Localization of calcium in skeletal and cardiac muscle

Summary The requirement of calcium (Ca²⁺) in the excitation-contraction coupling of both skeletal and cardiac muscle is well established. However, the exact location of the intracellular storage sites of Ca²⁺ is not firmly established. We report here on the ultrastructural distribution of Ca²⁺ in white and red skeletal muscle and in cardiac muscle of the rat using combined phosphate-pyroantimonate (PPA) and oxalate-pyroantimonate (OPA) procedures. The methods are based on (a) stabilization and/or trapping of Ca²⁺ during the primary fixation step in glutaraldehyde by potassium phosphate or oxalate; (b) subsequent wash-out of all non-trapped cations such as Na⁺ and Mg²⁺ in potassium phosphate or oxalate; (c) conversion of the complexed or trapped Ca²⁺ into an electron-dense calcium pyroantimonate salt in 100 m-thick tissue sections; and (d) wash-out of the excess potassium pyroantimonate at alkaline pH.With the OPA procedure, mitochondria of all muscle types showed little precipitate. The junctional sarcoplasmic reticulum was stongly reactive in relaxed white skeletal muscle, negative in contracted white fibres and negative in red skeletal and cardiac muscle, independent of the state of relaxation-contraction. Other organelles were essentially free of deposits.With the PPA method, the precipitate was almost exclusively confined to the sarcolemma and its T-tubular invaginations in cardiac and slow skeletal muscle, and was absent in fast skeletal muscle. Apart from occasional deposits in mitochondria, all other organelles were free of precipitate. The sarcolemma-associated deposits were clearly confined to the inner leaflet of the lipid bilayer. The amount of precipitate varied within the contraction cycle, relaxed cells possessing the highest density.Exposure of the tissue to La³⁺ resulted in the complete absence of sarcolemma-bound precipitate suggesting that the Ca²⁺ is exchangeable. Furthermore, these cytological data suggest a basic difference in Ca²⁺ storage between white skeletal muscle on the one hand, and red skeletal and cardiac muscle on the other.     

Localization of GABA-like immunoreactivity in the ectostriatum of domestic chicks: GABA immunocytochemistry combined with Golgi impregnation

Brain Cell Biology - The distribution of GABA-like immunoreactivity (GABA-LI) in the ectostriatal core (Ec) of domestic chicks (one to two days old) was investigated using (1) preembedding GABA...     

Cladistic analysis of the Cirripedia Thoracica

We present a cladistic analysis of the Cirripedia Thoracica using morphological characters and the Acrothoracica and Ascothoracida as outgroups. The list of characters comprised 32 shell and soft body features. The operational taxonomic units (OTUs) comprised 26 well-studied fossil and extant taxa, principally genera, since uncertainty about monophyly exists for most higher ranking taxonomic units. Parsimony analyses using PAUP 3.1.1 and Hennig86 produced 189 trees of assured minimal length. We also examined character evolution in the consensus trees using MacClade and Clados. The monophyly of the Balanomorpha and the Verrucomorpha sensu stricto is confirmed, and all trees featured a sister group relationship between the ‘living fossil Neoverruca and me Brachylepadomorpha. In the consensus trees the sequential progression of ‘pedunculate‘sister groups up to a node containing Neolepas also conforms to current views, but certain well-established taxa based solely on plesiomorphies stand out as paraphyletic, such as Pedunculata (= Lepadomorpha); Eolepadinae, Scalpellomorpha and Chthamaloidea. The 189 trees differed principally in the position of shell-less pedunculates, Neoverruca, the scalpelloid Capitulum, and the interrelationships within the Balanomorpha, although the 50% majority rule consensus tree almost fully resolved the latter. A monophyletic Sessilia comprising both Verrucomorpha and Balanomorpha appeared among the shortest trees, but not in the consensus. A tree with a monophyletic Verrucomorpha including Neoverruca had a tree length two steps longer than the consensus trees. Deletion of all extinct OTUs produced a radically different tree, which highlights the importance of fossils in estimating cirripede phylogeny. Mapping of our character set onto a manually constructed cladogram reflecting die most recent scenario of cirripede evolution resulted in a tree length five steps longer than any of our shortest trees. Our analysis reveals that several key questions in cirripede phylogeny remain unsolved, notably the position of shell-less forms and the transition from ‘pedunculate‘to ‘sessile‘barnacles. The inclusion of more fossil species at this point in our understanding of cirripede phylogeny will only result in even greater levels of uncertainty. When constructing the character list we also identified numerous uncertainties in the homology of traits commonly used in discussing cirripede evolution. Our study highlights larval ultrastructure, detailed studies of early ontogeny, and molecular data as the most promising areas for future research.     

