Modifications of drug metabolism by disulfiram and diethyldithiocarbamate. II. D-Glucuronic acid pathway.

Hepatic enzymes connected with the formation and metabolism of free D-glucuronic acid were affected in rats after treatment with disulfiram or diethyldithiocarbamate (300 mg/kg, intragastrically, per day, 4 X). The activities of UDPglucose dehydrogenase, UDPglucuronic acid pyrophosphatase, UDPglucuronosyltransferase and L-gulonate dehydrogenase were enhanced, while those of glucose-6-phosphate dehydrogenase, beta-glucuronidase and D-glucuronolactone dehydrogenase were inhibited. These changes were more pronounced with disulfiram than diethyldithiocarbamate. Treatment with phenobarbital (80 mg/kg, i.p., per day, 4 X) enhanced UDP glucuronosyl-transferase, but brought about different effects on the other enzymes. Concurrent administration of phenobarbital with disulfiram or diethyldithiocarbamate led to potentiation or antagonism of the primary effects of each compound when given alone. The results suggest that activation of the D-glucuronic acid pathway may proceed in various ways, and that it is not necessarily followed by a simultaneous induction of the microsomal mixed-function oxygenase activity.     

Effect of clofibrate treatment on carnitine acyltransferases in different subcellular fractions of rat liver

The subcellular distribution of carnitine acetyl-, octanoyl-, and palmitoyltransferase in the livers of normal and clofibrate-treated male rats was studied with isopycnic sucrose density gradient fraction.In normal liver 48% of total carnitine acetyltransferase activity was peroxisomal, 36% of the activity located in mitochondria and 16% in a membranous fraction containing microsomes. Carnitine octanoyltransferase and carnitine palmitoyltransferase were confined almost totally (77–81%) to mitochondria in normal liver.Clofibrate treatment increased the total activity of carnitine acetyltransferase over 30 times, whereas the total activities of the other two transferases were increased only 5-fold.From the three different subcellular carnitine acetyltransferases the mitochondrial one was not responsive to clofibrate treatment, i.e. the rise in mitochondrial activity was over 70-fold as contrasted to the 6- and 14-fold rises in peroxisomal and microsomal activities, respectively. After treatment mitochondria contained 79% of total activity.It is concluded that the clofibrate-induced increase of carnitine acetyltransferase activity is not due to the peroxisomal proliferation that occurs during clofibrate treatment. The rise in peroxisomal activity contributed only 8% to the total increase.After clofibrate treatment the greatest part of carnitine octanoyl- and palmitoyltrnasferase activities were located in mitochondria but a considerable amount of both activities was found also in the soluble fraction of liver.     

Century‐long apparent decrease in intrinsic water‐use efficiency with no evidence of progressive nutrient limitation in African tropical forests

Forests exhibit leaf‐ and ecosystem‐level responses to environmental changes. Specifically, rising carbon dioxide (CO₂) levels over the past century are expected to have increased the intrinsic water‐use efficiency (iWUE) of tropical trees while the ecosystem is gradually pushed into progressive nutrient limitation. Due to the long‐term character of these changes, however, observational datasets to validate both paradigms are limited in space and time. In this study, we used a unique herbarium record to go back nearly a century and show that despite the rise in CO₂ concentrations, iWUE has decreased in central African tropical trees in the Congo Basin. Although we find evidence that points to leaf‐level adaptation to increasing CO₂—that is, increasing photosynthesis‐related nutrients and decreasing maximum stomatal conductance, a decrease in leaf δ¹³C clearly indicates a decreasing iWUE over time. Additionally, the stoichiometric carbon to nitrogen and nitrogen to phosphorus ratios in the leaves show no sign of progressive nutrient limitation as they have remained constant since 1938, which suggests that nutrients have not increasingly limited productivity in this biome. Altogether, the data suggest that other environmental factors, such as increasing temperature, might have negatively affected net photosynthesis and consequently downregulated the iWUE. Results from this study reveal that the second largest tropical forest on Earth has responded differently to recent environmental changes than expected, highlighting the need for further on‐ground monitoring in the Congo Basin.     

Offspring phenotype is shaped by the nonsperm fraction of semen

In a large majority of animal species, the only contribution of males to the next generation has been assumed to be their genes (sperm). However, along with sperm, seminal plasma contains a wide array of extracellular factors that have many important functions in reproduction. Yet, the potential intergenerational effects of these factors are virtually unknown. We investigated these effects in European whitefish (Coregonus lavaretus) by experimentally manipulating the presence and identity of seminal plasma and by fertilizing the eggs of multiple females with the manipulated and unmanipulated semen of several males in a full‐factorial breeding design. The presence of both own seminal plasma and foreign seminal plasma inhibited sperm motility, and the removal of own seminal plasma decreased embryo survival. Embryos hatched significantly earlier after both semen manipulations than in control fertilizations; foreign seminal plasma also increased offspring aerobic swimming performance. Given that our experimental design allowed us to control potentially confounding sperm‐mediated (sire) effects and maternal effects, our results indicate that seminal plasma may have direct intergenerational consequences for offspring phenotype and performance. This novel source of offspring phenotypic variance may provide new insights into the evolution of polyandry and mechanisms that maintain heritable variation in fitness and associated female mating preferences.     

