Relationships between blood groups, isozymes and halothane reaction in pigs from a selection experiment

German Landrace pigs (n = 1500) were halothane-tested and blood samples were taken for the determination of A-O and H blood types as well as for the determination of PHI and 6-PGD isozymes. The pigs originated from two generations (7th and 8th) of a selection experiment 'selection for activity of NADPH-generating enzymes in backfat of pigs'. The selection lines are E-, E+ (selection for low and high enzyme activity), U- (selection for low ultrasonic backfat thickness) and K (control). Preliminary results show an average proportion of halothane-susceptible animals of 49%. The frequencies of halothane-positive pigs amount to 60%, 46%, 70% and 30% in lines E-, E+, U- and K, respectively. The investigation shows a non-random combination of the marker genes caused by linkage disequilibrium, especially in line E-. Recombination frequencies between the loci vary from 0% to 18%.     

Patterns of ion distribution in selected NaCl tolerant and normal lines of four grass species

Selected NaCl tolerant and unselected control lines ofHolcus lanatus L.,Lolium perenne L.,Dactylis glomerata L., andFestuca rubra L. were grown in sand culture at 0, 100, 200, 250, and/or 300 ml m^-3NaCl for seven weeks. The tolerant lines of all four species produced significantly greater both shoot and root dry matter at all NaCl treatments compared with the unselected control lines. Na⁺, K⁺, Cl^-, Ca²⁺, and Mg²⁺ contents of leaf, stalk, and roots of each species were determined. The tolerant lines ofH. lanatus contained less Na⁺ and less Ca²⁺ but higher K⁺ in shoots, compared with the unselected line. By contrast theL. perenne tolerant line had higher Na⁺ and Cl^- contents at 250, and 300 mol m^-3 NaCl in shoots than the unselected line suggesting a halophytic nature of the tolerant line.D. glomerata accumulated greater quantities of ions compared with the other species examined. The tolerant line contained significantly less Cl^- but more K⁺ in its shoots than the unselected line. Na⁺, Cl^-, and K⁺ contents in the shoots of the tolerant line ofF.rubra were higher than in the unselected line shoots. Therefore selection for NaCl tolerance may provide useful material for examining the basis of tolerance.     

Detection of<Emphasis Type="Italic">pap</Emphasis>-,<Emphasis Type="Italic">sfa</Emphasis>- and<Emphasis Type="Italic">afa</Emphasis>-specific DNA sequences in<Emphasis Type="Italic">Escherichia coli</Emphasis> strains isolated from extraintestinal material

P-fimbriae, S-fimbriae and AFA-adhesins are virulence factors responsible for adherence ofEscherichia coli strains to extraintestinal host-cell surface. Detection ofpap-,sfa- andafa-specific sequences performed by PCR revealed 74%pap ⁺, 65%sfa ⁺, and 8.3%afa ⁺ strains in a group of 84 extraintestialE. coli isolates. Detection in a group of fecal strains showed 29%pap ⁺, 21%sfa ⁺ and 4%afa ⁺ strains.pap together withsfa were found as the most frequent combination (56%) among extraintestinal isolates probably due to localization ofpap-andsfa-operons on a common pathogenicity island. The occurrence ofafa-specific sequence among 56 urine strains was 11%, although noafa ⁺ strain was detected among 28 gynecological isolates. No strains with detected adhesin operons were found among twenty (24%) extraintestinalE. coli strains.     

