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Biomedical subjects

J Zachar

Publications and source records attributed to J Zachar.

At least 37 records · Page 2Linked to original sources
PubMed1989

Na+-Ca2+ exchanger in the soluble fraction of crayfish striated muscle.

Proteins with Na+-Ca2+ exchange activity from the soluble fraction of crayfish striated muscle were inserted into asolectin proteoliposomes. A pH dependent calcium uptake with an optimum at the alkaline side and inhibition in the presence of sodium or strontium ions in the external medium was observed. When expressed per tissue wet weight the capacity for Na+-Ca2+ exchange of proteoliposomes with inserted soluble proteins was by one half higher than that of the membrane fraction and more than twice higher in comparison with the reconstituted membrane bound exchanger. Using polyacrylamide gel electrophoresis two most prominent proteins with Mr over 200 and 43 kDa could be detected in proteoliposomes with the highest Na+-Ca2+ exchange. It is assumed that protein(s) with Mr 43 kDa could represent the soluble Na+-Ca2+ exchanger in crayfish striated muscle soluble fraction.

Animals↗
PubMed1987

Calcium channels in crayfish muscle fibre fragments studied by means of the Vaseline gap technique.

Calcium currents in crayfish muscle fibres were studied by means of the vaseline gap voltage clamp technique. Overlapping potassium currents were fully suppressed using fibre fragments equilibrated in K+-free intracellular solution. The design of the recording chamber tailored to crayfish muscle fibres is described in detail. Ca currents recorded has a two-component time course. The transient (ICa, T) component (peaking in about 10 ms) attained, on average, maximal overall density of 26.4 microA/cm2 at depolarization to -4.6 mV from a holding potential of -80 mV. The steady (ICa, S) component attained 16.7 microA/cm2 (evaluated at the end of a 70 ms pulse) at +13.8 mV. The average overall surface area of the clamped membrane surface (including invaginated parts) was about 0.07 cm2. The ICa, S component could be separated from ICa, T using short inactivating prepulses. Voltage and time dependence of the transient component inactivation, as well as its recovery from inactivation, were in agreement with a Ca-dependent mechanism. Independent behaviour of the two Ca current components and differences in their properties support the hypothesis concerning the existence of two populations of Ca channels in the surface membrane of the crayfish muscle.

Animals↗
PubMed1987

Na+-Ca2+ exchange in plasma membranes of crayfish striated muscle.

Na+-Ca2+ exchange rates and some physico-chemical properties of the exchanger were studied in crayfish striated muscle membranes enriched in plasma membranes prepared by differential centrifugation of muscle microsomal fraction on discontinuous sucrose density gradient. The lightest subfraction with the highest Na+, K+-ATPase and Mg2+-ATPase activities also showed the highest Na+-Ca2+ exchange rates. A number of physico-chemical characteristics of the Na+-Ca2+ exchanger found in the present experiments were similar to those reported for excitable membranes of mammals, except for the temperature optimum (20 degrees C for the crayfish).

Animals↗
PubMed1987

Effects of sulfhydryl reagents on Na+-Ca2+ exchange in rat brain microsomal membranes.

Effects of six thiol reagents with different physico-chemical properties were tested on the Na+-dependent 45Ca2+ transport into the rat brain microsomal membrane vesicles. The mercurials p-chlormercuribenzoate and Mersalyl effectively inhibited 45Ca2+ uptake with IC50 values in the order of 10(-4) mol X l-1 in the medium. N-ethylmaleimide and its more lipophilic analog N-(4-(2-benzoxazolyl)phenyl)maleimide were much less effective at the same concentrations. 2,2'-dithiodipyridine markedly reduced 45Ca2+ uptake already at concentrations below 10(-4) mol X l-1, whereas 5,5'-dithiobis-2-nitrobenzoate in a concentration range 10(-6)-10(-3) mol X l-1 was a weak inhibitor. Inhibitory effects of the most potent inhibitors p-chlormercuribenzoate and 2,2'-dithiodipyridine were readily reversed by 1 mmol X l-1 dithiothreitol. The results suggest that free SH groups of membrane polypeptides are involved in the functioning of the Na+-Ca2+ exchanger in the nerve tissue cell membranes.

Animals↗
PubMed1986

Inhibition of Na+-Ca2+ exchange by calcium antagonists in rat brain microsomal membranes.

Na+-Ca2+ exchange rates were studied in native and/or pronase pretreated rat brain microsomal membranes in the presence of calcium channel antagonists verapamil, nimodipine and nifedipine. In native membranes all the substances used inhibited Na+-Ca2+ exchange. A relatively stronger inhibition was observed in membranes pretreated with pronase. The values of Ki for nimodipine and nifedipine did not change and it fell to about one half for verapamil. Lineweaver-Burk's plots have revealed that the verapamil inhibition in native membranes as well as in pronase pretreated ones was of a non-competitive type; Km for calcium oscillated around 15 mumol.l-1. It is suggested that the inhibition strength depends on the access of inhibitors to the membrane binding sites as well as on the solubility of inhibitors in membrane lipids.

