Effects of Sulphur-containing Compounds on Lettuce Fruit Germination

The germination of Lactuca sativa L. cv. Arctic King fruit ona 0.5 per cent agar substrate was inhibited in light at temperaturesabove 31?C (? 1.0?). This precisely repeatable value, termedthe upper temperature cut-off point, is characteristic for thislettuce variety. The position of this cut-off point is raisedby the presence of thiourea (500–1000 mg/1) and a numberof other thiol compounds. This effect is not shown by all thiolcompounds and is dependent on the configuration of the restof the molecule. The presence of a hydroxyl group in the testcompound often results in the temperature cut-off point beinglowered but again this effect depends on the configuration ofthe rest of the molecule. It is postulated that the stimulatingeffects of active thiol compounds are related to hydrophobicinteractions with a macro-molecule involved in the initiationof germination.     

Dibucaine effects on structural and elastic properties of lipid bilayers

In this work we report the interaction effects of the local anesthetic dibucaine (DBC) with lipid patches in model membranes by Atomic Force Microscopy (AFM). Supported lipid bilayers (egg phosphatidylcholine, EPC and dimyristoylphosphatidylcholine, DMPC) were prepared by fusion of unilamellar vesicles on mica and imaged in aqueous media. The AFM images show irregularly distributed and sized EPC patches on mica. On the other hand DMPC formation presents extensive bilayer regions on top of which multibilayer patches are formed. In the presence of DBC we observed a progressive disruption of these patches, but for DMPC bilayers this process occurred more slowly than for EPC. In both cases, phase images show the formation of small structures on the bilayer surface suggesting an effect on the elastic properties of the bilayers when DBC is present. Dynamic surface tension and dilatational surface elasticity measurements of EPC and DMPC monolayers in the presence of DBC by the pendant drop technique were also performed, in order to elucidate these results. The curve of lipid monolayer elasticity versus DBC concentration, for both EPC and DMPC cases, shows a maximum for the surface elasticity modulus at the same concentration where we observed the disruption of the bilayer by AFM. Our results suggest that changes in the local curvature of the bilayer induced by DBC could explain the anesthetic action in membranes.     

Urethane anesthesia blocks the development and expression of kindled seizures

The effect of anesthetic and subanesthetic doses of urethane on the development of amygdala kindled seizures and on the expression of previously kindled seizures was studied in hooded rats. An anesthetic dose of urethane (1.5 g/kg) almost completely eliminated evoked afterdischarge and completely eliminated convulsive behavior in both groups. It also eliminated the seizure response to pentylenetetrazol. Subanesthetic doses of urethane (0.25 and 0.5 g/kg) strongly attenuated the expression of previously kindled seizures. These results suggest that urethane may not be an appropriate anesthetic for the study of epileptiform phenomena.     

Attenuation of phospholipid signaling provides a novel mechanism for the action of valproic acid

Valproic acid (VPA) is used to treat epilepsy and bipolar disorder and to prevent migraine. It is also undergoing trials for cancer therapy. However, the biochemical and molecular biological actions of VPA are poorly understood. Using the social amoeba Dictyostelium discoideum, we show that an acute effect of VPA is the inhibition of chemotactic cell movement, a process partially dependent upon phospholipid signaling. Analysis of this process shows that VPA attenuates the signal-induced translocation of PH(Crac)-green fluorescent protein from cytosol to membrane, suggesting the inhibition of phosphatidylinositol-(3,4,5)-trisphosphate (PIP(3)) production. Direct labeling of lipids in vivo also shows a reduction in PIP and PIP(2) phosphorylation following VPA treatment. We further show that VPA acutely reduces endocytosis and exocytosis-processes previously shown to be dependent upon PIP(3) production. These results suggest that in Dictyostelium, VPA rapidly attenuates phospholipid signaling to reduce endocytic trafficking. To examine this effect in a mammalian model, we also tested depolarization-dependent neurotransmitter release in rat nerve terminals, and we show that this process is also suppressed upon application of VPA and an inhibitor of phosphatidylinositol 3-kinase. Although a more comprehensive analysis of the effect of VPA on lipid signaling will be necessary in mammalian systems, these results suggest that VPA may function to reduce phospholipid signaling processes and thus may provide a novel therapeutic effect for this drug.     