Ethylene-insensitive related phenotypes exist naturally in a genetically variable population of Dianthus barbatus

Within the genetically variable population of Dianthus barbatus’’Dagan’’, we identified genetic lines with flowers possessing ethylene insensitivite-related phenotypes. The phenotypes are: Caf ⁺ , with a fresh-looking corolla attached to a mature ovary; Inr ^– , with flowers whose petals do not inroll during flower senescence; and Rfi ⁺ , whose flower petals inroll, but recover from inrolling. The frequencies of Caf ⁺ , Inr ^- and Rfi ⁺ were 33%, 25% and 75% in the population inspected, respectively. These relatively high frequencies were probably due to continuous selection over the years for flowers with open and long-lasting corollas. By examining the distribution of the genetic lines which express two of these phenotypes, we determined that Caf ⁺ , Inr ^– and Rfi ⁺ are independent traits. However, these traits might be associated with male sterility. Exposure of a random sample of the population to exogenous ethylene, followed by examination of the resulting inrolling and wilting, revealed five different ethylene response groups. In one group ethylene enhanced both inrolling and wilting, and most genetic lines within this group exhibited a Caf ^– phenotype. In two other groups ethylene enhanced either the inrolling or the wilting, and both of the Caf ⁺ and Caf ^– phenotypes were included. Two other groups were completely non-responsive to ethylene, but in one of them the flower life span was twice as long as in the other, and all the genetic lines exhibited the Caf ⁺ phenotype. We concluded that the Caf ⁺ phenotype is most-likely related to ethylene insensitivity and that the inrolling and wilting are controlled by ethylene, probably via different pathways. Received: 25 June 2000 / Accepted: 15 November 2000     

Fluorescent labeling of (Na + K)-ATPase by 5-iodoacetamidofluorescein

5-Iodoacetamidofluorescein (5-IAF) covalently labels dog kidney (Na⁺ + K⁺)-ATPase with approximately 2 moles incorporated per mole of enzyme. ATPase and K⁺-phosphatase activities are fully retained after reaction, and the kinetic parameters for Na⁺, K⁺, Mg²⁺, ATP and p-nitrophenyl phosphate are likewise not significantly affected. The fluorescence of the bound 5-IAF is increased by ATP, Na⁺, and Mg²⁺, and decreased by K⁺. These fluorescence changes likely reflect ligand-induced stabilization of the E₁ or E₂ states of the enzyme.     

Positive selection of Tcrb-V10b+ T cells

The Tcrb-V10b⁺ T cell population has been examined with a newly established antibody, KT10b, specific for Tcrb-V10b but not Tcrb-V10a. H-2E⁺ mice have higher levels of Tcrb-V10b⁺ T cells (4.3%–11.%) than H-2E^– mice (2.2%–4.9%). This difference appears to be determined by levels of Tcrb-V10b⁺ T cells in the CD4 population. F₁ mice between H-2E⁺ and H-2E^– mice dominantly express higher levels of Tcrb-V10b⁺ T cells. [NOD (E–) x (NOD x A (E⁺))F₁] backcross mice show positive selection of Tcrb-V10b⁺ CD4⁺ T cells by H-2E. On the other hand other backcross analyses reveal positive selection of Tcrb-V10b⁺ CD8⁺ T cells by certain major histocompatibility class I molecules. Involvement of non-H-2 antigens in these positive selections remain to be determined. Address correspondence and offprint requests to: K. Tomonari.     

Characterization of whole-cell k<Superscript>+</Superscript> currents across the plasma membrane of pollen grain and tube protoplasts of <Emphasis Type="Italic">Lilium longiflorum</Emphasis>

Outward and inward currents, mainly carried by K⁺, were detected in protoplasts of pollen grains (PG) and pollen tubes (PT) of Lilium longiflorum Thunb. by using the whole-cell configuration of the patch-clamp technique. The outward K⁺ current (I_{K+ out}) was similar in both protoplast types, while the inward K⁺ current (I_{K+ in}) was higher in pollen tube protoplasts. In PT but not in PG protoplasts, inward K⁺ currents were already detectable at negative membrane voltages usually monitored in lily pollen. I_{K+ in} consisted of a slow and a fast current component, as revealed by fitting a sum of two exponential functions to the time-dependent current. The contribution of the fast component to the total inward current was higher in PT than in PG protoplasts, which was even more evident at acidic pH of the external medium. Therefore, based on the measured characteristics, the I_{K+ in} of PT protoplasts may contribute to the endogenous K⁺ currents surrounding a growing pollen tube. Abbreviations: BS, bath solution; BTP, bis-Tris-propane; MES, 2-N-morpholinoethane sulfonic acid; V_act, activation voltage; V_M, membrane voltage; E_rev, reversal potential; I_{K+ in}, inward K⁺ current; I_{K+ out}, outward K⁺ current; PG, pollen grain; PT, pollen tube; PM, pipette medium     

