RESUMEN

The distribution of cytoskeleton elements (microtubules and actin filaments) and SGLT1 or GLUT2 glucose transporter in enterocyte of rat intestine and Caco2 cell during hexose absorption has been considered. The alteration of SGLT1 and GLUT2 transporter distribution in absorptive cell of intestine villus depending on maltose concentration has been determined using the confocal microscope. The colocalization of the transporters and actin has been revealed. The increase of vesicles number close to microtubules in the apical part of cell during absorption of high hexoze concentration has been found by electron microscope. The fact together with the transporter and actin as well as actin and α-tubulin colocalizations can prove the participation of cytoskeleton elements in glucose transporter movement to apical membrane of the cells studied.

Asunto(s)

Citoesqueleto/metabolismo , Transportador de Glucosa de Tipo 2/metabolismo , Maltosa/metabolismo , Transportador 1 de Sodio-Glucosa/metabolismo , Actinas/metabolismo , Animales , Transporte Biológico , Células CACO-2 , Citoesqueleto/ultraestructura , Enterocitos/metabolismo , Enterocitos/ultraestructura , Expresión Génica , Transportador de Glucosa de Tipo 2/genética , Humanos , Absorción Intestinal/fisiología , Intestino Delgado/metabolismo , Intestino Delgado/ultraestructura , Masculino , Cultivo Primario de Células , Ratas , Ratas Wistar , Transportador 1 de Sodio-Glucosa/genética , Tubulina (Proteína)/metabolismo

RESUMEN

The results of this work concerning ultrastructural changes of U-937 cells in a state of apoptosis are largely in consistent with the same information available in the literature. However, we have got the original data on the ultrastructural changes of cell organelles and immune localization and distribution of proteasomes. It has been demonstrated that Golgi apparatus is located close to the plasma membrane in the case of apoptosis induced by incubating the cells in a hypertonic suchrose solution (200-400 mM). The fact can be considered as an indirect indication of depolymerization of cytoskeletal elements, in particular, MTs maintaining Golgi apparatus in a cell centre. In the later stages of apoptosis, the distances between Golgi cisterna are significantly increased. It can be explained by hydrolysis of golgins binding cisterna between each other. Mitochondria are not significantly changed in these cells. They have regularly disposed crista and sufficiently dense matrix with a few vacuoles. Proteasomes as rod-shaped osmiophilic particles (12 x 30 nm) have been revealed during each apoptosis stage both in nuclei and cytopl;asm of cells studied. The particles form aggregates of different densitities and sizes unlimited by membrane. It has been proposed that the particle aggregates revealed in the work are analogous to "processing bodies" or aggresomes described in the literature. They can be detected in cells under conditions of suppressed nucleus transcriptional processes in the nucleus and participate in storing and degradation of various mRNAs, RNP and proteins. The changes of intracellular contents of Na and K in a single cell during apoptosis induced by osmotic shock have been revealed using method of X-ray microanalysis. It has been demonstrated the increase in the ratio of intracellular contents Na+/K+ in the most of apoptotic cells in comparing with control cells.

Asunto(s)

Apoptosis/efectos de los fármacos , Soluciones Hipertónicas/farmacología , Sacarosa/farmacología , Citoesqueleto de Actina/química , Citoesqueleto de Actina/efectos de los fármacos , Cationes Monovalentes , Núcleo Celular/ultraestructura , Aparato de Golgi/ultraestructura , Humanos , Transporte Iónico , Microscopía Electrónica , Mitocondrias/ultraestructura , Potasio/metabolismo , Complejo de la Endopetidasa Proteasomal/ultraestructura , Análisis de la Célula Individual , Sodio/metabolismo , Células U937 , Espectroscopía de Absorción de Rayos X/métodos