Characterisation of the High Temperature Inhibition of Germination of Lettuce (Lactuca sativa)

Lactuca sativa L., lettuce (Arctic King) fruit germination is inhibited in the light at temperatures above 27^°C(±1.5^°). This value, which is precise and repeatable is referred to as the upper temperature cut-off point, and is characteristic for a particular species. It was found, that this cut-off point is shifted upwards by applying kinetin solutions (0.1–1 mg/1) or downwards by applying (±)-abscisic acid solutions (over 1 mg/1), the amount of shift being proportional to the level of active compound. In the presence of both compounds a balance was achieved. Effects of stimulators and inhibitors can most explicitly be described by reference to the direction and magnitude of the shift in the upper temperature cut-off point, which results from their application.     

The effect of anesthetics on hydrogen bonds An infrared study at low anesthetic concentrations

It is shown that a striking parallelism exists between the anesthetic potency of general halocarbon anesthetics and their influence on the hydrogen bond association constants in N-H…OC type hydrogen bonds, important for shaping the ion channels. It is further shown that the effect of potent anesthetics (which contain an acidic hydrogen) on the free/associated ratio in such hydrogen bonds is still significant at clinical anesthetic concentrations. It is argued that the results are in keeping with a pluralistic theory of anesthesia based on both hydrophobic and polar interactions.     

Volatile anesthetics affect nutrient availability in yeast

Volatile anesthetics affect all cells and tissues tested, but their mechanisms and sites of action remain unknown. To gain insight into the cellular activities of anesthetics, we have isolated genes that, when overexpressed, render Saccharomyces cerevisiae resistant to the volatile anesthetic isoflurane. One of these genes, WAK3/TAT1, encodes a permease that transports amino acids including leucine and tryptophan, for which our wild-type strain is auxotrophic. This suggests that availability of amino acids may play a key role in anesthetic response. Multiple lines of evidence support this proposal: (i) Deletion or overexpression of permeases that transport leucine and/or tryptophan alters anesthetic response; (ii) prototrophic strains are anesthetic resistant; (iii) altered concentrations of leucine and tryptophan in the medium affect anesthetic response; and (iv) uptake of leucine and tryptophan is inhibited during anesthetic exposure. Not all amino acids are critical for this response since we find that overexpression of the lysine permease does not affect anesthetic sensitivity. These findings are consistent with models in which anesthetics have a physiologically important effect on availability of specific amino acids by altering function of their permeases. In addition, we show that there is a relationship between nutrient availability and ubiquitin metabolism in this response.     

Contrasting membrane localization and behavior of halogenated cyclobutanes that follow or violate the Meyer-Overton hypothesis of general anesthetic potency.

The membrane localization and properties of two halogenated cyclobutanes were examined using 2H and 19F NMR. The common predictors of potency indicate that these two compounds will have anesthetic activity; however, 1,2-dichlorohexafluorocyclobutane (c(CCIFCCIFCF2CF2)) is not an effective anesthetic, whereas 1-chloro-1,2,2-trifluorocyclobutane (c(CCIFCF2CH2CH2)) is an effective general anesthetic. Using 2H NMR, the effect of these compounds on the acyl chain packing in palmitoyl (d31) oleoylphosphatidylcholine membranes was examined. The addition of the anesthetic c(CCIFCF2CH2CH2) results in small increases in the segmental order near the headgroup, whereas segments deeper in the bilayer show decreases in order. These results are consistent with those obtained previously for halothane, isoflurane, and enflurane. On the addition of the nonanesthetic c(CCIFCCIFCF2CF2), the segmental order in vitually unchanged, except for a slightly changed order near the segents 10-12 of the palmitoyl chains. These results, and the 19F chemical shifts, indicate that the anesthetic c(CCIFCF2CH2CH2) exhibits a preference for the membrane interface, as do the other general anesthetics, whereas the nonanesthetic c(CCIFCIFCF2CF2) resides within the membrane hydrocarbon core. The compound c(CCIFCCIFCF2CF2) and other nonanesthetic halocarbons have lower molecular dipole moments compared to effective anesthetic halocarbons, which may account for their altered distribution within the membrane. These data strongly suggest that preferential localization of a halocarbon within the membrane interface is a predictor of anesthetic potency. Furthermore, the data indicate that the properties and forces in the membrane interface deserve consideration as mediators of anesthetic activity.     

