Structural asymmetry in the frog retinal ganglion cell layer

The density of distribution and topographical features of small and large ganglion cells were investigated in total silver-impregnated preparations of the retina from two species of frogs (Rana ridibunda andR. temporaria). A horizontal band of increased density of ganglion cells was located in both species above the nasotemporal axis passing through the blind spot. Outside this band the density of the small cell population was maximal in the central zone of the retina, decreasing toward the periphery. In the upper halves of the retina the density of small cells was on average 26% greater than in the lower halves. Large ganglion cells, on the other hand, were more densely distributed in the lower half of the retina than in the upper half; this difference was particularly marked inR. temporaria (by 116%). The large cells were asymmetrically distributed relative to the dorsoventral axis also: In the nasal quadrants their density was 40–55% greater than in the temporal. Large cells were more densely distributed in the middle zone of the retina. Signs of asymmetry in the organization of the retinal output raster may be of adaptive ecologic importance and may determine the characteristics of formation of visually controlled food and avoidance behavioral reflexes.Research Institute of Applied Mathematics and Cybernetics, N. I. Lobachevskii State University, Gorkii. Translated from Neirofiziologiya, Vol. 17, No. 2, pp. 198–204, March–April, 1985.     

人羊膜上皮细胞和胚胎干细胞

人胚胎干细胞有着巨大的医学应用前景,但人胚胎干细胞要求的生长条件很高,体外很难模拟其生长的体内环境,因此控制人胚胎干细胞的生长常不理想,而使用鼠胚胎成纤维细胞(MEF)作为滋养层则存在动物源性污染的问题。该文阐述人羊膜上皮细胞(HAEC)的特点及其作为滋养层培养胚胎干细胞的现状,并探讨基因组DNA甲基化修饰在胚胎干细胞分化过程中的作用,为建立更优化的培养系统提供依据。     

Ultrastructure and elemental composition of frog bladder granular epithelial cells in normal state and upon stimulation of water transport

Changes in the frog urinary bladder granular cell ultrastructure were analysed in parallel with those in element composition of these cells after induction of water transport across the urinary bladder wall. Two ultrastructural (ultrathin section and freeze-fracture) methods were used in addition to two methods of object preparation for electron microprobe analysis--freeze-drying and freeze-substitution. It has been shown that arginin-vasotocin stimulation of osmotic water flow across the urinary bladder wall causes certain morphological changes in the granular cells: decrease in electron density of the cytoplasm, depolymerization of the apical submembrane layer of actin microfilaments, increase in the number of sites of specific granules and apical membrane fusion, emergency of intramembrane particle aggregates in the apical membrane P-face. The quantitative electron microprobe analysis made it possible to reveal a statistically significant increase in sodium and calcium concentration and fall in that of potassium and chlorine in granular cells after water transport stimulation. A concentration gradient of sodium and potassium ions was seen to appear along the apical-basal axis in the cytoplasm of granular cells. Possible association between the obvious morphological transformations in granular cells and changes in their elemental composition has been discussed, in addition to some regulatory significance of calcium concentration increase in granular cells after arginin-vasotocin-induced osmotic water transport.     

Characterization of the nuclear deformation caused by changes in endothelial cell shape

We investigated the mechanotransduction pathway in endothelial cells between their nucleus and adhesions to the extracellular matrix. First, we measured nuclear deformations in response to alterations of cell shape as cells detach from a flat surface. We found that the nuclear deformation appeared to be in direct and immediate response to alterations of the cell adhesion area. The nucleus was then treated as a neo-Hookean compressible material, and we estimated the stress associated with the cytoskeleton and acting on the nucleus during cell rounding. With the obtained stress field, we estimated the magnitude of the forces deforming the nucleus. Considering the initial and final components of this adhesion-cytoskeleton-nucleus force transmission pathway, we found our estimate for the internal forces acting on the nucleus to be on the same order of magnitude as previously measured traction forces, suggesting a direct mechanical link between adhesions and the nucleus.     

