Dihydroteugin,a neo-clerodane diterpenoid from Teucrium chamaedrys

From the aerial part of Teucrium chamaedrys a new neo-clerodane diterpenoid, dihydroteugin, has been isolated, besides the previously known diterpenoids teucrin A and teugin. The structure of dihydroteugin, (12S)-15,16-epoxy-2β,6β-dihydroxy-neo-cleroda-13(16), 14-diene-18,19:20,12-diolide, was established by chemical and spectroscopic means and by partial synthesis from teugin.     

Interaction of beta-lactam antibiotics with penicillin-binding proteins from Bacillus megaterium

The binding properties of 25 beta-lactam antibiotics to Bacillus megaterium membranes have been studied. The affinities of the antibiotics for the penicillin-binding proteins (PBPs) are also reported. We found that PBP 4 has the highest affinity for nearly all the antibiotics studied whereas PBP 5 has the lowest affinity. Both PBP 4 and PBP 5 appear to be dispensable for the maintenance of bacterial growth and survival and appear to be DD-carboxypeptidases. Only the beta-lactam cefmetazol bound preferentially to PBP 5 and has been used to study the inhibition of DD-carboxypeptidase. Comparative studies with beta-lactam that simultaneously result in (a) binding to PBPs 1 and 3, (b) inhibition of cell growth and (c) lysis, stressed the importance of PBPs 1 and 3 for cell growth and survival.     

Interfering with calcium release suppresses I gamma, the "hump" component of intramembranous charge movement in skeletal muscle.

Four manifestations of excitation-contraction (E-C) coupling were derived from measurements in cut skeletal muscle fibers of the frog, voltage clamped in a Vaseline-gap chamber: intramembranous charge movement currents, myoplasmic [Ca2+] transients, flux of calcium release from the sarcoplasmic reticulum (SR), and the intrinsic optical transparency change that accompanies calcium release. In attempts to suppress Ca release by direct effects on the SR, three interventions were applied: (a) a conditioning pulse that causes calcium release and inhibits release in subsequent pulses by Ca-dependent inactivation; (b) a series of brief, large pulses, separated by long intervals (greater than 700 ms), which deplete Ca2+ in the SR; and (c) intracellular application of the release channel blocker ruthenium red. All these reduced calcium release flux. None was expected to affect directly the voltage sensor of the T-tubule; however, all of them reduced or eliminated a component of charge movement current with the following characteristics: (a) delayed onset, peaking 10-20 ms into the pulse; (b) current reversal during the pulse, with an inward phase after the outward peak; and (c) OFF transient of smaller magnitude than the ON, of variable polarity, and sometimes biphasic. When the total charge movement current had a visible hump, the positive phase of the current eliminated by the interventions agreed with the hump in timing and size. The component of charge movement current blocked by the interventions was greater and had a greater inward phase in slack fibers with high [EGTA] inside than in stretched fibers with no EGTA. Its amplitude at -40 mV was on average 0.26 A/F (SEM 0.03) in slack fibers. The waveform of release flux determined from the Ca transients measured simultaneously with the membrane currents had, as described previously (Melzer, W., E. Ríos, and M. F. Schneider. 1984. Biophysical Journal. 45:637-641), an early peak followed by a descent to a steady level during the pulse. The time at which this peak occurred was highly correlated with the time to peak of the current suppressed, occurring on average 6.9 ms later (SEM 0.73 ms). The current suppressed by the above interventions in all cases had a time course similar to the time derivative of the release flux; specifically, the peak of the time derivative of release flux preceded the peak of the current suppressed by 0.7 ms (SEM 0.6 ms). The magnitude of the current blocked was highly correlated with the inhibitory effect of the interventions on Ca2+ release flux.(ABSTRACT TRUNCATED AT 400 WORDS)     

The relationship between Q gamma and Ca release from the sarcoplasmic reticulum in skeletal muscle

