Halothane inhibits the neurotoxin stimulated [14C]guanidinium influx through 'silent' sodium channels in rat glioma C6 cells

We have investigated the effect of pharmacological agents on [14C]guanidinium ion influx through sodium channels in C6 rat glioma and N18 mouse neuroblastoma cells. The sodium channels of the N18 cells can be activated by aconitine alone, indicating that they are voltage-dependent channels. In contrast, sodium channels in the C6 cells require the synergistic action of aconitine and scorpion toxin for activation and are therefore characterized as so-called silent channels. The general anesthetic halothane used at clinical concentrations, specifically inhibited the ion flux through the silent sodium channel of C6 rat glioma cells. The voltage-dependent channels of the N18 cells were insensitive to halothane at the concentrations tested.     

Volatile anesthetics inhibit the ion flux through Ca2+-activated K+ channels of rat glioma C6 cells

Ca2+-activated K+ channels in rat glioma C6 cells were investigated using monolayers of these cells in petri dishes. The ion flux through the channels was studied with 86Rb+ after addition of a Ca2+-ionophore to the incubation medium. Both the influx and efflux of 86Rb+ through these Ca2+-activated K+ channels were inhibited by the general anesthetic halothane (at clinical concentrations). Other volatile anesthetics such as isoflurane, enflurane and methoxyflurane also inhibited the Ca2+-activated K+ channels at clinical concentrations. Inhibition of these channels by general anesthetics could have profound effects on signal transmission in the brain.     

Separation of Crotalus atrox (western diamondback rattlesnake) alpha-proteinase from serine proteinase and hemorrhagic factor activities

A method for obtaining Crotalus atrox alpha-proteinase (EC 3.4.24.1) in a pure form has been developed. Fractionation of the crude venom on DEAE-Sepharose, followed by gel filtration on Bio-Gel P-150 and chromatography on CM-Sepharose, yielded an alpha-proteinase preparation which showed a single band on disc and sodium dodecyl sulfate-polyacrylamide gel electrophoresis and had an activity on casein approximately twice that previously reported. The enzyme is a nonglycosylated single-chain polypeptide with a molecular weight of 26,738 and a pI of 8.15. Proteolytic activity on casein, alpha 1-antichymotrypsin, and Cl-inhibitor was abolished by treatment of alpha-proteinase with 1 mM EDTA, but full activity was retained in the presence of 1 mM phenylmethylsulfonyl fluoride. Caseinolytic activity was increased by 33 and 55% in the presence of 10 mM Mg2+ and Ca2+, respectively. Pure alpha-proteinase is devoid of esterolytic activity on H-D-Pro-Phe-Arg-p-nitroanilide (S-2302), benzoyl-L-arginine ethyl ester, and benzoyl-L-tyrosine ethyl ester. The final preparation has no hemorrhagic factor activity.     

Butyrate induced accumulation of a 2.3 kb polyadenylated H1(0) histone mRNA in HeLa cells.

Sodium butyrate was used to induce the accumulation of human H1(0) mRNA in HeLa cells. The length of this mRNA (2,300 nucleotides) was determined by Northern blot hybridization and S1 nuclease analysis using a human H1(0) gene probe. The mRNA shows long 5' and 3' non coding segments and it is polyadenylated. The signal for this step of mRNA maturation (cleavage and polyadenylation) appears to be the hexanucleotide AAUAAA in analogy to most (other than histone) mRNA species. Thus, the mode of maturation of H1(0) mRNA differs, on one hand, from that of the cell cycle dependent mRNA species, where it is based on a specific stem-and-loop structure. On the other hand, the 3' end of H1(0) mRNA varies from H5 mRNA, which is characterized by two unique dyad symmetry structures at its 3' end.     