RESUMEN

Investigation of the structure of ordered protein aggregates--amyloid fibrils, the influence of the native structure of the protein and the environment on the process of fibrillation is currently the subject of intensive research. The present work is devoted to the study of the kinetics of insulin amyloid fibrils formation at low pH values (which are produced at many stages of the isolation and purification of the protein) using a fluorescent probe thioflavin T (ThT). It has been shown that the increase of fluorescence intensity of ThT during the formation of amyloid fibrils is described by a sigmoidal curve, in which 3 areas can be distinguished: the lag phase, the growth and the plateau, which characterize the various stages of fibril formation. Despite the variation in the length of the lag phase at the same experimental conditions (pH and temperature), we have found its reduction with stirring the solution and seeding. Data obtained using electron microscopy showed that the formed fibrils are long, linear filament having a diameter of -20 nm. With increasing incubation time fibril diameter did not change while their length increases to 2-3 µm, which was accompanied by a significant increase in the number of aggregates of fibrils. All the experimental data shows that, regardless of the kinetics of the formation of amyloid fibrils, their properties after the fibrillation process are identical. The results of this work together with the previously studies of insulin amyloid fibrils might be important for clarification the mechanism of their formation, as well as for the treatment of amyloidosis associated with the aggregation of insulin.

Asunto(s)

Amiloide/química , Insulina/química , Amiloide/ultraestructura , Benzotiazoles , Colorantes Fluorescentes , Concentración de Iones de Hidrógeno , Cinética , Agregado de Proteínas , Soluciones , Temperatura , Termodinámica , Tiazoles

RESUMEN

Distribution of SGLT1 and GLUT2 hexose transporters as well as that of fibrillar actin and tight junction proteins in cultured Caco2 cells incubated in medium with different hexose concentrations has been considered. Glucose absorption by the cells from incubation medium has been determined. Fibrillar actin was concentrated in the microvilli and closely to tight junction. The actin distribution was not dependent on the glucose concentration. There was no SGLT1 association with brush border actin and the transporter localization was not dependent on the concentration of hexose. GLUT2 was localized in the basal part of Caco2 cells after low concentration hexose load (2.5 mM). The transporter was colocalized with microvilli actin in the apical part of the cells after high concentration hexose load (25 mM). The tight junction proteins, occludin and claudin 1, 3, 4 were not dependent on glucose concentration. Claudin 2 was not detected in Caco2 cells. Caco2 cell culture can be used as a model for studying of hexose transport in small intestine epithelium.

Asunto(s)

Glucosa/metabolismo , Glucosa/farmacología , Mucosa Intestinal/citología , Mucosa Intestinal/metabolismo , Microvellosidades/metabolismo , Actinas/metabolismo , Transporte Biológico , Células CACO-2 , Claudinas/metabolismo , Expresión Génica/efectos de los fármacos , Transportador de Glucosa de Tipo 2/genética , Transportador de Glucosa de Tipo 2/metabolismo , Humanos , Técnicas In Vitro , Proteínas de la Membrana/metabolismo , Ocludina , Transportador 1 de Sodio-Glucosa/genética , Transportador 1 de Sodio-Glucosa/metabolismo , Uniones Estrechas/metabolismo

RESUMEN

The ultrastructural and immunocytochemical study of rat male germ cells on different developing stages has been made. The investigation of morphological changes of spermatogenic cells has demonstrated the presence of tight connections between chromatoid bodies (CBs) and other cell organelles, particularly with the nucleus and Golgi apparatus; has revealed the association of manchette noncentrosomal microtubules (MT) with spermatid perinuclear ring plasma membrane (PM) in the zone of the adhesion intercellular contact--zonula adhaerens (ZA). The comparison of the results obtained in this work with available literary data has given possibility to analyze expected pathways of noncentrosomal MT nucleation in the late spermatids. This paper puts the supposition that noncentrosomal MTs are nucleated on the sites of perinuclear ring ZA. The immunocytochemical analysis discovered two novel proteins for these cells--BASP1 and MARCKS. It has been shown that these proteins present in the CBs in the early spermatids. During the spermatozoid differentiation these proteins are revealed along the outer dense fibers (ODFs) of the sperm tail. BASP1 and MARCKS are supposed to involve in the processes of calcium accumulation in the CBs and ODFs. Calcium ions seem to play the significant role in RNA processing and protein synthesis in spermatids. Calcium is also necessary for the mobility of sperms which is mainly determined by ODFs.