AFM study of interaction forces in supported planar DPPC bilayers in the presence of general anesthetic halothane

In spite of numerous investigations, the molecular mechanism of general anesthetics action is still not well understood. It has been shown that the anesthetic potency is related to the ability of an anesthetic to partition into the membrane. We have investigated changes in structure, dynamics and forces of interaction in supported dipalmitoylphosphatidylcholine (DPPC) bilayers in the presence of the general anesthetic halothane. In the present study, we measured the forces of interaction between the probe and the bilayer using an atomic force microscope. The changes in force curves as a function of anesthetic incorporation were analyzed. Force measurements were in good agreement with AFM imaging data, and provided valuable information on bilayer thickness, structural transitions, and halothane-induced changes in electrostatic and adhesive properties.     

AFM study of interaction forces in supported planar DPPC bilayers in the presence of general anesthetic halothane

In spite of numerous investigations, the molecular mechanism of general anesthetics action is still not well understood. It has been shown that the anesthetic potency is related to the ability of an anesthetic to partition into the membrane. We have investigated changes in structure, dynamics and forces of interaction in supported dipalmitoylphosphatidylcholine (DPPC) bilayers in the presence of the general anesthetic halothane. In the present study, we measured the forces of interaction between the probe and the bilayer using an atomic force microscope. The changes in force curves as a function of anesthetic incorporation were analyzed. Force measurements were in good agreement with AFM imaging data, and provided valuable information on bilayer thickness, structural transitions, and halothane-induced changes in electrostatic and adhesive properties.     

Interpretation of diagnostic data: 4. How to do it with a more complex table.

A more complex table is especially useful when a diagnostic test produces a wide range of results and your patient''s levels are near one of the extremes. The following guidelines will be useful: Identify the several cut-off points that could be used. Fill in a complex table along the lines of Table I, showing the numbers of patients at each level who have and do not have the target disorder. Generate a simple table for each cut-off point, as in Table II, and determine the sensitivity (TP rate) and specificity (TN rate) at each of them. Select the cut-off point that makes the most sense for your patient''s test result and proceed as in parts 2 and 3 of our series. Alternatively, construct an ROC curve by plotting the TP and FP rates that attend each cut-off point. If you keep your tables and ROC curves close at hand, you will gradually accumulate a set of very useful guides. However, if you looked very hard at what was happening, you will probably have noticed that they are not very useful for patients whose test results fall in the middle zones, or for those with just one positive result of two tests; the post-test likelihood of disease in these patients lurches back and forth past 50%, depending on where the cut-off point is. We will show you how to tackle this problem in part 5 of our series. It involves some maths, but you will find that its very powerful clinical application can be achieved with a simple nomogram or with some simple calculations.     

Background and interpretation of the 'Marine Trophic Index' as a measure of biodiversity

Since the demonstration, in 1998, of the phenomenon now widely known as 'fishing down marine food webs', and the publication of a critical rejoinder by Food and Agricultural Organization (FAO) staff, a number of studies have been conducted in different parts of the world, based on more detailed data than the global FAO fisheries statistics originally used, which established the validity and ubiquity of this phenomenon. In this contribution, we briefly review how, rather than being an artefact of biased data, this phenomenon was in fact largely masked by such data, and is in fact more widespread than was initially anticipated. This is made visible here by comparing two global maps of trophic level (TL) changes from the early 1950s to the present. The first presents the 50-year difference of the grand mean TL values originally used to demonstrate the fishing down effect, while the second is based on means above a cut-off TL (here set at 3.25), thus eliminating the highly variable and abundant small pelagic fishes caught throughout the world. Based on this, we suggest that using mean TL as 'Marine Trophic Index' (MTI), as endorsed by the Convention on Biological Diversity, always be done with an explicitly stated cut-off TL (i.e. cutMTI), chosen (as is the case with our proposed value of 3.25) to emphasize changes in the relative abundance of the more threatened, high-TL fishes. We also point out the need to improve the taxonomic resolution, completeness and accuracy of the national and international fisheries catch data series upon which the cutMTI is to be based.     