Reversal by DNA amplifications of an unusual mutation blocking alkane and alcohol utilization in Pseudomonas putida

Summary We analyzed the reversion of strains carrying alk208, a mutation in the alkBAC (alkane utilization) region of the Pseudomonas CAM-OCT plasmid. Reversion of alk208 was stimulated 25 to 75-fold by small doses of UV-irradiation. All alkane hydroxylase-positive (AlkB⁺) revertants proved to be aliphatic alcohol dehydrogenasepositive (AlkC⁺) as well, whereas AlkC⁺ revertants could be either AlkB⁺ or AlkB^-. Most of the AlkB^- AlkC⁺ partial revertants produced AlkC^- segregants at measurable frequencies. UV-irradiation substantially increased the rate of AlkC^- segregation. Most segregants reverted to AlkB⁺ or AlkC⁺ at frequencies similar to the original alk208 strain. Dot blot hybridization analyses using cloned probes from various regions of CAM-OCT revealed that the partial revertants contained specific amplications of alk DNA. The endpoints of these amplifications mapped in at least two regions. AlkC^- segregants had lost the DNA amplifications.     

Differential MHC expression requirements for positive selection of separate TCR Vb families

 Positive selection has been proposed to be involved in protection from diabetes. We examined positive selection by fluorescence-activated cell sorter analyses in thymocytes of protected and susceptible E-transgenic and non-transgenic NOD mice. Three Vb families showed positive selection in E-transgenic mice. Vb6⁺CD4⁺ and Vb10⁺CD4⁺ thymocytes were found at higher frequencies in both protected NOD-Ea and susceptible NOD-DY mice. The increased frequencies of Vb13⁺CD8⁺ thymocytes were found in protected NOD-Ea mice only, and not in susceptible NOD-DY transgenic mice. These three Vb families were further examined in bone-marrow chimeras between NOD-Ea and non-transgenic NOD mice, where we could examine the contribution of E-expressing bone-marrow-derived cells in positive selection. We find that NOD-Ea→NOD-Ea chimeras have an increased positive selection of Vb13⁺CD8⁺ cells and that positive selection is more efficient when both thymic epithelium and bone-marrow-derived cells express the E molecule. This was also seen for Vb6⁺CD4⁺ cells. However, for Vb6, bone-marrow-derived cells alone were also capable of positive selection. Positive selection of Vb10⁺CD4⁺ cells was restricted to E-expressing thymic epithelium only. For Vb13⁺CD8⁺ cells, we found that positive selection is most efficient with E-expression on both thymic epithelium and bone-marrow-derived cells, although positive selection also occurs with E-positive epithelium only. For Vb6⁺ CD4⁺ cells, the dominating selecting cells are bone-marrow-derived cells, and Vb10⁺CD4⁺ cells seem to be selected exclusively by the thymic epithelium. Thus, the conditions for positive selection seem to vary considerably between different Vb families. Received: 8 April 1998 / Revised: 29 June 1998     

Mechanism of Cl- sensitivity in internal ion receptors of the leech; an inward current gated off by Cl- in the nephridial nerve cells

