Localization Of ATPase Activity Research Articles

Localization of ATPase Activity on the Plasmalemma in the Dorsal Tissue of Rice Caryopsis and its Relation to Assimilate Transport

Localization of ATPase Activity on the Plasmalemma in the Dorsal Tissue of Rice Caryopsis and its Relation to Assimilate Transport

Cytochemical localization of ATPase activity in the leaves of a seagrass and a submerged halophyte

Flowering plants that grow submerged in seawater are known as seagrasses, while those that grow submerged in water of generally lower and varying salinity as in estuaries, are known as submerged halophytes. Knowledge of salt tolerance mechanisms in both groups of plants is important to our understanding of their biology. Recently, high ATPase activity was reported to be associated with the copiously-invaginated plasma membrane of leaf epidermal cells of a seagrass (Zostera marina), suggesting that ATPase played a role in salt regulation. In the present study, the seagrass Halophila ovalis and the submerged halophyte Ruppia maritima growing at salinities of 35°/00 and 25°/00 respectively, were used. The subcellular distribution of ATPase in leaf epidermal cells of both submerged aquatics was accomplished using a lead precipitation technique specific for ATPase activity. Lead deposits in the cells were an indication of sites of ATPase activity. Transmission electron microscopy (TEM) and energy dispersive X-ray (EDX) microanalysis using a Joel 6100 SEM with a Noran Voyager 2100 EDX microanalyser, were used to study lead distribution. Ultrathin sections for the TEM were examined unstained.

Localization of ATPase activity in dendritic spines of the cerebral cortex

An ATPase activity has been demonstrated in dendritic spines of the adult rat cerebral cortex using cerium to capture inorganic phosphate that is liberated during the enzymatic hydrolysis of ATP. Small pieces of cerebral cortex were fixed and incubated in a standard incubation medium containing both Ca2+ and Mg2+ at pH 7.2; other modifications of the incubation medium are described below. Electron microscopic examination of the cerium phosphate reaction product showed an electron dense precipitate localized in the cytoplasm of the spine behind the postsynaptic density. Whereas the postsynaptic density, itself, is not reactive, dense reaction product is seen immediately underneath the postsynaptic density and extending into the subsynaptic web. Reaction product is also associated with membranous cisternae within the dendritic spine. The reaction occurred in the presence of Ca2+ and Mg2+ and either of these two ions alone. However, virtually no reaction product is seen when the tissue was incubated in a medium devoid of Ca2+ and Mg2+, or in a medium containing Mg2+ and EGTA, suggesting that trace Ca2+ is necessary, but not sufficient for the reaction. Addition of p-chloromercurobenzoate, which selectively blocks SH groups, inhibited the reaction in the presence of Ca2+ and Mg2+, or both of these ions. The effect of pH on the reaction was determined using a lead precipitation method. The reaction occurred at pH 9.2 in the presence of Ca2+ alone. In the presence of Mg2+ alone, the reaction product appeared somewhat reduced at this pH. The presence of an ATPase activity, which is dependent upon Ca2+ in dendritic spines where actin and actin-binding proteins have also been localized, suggests that this activity may be involved in the dynamics of cytoskeletal function leading to shape changes in dendritic spines and synapses, as seen with various physiological and behavioral paradigms.

Ultrastructural localization of ATPase activity in cotton fiber during elongation

The ultrastructural distribution of potassium chloride stimulated adenosine triphosphatase activity was investigated in the outer integument of a linted cultivar of cotton and a lintless (naked seed) mutant from one day preanthesis to eight days postanthesis by using a heavy metal simultaneous capture reaction technique. No enzyme activity other than in mitochondria was observed in the lintless mutant. In the linted cultivar no ATP-specific enzyme activity was seen in non-elongating epidermal cells, subepidermal cells of the outer integuments or any controls. As fiber initials started elongating, enzyme activity gradually appeared on the tonoplasts of enlarging vacuoles. Heavier lead phosphate deposits were observed on the membrane of small vacuoles compared to the tonoplast. This activity continued at least to eight days postanthesis. The enzyme inhibitor, N,N-dicyclohexylcarbodiimide inhibited, while KCl stimulated, tonoplast ATPase activity. The gradual increase of ATPase activity on the tonoplast of expanding fibers, but not on the tonoplasts of non-fiber cells, suggests the active transport of osmotically active compounds, presumably potassium and malate, into the vacuoles of expanding fibers. Fusion of smaller vacuoles with the large central vacuole indicates that these structures contribute additional membrane components along with their enzyme activity to the tonoplast of expanding fibers. The occurrence of ATPase activity, of ER-derived vesicular structures, and the organized pattern of deposition of these structures on the tonoplast indicate ER-originated ATPase activity. This study supports the theory of osmoregulation in cotton fiber where ATPase provides the energy for active accumulation of osmotically active compounds, (K+, malate) into the vacuoles, thereby generating and maintaining the turgor pressure required for fiber expansion.