Asunto(s)

Proteínas de Unión a Calmodulina/genética , Proteínas del Citoesqueleto/genética , Citoesqueleto/ultraestructura , Péptidos y Proteínas de Señalización Intracelular/genética , Proteínas de la Membrana/genética , Proteínas del Tejido Nervioso/genética , Espermátides/ultraestructura , Espermatogénesis/fisiología , Uniones Adherentes/fisiología , Uniones Adherentes/ultraestructura , Animales , Calcio/fisiología , Diferenciación Celular , Núcleo Celular/fisiología , Núcleo Celular/ultraestructura , Citoesqueleto/fisiología , Regulación del Desarrollo de la Expresión Génica , Aparato de Golgi/fisiología , Aparato de Golgi/ultraestructura , Humanos , Inmunohistoquímica , Masculino , Microscopía Electrónica , Sustrato de la Proteína Quinasa C Rico en Alanina Miristoilada , Ratas , Ratas Wistar , Cola del Espermatozoide/fisiología , Cola del Espermatozoide/ultraestructura , Espermátides/fisiología

RESUMEN

Continuous cell lines originating from the kidney collecting duct represent a powerful tool for the modeling of water and ions reabsorbtion processes. Present review considers the basic methodical approaches being utilized to study vasopressin-induced water transport mechanisms in the cell culture conditions--microscopical methods, electrophysiological measurements, various ways of evaluation of water flow across the cell monolayer, transfections of native and mutant proteins, GFP-technology. The results of the highest significance for the understanding of collecting ducts function which were obtained with usage of these methods are analyzed in the review.

Asunto(s)

Túbulos Renales Colectores/fisiología , Modelos Biológicos , Animales , Línea Celular , Perros , Humanos , Túbulos Renales Colectores/citología

RESUMEN

SGLT1 and GLUT2 hexose transporter distribution into enterocytes of small intestine isolated loop and Caco2 cell culture after absorption of high and low hexose concentrations has been considered. SGLT1 was found along intestine villus edge in isolated loop. After high concentration hexose load GLUT2 appeared to be situated in the apical parts of enterocytes. It is evident that GLUT2 participates in hexose transport across apical membrane. Cultured Caco2 cells form microvilli and cell junction complex typical for enterocyte. Glucose and galactose absorption by the cells from incubation medium has been observed. SGLT1 transporter is situated in the apical parts and around the nuclens of Caco2 cells and combined into globules. After low concentration hexose load, CLUT2 transporter is localized in the basal parts of Caco2 cells. Caco2 cell culture can be used as a model for studying of hexose transport in small intestine epithelium.

Asunto(s)

Enterocitos/metabolismo , Transportador de Glucosa de Tipo 2/metabolismo , Hexosas/metabolismo , Absorción Intestinal , Intestino Delgado/metabolismo , Transportador 1 de Sodio-Glucosa/metabolismo , Animales , Células CACO-2 , Humanos , Intestino Delgado/citología , Masculino , Microscopía Confocal , Microscopía Electrónica , Ratas , Ratas Wistar

RESUMEN

Alpha-Crystallin type heat shock protein (alpha-HSP) IbpA from Acholeplasma laidlawii was expressed in Escherichia coil and isolated from cell extract on Ni-sepharose column. Recombinant IbpA, like other alpha-HSPs, spontaneously formed oligomeres in vitro. High resolution electron microscopy revealed regular structures with 15 nm in diameter. Evaluation of molecular mass of IbpA oligomers was performed by gel filtration. Most of oligomers consist of 24 subunits. Recombinant IbpA prevents heat denaturation of soluble proteins in cell extract of E. coli and displays a mild positive effect on thermotolerance of E. coli cells during severe heat shock. We investigated a localization of IbpA in A. laidlawii cell by immunocytochemistry. We suppose that IbpA may protect various intracellular structures from damage during heat shock.

Asunto(s)

Acholeplasma laidlawii/metabolismo , Proteínas Bacterianas/metabolismo , Multimerización de Proteína , alfa-Cristalinas/metabolismo , Acholeplasma laidlawii/genética , Proteínas Bacterianas/genética , Escherichia coli/genética , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , alfa-Cristalinas/genética