Summary The nephridial nerve cells of the leech, Hirudo medicinalis, 34 sensory cells, each associated with one nephridium, are sensitive to changes in extracellular Cl^- concentration, an important factor in ion homeostasis. Using single-electrode current- and voltage clamp and ion substitution techniques, the specificity and mechanism of Cl^- sensitivity of the nephridial nerve cell was studied in isolated preparations. Increase of the normally low external Cl^- concentration leads to immediate and sustained hyperpolarization, decrease of the frequency of bursts and decrease of membrane conductance. The response is halogen specific: Cl^- can be replaced by Br^–, but not by organic mono- or divalent anions or inorganic divalent anions.At physiological Cl^- concentrations (36mM extra-cellular Cl^-), the nephridial nerve cell has a high resting conductance for Cl^- and the membrane potential is governed by Cl^-. In high extracellular Cl^- concentrations (110–130 mM), membrane conductance is low, most likely due to the gating off of Cl^- channels. Under these conditions, membrane potential is dominated by the K⁺ distribution and the nephridial nerve cell hyperpolarizes towards E_K.Abbreviations NNC nephridial nerve cell - V _m membrane potential - E _Cl(k) equilibrium potential for Cl (K) - IV-curve current-voltage relationship     

An assessment of thermodynamic reaction constants for simulating aqueous environmental monomethylmercury speciation

ABSTRACT

Monomethylmercury (CH₃Hg ⁺) is both the most ecologically significant and the least well characterized species of mercury in environmental settings. Our understanding of the environmental speciation behavior of this compound is limited both as the result of lesser available laboratory data (when compared to inorganic mercury) as well as the uncertainties associated with our understanding of the properties of environmental ligands. A careful examination and synthesis of data reported in the technical literature led to the following findings: (1) a 25°C, zero ionic strength bicarbonate ion complexation constant estimate is remarkably close to an earlier reported value at 0.4 M: CH₃Hg⁺ + HCo₃^-?CH₃HgHCO₃,log₁₀K = 2.6 (±0.22, 1 SD), (2) three 25°C zero ionic strength reaction constants reported by DeRobertis et al.(1998) were confirmed to within ~±0.1 log₁₀K units: CH₃Hg ⁺+ OH^-?CH₃HgOH, log₁₀K = 9.47; 2CH₃Hg ⁺ + H₂O?(CH₃Hg)₂OH ⁺ + H⁺, log₁₀K =?2.15; CH₃Hg ⁺+ Cl^-?CH₃HgCl, log₁₀K = 5.45, (3) “best estimate” literature complexation constants corrected to zero ionic strength include: CH₃Hg ⁺ + F^-?CH₃HgF, log₁₀K = 1.75 (20°C corr. Schwart-zenbach and Schellenberg, 1965); CH₃Hg ⁺ + Br^-?CH₃HgBr, log₁₀K = 6.87 (20°C corr. Schwartzenbach and Schellenberg, 1965); CH₃Hg ⁺ +1^-?CH₃HgI, log₁₀K = 8.85 (20°C corr. Schwartzenbach and Schellenberg, 1965); and CH₃Hg ⁺+ SO₄^2-?CH₃HgSO₄^-,log₁₀K = 2.64 (25°C, DeRobertis et al., 1998), (4) literature reported values for simulating monomethylmercury complexation with the carbonate ion may be too low: CH₃Hg ⁺+ CO₃^2-?CH₃HgCO₃^-, log₁₀K = 6.1 (Rabenstein et al., 1976; Erni, 1981), and (5) ‘‘best estimate’’ constants for simulating methyl mercury complexation with reduced environmental sulfur species include: CH₃Hg ⁺ + S^2-?CH₃HgS ^-, log₁₀K = 21.1; CH³Hg ⁺+ SH ^-? CH3HgSH, log₁₀K = 14.5 (H ⁺ + SH^-?CH₂S, log₁₀K = 6.88; Dyrssen and Wedborg, 1991); CH₃Hg ⁺ + RS^-?CH₃HgSR, log₁₀K = 16.5 (H + + RS^-?RSH, log₁₀K = 9.96; Qian et al., 2002); and CH₃Hg ⁺+ CH₃HgS^{1 -}?(CH₃Hg)₂S, log₁₀K = 16.32 (Schwartzenbach and Schellenberg, 1965; Rabenstein et al., 1978; and Erni, 1981).     

