribution was irregular in its distribution profile.

The neuronal composition of area 17 of rat visual cortex. I. The pyramidal cells.

The pyramidal cells in area 17 of rat visual cortex have been examined by light microscopy using Golgi preparations and semithin plastic sections, and by electron microscopy. Pyramidal cells have cell bodies in layers II-VIa. The pyramidal cells in the lower portion of layer II/III are typical examples of this neuronal type in that they have pyramidal-shaped cell bodies, apical dendrites which ascend to layer I, and a skirt of basal dendrites. The pyramidal cells in upper layer II/III are similar in form but have shorter apical dendrites, while the most superficial pyramidal cells lack apical dendrites and instead have two or more primary dendrites that emanate from the upper surface of their somata. In layer V the pyramidal cells are of two sizes, medium and large, and both have a typical morphology, although the larger neurons have thicker apical dendrites and better-developed axon hillocks than the medium-sized pyramids. The medium-sized pyramidal cells of layer V outnumber the large ones to a ratio of 2.5:1. In layer IV a few typical medium-sized pyramidal cells are present, but the majority are small and can be regarded as star pyramids for they have dendrites radiating in all directions. No clearly identified spiny stellate cells have been encountered in layer IV. The pyramidal cells of layer VIa are also small, and most of them have apical dendrites which only ascend as far as layer IV. In addition to these varieties, both inverted and horizontally inclined pyramidal cells have been encountered. In electron micrographs it is apparent that although all of the pyramidal cells have symmetric axosomatic synapses, the frequency with which these synapses occur varies. The cell bodies of the various forms of pyramidal cells do not show a standard cytology. The medium-sized pyramidal cells of layer II/III usually have rounded nuclei, while the nuclei of the small pyramidal cells of layers IV and VIa are somewhat more irregular, and the large pyramidal cells of layer V have deeply indented nuclear envelopes. The appearance of the perikaryal cytoplasm also varies. The larger pyramidal cells have numerous mitochondria and well-developed Nissl bodies in their perikaryal cytoplasm, but the smaller cells have much-less-pronounced mitochondria and their rough endoplasmic reticulum is only organized into stacks at the bases of dendrites. Pyramidal cells account for about 87% of profiles of neuronal cell bodies with nuclei in layer II/III, 90% in layer IV, 89% in layer V, and 97% in layer VIa.

The neuronal composition of area 17 of rat visual cortex. II. The nonpyramidal cells.

In the preceding article the characteristics of the various types of pyramidal cells present in area 17 of rat visual cortex were described (Peters and Kara, '85). In the present article the nonpyramidal cell population of this cortex is considered. It is known from Golgi preparations that in layers II-VIa there are bipolar cells, smooth or sparsely spinous multipolar and bitufted cells with either unmyelinated local plexus or myelinated axons, and chandelier cells. Each of these cell types has been previously examined in Golgi-electron microscopic preparations. The question now being asked is whether the information about the characteristics of these different types of nonpyramidal cells derived from the Golgi-electron microscopic studies can be used to identify the cell bodies of nonpyramidal cells in tissue prepared for conventional electron microscopy. If this can be done then the neuronal composition of area 17 can be determined. It has been found that the cell bodies of bipolar cells can be readily identified because they are elongate and have nuclei with a vertical infolding and few axosomatic synapses, which are of both the symmetric and asymmetric varieties. Evidence is presented to show that there are two types of bipolar cells, small ones and large ones, the large ones being distinguished by their well-developed endoplasmic reticulum in which the cisternae are arranged parallel to the cell surface. Bipolar cells account for 6% of the neuronal profiles in layer II/III, 3% in layer IV, 5% in layer V, and 2% in layer VIa. The cell bodies of other types of nonpyramidal cells in layers II-VIa cannot be distinguished from each other in thin sections, because recognition of the different cell types depends upon the characteristics and distribution of their dendrites and axons. However, it is evident that in this group of neurons there are some with small cell bodies and others with large cell bodies, and in both size groups there are varieties of neurons which can be recognized from the characteristics of their perikaryal cytoplasm. All of these neurons have both symmetric and asymmetric axosomatic synapses. The greatest number of these nonpyramidal cells which are not bipolar in form is found within layer II/III, where they account for 7% of all neuronal profiles. These neurons comprise 4% of all neuronal profiles in layer IV, 6% in layer V, and 2% in layer VIa. Layers I and VIb contain only nonpyramidal cells, but these are different from the ones in layers II-VIa.(ABSTRACT TRUNCATED AT 400 WORDS)

Distribution patterns of cytoplasmic microtubules in epidermal keratinocytes.