How to boost antioxidants by lipophilization?

Covalent modification of antioxidants through lipophilization is an important field of research aiming at developing antioxidants with improved efficacy. However, due to insufficient knowledge on how hydrophobicity affects antioxidant activity, lipophilization strategies have been largely based on empirism. Often, the resulting lipophilized antioxidants were not optimal. Here we described how the body of knowledge regarding hydrophobicity has been dramatically redefined as unexpected results were recently published. Using a broad range of lipophilized antioxidants assessed in dispersed lipids models and cultured cells, it has been demonstrated that the antioxidant activity increases progressively with increasing chain length up to a critical point, beyond which the activity of the compounds dramatically decreases. Taking into account this nonlinear phenomenon, also known as cut-off effect, antioxidant drug designers now have to seek the critical chain length to synthesize the optimal drug in a rational manner. Here, we briefly presented three putative mechanisms of action to try to account for the cut-off effect.     

Methoxyflurane acts at the substrate binding site of cytochrome P450 LM2 to induce a dependence on cytochrome b5

Rabbit cytochrome P450 isozyme 2 requires cytochrome b5 to metabolize the volatile anesthetic methoxyflurane but not the substrate benzphetamine [E. Canova-Davis and L. Waskell (1984) J. Biol. Chem. 259, 2541-2546]. To determine whether the requirement for cytochrome b5 for methoxyflurane oxidation is mediated by an allosteric effect on cytochrome P450 LM2 or cytochrome P450 reductase, we have investigated whether this anesthetic can induce a role for cytochrome b5 in benzphetamine metabolism. Using rabbit liver microsomes and antibodies raised in guinea pigs against rabbit cytochrome b5, we found that methoxyflurane did not create a cytochrome b5 requirement for benzphetamine metabolism. Methoxyflurane also failed to induce a role for cytochrome b5 in benzphetamine metabolism in the purified, reconstituted mixed function oxidase system. Studies of the reaction kinetics established that in the absence of cytochrome b5, methoxyflurane and benzphetamine are competitive inhibitors, and that in the presence of cytochrome b5, benzphetamine and methoxyflurane are two alternate substrates in competition for a single site on the same enzyme. These results all indicate that the methoxyflurane-induced cytochrome b5 dependence of the mixed function oxidase cytochrome P450 LM2 system is a direct result of the interaction between methoxyflurane and the substrate binding site of cytochrome P450 LM2 and suggest the focus of future studies of this question.     

Structural basis for the inhibition of firefly luciferase by a general anesthetic.

The firefly luciferase enzyme from Photinus pyralis is probably the best-characterized model system for studying anesthetic-protein interactions. It binds a diverse range of general anesthetics over a large potency range, displays a sensitivity to anesthetics that is very similar to that found in animals, and has an anesthetic sensitivity that can be modulated by one of its substrates (ATP). In this paper we describe the properties of bromoform acting as a general anesthetic (in Rana temporaria tadpoles) and as an inhibitor of the firefly luciferase enzyme at high and low ATP concentrations. In addition, we describe the crystal structure of the low-ATP form of the luciferase enzyme in the presence of bromoform at 2.2-A resolution. These results provide a structural basis for understanding the anesthetic inhibition of the enzyme, as well as an explanation for the ATP modulation of its anesthetic sensitivity.     