Concomitant loss of cell surface fibronectin and laminin from transformed rat kidney cells

Both fibronectin and laminin were found by immunofluorescence as a matrix at the surface of normal rat kidney cells. These matrices were absent from the surface of virally transformed rat kidney cells. Soluble fibronectin and laminin were detected in the culture media of the transformed as well as the normal cells. Culture supernates of the transformed cells contained even more fibronectin than the supernates of the transformed cells contained even more fibronectin than the supernates of the normal cells while laminin was present in similar amounts in both culture media. This shows that the loss of fibronectin and laminin from the surface of the transformed cells is caused by failure of the cells to deposit these proteins into an insoluble matrix and not caused by inadequate production. Fibronectins isolated from culture media of the normal and transformed cells were similar in SDS polyacrylamide gel electrophresis. Laminin isolated from culture media by affinity chromatography on heparin-Sepharose followed by immunoprecipitation was composed of three main polypeptides, one with a molecular weight of 400,000 and two with a molecular weight close to 200,000 in both cell types. Fibronectins from both cell types were equally active in promoting cell attachment. Rat fibronectin from transformed cells, like normal cells, when applied to culture dishes coated with fibronectin, readily attached and spread on the substratum, requiring approximately the same amount of fibronectin as the normal cells. On the basis of these results it seem that the failure of the transformed cells to incorporate fibronectin into an insoluble cell surface matix is not a consequence of a demonstrable change in the functional characteristics of the fibronectin molecule or in the ability of the cells to interact with fibronectin. It may depend on as yet unidentified interactions of the cell surface. Similar interactions may be needed for the deposition of laminin into the matrix, because laminin was also absent from the surface of transformed cells, despite its being synthesized by these cells.     

The impact of acidification on diatoms and chemistry of Dutch moorland pools

Summary Moorland pools are shallow oligotrophic soft water lakes on poorly buffered sandy soils. Diatom assemblages of samples from 16 pools taken in 1920 and 1978 were compared by analysis of pH-spectra, diversity, dissimilarity and multivariate statistical techniques.The pH-spectra of pools in the southern (S) and central (C) part of the country indicate a fall in pH from 4.5–6.0 in the old samples to 3.7–4.6 in the recent ones. The pH-spectra of the northern pools (N) do not indicate a significant shift from the original pH (ca 4.5).The number of species in the count and the diversity (indices of Simpson and Shannon) decreased significantly in S+C, and that goes also for the dissimilarity index of Dyer. No changes were found in N.The first component (PC 1) of the principal component analysis explains 61% of total variance. PC 1 is correlated with log [SO₄] (r=0.83, p<0.001) and even better (r=0.95, p<0.001) with the relative sulphate concentration,i.e. the ratio of sulphate to all major anions (sulphate, chloride, bicarbonate). All old samples have low scores on PC 1, recent samples have low scores on the second (PC 2) and third (PC 3) principal component. Old samples have high scores on PC 2 and PC 3, explaining 9 and 6% of total variance, respectively.The orginal variation, caused by regional factors, is replaced by a SO₄ ^2– controlled variation. PC 1 is nearly completely determined by the relative abundance ofEunotia exigua. This species, which is known to be very resistant to pollution by sulphur, aluminium and heavy metals, increased largely from 1920 to 1978.In spite of the rather homogeneous distribution of wet sulphate deposition in the Netherlands, substantial differences in SO₄ ^2– content in the pools are observed, being lowest in N (0.13–0.48 meq.l^–1) and highest in S+C (0.38–1.65 meq.l^–1). Sulphate is positively correlated with calcium, aluminum and magnesium but negatively with factors that characterize humic acid waters (e.g. permanganate-consumption, iron and the ratio of univalent to divalent cations). Sulphate concentration depends on the intensity of sulphate reduction, accumulation by dry deposition in surrounding forests of Scots pine, drought and atmospheric deposition.     