RESUMEN

Localization of the protein FtsZ in Mycoplasma hominis cells was determined. Ultra thin sections were treated by rabbit polyclonal antibodies against FtsZ M. hominis: a conjugate of protein A with colloidal gold particles was used instead of secondary antibodies. Considerable polymorphism of cells was seen on electron microscopy pictures of M. hominis cells, which is typical for mycoplasmas. Among a wide variety of cell shapes we distinguished dumbbell-shaped dividing cells, and the cells connected with each other with the aid of thin membrane tubules (former constrictions). Dominants distribution of the label in the constriction area of dividing M. hominis cells and in the area of the thin membrane tubules was observed. We revealed the cross septum in the mycoplasma cells for the first time, as well as the gold labeling of this structure. Furthermore, in some rounded and oval cells colloidal gold particles labeled the whole plasma membrane in ring-shaped manner. Probably, the label in these cases marks a submembrane contractile ring (Z-ring). The facts mentioned above confirm that FtsZ of M. hominis plays an active role in the mycoplasma cytokinesis. In a series of cases spiral-like distribution of gold particles was observed. Probably, FtsZ protofilaments in M. hominis cells can form spiral structures similar to Z-spirals of Bacillus subtilis and Escherichia coli. Its presence in mycoplasma cells may be considered as an important argument in favour of model of Z-ring assembling through reorganization of Z-spirals. FtsZ also may participate in maintenance of mycoplasma cell shape (membrane localization).

Asunto(s)

Proteínas Bacterianas/metabolismo , Proteínas del Citoesqueleto/metabolismo , Mycoplasma hominis/metabolismo , Proteínas Bacterianas/ultraestructura , Proteínas del Citoesqueleto/ultraestructura , Microscopía Inmunoelectrónica , Mycoplasma hominis/ultraestructura

RESUMEN

Alterations in the structural organization of MDCK cells under arginin-vasopressin (AVP) action were studied by electron and fluorescent microscopy. Electron-microscopical evidence was obtained that MDCK cells within monolayer form structurally distinct apical and basolateral surfaces separated by well-developed intercellular contact zones. It was proved that AVP specifically bound to the receptors on the basolateral surface of the cells, and was internalized from the surface in 10-15 min. AVP action resulted in fragmentation of Golgi apparatus and swelling of Golgi cisterns caused by initiation of osmotic water flow across the monolayer. Significant depolymerization of cells actin cytoskeleton was revealed under AVP or forskolin (an adenylate cyclase activator) exposure. Functional role and regulatory mechanisms of described structural alterations are discussed.

Asunto(s)

Arginina Vasopresina/farmacología , Células Epiteliales/efectos de los fármacos , Actinas/metabolismo , Adenilil Ciclasas/metabolismo , Animales , Arginina Vasopresina/metabolismo , Transporte Biológico , Línea Celular , Colforsina/farmacología , Perros , Células Epiteliales/ultraestructura , Aparato de Golgi/metabolismo , Aparato de Golgi/ultraestructura , Microscopía Electrónica , Microscopía Fluorescente , Ósmosis , Receptores de Vasopresinas/metabolismo , Agua/metabolismo

RESUMEN

This review summarizes modem data on Golgi apparatus of parasitic protists and demonstrates how the parasitic lifestyle determines functional and structural peculiarities of secretory systems in unrelated groups of unicellular parasites, in comparison to ones of "model systems", mammalian and yeast cells. The review covers the most well-studied protists, predominantly of high medical importance, belonging to following taxons: Parabasalia (Trichomonas), Diplomonada (Giardia), Entamoebidae (Entamoeba), parasitic Alveolata of the phyllum Apicomplexa (Toxoplasma and Plasmodium), and Kinetoplastida (Trypanosoma and Leishmania). Numerous recent publications demonstrated that studies on intracellular traffic in the mentioned above parasites essentially advanced our knowledge of Golgi function, traditionally based on research of cultured mammalian and yeast cells. Morphology of Golgi organelle in eukaryotes from various taxonomic groups has been compared. Within three of total six the highest taxons of Eukatyota (Adl et al., 2005) there exist at minimum eight groups represented by species lacking Golgi dictiosomes. However, biochemical and (or) molecular (genomic) evidences indicate that the organelle with functions of Golgi was present in every studied so far lineage of eukaryotes. Loss of Golgi organelle is a secondary event, which has been proven by identification of Golgi genes in the genomes of Golgi-lacking lineages. This loss might have occurred independently several times in the course of evolution. Neither the number of stacks, nor the size of the organelle correlates with intensity of secretion, or the position of the species on the evolutionary tree (in terms of presumably early/lately diverged lineages).