Ketamine inhibits fetal ACTH responses to cerebral hypoperfusion

The present study tested the effect of ketamine on the fetal reflex responses of late-gestation sheep to brachiocephalic occlusion (BCO), a stimulus that mimics the reduction in cerebral blood flow that results from severe fetal hypotension. Ketamine, a dissociative anesthetic and known noncompetitive antagonist of N-methyl D-aspartate (NMDA) receptors, has previously been shown to impair chemoreceptor responsiveness. Studies from this laboratory suggest that fetal reflex ACTH responses to hypotension are largely mediated by chemoreceptors; therefore, we hypothesized that ketamine would inhibit the reflex hormonal response to BCO. Chronically catheterized fetal sheep were subjected to acute cerebral hypoperfusion through occlusion of the brachiocephalic artery. Fetal blood pressure and heart rate were continuously recorded, and fetal blood samples drawn during the experiment were analyzed with specific hormone assays. Our results demonstrate that ketamine attenuates hemodynamic responses to cerebral hypoperfusion and is a potent inhibitor of ACTH and proopiomelanocortin (POMC)/pro-ACTH release. These data support the hypothesis that fetal reflex responses hypotension are chemoreceptor mediated. Given the potency with which ketamine inhibits ACTH response to fetal hypotension, we suggest that the use of ketamine or other anesthetic or analgesic drugs that block or otherwise interact with the NMDA-glutamate pathways, in late pregnancy or in preterm newborns be reconsidered.     

Uptake of dibucaine into large unilamellar vesicles in response to a membrane potential

Local amine anesthetics appear to exert their effects in the charged (protonated) form on the cytoplasmic side of excitable membranes. Two features of interest are the mechanism whereby these drugs move across the membrane to the inner monolayer and the actual membrane concentrations achieved. In this work, we have investigated the influence of a K+ diffusion potential, delta psi, on the transmembrane distribution and concentration of the local anesthetic dibucaine employing large unilamellar vesicle systems. It is demonstrated that egg phosphatidylcholine large unilamellar vesicles exhibiting a delta psi (interior negative) actively accumulate dibucaine to achieve high interior concentrations. 31P and 13C nuclear magnetic resonance studies show that the internalized drug is localized to the vesicle inner monolayer, and suggest that the protonated form of the anesthetic is the species that is actively transported. The inner monolayer anesthetic concentrations thus achieved can be an order of magnitude or more larger than predicted on the basis of anesthetic lipid-water partition coefficients. It is suggested that these effects may be related to the mechanisms whereby local anesthetics are localized and concentrated at their sites of action in nerve membranes.     

Further implications of a theory of immunity.

A theory of immunity presented previously showed that many immune phenomena could be explained in terms of a simple model involving interactions between (i) the receptors on immunologically competent cells, (ii) antigenic determinants, (iii) natural antibody and (iv) complement. Several features of the theory subsequently gained experimental support. The present paper extends the analysis and predicts that two mechanisms operate in the induction of immunological tolerance to self antigenic determinants. High specificity anti-self cells are destroyed by a mechanism involving complement. In contrast, low specificity anti-self cells are stimulated by self determinants and increase in numbers. This increases the concentration of the natural antibody secreted by these cells which acts as a “blocking” antibody preventing their continued stimulation by self determinants.The expanded clones of low specificity anti-self cells, which may be of high specificity for “near-self” determinants, (i) are responsible for the greater immunological responsiveness between non-identical members of the same species than between members of different species (“alloaggression”), and (ii) provide a barrier opposing the progressive evolution of the surface determinants of a pathogen into forms identical with the surface determinants of its host.Following exposure to a given self or not-self antigenic determinant, the distribution curve for cells of varying specificities for the determinant shows a sharp cut-off point between high and low specificity cells. The position of this cut-off point is critical in determining the subsequent response of the organism to the determinant. Variables affecting the cut-off point include, antibody present prior to the exposure of cells to the determinant, complement, complement inhibitors, the cell membrane and certain drugs. Autoimmune diseases are improved by drugs (e.g. chloroquine) which move the position of the cut-off point towards cells of low specificities for self determinants.     

Effect of general anesthesia on syncytiotrophoblast plasma membranes from human placenta

This investigation shows that a general anesthetic produces similar effects in vivo and in vitro. Anesthesia with a barbituric drug, Thiopental, induces an increase in membrane fluidity and a decrease in the activity of acetylcholinesterase in syncytiotrophoblast plasma membranes (SPM) obtained from placentas after caesarean section. The same effects can be reproduced in vitro after anesthetic addition to the isolated plasma membranes. Morphological and freeze-fracturing studies also suggest that membrane protein components are affected by anesthetics.