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A Hassonvoloch - One of the best experts on this subject based on the ideXlab platform.

  • identification and distribution of chondroitin sulfate in the three electric organs of the electric eel Electrophorus electricus l
    Comparative Biochemistry and Physiology B, 2007
    Co-Authors: Maisa L S Souza, A Hassonvoloch, Nilson Nunestavares, Cristiano F Freitas, Mariaaparecida O Domingos, Luiz Eurico Nasciutti, Luizclaudio F Silva
    Abstract:

    The electrogenic tissue of the electric eel Electrophorus electricus (L.) is distributed in three well-defined electric organs, the Main electric organ, Sach's organ and Hunter's organ. Sulfated glycosaminoglycan (GAG) composition was characterized in the three electric organs of the electric eel. Sulfated GAGs were analyzed in the electric organs using metachromatic staining, biochemical analysis including electrophoresis before and after specific enzymatic or chemical degradations, and immunostaining with an antibody against chondroitin sulfate (CS). Our results showed in the three electric organs that CS was the main sulfated GAG species detected, accompanied by small and diminutive amounts of CS/dermatan sulfate hybrid chains and heparan sulfate (HS), respectively. However, HS was not detected in the Sach's organ. CS was predominantly detected in the innervated membrane face of the electroplaques in the three electric organs. Our findings extend previous observations on the GAG composition in the electric organs of E. electricus and provide new information regarding the tissue distribution and location of CS.

  • purification and amino acid sequence of fructose 1 6 bisphosphate aldolase from the electric organ of Electrophorus electricus l
    Zeitschrift für Naturforschung C, 2006
    Co-Authors: Salvatore G Desimone, Celia Maria Batista E Silva, Christiane Cardoso M De Salles, A Hassonvoloch
    Abstract:

    A soluble fructose-1,6-bisphosphate aldolase enzyme has been purified 50.2-fold (2.36%) at the homogeneity from the electric organ of Electrophorus electricus by one step of DEAE-52 anion exchange chromatography followed by Superose-12 gel filtration-FPLC. Like other aldolase enzymes the E. electricus protein is a dimer with two identical subunits of 45 kDa. The N-terminal (20 residues) revealed a high homology with S. aurata (75%, goldfish), R. ratus and M. musculus (mouse, 80%) enzymes.

  • inhibition of acetylcholinesterase from Electrophorus electricus l by tricyclic antidepressants
    The International Journal of Biochemistry & Cell Biology, 2002
    Co-Authors: Nilson Nunestavares, Nery A Da Matta, C Batista M E Silva, Glauce Maria Nunes Araujo, S R W Louro, A Hassonvoloch
    Abstract:

    The effects of tricyclic antidepressants drugs (TCA) amitriptyline, imipramine and nortriptyline, on purified Electrophorus electricus (L.) acetylcholinesterase (AChE; acetylcholine hydrolase, EC 3.1.1.7) were studied using kinetic methods and specific fluorescent probe propidium. The antidepressants inhibited AChE activity by a non-competitive mechanism. Inhibition constants range from 200 to 400 microM. Dimethylated amitriptyline and imipramine were more potent inhibitors than the monomethylated nortriptyline. Fluorescence measurements using bis-quaternary ligand propidium were used to monitor ligand-binding properties of these cationic antidepressants to the AChE peripheral anionic site (PAS). This ligand exhibited an eight-fold fluorescence enhancement upon binding to the peripheral anionic site of AChE from E. electricus (L.) with K(D)=7 x 10(-7)M. It was observed that TCA drugs displaced propidium from the enzyme. On the basis of the displacement experiments antidepressant dissociation constants were determined. Similar values for the inhibition constants suggest that these drugs have similar affinity to the peripheral anionic site. The results also indicate that the catalytic active center of AChE does not participate in the interaction of enzyme with tricyclic antidepressants. These studies suggest that the binding site for tricyclic antidepressants is located at the peripheral anionic site of E. electricus (L.) acetylcholinesterase.

  • choline acetyltransferase detection in normal and denervated electrocyte from Electrophorus electricus l using a confocal scanning optical microscopy analysis
    Anais Da Academia Brasileira De Ciencias, 2000
    Co-Authors: Nilson Nunestavares, Narcisa L Cunhaesilva, A Hassonvoloch
    Abstract:

    Acetylcholine is the neurotransmitter responsible for the transmission of impulses from cholinergic neurons to cells of innervated tissues. Its biosynthesis is catalyzed by the enzyme Choline acetyltransferase that is considered to be a phenotypically specific marker for cholinergic system. It is well known that the regulation of Choline acetyltransferase activity under physiological and pathological conditions is important for development and neuronal activities of cholinergic functions. We observed the distribution of Choline acetyltransferase in sections from the normal and denervated main electric organ sections of Electrophorus electricus (L.) by immunofluorescence using a anti-Choline acetyltransferase antibody. The animals were submitted to a surgical procedure to remove about 20 nerves and after 30 and 60 days, they were sacrificed. After 30 days, the results from immunohistochemistry demonstrated an increase on the Choline acetyltransferase distribution at denervated tissue sections when compared with the sections from the normal contralateral organ. A very similar labeling was observed between normal and denervated tissue sections of the animals after 60 days. However, Choline acetyltransferase activity (nmolesACh/ min/ mg of protein) in extracts obtained from electrocyte microsomal preparation, estimated by Fonnun's method (Fonnun 1975), was 70% lower in the denervated extracts.