Ultracytochemical localization and characterization of membrane-bound ATPases in lateral buds from intact and decapitated plants of an aquatic fern,Marsilea drummondii A. Br.

The electron-microscopic localization of ATPase activity was studied in the median lateral bud-bearing nodes in intact and decapitated sporophytes ofMarsilea drummondii. ATPase activity was confined to the plasma membrane of xylem transfer cells, phloem parenchyma cells and sieve tubes belonging to the stele of the main axis. No activity was found in the bud branch. One of the membrane-bound ATPase components is a Mg2+ ATPase, which is stimulated by K+ and functions optimally at pH 6.5 with ATP as substrate. Numerous inhibitors were tested which restricted or completely inhibited ATPase sites in transfer cells and sieve tubes. The use of CCCP, DCCD, DES, NEM or PCMBS allowed a distinction to be made between these conducting elements which are differentially affected by the inhibitors. The structurally polarized transfer cells remain functional on the side abutting the xylem tracheids but are inoperative on the opposite side,i.e., in phloem loading. The Mg2+-K+ ATPase in the bud trace was rapidly stimulated (in one hour) after decapitation. These results are consistent with a transport role for this ATPase, and support the view that its stimulation is a prerequisite for the resumption of bud activity.

Effects of denervation on the distribution of myosin isozymes in skeletal muscle fibers

The pattern of distribution of myosin isozymes was examined with respect to the fiber population in the denervated rat diaphragm. Myosin was demonstrated by indirect immunofluorescence using antibodies against chicken myosin and by localization of ATPase activity. In the normal diaphragm, a heterogeneous fast-twitch muscle, the component fibers react with antibodies against fast or slow myosin, but usually not with both. By 8 weeks after removal of the nerve supply, all fibers reacted with antibodies specific for fast myosin, and many reacted with anti-slow myosin as well. This indicates either that multiple forms of myosin coexist within individual muscle fibers or that a unique myosin(s) is present which has determinants in common with both fast and slow isozymes. This pattern was observed for at least 24 weeks after denervation. At all stages, the localization of alkali-stable and acid-stable ATPase activity correlated well with the response to anti-fast and anti-slow myosin, respectively. We conclude that the normal adult neuromuscular relationship is required for the preferential distribution of fast and slow myosins into different populations of fibers. New myosin is nevertheless synthesized in the absence of the nervous system, and characteristics of both fast and slow isozymes are evident even after prolonged denervation.

The validity of the lead precipitation technique for the localization of ATPase activity in plant cells

The claim that osmium-containing deposits which lack lead are frequently and incorrectly interpreted as enzymatic reaction products in lead precipitation techniques for ATPase localization in plants is without foundation. Proper controls clearly demonstrate the enzymatic origin of membrane-located deposits and the presence of lead is confirmed by analytical electron microscopy.

Cytochemical localization of ATPase activity in phloem tissues ofRicinus communis

Standard lead precipitation procedures have been used to examine the localization of ATPase activity in phloem tissues ofRicinus communis. Reaction product was localized on the plasma membrane of the companion cells associated with sieve elements and of parenchyma cells in phloem tissues from the leaf, petiole, stem and root. ATPase activity was also present on the plasma membrane and dispersed P-protein of sieve elements in petiole, stem and root tissue, but was absent from the plasma membrane of these cells in the leaf minor veins. Substitution ofβ-glycerophosphate for ATP produced no change in the localization of reaction product in leaf tissue. These findings are discussed in relation to current theories on the mechanism of sugar transport and phloem loading.

