Ca2+- and voltage-dependent K+ conductance in dispersed parathyroid cells

The membrane ionic conductances of dispersed parathyroid cells kept in primary culture were studied using the "whole-cell" and "inside-out excised patch" variants of the patch-clamp technique. The major component of the total current was a voltage-dependent outward K+ current without an appreciable inward current. The amplitude of the K+ current was markedly reduced when free internal Ca2+ was buffered by addition of 10 mM EGTA. Recordings of single-channel current in excised membrane patches revealed the presence of K+ channels with large unitary conductance (200 pS in symmetrical 130 mM K+ solutions) which were also activated by depolarization when internal Ca2+ concentration was about 10(-5)-10(-6) M. At any membrane voltage these channels were closed most of the time at internal Ca2+ concentrations lower than 10(-10) M. These results demonstrate the existence of a Ca2+- and voltage-dependent K+ permeability in parathyroid cells which may participate in the unusual membrane potential changes induced by alterations of external Ca2+ and, possibly, in the regulation of parathormone secretion.     

Immobilized cells: behaviour of carrageenan entrapped yeast during continuous ethanol fermentation

Summary Data of cell concentration, viability and microscopic observation of cell distribution inside carrageenan immobilized yeast beads are reported. Results were obtained from a continuous packed-bed reactor performing alcoholic fermentation and the main observations made on cell activity are in agreement with the fermentation profiles inside the fermenter.     

Plasma norepinephrine and heart rate dynamics during recovery from submaximal exercise in man

The time course of heart rate (HR) and venous blood norepinephrine concentration [NE], as an expression of the sympathetic nervous activity (SNA), was studied in six sedentary young men during recovery from three periods of cycle ergometer exercise at 21% +/- 2.8%, 43% +/- 2.1% and 65% +/- 2.3% of VO2max respectively (mean +/- SE). The HR decreased mono-exponentially with tau values of 13.6 +/- 1.6 s, 32.7 +/- 5.6 s and 55.8 +/- 8.1 s respectively in the three periods of exercise. At the low exercise level no change in [NE] was found. At medium and high exercise intensity: (a) [NE] increased significantly at the 5th min of exercise (delta [NE] = 207.7 +/- 22.5 pg.ml-1 and 521.3 +/- 58.3 pg.ml-1 respectively); (b) after a time lag of 1 min [NE] decreased exponentially (tau = 87 s and 101 s respectively); (c) in the 1st min HR decreased about 35 beats.min-1; (d) from the 2nd to 5th min of recovery HR and [NE] were linearly related (100 pg.ml-1 delta [NE] congruent to 5 beats.min-1). In the 1st min of recovery, independent of the exercise intensity, the adjustment of HR appears to have been due mainly to the prompt restoration of vagal tone. The further decrease in HR toward the resting value could then be attributed to the return of SNA to the pre-exercise level.     

The use of functional analysis of the ribosome as a tool to determine archaebacterial phylogeny

Forty different antibiotics with diverse kingdom and functional specificities were used to measure the functional characteristics of the archaebacterial translation apparatus. The resulting inhibitory curves, which are characteristic of the cell-free system analyzed, were transformed into quantitative values that were used to cluster the different archaebacteria analyzed. This cluster resembles the phylogenetic tree generated by 16S rRNA sequence comparisons. These results strongly suggest that functional analysis of an appropriate evolutionary clock, such as the ribosome, is of intrinsic phylogenetic value. More importantly, they indicate that the study of the nexus between genotypic and phenotypic (functional) information may shed considerable light on the evolution of the protein synthetic machinery.     

Transposon-generated Tn10 insertion mutations at thearo genes ofEscherichia coli K-12

We have obtained a set ofEscherichia coli K-12 derivatives with transposon-generated Tn10 insertion mutations at thearo genes of their aromatic biosynthetic pathway. Bacteriophage NK561 (Tn10) has been used for transposon mutagenesis ofE. coli, strain BW545. Tetracycline (Tc)-resistant derivatives were screened by their Aro^– phenotype by growth on a minimal medium with adequate requirements. Sixaro mutant types were mapped; two strains werearoA, twoaroD, onearoB oraroE, and onearoC. A selective medium and ad-cycloserine enrichment in the presence of tetracycline were used to select for Aro^–, Tc-sensitive derivatives. The reversion index to aromatic-independent colonies of some derivatives was less than 2 × 10^–11 per bacterium per generation. P1 transduction experiments transferred an aroA::Tn10 insertion fromE. coli BW545 to an enterotoxigenicE. coli strain from porcine origin. Derivatives of this strain beingaro, Tc-sensitive and not reverting toaro ⁺ at a detectable frequency, and many others transduced at will, may prove their usefulness as live vaccines.     