Correlated response in male and female sterility to selection for pupa weight in Tribolium castaneum

Summary The correlated responses in male and female sterility to 50 generations of individual selection for pupa weight in Tribolium were analyzed. Two replicate lines (S-lines) were selected for heavier pupa weight and stabilizing selection for pupa weight was practiced in two replicate control lines (C-lines). There was close agreement between replicates in both sets of lines for direct and correlated responses. The rate of inbreeding has been constant for all lines (approximately 0.5% per generation).Regression of generation means for pupa weight on generation of selection indicated a significant linear regression in the direct response for both lines. The linear increases of 46 and 55 g. per generation in the S-lines accounted for 98% of the variation among generations and the linear decreases of 5 and 10 g. per generation in the C-lines accounted for 70–90% of the variation in the generation means.Maximum likelihood estimators were used to calculate the frequency of male and female sterility for each generation and line. Average sterility in the base population ranged from about 4 to 12% for both sexes. Polynomial regressions of percent sterility on generation of selection showed that quadratic and higher order regressions were occasionally significant but accounted for a relatively small fraction of the total variation. In the two S-line replicates, linear regression coefficients of percent sterility on generation number were 0.16±.09 and 0.20±.07 for males and 0.72±.08 and 0.54±.08 for females, suggesting a larger correlated response in female than in male sterility. In the C-lines, linear regression coefficients were 0.02±.08 and –.12±.05 for males in the two replicates and –.05±.05 and –.05±.05 for females. Estimates of realized genetic correlations between pupa weight and sterility in the S-lines ranged from 0.04 to 0.14 for males and from 0.14 to 0.37 for females when the heritability of sterility was allowed to take on values from 0.05 to 0.25.Supported by NSF Grants G-1238 and GB-5987, NIH Grant GM-16074 and NIH Fellowship 1 FO2 GM4 5130-01.     

In Xenopus laevis, the product of a developmentally regulated mRNA is structurally and functionally homologous to a Saccharomyces cerevisiae protein involved in translation fidelity.

We have performed a differential screen of a Xenopus egg cDNA library and selected two clones (Cl1 and Cl2) corresponding to mRNA which are specifically adenylated and recruited into polysomes after fertilization. Sequence analysis of Cl1 reveals that the corresponding protein is 67.5% identical (83% similar) to the product of the Saccharomyces cerevisiae SUP45 (also called SUP1 or SAL4) gene. This gene, when mutated, is an omnipotent suppressor of nonsense codons. When expressed in a sup45 mutant, the Xenopus Cl1 cDNA was able to suppress sup45-related phenotypes, showing that the structural homology reflects a functional homology. Our discovery of a structural and functional homolog in Xenopus cells implies that the function of SUP45 is not restricted to lower eukaryotes and that the SUP45 protein may perform a crucial cellular function in higher eukaryotes.     

A major difference between the divergence patterns within the lines-1 families in mice and voles

L1 retroposons are represented in mice by subfamilies of interspersed sequences of varied abundance. Previous analyses have indicated that subfamilies are generated by duplicative transposition of a small number of members of the L1 family, the progeny of which then become a major component of the murine L1 population, and are not due to any active processes generating homology within preexisting groups of elements in a particular species. In mice, more than a third of the L1 elements belong to a clade that became active approximately 5 Mya and whose elements are > or = 95% identical. We have collected sequence information from 13 L1 elements isolated from two species of voles (Rodentia: Microtinae: Microtus and Arvicola) and have found that divergence within the vole L1 population is quite different from that in mice, in that there is no abundant subfamily of homologous elements. Individual L1 elements from voles are very divergent from one another and belong to a clade that began a period of elevated duplicative transposition approximately 13 Mya. Sequence analyses of portions of these divergent L1 elements (approximately 250 bp each) gave no evidence for concerted evolution having acted on the vole L1 elements since the split of the two vole lineages approximately 3.5 Mya; that is, the observed interspecific divergence (6.7%-24.7%) is not larger than the intraspecific divergence (7.9%-27.2%), and phylogenetic analyses showed no clustering into Arvicola and Microtus clades.