Genome-wide association study for growth and fatness traits in Chinese Sujiang pigs

Growth and fatness traits are complex and economically important traits in the pig industry. The molecular basis underlying porcine growth and fatness traits remains largely unknown. To uncover genetic loci and candidate genes for these traits, we explored the GeneSeek GGP Porcine 80K SNP chip to perform a GWAS for seven growth and fatness traits in 365 individuals from the Sujiang pig, a recently developed breed in China. We identified two, 17, one and 11 SNPs surpassing the suggestively significant threshold (P < 1.86 × 10⁻⁵) for body weight, chest circumference, chest width and backfat thickness respectively. Of these SNPs, 20 represent novel genetic loci, and five and four SNPs were respectively associated with chest circumference and backfat thickness at a genome-wide significant threshold (P < 9.31 × 10⁻⁷). Eight SNPs had a pleiotropic effect on both chest circumference and backfat thickness. The most remarkable locus resided in a region between 72.95 and 76.27 Mb on pig chromosome 4, harboring a number of previously reported quantitative trait loci related to backfat deposition. In addition to two reported genes (PLAG1 and TAS2R38), we identified four genes including GABRB3, ZNF106, XKR4 and MGAM as novel candidates for body weight and backfat thickness at the mapped loci. Our findings provide insights into the genetic architecture of porcine growth and fatness traits and potential markers for selective breeding of Chinese Sujiang pigs.     

Divalent cations and the phosphatase activity of the (Na + K)-dependent ATPase

Phosphatase activity of a kidney (Na + K)-ATPase preparation was optimally active with Mg²⁺ plus K⁺. Mn²⁺ was less effective and Ca²⁺ could not substitute for Mg²⁺. However, adding Ca²⁺ with Mg²⁺ or substituting Mn²⁺ for Mg²⁺ activated it appreciably in the absence of added K⁺, and all three divalent cations decreased apparent affinity for K⁺. Inhibition by Na⁺ decreased with higher Mg²⁺ concentrations, when Ca²⁺ was added, and when Mn²⁺ was substituted for Mg²⁺. Dimethyl sulfoxide, which favorsE ₂ conformations of the enzyme, increased apparent affinity for K⁺, whereas oligomycin, which favorsE ₁ conformations, decreased it. These observations are interpretable in terms of activation through two classes of cation sites. (i) At divalent cation sites, Mg²⁺ and Mn²⁺, favoring (under these conditions)E ₂ conformations, are effective, whereas Ca²⁺, favoringE ₁, is not, and monovalent cations complete. (ii) At monovalent cation sites divalent cations compete with K⁺, and although Ca²⁺ and Mn²⁺ are fairly effective, Mg²⁺ is a poor substitute for K⁺, while Na⁺ at these sites favorsE ₁ conformations. K⁺ increases theK _m for substrate, but both Ca²⁺ and Mn²⁺ decrease it, perhaps by competing with K⁺. On the other hand, phosphatase activity in the presence of Na⁺ plus K⁺ is stimulated by dimethyl sulfoxide, by higher concentrations of Mg²⁺ and Mn²⁺, but not by adding Ca²⁺; this is consistent with stimulation occurring through facilitation of an E₁ to E₂ transition, perhaps an E₁-P to E₂-P step like that in the (Na + K)-ATPase reaction sequence. However, oligomycin stimulates phosphatase activity with Mg²⁺ plus Na⁺ alone or Mg²⁺ plus Na⁺ plus low K⁺: this effect of oligomycin may reflect acceleration, in the absence of adequate K⁺, of an alternative E₂-P to E₁ pathway bypassing the monovalent cation-activated steps in the hydrolytic sequence.     