Localization of ATPase activity to the glial-like cells of the pars intermedia.

Abstract An adenosine triphosphatase was histochemically localized in the pars intermedia of the pituitary of the frog, Rana berlandieri forrei . The reaction product was found primarily between membranes of stellate (glial-like) cells and secretory cells and could be partially inhibited by ouabain. It was also present between two stellate cell membranes and in other intermembrane spaces bounded on at least one side by a stellate cell. In no instance was reaction product seen between membranes of two secretory cells or on the surface of stellate cells not contacted by other cellular elements. Thus, it is suggested that stellate cells contain an ATPase, possibly a Na + K + -activated ATPase, which might then allow these cells to serve as regulators of the extracellular ionic environment. Furthermore, since ouabain or removal of K + is inhibitory to melanophore-stimulating hormone (MSH) secretion from isolated pars intermedia in vitro , it is also possible that these cells may play a functional role in the control of MSH secretion.

Histochemical Evidence for the Occurrence of Oligomycin-sensitive Plasmalemma ATPase in Corn Roots

A cytochemical study has been made on the localization of ATPase activity in corn (Zea mays L.) roots. Light microscopy shows washing for 4 hours to increase the general ATPase activity in the peripheral layers of the root cortex; oligomycin and N,N-dicyclohexylcarbodiimide inhibit this activity, oligomycin being more effective. Ultrastructural studies of ATPase location show oligomycin treatment to inhibit both mitochondrial and plasmalemma ATPase, but only in the epidermis and outer cortex. Studies with lipid-soluble dyes indicate that oligomycin might not penetrate very deeply into root tissue in the time span of these experiments. It is suggested that the strong inhibition of ion absorption by oligomycin without a corresponding decline in ATP content is probably due to inhibition of ion absorption in the peripheral cell layers, thus limiting the supply of ion for symplastic transport to the uninhibited tissues.

Analysis of pathways of transport of materials into glandular cells

Movement of materials in the metabolic component of the microcirculatory system of the exocrine part of the pancreas inRanatemporaria takes place from the blood capillary to the pericapillary space, then into the intercellular spaces, and from them into the glandular cells. An ultrastructural study of the localization of ATPase activity in the metabolic component confirmed these data: A high concentration of lead phosphate granules was observed in the endothelium of the blood capillaries, on the fibrillary structures of the interstices of the pericapillary space, on the lateral plasma membrane of the exocrine pancreacytes, and on the cytoplasmic outgrowths forming pinocytotic vacuoles.

Postmortem electron-cytochemical investigation of brain ATPase activity

The localization of ATPase activity in the rat and human brains at various times after death was determined by a method using lead. This activity was discovered in the cytoplasm of the cells, the chromatin and nucleolus, and also in synaptic terminals. The reaction product in the blood capillaries was localized in the basal layer and on the endothelial cell membranes. The results demonstrate the preservation of a high level of brain ATPase activity after death.

Localization of Atpase Activity in the Sarcoplasmic Reticulum of Myocardium

Transcellular exchange of various ions takes place via adenosine triphosphatases (ATPases) which are embedded in the cell and its organelle membranes. Due to heterogeneity in the functional roles of myocardial cell membranes, it is generally agreed that different kinds of ATPases exist in the cell. Biochemical evidence indicates that a major ATPase is involved with the Na+-K+ pump and is located in the myocardial cell membrane. Another ATPase, Ca transport ATPase, is localized in the sarcoplasmic reticulum. Little is presently known of the cytochemical localization and distribution of ATPases and their correlation with myocardial cell membranes. In this study an attempt was made to demonstrate the cytochemical distribution of Ca++ ATPase in the myocardium.

Fine structure and localization of adenosine triphosphatase in the halophyte Suaeda maritima.

The fine structure and localization of ATP-ase activity has been investigated in cells of the halophyte,Suaeda maritima. No significant differences were observed in the fine structure ofSuaeda as compared with that of mesophytes, nor were there any differences found betweenSuaeda cells grown in the presence or absence of high levels of sodium chloride. ATP-ase activity was localized predominantly in the plasmalemma and the cell walls of leaf cells in particular showed an interesting crenated appearance; these featured are discussed in relation to the ionic relationships of halophytes. The cell vacuoles contained a variety of inclusion bodies and showed evidence of considerable autophagic activity.