Distribution of actin microfilaments in frog skin epithelial granular cells

Summary— Microfilaments were localised by immunofluorescence and immunogold cytochemistry to examine their distribution in granular cells of the isolated frog skin epithelium. Strongly fluorescent bundles of actin were observed beneath the plasma membrane with little evidence for actin in the central regions. Higher resolution offered by cytochemistry revealed that bundles of actin filaments comprised a substantial portion of the cortical cytoskeleton. Quantitative analysis of the frequency of gold label revealed an extremely rich array of filaments beneath the apical membrane of granular cells, with markedly less babel along the basolateral membrane and in the central cytoplasm. Treating cells with cytochalasin B or arginine vasopressin caused an apparent disruption of the apical actin fibres, concurrent with a decrease in gold label density. Assumably these signs are indicative of depolymerization of the filaments. Although the significance of this distribution is unknown, the apical polarisation of actin is consistent with a role in regulating the Na⁺ permeability of the apical membrane. The data are discussed in relation to possible roles of the cytoskeleton in the regulation of transepithelial sodium transport by vasopressin.     

Changes in cell attachment by RSV transformation in rat liver epithelial cells.

BRL, a non-malignant rat liver epithelial-like cell line, possessed the ability to adhere through fibronectin to a solid substrate. Oncogenical transformation of these BRL cells with RSV induced a significant decrease in the fibronectin molecules in the extracellular matrix and reduction in its ability to adhere to fibronectin. The alpha 5 and beta 1 subunits of integrin (fibronectin receptor) were quantitatively diminished during RSV transformation in BRL cells. These results suggest that adhesive reduction of BRL cells to a substrate by RSV transformation may be caused by a decrease in cell surface fibronectin and fibronectin receptor molecules.     

Positioning and capture of cell surface-associated microtubules in epithelial tendon cells that differentiate in primary embryonic Drosophila cell cultures

Using primary embryonic Drosophila cell cultures, we have investigated the assembly of transcellular microtubule bundles in epidermal tendon cells. Muscles attach to the tendon cells of previously undescribed epidermal balls that form shortly after culture initiation. Basal capture of microtubule ends in cultured tendon cells is confined to discrete sites that occupy a relatively small proportion of the basal cell surface. These capturing sites are associated with hemiadherens junctions that link the ends of muscle cells to tendon cell bases. In vivo, muscle attachment and microtubule capture occur across the entire cell base. The cultured tendon cells reveal that the basal ends of their microtubules can be precisely targeted to small, pre-existing, structurally well-defined cortical capturing sites. However, a search and capture targeting procedure, such as that undertaken by kinetochore microtubules, cannot fully account for the precision of microtubule capture and positioning in tendon cells. We propose that cross-linkage of microtubules is also required to zip them into apicobasally oriented alignment, progressing from captured basal plus ends to apical minus ends. This involves repositioning of apical minus ends before they become anchored to an apical set of hemiadherens junctions. The proposal is consistent with our finding that hemiadherens junctions assemble at tendon cell bases before they do so at cell apices in both cultures and embryos. It is argued that control of microtubule positioning in the challenging spatial situations found in vitro involves the same procedures as those that operate in vivo.     

Disruption of keratin filaments in embryonic epithelial cell types.

The murine keratins Endo B and Endo A, which are homologs of the human keratins K18 and K8, constitute the intermediate filaments (IFs) that are found in all simple epithelia of the adult and in the first epithelial derivatives of the early embryo. The cellular role of simple epithelial keratins in development and differentiation was investigated by inducing filament collapse in HR9 endoderm and F9 embryonal carcinoma cells in which mutant Endo B protein was constitutively expressed. By immunolocalization techniques a perturbation of the keratin network was revealed as well as concomitant disruption of vimentin IFs and displacement of surface desmosomal proteins, demonstrating an intimate structural association of Endo B/A filaments with these cellular components. In aggregates of differentiating F9 cells displaying altered Endo A/B IFs, the formation of a compact, polarized visceral endoderm layer was significantly compromised. These results indicate that an intact keratin network influences the three-dimensional formation of cell-cell or cell-substratum contacts in embryonic visceral endoderm.     

Overview of cell shape: cytoskeletons shape bacterial cells

An evolving hypothesis is that bacterial cell shape is determined by cytoskeletal elements that localize peptidoglycan synthetic machineries. In most bacteria FtsZ assembles into the Z ring which recruits the machinery necessary for cytokinesis. Most rod shaped cells require MreB which assembles into cables that run between the poles of the cell and distribute various components of peptidoglycan metabolism along the cell length. Cells with other shapes have additional cytoskeletal elements that either localize synthetic machineries or possibly influence their activity.     

Exocytosis upon osmotic swelling in human epithelial cells.

Upon osmotic swelling human epithelial cells exhibited significant increases in the membrane capacitance. Evidence for exocytosis includes its dependency on temperature, cytosolic Ca2+ and ATP as well as its sensitivity to guanosine 5'-O-(3-thio)triphosphate (GTP gamma S) and N-ethylmaleimide (NEM). A role of the osmotic exocytosis in the subsequent cell volume regulation is suggested.     