Asunto(s)

Eucariontes/citología , Aparato de Golgi/fisiología , Animales , Evolución Biológica , Transporte Biológico , Endosomas/metabolismo , Células Eucariotas/metabolismo , Aparato de Golgi/ultraestructura , Membranas Intracelulares/metabolismo , Proteínas Protozoarias/metabolismo , Levaduras/metabolismo

RESUMEN

This review is dedicated to the structure and function of Golgi apparatus (GA). It summarizes contemporary data published in numerous experimental papers and in several reviews. Possible ways of intra-Golgi transport of proteins, existent models of structural and functional organization of Golgi organelle, as well as the issues of its biogenesis, posttranslational modification and sorting of proteins and lipids, and mechanisms of their traffic-king are discussed. Special attention is paid to the role of coatomer proteins (COPI, COPII and clathrin), fusion proteins (SNAREs), and small GTPases (ARF, SARI) in the secretory pathway. In addition, the phenomena of ultrastructural alterations of GA due to various functional conditions and physiological stimuli are specifically accented. We included in this review our original data on a probable involvement of GA in water transport, and on the organization of atypical GA in microsporidia--intracellular parasitic protists.

Asunto(s)

Aparato de Golgi , Animales , Proteína Coatómero/fisiología , GTP Fosfohidrolasas/fisiología , Aparato de Golgi/química , Aparato de Golgi/metabolismo , Aparato de Golgi/ultraestructura , Metabolismo de los Lípidos , Microscopía Electrónica , Microsporidios/ultraestructura , Células Vegetales , Transporte de Proteínas , Proteínas SNARE/fisiología , Agua/metabolismo

RESUMEN

To elucidate mechanisms providing transport of sugars across intestinal epithelium, on taking into account the current hypotheses (active transport, participation of paracellular transport and passive component of transcellular transport), it was important to reveal structural changes of tight junctions and distribution of the carriers of facilitated diffusion of GLUT2 and protein kinase C during absorption of glucose. On using confocal and electron microscopy, ultrastructural and immunocytochemical studies of enterocytes after perfusion of isolated rat small intestine fragment with 75 mM glucose (chronic experiment) have shown: 1) fluorescent labels of transporter GLUT2 and PKCbetaII are located in the apical area of enterocytes situated at the upper half of the villus. Antibodies against GLUT2, conjugated with gold, are revealed at the microvilli or apical membrane and in the area of terminal network; 2) no ultrastructural changes of the tight junction are detected on ultrathin sections and freeze--fracture replics. At the same time, fluorescent and gold labels against actin are concentrated in the vicinity of the lateral membrane in the tight junction area. The results obtained can serve a confirmation of a hypothesis that at high glucose concentrations GLUT2 participates in its transfer across the apical membrane.

Asunto(s)

Enterocitos/metabolismo , Transportador de Glucosa de Tipo 2/metabolismo , Glucosa/metabolismo , Intestino Delgado/metabolismo , Proteína Quinasa C/metabolismo , Animales , Transporte Biológico , Desmosomas/metabolismo , Desmosomas/ultraestructura , Difusión , Enterocitos/ultraestructura , Inmunohistoquímica , Absorción Intestinal , Intestino Delgado/ultraestructura , Microscopía Confocal , Microscopía Electrónica de Transmisión , Proteína Quinasa C beta , Ratas , Ratas Wistar , Uniones Estrechas/metabolismo , Uniones Estrechas/ultraestructura

RESUMEN

In chronic experiments on Wistar rats, glucose and galactose absorption in the isolated loop of the small intestine considerably decreased in presence of both phloridzine am phloritine (inhibitors of the glucose transporters SGLT1 and GLUT2). The load of the isolated loop with glucose or galactose solutions scarcely influenced the absorption of 2-deoxi-D-glucose (substrate for GLUT2). According to the immunocytochemical analysis by means of confocal microscopy, after the load of the isolated loop with glucose (75 mM) the labels to GLUT2 and proteinkinase C (PKC betalI) were concentrated mainly in the apical part of the enterocytes, whereas after the load with the Ringer solution--in the basal part of the enterocytes. It was shown on the mathematical model that the part of the facilitated diffusion in the total glucose absorption was considerably lesser in comparison with the active transport mediated by SGLT1. Thus the findings support the hypothesis about a recruitment of the transporter GLUT2 into the apical membrane of the enterocytes and its involvement in glucose transfer across this membrane. However, under natural conditions, the active transport is the main mechanism of glucose absorption, whereas the facilitated diffusion plays a certain role only at high carbohydrate loads.