Asymmetric membrane currents and fluxes of Ca2+ release were determined in skeletal muscle fibers voltage clamped in a Vaseline-gap chamber. The conditioning pulse protocol 1 for suppressing Ca2+ release and the "hump" component of charge movement current (I gamma), described in the first paper of this series, was applied at different test pulse voltages. The amplitude of the current suppressed during the ON transient reached a maximum at slightly suprathreshold test voltages (-50 to -40 mV) and decayed at higher voltages. The component of charge movement current suppressed by 20 microM tetracaine also went through a maximum at low pulse voltages. This anomalous voltage dependence is thus a property of I gamma, defined by either the conditioning protocol or the tetracaine effect. A negative (inward-going) phase was often observed in the asymmetric current during the ON of depolarizing pulses. This inward phase was shown to be an intramembranous charge movement based on (a) its presence in the records of total membrane current, (b) its voltage dependence, with a maximum at slightly suprathreshold voltages, (c) its association with a "hump" in the asymmetric current, (d) its inhibition by interventions that reduce the "hump", (e) equality of ON and OFF areas in the records of asymmetric current presenting this inward phase, and (f) its kinetic relationship with the time derivative of Ca release flux. The nonmonotonic voltage dependence of the amplitude of the hump and the possibility of an inward phase of intramembranous charge movement are used as the main criteria in the quantitative testing of a specific model. According to this model, released Ca2+ binds to negatively charged sites on the myoplasmic face of the voltage sensor and increases the local transmembrane potential, thus driving additional charge movement (the hump). This model successfully predicts the anomalous voltage dependence and all the kinetic properties of I gamma described in the previous papers. It also accounts for the inward phase in total asymmetric current and in the current suppressed by protocol 1. According to this model, I gamma accompanies activating transitions at the same set of voltage sensors as I beta. Therefore it should open additional release channels, which in turn should cause more I gamma, providing a positive feedback mechanism in the regulation of calcium release.     

Fibroblastlike primary cells from human colon adenocarcinoma explants: Collagen biosynthesis

Summary Fibroblastlike primary cells have been obtained from human colon adenocarcinoma explants. Such cells disappear during cell culture and thus have not been previously studied. These cells have a number of altered phenotypic characteristics: a) morphology; b) growth behavior and adherence to culture substrate (they required 3 h for 90% attachment and only presented a flattened morphology 40 h after platting); and c) collagen metabolism. Increased protein biosynthesis (about double than control colon-derived fibroblasts) and maintained ability for collagen biosynthesis have been observed for the tumor-associated fibroblastlike cells. Thus, the collagen to noncollagenous proteins ratio was decreased for these cells. They exhibited an altered type I:type III collagen (5:1 instead of 3:1 in colon fibroblasts) and procollagen (2:1 against 5:1 in colon fibroblasts) ratios as well as a decreased secretion of collagen with an abnormal deposition of procollagens in the cell layer. These studies show a permanent phenotypic alteration in the tumor-associated fibroblastlike cells.     

Uridine transport in basolateral plasma membrane vesicles from rat liver

Summary The characteristics of uridine transport were studied in basolateral plasma membrane vesicles isolated from rat liver. Uridine was not metabolized under transport measurement conditions and was taken up into an osmotically active space with no significant binding of uridine to the membrane vesicles. Uridine uptake was sodium dependent, showing no significant stimulation by other monovalent cations. Kinetic analysis of the sodium-dependent component showed a single system with Michaelis-Menten kinetics. Parameter values were K _M 8.9 m and V _max 0.57 pmol/mg prot/sec. Uridine transport proved to be electrogenic, since, firstly, the Hill plot of the kinetic data suggested a 1 uridine: 1 Na⁺ stoichiometry, secondly, valinomycin enhanced basal uridine uptake rates and, thirdly, the permeant nature of the Na⁺ counterions determined uridine transport rates (SCN^– > NO ₃ ^– > Cl^– > SO ₄ ^2– ). Other purines and pyrimidines cis-inhibited and trans-stimulated uridine uptake.This work has been partially supported by grant PM90-0162 from D.G.I.C.Y.T. (Ministerio de Educación y Ciencia, Spain). B.R.-M. is a research fellow supported by the Nestlé Nutrition Research Grant Programme.     