The distribution of cytoplasmic microtubules in cultured guinea-pig keratinocytes was investigated using immunofluorescence (IF) microscopy with monospecific anti-tubulin antibodies and electron microscopy (EM). In culture, adherent cells displayed networks of thin fluorescent fibres, while a homogeneous and/or granular cytoplasmic IF was shown in the cells of upper layers as well as in trypsinized cells. By EM many microtubules were shown in adherent cells but there were fewer or none in the upper layers. An increase in calcium ion (Ca2+) concentration and the addition of an ionophore (X537A) to the culture medium caused disassembly of microtubules. This effect was cancelled by a calmodulin inhibitor. Cryostat sections of normal human and guinea-pig epidermis stained with anti-tubulin antibodies showed a homogeneous and/or granular cytoplasmic IF from basal to granular layers but no detectable IF was seen in the horny layer. These results suggest that keratinocytes contain a cellular pool of tubulin in various states of polymerization and that microtubule disassembly may occur during differentiation, probably being regulated by Ca2+-calmodulin complexes.

Rat C-6 glioma cells maintained in an organ culture system: a study of kinetic parameters.

Rat C-6 glioma cells were grown on a sponge foam matrix in an organ culture system and the cell cycle parameters, including the growth fraction (GF), were assessed after autoradiography. The zones of growth consisted of a compact upper layer (UL) at the gaseous interface, a central necrotic layer and a deeper lower layer (LL) which invaded the matrix. The fraction of continuously labeled mitoses (FCLM) was similar in both the UL and LL cells. The derivatives of the FCLM curves obtained in three experiments gave an average modal TG2 of 5 hr. A mathematical model relating GF, TG2, TC and labeling index as a function of time, LI(t), was devised for cells in a steady state exposed continuously to tritiated thymidine and was applied to data obtained from UL cells. A mean GF of 9% (range: 8-10%) and a mean cell cycle time (TC') of 27 hr (range: 13-47 hr) were obtained. The mean TS was calculated to be 11 hr (range: 8-16 hr) by the method of grain counts per mitotic figure or grain index (GI). Knowledge of TS permitted alternative calculation of the cell cycle time from the equation Ts/TC = LI(0)/GF:this gave a mean cell cycle time (TC") of 29 hr (range: 20-45hr). Except for the GF, the cell kinetics were comparable to those of the same cell line grown in monolayer culture. The GF in the in vitro system described is in the lower range reported in some human malignant gliomas in vivo.

Alcian blue staining during the formation of mesoblast in the primitive streak stage chick blastoderm.

The distribution of alcian blue (AB) positivity, and its sensitivity to streptococcal and testicular hyaluronidase, were studied in primitive streak stage chick blastoderms. Accumulation of hyaluronate was observed in deep layer (DL) cells and on laterally migrating middle layer (ML) cells. During the formation of the middle layer, a first stage, namely de-epithelialization of the upper layer cells, is recognized and correlated with the absence of hyaluronate. A second stage, namely migration of the de-epithelialized upper layer cells laterally to the edge of the area pellucida, is correlated with the accumulation of AB-positivity. The AB-staining also demonstrated the accumulation of both sulphated and not-sulphated mucopolysaccharides, where a basal lamina is present.

Somatic sensory cortex (SmI) of the prosimian primate Galago crassicaudatus: Organization of mechanoreceptive input from the hand in relation to cytoarchitecture

Mechanoreceptive input from the hand to the somatic sensory cortex (SmI) of the prosimian primate Galago crassicaudatus was examined with microelectrode mapping methods. In anesthetized animals, low threshold cutaneous input from the hand projects to SmI cortex in a single, complete, somatotopically organized pattern. Within this single pattern, cells with receptive fields on the glabrous skin of the palm, digits and digit tips are located in the rostral half, and cells with RFs on the hairy skin of the dorsal hand and digits are located in the caudal half of the hand areas. The cutaneous hand area is coextensive with the densely granular architectonic region of SmI. Studies of single cells in this region of awake galagos reveal the same pattern of cutaneous input and, in addition, demonstrate the presence of cells responding to joint movement not detected in anesthetized animals. Cells responsive to joint movement are arranged in vertically oriented columns located adjacent to cutaneous columns with receptive fields on the same part of the hand. In anesthetized animals, cells rostral to the granular region, in an area typified by increasing numbers of pyramidal cells in layer V and decreasing numbers of granular cells in upper layers, respond to high threshold stimulation of large areas of the hand. The few cells isolated in this area in awake animals respond to either active or passive hand movements. In such animals, cells caudal to the granular region, in an area characterized as agranular and alaminar cortex, respond to either passive stimulation of single or multiple joints or to active hand movements. These results, together with similar findings in a related prosimian, Nycticebus coucang, emphasize the generality of a single cutaneous hand area in SmI of prosimian species. The demonstration of multiple hand areas corresponding to multiple cytoarchitectonic subdivisions in SmI of Old and New World simians illustrates the increased degree of SmI differentiation from the prosimian to the simian grade of organization. The present results further suggest that determination of the homologues of multiple areas or subdivisions within and surrounding SmI in primates will require comparisons of somatotopy, submodality, sulcal patterns, cytoarchitecture, and connectivity in representative members of prosimian and simian families.