  • purification and partial characterization of creatine kinase from electric organ of Electrophorus electricus l
    The International Journal of Biochemistry & Cell Biology, 2000
    Co-Authors: C Batista M E Silva, Nery A Da Matta, Salvatore Giovannidesimone, Nunes N Tavares, A Hassonvoloch
    Abstract:

    The present investigation deals with the purification and the partial characterization of the soluble creatine kinase (CK) isoenzyme, isolated from the electric organ electrocyte of Electrophorus electricus (L.). Purification was performed by precipitation of the enzyme in the crude extract with ammonium sulfate (80%). The precipitate obtained was analyzed on an ion exchange column of diethylaminoethyl cellulose-52 (DEAE) followed by gel filtration on Superose 12 in a Fast Protein Liquid Chromatography (FPLC) system. Electrophoretic mobility of the active peak confirmed previous results identifying the hybrid isoenzyme MB in the eletrocyte cytoplasm. Electrocyte CK is a dimeric enzyme with two identical subunits of approximately 40 kDa as estimated by sodium dodecyl sulfate‐polyacrylamide gel electrophoresis (SDS‐PAGE). The sequence analysis of the N-terminal peptide (14 amino acids) of the 40 kDa subunit showed homology with other CK enzymes from electric fish (Torpedo) and human muscle type CK. # 2000 Elsevier Science Ltd. All rights reserved.

Shit F Chew - One of the best experts on this subject based on the ideXlab platform.

  • voltage gated na channel isoforms and their mrna expression levels and protein abundance in three electric organs and the skeletal muscle of the electric eel Electrophorus electricus
    PLOS ONE, 2016
    Co-Authors: Biyun Ching, Jia M Woo, Kum C Hiong, Mel V Boo, Wai P Wong, Shit F Chew
    Abstract:

    This study aimed to obtain the coding cDNA sequences of voltage-gated Na+ channel (scn) α-subunit (scna) and β-subunit (scnb) isoforms from, and to quantify their transcript levels in, the main electric organ (EO), Hunter's EO, Sach's EO and the skeletal muscle (SM) of the electric eel, Electrophorus electricus, which can generate both high and low voltage electric organ discharges (EODs). The full coding sequences of two scna (scn4aa and scn4ab) and three scnb (scn1b, scn2b and scn4b) were identified for the first time (except scn4aa) in E. electricus. In adult fish, the scn4aa transcript level was the highest in the main EO and the lowest in the Sach's EO, indicating that it might play an important role in generating high voltage EODs. For scn4ab/Scn4ab, the transcript and protein levels were unexpectedly high in the EOs, with expression levels in the main EO and the Hunter's EO comparable to those of scn4aa. As the key domains affecting the properties of the channel were mostly conserved between Scn4aa and Scn4ab, Scn4ab might play a role in electrogenesis. Concerning scnb, the transcript level of scn4b was much higher than those of scn1b and scn2b in the EOs and the SM. While the transcript level of scn4b was the highest in the main EO, protein abundance of Scn4b was the highest in the SM. Taken together, it is unlikely that Scna could function independently to generate EODs in the EOs as previously suggested. It is probable that different combinations of Scn4aa/Scn4ab and various Scnb isoforms in the three EOs account for the differences in EODs produced in E. electricus. In general, the transcript levels of various scn isoforms in the EOs and the SM were much higher in adult than in juvenile, and the three EOs of the juvenile fish could be functionally indistinct.