Asunto(s)

Enterocitos/metabolismo , Glucosa/metabolismo , Intestino Delgado/metabolismo , Animales , Transporte Biológico , Membrana Celular/metabolismo , Desoxiglucosa/metabolismo , Difusión , Galactosa/metabolismo , Transportador de Glucosa de Tipo 2/antagonistas & inhibidores , Transportador de Glucosa de Tipo 2/fisiología , Inmunohistoquímica , Técnicas In Vitro , Absorción Intestinal , Masculino , Microscopía Confocal , Modelos Biológicos , Floretina/farmacología , Florizina/farmacología , Ratas , Ratas Wistar , Transportador 1 de Sodio-Glucosa/antagonistas & inhibidores , Transportador 1 de Sodio-Glucosa/fisiología

RESUMEN

This review is dedicated to the structure and function of Golgi apparatus (GA). It summarizes contemporary data published in numerous experimental papers and in several reviews. Possible ways of intra-Golgi transport of proteins, existent models of structural and functional organization of Golgi organelle, as well as the issues of its biogenesis, posttranslational modification and sorting of proteins and lipids, and mechanisms of their trafficking are discussed. Special attention is paid to the role of coatomer proteins (COPI, COPII and clathrin), fusion proteins (SNAREs), and small GTPases (ARF, SARI) in the secretory pathway. In addition, the phenomena of ultrastructural alterations of GA due to various functional conditions and physiological stimuli are specifically accented. We included in this review our original data on a probable involvement of GA in water transport, and on the organization of atypical GA in microsporidia--intracellular parasitic protists.

Asunto(s)

Aparato de Golgi/fisiología , Aparato de Golgi/ultraestructura , Animales , Proteínas Portadoras/metabolismo , Clatrina/metabolismo , Proteína Coat de Complejo I/metabolismo , GTP Fosfohidrolasas/metabolismo , Gryllidae/parasitología , Metabolismo de los Lípidos , Microsporidios/citología , Microsporidios/ultraestructura , Proteínas/metabolismo , Proteínas SNARE/metabolismo , Agua/metabolismo

RESUMEN

Morpho-physiological characteristics of the transport of cyclic nonapeptide arginine vasopressin (AVP) across the rat intestinal epithelium was studied in experiments in vitro. A partial absorption of physiologically active AVP was followed when filling the isolated intestinal lumen by hormone solution. By methods of immunoelectron and immunofluorescence confocal microscopy, using polyclonal anti-AVP antibodies, cytoplasmic localization of AVP label was shown in enterocytes. The AVP label was also observed in the intercellular space in the basal area of epithelium. No label was revealed in the intercellular junctions, and no predominant label accumulation was found in any cytoplasmic structures of the epithelial cells. The obtained results are considered as evidence for the transcellular pathway of partial AVP absorption in rat small intestine.

Asunto(s)

Arginina Vasopresina/metabolismo , Mucosa Intestinal/metabolismo , Intestino Delgado/metabolismo , Animales , Transporte Biológico , Citoplasma/metabolismo , Enterocitos/metabolismo , Espacio Extracelular/metabolismo , Inmunohistoquímica , Técnicas In Vitro , Absorción Intestinal , Microscopía Confocal , Microscopía Inmunoelectrónica , Ratas , Ratas Wistar

RESUMEN

To elucidate the mechanism of glucose absorption at high substrate concentrations, we studied structural and ultrastructural peculiarities of enterocytes arranged at different levels along the intestinal villus. The preparations were obtained from an isolated segment of the rat small intestine after its perfusion with maltose solutions with both low (25 mM) and high (100 mM) concentrations, respectively. Under conditions of chronic experiment at high substrate concentration, an enlargement of intercellular clefts, indicating glucose absorption, occurred in deeper areas of the villus. Besides, also in chronic experiment, we studied kinetics of maltose hydrolysis and derived glucose absorption in the isolated segment of the rat small intestine after its perfusion with maltose at superhigh (up to 200 mM) initial concentrations. Based on these data, a conclusion is made that active transport is the main mechanism of absorption of glucose derived from maltose hydrolysis, operating both at low disaccharide concentrations, and in the range of its superhigh (up to 200 mM) concentrations.