Rehabilitation following early malnutrition in the rat: Body weight, brain size, and cerebral cortex development

Sprague-Dawley rats were malnourished by giving their mothers an 8% casein diet starting at day 10 of gestation, while controls were fed a 24% casein diet. Starting at postnatal day 20 (P20), rehabilitation of the malnourished animals was attempted by: (1) feeding both mother and young a 24% casein diet, (2) leaving the pups with their mothers until they were 40 days old, and (3) reducing the litter size from 8 to 4 pups. Observations were made on aldehyde-perfused tissue from animals 20, 40 and 70 days old. The somatosensory cortex from one hemisphere was embedded in Araldite, and that from the other side was processed for Golgi staining. At 20 days of age the body weight of the malnourished animals was 21% that of the controls, but at 70 days it was no longer different. The anterior-posterior length, the width, and the height of the cerebral hemispheres were also significantly reduced at P20, but the differences had disappeared by P70. The thickness of area 3 of the cerebral cortex was measured in 1 μm sections. It was significantly reduced in the malnourished animals at P20, but at P40, following rehabilitation, the difference was no longer statistically significant. In tangential 1 μm sections the fraction of the volume of tissue occupied by neuropil was measured in layers II through IV. At P20 it was significantly reduced only in the upper half of layers II/III of the malnourished animals; at P40 this difference was no longer present. The mean volume of upper layer II/III cell bodies was estimated and found to be significantly reduced in the experimental animals at P20 but not at P40. In the Golgi preparations, pyramidal cells in upper layer II/III were studied. Their estimated volume, as well as the thickness of their basal dendrites, was significantly reduced in the 20 day malnourished animals, but not in the rehabilitated animals. These results show that animals severely malnourished until 20 days of age can reach normal body weight and attain cerebral hemispheres of normal size when proper nutrition is provided. The effects of malnutrition on the cerebral cortex of these animals are most apparent in upper layer II/III which, during the time of nutritional restriction, is the least developed of the cortical layers. However, when proper nutrition is provided, the cerebral cortex may attain normal morphology.

Morphological correlates of visual field transformation in the corpus callosum

Horseradish peroxidase (HRP) injections at the boundary of areas 17 and 18 in cat visual cortex resulted in retrograde labelling of neurones located in contralateral areas 17 and 18. The HRP-positive neurones were spread over a region located around the representation of the vertical meridian; they were most numerous in the locus for area centralis. Layer III pyramidal cells and stellate cells in upper layer IV were identified as the principal neurones of origin of the visual callosal pathway.

Responses to visual stimulation and relationship between visual, auditory, and somatosensory inputs in mouse superior colliculus.

The superior colliculus was studied in anesthetized mice by recording from single cells and from unit clusters. The topographic representation of the visual filed was similar to what has been found in other mammals, with the temporal part of the contralateral visual field projecting posteriorly and the inferior visual field projecting laterally. At the anterior margin of the tectum receptive fields recorded through the contralateral eye and invaded the ipsilateral visual hemifield for up to 35 degrees, suggesting that the entire visual field through one eye is represented on the contralateral superior colliculus. Cells located closest to the tectal surface had relatively small receptive fields, averaging 9 degrees in center diameter; field sizes increased steadily with depth. The prevailing cell type in the stratum zonal and superficial gray responded best to a small dark or light object of any shape moved slowly through the receptive-field center or to turning a small stationary spot on or off. Large objects or diffuse light were usually much less effective. Less than one-quarter of superficial layer cells showed directional selectivity to a moving object, the majority of these favoring up and nasal movement. The chief visual cell type in the stratum opticum and upper part of the intermediate gray resembled in the newness neurons described for many other vertebrates: they had large receptive fields and responded best to up and nasal movement of a small dark or light object, whose optimal size was similar to the optimum for upper-layer cells. If the same part of the receptive field was repeatedly stimulated there was a marked tendency to habituate. Only very few cels responded to the ipsilateral eye. Intermixed with visual cells in the upper part of the intermediate gray were cells that responded to somatosensory or auditory stimuli. Here bimodal and trimodal cells were also seen. In deeper layers somatosensory and auditory modalities tended to take over. These two modalities were not segregated into sublayers but rather seemed to be arranged in clusters. Responses to somatosensory and auditory stimuli were brisk, showing little habituation to repeated stimulation.