Evaluation of host and pathogen responses for screening soil amendments to control Sclerotium rolfsii

Three strains of Bradyrhizobium, 280A, 2209A and 32H1, that nodulated peanuts (Arachis hypogaea L.), were tested for their ability to grow and survive at elevated temperatures of up to 42°C in laboratory culture. Strain 32H1 was unable to grow at 37°C and was more sensitive to elevated temperatures than the other two strains. All three produced heat-shock proteins of molecular weights 17 kDa and 18 kDa. Two greenhouse experiments were conducted to determine the effect of high root temperature on nodulation, growth and nitrogen fixation of peanut. Two peanut varieties (Virginia cv NC7 and Spanish cv Pronto) were inoculated and exposed to root temperatures of 30°, 37° and 40°C. Nodulation and nitrogen fixation were strongly affected by root temperature but there was no variety × temperature interaction. At a constant 40°C root temperature no nodules were formed. Nodules were formed when roots were exposed to this temperature with diurnal cycling but no nitrogen fixation occurred. Highest plant dry weight, shoot nitrogen content and total nitrogen were observed at a constant root temperature of 30°C. Increasing root temperature to 37°C reduced average nitrogen content by 37% and total nitrogen by 49% but did not reduce nodulation. The symbiotic performance of the strains corresponded to their abilities to grow and survive at high temperature in culture.     

Platinum complexes of p- and m-aminobenzamidine. Synthesis, characterization, and cytotoxic activity.

Four platinum(II) aminobenzamidine complexes have been prepared and characterized by IR and 1H and 13C NMR spectroscopy, and tested for their ability to interact with the nicked and closed circular forms of the pUC8 plasmid DNA. The results show that the complexes of formula [Pt(LH)2Cl2]2X have a cis- geometry with an amino-Pt bonding, where L is either p- or m-aminobenzamidine and where 2X is 2Cl- or PtCl4(2-). It was observed that these complexes significantly alter the electrophoretic mobility of nicked and closed circular forms of DNA and that the alteration in electrophoretic mobility due to Pt(II)-p-aminobenzamidine binding is higher than that due to Pt(II)-m-aminobenzamidine. No difference in mobility was observed whether the DNA interacted with complexes having as counteranion Cl- or PtCl4(2-). The synthesized compounds were, in addition, assayed for antitumor activity in vitro against colon (CX-1), lung (LX-1), and mammary (MX-1) human tumor cells. The results show that these complexes inhibited the multiplication of the tumor cells and that they show higher specificity for lung cells.     

Ouabain-insensitive, Na-ATPase activity in pure suspensions of rat kidney proximal tubules

The present work was undertaken to evaluate the distribution of the Na-ATPase activity in the different components of the rat kidney cortex. Suspensions of glomeruli, proximal and distal tubules were prepared following a collagenase digestion of outermost kidney cortex slices and a separation on a Percoll gradient. It was found that the Na-ATPase activity is higher in the fraction enriched in proximal tubules. The fraction enriched in glomeruli and in distal tubules show also a Na-ATPase activity, but it is lower.     

Effects of eel (Anguilla anguilla L.) removals from selected sites of a stream on its subsequent densities

We assessed the effect of eel (Anguilla anguilla) removal from three sites of a Cantabrian stream upon its subsequent densities. In the first sample (Sept. 1986) numbers and densities were estimated as 43, 45 and 84 ind and 3490, 3030 and 3750 ind ha ⁻¹. Removal of these eels reduced the subsequent numbers and densities which, except on two occasions, were never reached again during the two years (eleven estimates) of study. Highest densities were recorded in the uppermost site in May and July, 1987, coincident with a strong drought and the lowest densities occurred in 1988 during a normal wet year. We hypothesize first that, because of a selective underground homing behaviour of eels, electro-fishing is inefficient and results in underestimates of the population. Second, seasonal variations of water discharge and droughts may not influence the homing behaviour of'eels until a threshold of dryness is reached. If this occurs, eels abandon their refuges and move towards the stream bottom. It seems that in Arroyo Chabatchos this threshold was exceeded in the summer of 1987 when the highest densities were estimated. The re-colonization of these sites experimentally depleted of eels, is a slow procces that lasts for, at least, two years.