  • na k atpase α subunit nkaα isoforms and their mrna expression levels overall nkaα protein abundance and kinetic properties of nka in the skeletal muscle and three electric organs of the electric eel Electrophorus electricus
    PLOS ONE, 2015
    Co-Authors: Biyun Ching, Jia M Woo, Kum C Hiong, Mel V Boo, Wai P Wong, Celine Yen Ling Choo, Shit F Chew
    Abstract:

    This study aimed to obtain the coding cDNA sequences of Na+/K+-ATPase α (nkaα) isoforms from, and to quantify their mRNA expression in, the skeletal muscle (SM), the main electric organ (EO), the Hunter’s EO and the Sach’s EO of the electric eel, Electrophorus electricus. Four nkaα isoforms (nkaα1c1, nkaα1c2, nkaα2 and nkaα3) were obtained from the SM and the EOs of E. electricus. Based on mRNA expression levels, the major nkaα expressed in the SM and the three EOs of juvenile and adult E. electricus were nkaα1c1 and nkaα2, respectively. Molecular characterization of the deduced Nkaα1c1 and Nkaα2 sequences indicates that they probably have different affinities to Na+ and K+. Western blotting demonstrated that the protein abundance of Nkaα was barely detectable in the SM, but strongly detected in the main and Hunter’s EOs and weakly in the Sach’s EO of juvenile and adult E. electricus. These results corroborate the fact that the main EO and Hunter’s EO have high densities of Na+ channels and produce high voltage discharges while the Sach’s EO produces low voltage discharges. More importantly, there were significant differences in kinetic properties of Nka among the three EOs of juvenile E. electricus. The highest and lowest Vmax of Nka were detected in the main EO and the Sach’s EO, respectively, with the Hunter’s EO having a Vmax value intermediate between the two, indicating that the metabolic costs of EO discharge could be the highest in the main EO. Furthermore, the Nka from the main EO had the lowest Km (or highest affinity) for Na+ and K+ among the three EOs, suggesting that the Nka of the main EO was more effective than those of the other two EOs in maintaining intracellular Na+ and K+ homeostasis and in clearing extracellular K+ after EO discharge.

Ernest Schoffeniels - One of the best experts on this subject based on the ideXlab platform.

  • solubilization of thiamine triphosphatase from the electric organ of Electrophorus electricus
    Biochimica et Biophysica Acta, 1991
    Co-Authors: Lucien Bettendorff, Isabelle Longree, Pierre Wins, Ernest Schoffeniels
    Abstract:

    Abstract The membrane-associated, anion-regulated thiamine triphosphatase from Electrophorus electricus electric organ can be solubilized by various neutral detergents. Polyoxyethylene ethers are the most effective. Anionic detergents readily inactivate the enzyme. A 6.4-fold increase in specific activity is obtained by successive treatment of crude membranes with octanoyl-N-methylglucamide, which solubilized other proteins, and Lubrol-PX with releases 60% of the thiamine triphosphatase (TTPase) activity. Solubilization by Lubrol-PX rapidly modifies kinetic parameters. The Km, Vmax and pH optimum are decreased. However, the solubilized TTPase may be kept at 0°C for many hours without further change in specific activity. At 35°C, the half-life is still 83 min at pH 5.0, but denaturation becomes rapid at pH ⩾ 7. Solubilization modifies anion effects on TTPase activity. The activating effect of nitrate is nearly lost, while inhibition by sulfate is no longer time-dependent.

Esperanza Recio-pinto - One of the best experts on this subject based on the ideXlab platform.

  • Molecular Cloning and Expression of a Kv1.1-like Potassium Channel from the Electric Organ of Electrophorus electricus
    The Journal of Membrane Biology, 2003
    Co-Authors: William B. Thornhill, M. B. Wu, Jhon-jairo Sutachan, I Watanabe, X Wu, Esperanza Recio-pinto
    Abstract:

    Electrocytes from the electric organ of Electrophorus electricus exhibited sodium action potentials that have been proposed to be repolarized by leak currents and not by outward voltage-gated potassium currents. However, patch-clamp recordings have suggested that electrocytes may contain a very low density of voltage-gated K^+ channels. We report here the cloning of a K^+ channel from an eel electric organ cDNA library, which, when expressed in mammalian tissue culture cells, displayed delayed-rectifier K^+ channel characteristics. The amino-acid sequence of the eel K^+ channel had the highest identity to Kv1.1 potassium channels. However, different important functional regions of eel Kv1.1 had higher amino-acid identity to other Kv1 members, for example, the eel Kv1.1 S4-S5 region was identical to Kv1.5 and Kv1.6. Northern blot analysis indicated that eel Kv1.1 mRNA was expressed at appreciable levels in the electric organ but it was not detected in eel brain, muscle, or cardiac tissue. Because electrocytes do not express robust outward voltage-gated potassium currents we speculate that eel Kv1.1 channels are chronically inhibited in the electric organ and may be functionally recruited by an unknown mechanism.

Carlos Alberto Chagas - One of the best experts on this subject based on the ideXlab platform.