Asunto(s)

Enterocitos/metabolismo , Glucosa/metabolismo , Intestino Delgado/metabolismo , Maltosa/metabolismo , Animales , Transporte Biológico Activo , Enterocitos/ultraestructura , Absorción Intestinal , Intestino Delgado/ultraestructura , Masculino , Maltosa/administración & dosificación , Microscopía Electrónica , Perfusión , Ratas , Ratas Wistar

RESUMEN

Microtubules (MTs) are necessary components of all eukaryotic cells. They fulfill various functions being involved in cell division, ciliar and flagellar beating, cell shape maintaining, organelle distribution in the cell, organization of other cytoskeletal elements. Dynamic features of MTs have been commonly studied in vitro or on undiffirentiated cultured cells by means of molecular and ultrastructural methods. It is generally accepted that the phenomenon of dynamic instability is the major mechanism of MT turnover in the cell. MTs radiate from the centrosome and take part in the distribution of cell organelles. In addition, epithelial, nerve, and skeletal muscle cells contain non-centrosomal MTs. A few hypothesis of their origin have been so far put forward. According to the capture-release hypothesis, MTs are first nucleated on the a centrosome, then release to be driven in various parts of the cell by molecular motors. Some alternative mechanisms of non-centrosomal MT formation are also proposed in literature. For example, the nucleation sites were reported not only in centrosomes but also in other parts of cells, such as the apical membranes of epithelial cells, the nuclear membrane of muscle cells, pigment granule aggregates of melanophores. On studying frog urinary bladder and large intestine epithelial cells the authors observed in these cells numerous non-centrosomal MTs. This makes epithelial cells, good models for analysing structural and dynamic features of non-centrosomal MTs in differentiated cells. For the urinary bladder the pool of specific granules may serve as MT organizing centers. Non-cenrosomal MTs of these cells have big diameters (35-38 nm) and form bundles oriented in the apical-basal axis of the cell. In addition, non-centrosomal MTs of these cells may participate in the transport of specific granules and giant vacuoles that appear under stimulated water flows through the cell.

Asunto(s)

Células Epiteliales/fisiología , Microtúbulos/fisiología , Animales , Anuros , División Celular , Membrana Celular/fisiología , Centrosoma/fisiología , Cilios/fisiología , Gránulos Citoplasmáticos/fisiología , Gránulos Citoplasmáticos/ultraestructura , Células Epiteliales/ultraestructura , Flagelos/fisiología , Microtúbulos/ultraestructura , Orgánulos/fisiología , Vejiga Urinaria/ultraestructura

RESUMEN

Three cell types have been revealed in the epithelium of the frog large intestine: granular, mitochondria-rich, and mucosal cells. Under a low water permeability (0.12 +/- 0.10 mkl/(min.cm2)) the distribution of intramembrane particles (IMP) in the apical cell membrane was the same as in the most cell plasma membranes studied with freeze-fracture method. Under rising osmotic permeability and water absorption (0.43 +/- 0.05 mkl/(min.cm2)) the IMP distribution did not change. In these conditions, the quantity of fusion sites between granule membranes and the apical membrane increased, and the intercellular spaces in basolateral epithelial region were diluted. A a low water permeability, in addition to usual microtubules, bundles of noncentrosomal microtubules with associated osmiophilic globules were revealed. A comparative analysis has been made of the present evidence and previously obtained data on the frog urinary bladder epithelium.

Asunto(s)

Absorción Intestinal , Mucosa Intestinal/citología , Intestino Grueso/fisiología , Agua/metabolismo , Animales , Técnica de Fractura por Congelación , Inmunohistoquímica , Mucosa Intestinal/metabolismo , Mucosa Intestinal/ultraestructura , Microscopía Electrónica , Concentración Osmolar , Rana temporaria

RESUMEN

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.

Asunto(s)

Urotelio/ultraestructura , Agua/metabolismo , Animales , Microanálisis por Sonda Electrónica , Técnica de Fractura por Congelación , Microscopía Electrónica , Ósmosis , Rana temporaria , Urotelio/citología , Urotelio/efectos de los fármacos , Urotelio/metabolismo , Vasopresinas/farmacología