Intestinal mucosae enzymatic and histochemical changes during infectious diarrhea in calves.

Two, healthy, 4-day-old calves were fed an enteropathogenic agent and fatal diarrhea ensued. Biopsies and mucosal scrapings from the ileum and jejunum were taken before introduction of the agent and at 24, 48 and 72 hours thereafter. The biopsies and scrapings were used to analyze the changes in cytochemical localization and biochemical activity of a variety of phosphataases. Within 24 hours, adenosine triphosphate phosphohydrolase activity was almost totally inhibited while the activity of nonspecific phosphatase and nucleoside phosphatase was only partially inhibited and that of Na+-K+ ATPase not effected at all. In preinoculation control sections, the localization of ATPase activity was heaviest in the apices of absorptive epithelial cells located at the base of the villi. This reaction was completely lost within 24 hours after the inoculation with virus. The enzymatic inhibition occurred 12 to 24 hours before diarrhea became evident.

Fine structure and surface adenosinetriphosphatase activity of a human hepatoma.

A study has been made of the fine structure of a human hepatoma and of the subcellular distribution of its surface ATPase activity. Many cells showed close resemblances to normal liver parenchymal cells; organelles were polarized around structures resembling bile canaliculi, cisternae of rough-surfaced endoplasmic reticulum were well developed and glycogen stores were plentiful. Mitochondria were abnormal, showing swelling, pallor of the matrix, loss of dense granules and abnormal cristae. ATPase activity was present at the plasma membranes fronting adjacent tumor cells but was particularly prominent at the sites which resembled bile canaliculi, where its presence could be related to microvilli. The similarities in fine structure and localization of ATPase activity between this tumor and normal liver are discussed in relation both to other human and to experimental animal hepatomas, and their possible functional significance is considered.

Endocytosis in thyroid follicle cells: III. An electron microscopic study of the cell surface and related structures

Endocytosis in thyroid follicle cells from untreated, suppressed and TSH-stimulated glands were studied. In untreated glands, images indicating a micropinocytic process resulting in the production of bristle-coated vesicles were observed at the apical surface. In suppressed glands, signs of apical micropinocytosis were only rarely encountered. After TSH stimulation, the coated vesicles reappeared. In addition, pictures indicating endocytosis through engulfment of colloid by large cytoplasmic protrusions were observed. Similar pictures were rarely encountered in normal glands. In all the experimental groups, signs of micropinocytosis by coated vesicles were observed along the lateral cell surface as well as along narrow infoldings in the basal plasma membrane. Along parts of the basal cell surface bordering on basement membrane, smooth vesicles suggesting a different form of endocytosis were observed. Differences in structure and thickness of the plasma membrane of the different surfaces of the cells were described, as well as differences in the localization of ATPase activity. Small vesicular structures, displaying ATPase activity were observed both in the basal portions of follicle cells and in the thyroid capillary walls. Heavy deposits of ATPase reaction products were found on the plasma membrane of parafollicular cells and in the interspace between a parafollicular and a follicle cell. Only scattered deposits were found between the lateral surfaces of adjacent follicle cells. Desmosomes were found between adjacent follicle cells as well as between parafollicular cells, but were not observed along the contact between a parafollicular and a follicle cell.

Localization of ATPase activity in striated muscle and probable sources of artifact.

ABSTRACT An attempt has been made to localize sites of ATPase activity in stretched fibrils from glycerinated rabbit muscle by precipitating the liberated phosphate with lead ions. As a control, the distribution of lead phosphate grains precipitated by adding phosphate solutions to fibrils incubated with lead (without ATP) has been studied. In these conditions, the grains are not uniformly distributed along the fibrils, but show many of the features of the pattern of distribution that is found after the fibres have been incubated in ATP and lead. It is suggested that differential lead binding by the protein filaments, or movement of the grains, may be responsible for this non-uniform distribution, and that consequently no conclusions concerning sites of ATPase activity within the sarcomere can be drawn from the results of methods using lead precipitation.