  • differences in the isodesmin pattern between the electric organs of Electrophorus electricus l
    Comparative Biochemistry and Physiology B, 1998
    Co-Authors: Manuel Luis Costa, Carlos Alberto Chagas, Claudia Mermelstein, Maira Monteiro Froes, Vivaldo Moura Neto
    Abstract:

    Desmin, the intermediate filament protein of muscle, is present in the electric organs of Electrophorus electricus L. as five isovariants, instead of the one to two isovariants found in muscle. We analyzed the isodesmin pattern in the three different electric organs using densitometry of Coomassie blue-stained bands in electrofocusing polyacrylamide gel electrophoresis. We were able to compare the relative amount of each of the five desmin isovariants in an isodesmin pattern characteristic of each electric organ. These patterns proved to be, in some cases, statistically different. Desmin in each electric organ could have slightly different functions in order to correlate with the organ-specific isovariant patterns.

  • desmin and actin filaments in membrane cytoskeletal preparations of the electric tissue of Electrophorus electricus l
    Archives of Histology and Cytology, 1997
    Co-Authors: Claudia Mermelstein, M C R Cordeiro, Marlene Benchimol, Carlos Alberto Chagas, Marilia Taffarel, Vivaldo Moura Neto
    Abstract:

    The electrocyte of the electric organ of the electric eel, Electrophorus electricus, L was investigated by light and electron microscopy as well as immunoelectron microscopy, in order to clarify the fine structures and distribution of cytoskeleton filaments and their relations to proteins, especially desmin and actin. Cytoskeleton-enriched fractions of the electrocytes were analysed with SDS-PAGE. It was verified that a meshwork of filaments was distributed in the electrocytes, more abundantly in the anterior than in the posterior part of the cell, and that this could be associated with membrane invaginations. Desmin and actin were the components of this meshwork, suggesting that desmin intermediate filaments and actin filaments might play a role in the maintenance of the morphology of electrocytes and, as an intracellular filamentous meshwork, they may contribute to the organization of the components of membranes and papillae formation on the anterior face of the electrocytes.

  • desmin filaments in the electrocytes of the electric organ of the electric eel Electrophorus electricus
    Cell and Tissue Research, 1996
    Co-Authors: M C R Cordeiro, Marlene Benchimol, M V C Faria, Carlos Alberto Chagas, Claudia Mermelstein, Moura V Neto
    Abstract:

    Desmin protein is an abundant constituent of the intermediate filaments in the electrocytes of the electric organ of the electric eel Electrophorus electricus. Polyclonal antibodies were raised against purified desmin from the electric organ and used for immunolabeling of the protein in reconstituted filaments. In thick sections of the main electric organ that has been stained with fluorescein-labeled desmin-specific antibodies, light microscope revealed a diffuse meshwork of desmin filaments dispersed in the cytoplasm of electrocytes. In the region under the membrane, the immunostaining was slightly more intense than elsewhere. The meshwork of intermediate filaments composed of desmin was examined by electron microscopy of the main electric organ. Immuno-gold labeling demonstrated a widespread meshwork of desmin filaments in the cytoplasm and in close association with the plasma membrane. These observations suggest that intermediate filaments play a role in the maintenance of the morphology of electrocytes and, as an intracellular meshwork spanning the width of the cell, they may contribute to the organization of the intracellular compartments.

  • microheterogeneity of desmin in the electric organ and dorsal muscle of the electric eel Electrophorus electricus
    Comparative Biochemistry and Physiology Part A: Physiology, 1995
    Co-Authors: M C R Cordeiro, Moura V Neto, Marlene Benchimol, M V C Faria, Carlos Alberto Chagas
    Abstract:

    Abstract Two-dimensional gel electrophoresis of purified desmin from the electric organ of Electrophorus electricus exhibited five isoforms as opposed to four from the dorsal muscle of the eel. Differences in isoforms may indicate different degrees of desmin phosphorylation in these tissues. The similarities detected in the two less acidic isodesmin from the electric organ and from the dorsal muscle of Electrophorus electricus suggest that the electric organ has conserved some characteristics seen in the dorsal muscle, favoring previous suggestions that considered the electric tissue as a differentiated form of striated muscle.

  • heterogeneity of purified actin in the electric organ of the electric eel Electrophorus electricus
    Journal of Experimental Zoology, 1991
    Co-Authors: Lilian Ayressa, Vivaldo Moura Neto, Mecia M Oliveira, Carlos Alberto Chagas
    Abstract:

    Actin extracted from the electric organ and epaxial muscle of the electric eel Electrophorus electricus was characterized by several biochemical techniques. Isoelectric focusing gels revealed two isoforms of actin in the electric organ, in contrast to the unique form of actin present in the epaxial muscle of this eel and the rabbit leg muscle, which were used as controls. Proteolytic digestion and immunoblotting analysis confirmed that these isoforms were composed of actin. When actin from the electric organ was mixed with actin from the epaxial muscle or from rabbit muscle and subjected to isoelectric focusing gel, it was possible to identify an alpha-actin isoform in the electric organ as well as a more basic actin that migrated in the same manner as cytoplasmic actin or gamma actin from smooth muscle. The data are interpreted in terms of the differentiation of the electric organ of E. electricus.