Inhibition of the cardiac inward rectifier potassium currents by KB-R7943

KB-R7943 (2-[2-[4-(4-nitrobenzyloxy)phenyl]ethyl]isothiourea) was developed as a specific inhibitor of the sarcolemmal sodium–calcium exchanger (NCX) with potential experimental and therapeutic use. However, KB-R7943 is shown to be a potent blocker of several ion currents including inward and delayed rectifier K⁺ currents of cardiomyocytes. To further characterize KB-R7943 as a blocker of the cardiac inward rectifiers we compared KB-R7943 sensitivity of the background inward rectifier (I_K1) and the carbacholine-induced inward rectifier (I_KACh) currents in mammalian (Rattus norvegicus; rat) and fish (Carassius carassius; crucian carp) cardiac myocytes. The basal I_K1 of ventricular myocytes was blocked with apparent IC50-values of 4.6 × 10^− 6 M and 3.5 × 10^− 6 M for rat and fish, respectively. I_KACh was almost an order of magnitude more sensitive to KB-R7943 than I_K1 with IC₅₀-values of 6.2 × 10^− 7 M for rat and 2.5 × 10^− 7 M for fish. The fish cardiac NCX current was half-maximally blocked at the concentration of 1.9–3 × 10^− 6 M in both forward and reversed mode of operation. Thus, the sensitivity of three cardiac currents to KB-R7943 block increases in the order I_K1 ~ I_NCX < I_KACh. Therefore, the ability of KB-R7943 to block inward rectifier potassium currents, in particular I_KACh, should be taken into account when interpreting the data with this inhibitor from in vivo and in vitro experiments in both mammalian and fish models.     

Integrins regulate centrosome integrity and astrocyte polarization following a wound

In response to a wound, astrocytes in culture extend microtubule‐rich processes and polarize, orienting their centrosomes and Golgi apparatus woundside. β1 Integrin null astrocytes fail to extend processes toward the wound, and are disoriented, and often migrate away orthogonal, to the wound. The centrosome is unusually fragmented in β1 integrin null astrocytes. Expression of a β1 integrin cDNA in the null background yields cells with intact centrosomes that polarize and extend processes normally. Fragmented centrosomes rapidly assemble following integrin ligation and cell attachment. However, several experiments indicated that cell adhesion is not necessary. For example, astrocytes in suspension expressing a chimeric β1 subunit that can be activated by an antibody assemble centrosomes suggesting that β1 activation is sufficient to cause centrosome assembly in the absence of cell adhesion. siRNA knockdown of PCM1, a major centrosomal protein, inhibits cell polarization, consistent with the notion that centrosomes are necessary for polarity and that integrins regulate polarity via centrosome integrity. Screening inhibitors of molecules downstream of integrins indicate that neither FAK nor ILK is involved in regulation of centrosome integrity. In contrast, blebbistatin, a specific inhibitor of non‐muscle myosin II (NMII), mimics the response of β1 integrin null astrocytes by disrupting centrosome integrity and cell polarization. Blebbistatin also inhibits integrin‐mediated centrosome assembly in astrocytes attaching to fibronectin, consistent with the hypothesis that NMII functions downstream of integrins in regulating centrosome integrity. © 2012 Wiley Periodicals, Inc. Develop Neurobiol 73: 333–353, 2013.     

Snowflake Vitreoretinal Degeneration (SVD) Mutation R162W Provides New Insights into Kir7.1 Ion Channel Structure and Function

Snowflake Vitreoretinal Degeneration (SVD) is associated with the R162W mutation of the Kir7.1 inwardly-rectifying potassium channel. Kir7.1 is found at the apical membrane of Retinal Pigment Epithelial (RPE) cells, adjacent to the photoreceptor neurons. The SVD phenotype ranges from RPE degeneration to an abnormal b-wave to a liquid vitreous. We sought to determine how this mutation alters the structure and function of the human Kir7.1 channel. In this study, we expressed a Kir7.1 construct with the R162W mutation in CHO cells to evaluate function of the ion channel. Compared to the wild-type protein, the mutant protein exhibited a non-functional Kir channel that resulted in depolarization of the resting membrane potential. Upon co-expression with wild-type Kir7.1, R162W mutant showed a reduction of I_Kir7.1 and positive shift in ‘0’ current potential. Homology modeling based on the structure of a bacterial Kir channel protein suggested that the effect of R162W mutation is a result of loss of hydrogen bonding by the regulatory lipid binding domain of the cytoplasmic structure.