Animals↗
PubMed1985

Na+-Ca2+ exchange in rat brain microsomal membranes pretreated with pronase and/or SDS.

The effects of pronase and/or SDS pretreatment on Na+-Ca2+ exchange were studied in rat brain microsomal membranes. Pronase in concentrations that liberated 11% of the membrane proteins stimulated the Na+-Ca2+ exchange. When about 24% of the proteins were split off, the results did not differ from those in control experiments. When 40% or more of the proteins were solubilized, Na+-Ca2+ exchange was abolished. Pronase pretreatment did not change the Km value for Ca2+, it increased Vmax only. The effect of pronase was partially blocked by Trasylol. Neuraminidase had no effect on Na+-Ca2+ exchange. SDS pretreatment of the membranes inhibited Na+-Ca2+ exchange: when 25% of membrane proteins were solubilized with SDS, the Na+-Ca2+ exchange was abolished while the same amount of proteins split off with pronase did not change the rate of Na+-Ca2+ exchange as related to membrane proteins. Ischaemia lasting for 2-4 h or complete hypoxia which should stimulate endogenous proteinases due to the rise of free intracellular calcium did not influence the Na+-Ca2+ exchange. A decrease in Na+-Ca2+ exchange rate was observed when proteins with molecular weight between 45,000 and 20,000 were split off from the membranes. It is assumed that the Na+-Ca2+ antiporter is a polypeptide from the group of proteins within the above molecular weights.

Animals↗
PubMed1983

Selectivity of sodium channels in denervated tonic muscle fibre membrane of the frog.

Ionic selectivity of sodium channels was examined under voltage clamp conditions in normal and denervated twitch fibres and denervated tonic fibres isolated from m. ileofibularis of the frog (R. temporaria). Membrane currents were recorded by means of the Hille-Campbell vaseline-gap voltage clamp method from muscle fibre segments exposed to a potassium-free artificial internal solution. Permeability ratio (PS/PNa) were determined from changes in the reversal potential after replacing all Na ions in the solution bathing the voltage clamped external membrane area with sodium substituting ions (S). The permeability sequence was: Na+ greater than Li+ greater than NH4+ greater than K+. No inward currents were observed for Ca2+. The permeability ratios were as follows. Denervated tonic fibres: 1:0.88:0.23:0.012; control twitch fibres: 1:0.94:0.22:0.076; denervated twitch fibres: 1:0.91:0.14:0.082. The permeability to Li+ ions deviates from independence to a greater extent in tonic than in phasic fibres. Our results are consistent with the Hille model of sodium channel selectivity, and they support the hypothesis that sodium channels formed in denervated tonic muscle fibres of the frog are of the same genetic origin as Na channels expressed under physiological conditions.

Animals↗
PubMed1983

NMR studies on interactions of ribonuclease Sa with Guo-3'-P.

Some features of the interaction of guanyloribonuclease Sa from Streptomyces aureofaciens with its competitive inhibitor Guo-3'-P were investigated by 1H and 31P NMR spectroscopy. The pH dependence of chemical shifts of C(2)-H protons of the histidine residue of the enzyme were analysed, in the absence and presence of Guo-3'-P. This analysis showed that only one of the two histidines of ribonuclease Sa is located in the active site of the enzyme. 31P NMR resonances of the nucleotide and of its complex with the enzyme indicated that this histidine interacts with the phosphate group of the substrate. The possible relationship between the observed perturbation of the NMR titration curve of the active site of histidine and a conformational change in the enzyme molecule at a pH of approximately 7.5 is also discussed.

Endoribonucleases↗
PubMed1980

A quantitative estimation of components in crayfish muscle fibres by stereological methods.

The volume and surface densities of structural components in isolated long-sarcomere fibres of crayfish (Astacus fluviatilis) were estimated by stereological methods. The relative volumes were as follows (in % of the fibre volume): sarcolemmal invaginations (SI) 0.74; T-system 1.1; whole sarcoplasmic reticulum (SR) 4.7; junctional SR cisternae 0.41; mitochondria 1.1; nuclei 0.57; myofibrils 81.6; peripheral basal lamina 1.1; junctional gap 0.043; lysosomes 0.029. The values for surface densities were (in microns2/100 microns3): peripheral sarcolemma 1.27; SI 7.21; T-system 40.2; whole SR 146.4; junctional SR cisternae 14.88; junctional area of SR cisternae 7.05; mitochondria 8.98; nuclei 0.75; myofibrils 163. The invaginated membranes (SI and T-system) made up 97.4% of the total area of surface membranes and increased the effective area of peripheral sarcolemma 39 times. Using the values of the total electrical capacity from previous investigations on Astacus fibres, the specific capacity of surface membranes was calculated as 1.08 microF/cm2.

Animals↗