Dissociation of intracellular lysosomal rupture from the cell death caused by silica

The relationship between intracellular lysosomal rupture and cell death caused by silica was studied in P388d(1) macrophages. After 3 h of exposure to 150 μg silica in medium containing 1.8 mM Ca(2+), 60 percent of the cells were unable to exclude trypan blue. In the absence of extracellular Ca(2+), however, all of the cells remained viable. Phagocytosis of silica particles occurred to the same extent in the presence or absence of Ca(2+). The percentage of P388D(1) cells killed by silica depended on the dose and the concentration of Ca(2+) in the medium. Intracellular lyosomal rupture after exposure to silica was measured by acridine orange fluorescence or histochemical assay of horseradish peroxidase. With either assay, 60 percent of the cells exposed to 150 μg silica for 3 h in the presence of Ca(2+) showed intracellular lysosomal rupture, was not associated with measureable degradation of total DNA, RNA, protein, or phospholipids or accelerated turnover of exogenous horseradish peroxidase. Pretreatment with promethazine (20 μg/ml) protected 80 percent of P388D(1) macrophages against silica toxicity although lysosomal rupture occurred in 60-70 percent of the cells. Intracellular lysosomal rupture was prevented in 80 percent of the cells by pretreatment with indomethacin (5 x 10(-5)M), yet 40-50 percent of the cells died after 3 h of exposure to 150 μg silica in 1.8 mM extracellular Ca(2+). The calcium ionophore A23187 also caused intracellular lysosomal rupture in 90-98 percent of the cells treated for 1 h in either the presence or absence of extracellular Ca(2+). With the addition of 1.8 mM Ca(2+), 80 percent of the cells was killed after 3 h, whereas all of the cells remained viable in the absence of Ca(2+). These experiments suggest that intracellular lysosomal rupture is not causally related to the cell death cause by silica or {"type":"entrez-nucleotide","attrs":{"text":"A23187","term_id":"833253","term_text":"A23187"}}A23187. Cell death is dependent on extracellular Ca(2+) and may be mediated by an influx of these ions across the plasma membrane permeability barrier damaged directly by exposure to these toxins.     

Sensitivity to weighting in life cycle impact assessment (LCIA)

The International Journal of Life Cycle Assessment - Weighting in life cycle assessment (LCA) incorporates stakeholder preferences in the decision-making process of comparative LCAs. Research...     

Synthesis and Biological Activity of Some 2-Aminopurine Carbonucleosides

Abstract

A series of new o ne t wo subtituted c arbonucleoside analogues (OTC) of purine was synthesized and evaluated against cytomegalovims and varicella-zoster virus in human embryonic lung (HEL) cells.     

INFLUENCE OF FRESHWATER FLOW REGIME ON FISH ASSEMBLAGES IN THE GREAT FISH RIVER AND ESTUARY

Summary

Two and a half years of data were collected from the lower Great Fish River, head region and estuary to determine the fish species composition within these areas. Gilchristella aestuaria, Liza dumerilii, Rhabdosargus holubi and Pomadasys commersonnii were the four most abundant species captured, with riverine flow rate having an important effect on both species composition and numbers of fishes in the different regions. Most marine species displayed a strong inverse relationship between catch per unit effort and elevated freshwater inputs. Euryhaline marine species dominated the catches at all sampling sites during low flows but were less common during high flow periods when catadromous species were most abundant. Based on the available evidence it is suggested that for most marine species in the river this decline in abundance is related to low conductivity levels following floods rather than avoidance of elevated flows. The impact of elevated suspensoid concentrations and lowered dissolved oxygen concentrations on freshwater and estuarine fish populations during major river flooding is also discussed.