Changes in physiological properties of frog taste cells following denervation

The structure of the taste organ and the physiological propertiesof the taste cell of the bullfrog (Rana catesbeiana) after theglossopharyngeal nerve transection were investigated. The frogtaste organ is composed of taste, supporting and basal cells.As nerve terminals within the taste organ degenerated, nerveresponses to mechanical, chemical and electrical stimuli graduallydeclined and finally ceased in 7 days during the summer and15 days during the winter. However, the taste cells still hadresting potentials and responded to four basic taste stimuliby generating receptor potentials with the various lengths ofduration, even 140 days after denervation. The glossopharyngealnerve transection affected the magnitude of resting potentialssignificantly. However, the taste, supporting and basal cellsmaintained their normal structures 140 days after surgery. Itis concluded, therefore, that neural dependency of the frogtaste organ is not as great as that in mammals.     

Star-shaped cells of the amacrine cell layer

Using silver impregnation techniques and light microscopy we have described a new type of interneuron in the adult chicken retina. These cells are located in the innermost level of the INL. Expansions arise from the basal portion of the perikaryon and ascend from there, ending in the inner third of the INL. Other expansions extend through the three first sublayers of the IPL. We compared the morphological differences shown by these cells in relation to the amacrine cells and interplexiform cells, all of which are situated on the same stratum.     

Membrane and junctional properties of dissociated frog lens epithelial cells

Summary Individual cells and cell pairs were isolated from frog lens epithelium. Individual cells were whole cell voltage clamped and the current-voltage relationship was determined. The cells had a mean resting voltage of –54.3 mV and a mean input resistance of 1.4 G. The current-voltage relationship was linear near the cell resting voltage, but showed decreased resistance with large depolarization or hyperpolarization. Junctional currents between pairs of cells were recorded using the dual whole cell voltage-clamp technique. The corrected junctional resistance was 15.5 M (64.5 nS). The junctional current-voltage relationship was linear. A combination of ATP and cAMP, in the electodes, stabilized junctional resistance. Currents recorded when uncoupling was nearly complete, showed evidence of single connexon gating events. A single-channel conductance of about 100 pS was prominent. Dye spread between isolated cell pairs was demonstrated using Lucifer Yellow CH in a whole cell configuration. Photodamage to the cells due to the dye was apparent. Dye loaded cells, in the presence of exciting light, showed decreased resting voltages, decreased input resistances and morphological changes. Glutathione (20mm) delayed this damage.     

Ultrastructural changes in the embryonic cells of the frog Microhyla ornata after cytochalasin H treatment

Biological effects of cytochalasin H (CH), a newly isolated mould metabolite, have been found to bring about disaggregation of embryonic cells and to inhibit cytokinesis. Disaggregation is known to be a phenomenon related to the cell surface. (The cells are held together by a mucopolysaccharide glycoprotein complex.) In the present work the fact that the mucopolysaccharide glycoprotein surface coat gets affected by CH treatment is confirmed by electron microscopy with the help of Lanthanum, a specific marker, which gets selectively absorbed to the cell surface material and renders it electron dense. The ultrastructural observations indicated the reduction of the cell surface material in treated embryos as compared to the controls. The reappearance of lanthanum-bound cell surface material in the recovered embryos was also observed. However, the exact mechanism of the action of CH on the cell surface remains to be clarified.     

Compression and dilation of the membrane-cortex layer generates rapid changes in cell shape

Rapid changes in cellular morphology require a cell body that is highly flexible yet retains sufficient strength to maintain structural integrity. We present a mechanism that meets both of these requirements. We demonstrate that compression (folding) and subsequent dilation (unfolding) of the coupled plasma membrane–cortex layer generates rapid shape transformations in rounded cells. Two- and three-dimensional live-cell images showed that the cyclic process of membrane-cortex compression and dilation resulted in a traveling wave of cortical actin density. We also demonstrate that the membrane-cortex traveling wave led to amoeboid-like cell migration. The compression–dilation hypothesis offers a mechanism for large-scale cell shape transformations that is complementary to blebbing, where the plasma membrane detaches from the actin cortex and is initially unsupported when the bleb extends as a result of cytosolic pressure. Our findings provide insight into the mechanisms that drive the rapid morphological changes that occur in many physiological contexts, such as amoeboid migration and cytokinesis.