An improved purification procedure and properties of casein kinase II from brain

A simple and short purification procedure applicable to casein kinase II has been developed, for fully characterizing the enzyme from calf cerebral cortex cytosol. The procedure consists of four chromatographic steps: DEAE-cellulose, phosphocellulose, phosvitin-Sepharose and ATP-agarose which yields 87% pure casein kinase II. The purified enzyme shows three major bands with apparent molecular masses of 42, 38, and 27 kDa by polyacrylamide gel electrophoresis in sodium dodecyl sulfate and is self-autophosphorylated on its 27 kDa polypeptide. The enzyme shows all the characteristics described for casein kinase II from other sources: it is independent of cyclic nucleotides, calcium/phospholipids, and double-stranded poly(I).poly(C); it can utilize both ATP and GTP as phosphoryl donors and can phosphorylate both casein and phosvitin but not histone. The kinetic studies establish that theK _m for ATP is 12.5 M and 25.1 M when using phosvitin and casein respectively as phosphoryl acceptors. TheK _m for phosvitin is 0.91 mg/ml and for casein 1.43 mg/ml, while theV _max is 315 nmol/min/per mg protein and 479 nmol/min/per mg protein for phosvitin and casein respectively. The activity of the kinase is highly stimulated by KCl or NaCl, and almost completely inhibited by heparin concentrations of 1 g/ml (92%). This inhibition is reduced to only 33% in the presence of optimal KCl concentrations (150 mM). Spermine stimulates enzyme activity, whilst hemin produces a slight inhibition.     

Study on the mutagenicity of nifurtimox and eight derivatives with the L-arabinose resistance test of Salmonella typhimurium

The mutagenicity of nifurtimox (nfx) and 8 nfx analogues has been investigated with the L-arabinose forward-mutation assay of Salmonella typhimurium. The nfx analogues tested were obtained by replacing the 3-methyl-4-yl-tetrahydro-1,4-thiazine-1,1-dioxide group of the parent compound with the following other groups: indazol-1-yl (1); pyrazol-1-yl (2); benzimidazol-1-yl (3); 1,2,4-triazol-4-yl (4); 1-methyl-3-methylthio-1,2,4-triazol-4-yl-5-thione (5); 3,5-bis(methylthio)-1,2,4-triazol-4-yl (6); 1-adamantyl (7); 4,6-diphenylpyridin-1-yl-2-one (8). The mutagenic activity of each chemical was determined by the standard plate-incorporation test, in the presence or absence of the S9 activation mixture. The 9 compounds were mutagenic and exhibited linear dose-mutagenic response relationships. They were direct-acting mutagens and showed a nearly 1000-fold range in mutagenic potency from chemical 1 to nfx. In most cases, the addition of S9 mixture to the test plates decreased the mutagenicity of compounds. This effect was particularly noticeable in the case of chemicals 1-3, 5 and 7 where a more than 70% decrease in mutagenic activity was observed in the presence of the S9 mixture. The mutagenic potency of compounds in the Ara test showed a negative linear correlation with previously reported antitrypanosomal activity. Thus, chemicals 6 and 8 with in vitro activities against Trypanosoma cruzi clearly superior to that of nfx showed 2 of the lowest mutagenic potencies in the Ara test and these were only somewhat higher than the mutagenicity of the reference drug.     

The link between the electrogenic and redox functions of the plasmalemma and the energy metabolism of the cell

A dependence of the plasmalemma redox activity, determined by the reduction of external electron acceptors (ferricyanide, nitro-blue tetrazolium), on the energy state of the cell, which was modified by light conditions or introduction of glucose into the media, was shown on leaves of Elodea canadensis Rich. Glucose (10 m M ) and light (40 W m^-2) caused hyperpolarization of the membrane potential and stimulated the redox activity of the plasmalemma. 3-(3,4-Dichlorophenyl)-1,1-dimethylurea (DCMU) completely inhibited the light activation of electrogenic and redox functions of the plasmalemma. The light saturation intensity for membrane potential and ferricyanide reductase activity was 10–30% of the light saturation of photosynthesis. Membrane potential, K⁺ transport and plasmalemma redox activity changed in parallel in response to light and darkness and when DCMU was added. Ferricyanide reductase activity is suggested to be a simple parameter for characterizing the energy state of the cell. The functional significance of the light-induced hyperpolarization of the membrane potential is discussed.