Potassium transport in red blood cells of frog Rana temporaria: demonstration of a K−Cl cotransport

Pathways of K⁺ movement across the erythrocyte membrane of frog Rana temporaria were studied using ⁸⁶Rb as a tracer. The K⁺ influx was significantly blocked by 0.1 mmol·l^-1 ouabain (by 30%) and 1 mmol·l^-1 furosemide (by 56%) in the red cells incubated in saline at physiological K⁺ concentration (2.7 mmol·l^-1). Ouabain and furosemide had an additive effect on K⁺ transport in frog red cells. The ouabain-sensitive and furosemide-sensitive components of K⁺ influx saturated as f(K⁺)_e with apparent K _m values for external K _e ⁺ concentration of 0.96±0.11 and 4.6±0.5 mmol·l^-1 and V _max of 0.89±0.04 and 2.8±0.4 mmol·l cells^-1·h^-1, respectively. The residual ouabain-furosemide-resistant component was also a saturable function of K _e ⁺ medium concentration. Total K⁺ influx was significantly reduced when frog erythrocytes were incubated in NO _- ³ medium. Furosemide did not affect K⁺ transport in frog red cells in NO ₃ ^- media. At the same K _e ⁺ concentration the ouabain-furosemide-insensitive K⁺ influx in Cl^- medium was significantly greater than that in NO _- ³ medium. We found no inhibitory effect of 1 mmol·l^-1 furosemide on Na⁺ influx in frog red cells in Cl^- medium. K⁺ loss from the frog erythrocytes in a K⁺-free medium was significantly reduced (mean 58%) after replacement of Cl^- with NO _- ³ . Furosemide (0.5 mmol·l^-1) did not produce any significant reduction in the K⁺ loss in both media. The Cl^--dependent component of K⁺ loss from frog red cells was 5.7±1.2 mmol·l^-1·h^-1. These results indicate that about two-thirds of the total K⁺ influx in frog erythrocytes is mediated by a K–Cl cotransport which is only partially blocked by furosemide.Abbreviations DMSO dimethyl sulphoxide - K _e ⁺ external concentration of K⁺ - K _m apparent Michaelis constant for external - K⁺ K _e ⁺ at V _max/2 - RBC red blood cell(s) - V _max maximal velocity of the unidirectional K⁺ influx - TRIS tris(hydroxymethyl)aminomethane     

Characterization of conformational changes in (Na,K) ATPase labeled with fluorescein at the active site

Conformational changes have been studied in (Na,K) ATPase labeled at or near the ATP binding region with fluorescein following incubation with fluorescein isothiocyanate (FITC). One or two fluorescein groups are bound per ATPase molecule. (Na,K) ATPase activity, phosphorylation from ATP, and nucleotide binding are abolished in labeled enzyme, but phosphorylation from inorganic phosphate or K-phosphatase activity are only partially inactivated. The fluorescein groups are incorporated only into the 96 KD catalytic chain of the (Na,K) ATPase, and presence of ATP during the incubation with FITC protects against the incorporation and inhibition of enzymic activity. Upon trypsin treatment of labeled membranes the fluorescein appears first in a 58 KD fragment and eventually is released into the medium. The fluorescein-labeled (Na,K) ATPase shows a large quenching of fluorescence (15–20%) on conversion of the E₁ or E₁ · Na conformation in cation-free or Na⁺-rich media to the E₂ · (K) form in K⁺ (or congeners Tl⁺, Rb⁺, Cs⁺, NH ₄ ⁺ ) rich media. Cation titrations suggest that K⁺ and Na⁺ ions compete at a single binding site and stabilize E₁ · Na or E₂ · (K) respectively;K _K0.23 mM,K _Na1.2 mM. The rate of the conformational transition E₂ · (K) E₁ · Na is slow,k=0.3 sec^–1, but contrary to previous experience [7, 8] ATP does not stimulate this rate. The rate of the transitions E₁ + K⁺ E₂ · (K) rises sharply with K⁺ concentration and shows saturation behavior, from which ak _max286 sec^–1 andK _k74 mM are deduced. The data support and extend the previous suggestion that K⁺ ions bound initially at a low-affinity (probably cytoplasm oriented) site in state E₁ are trapped in the occluded form E₂ · (K) by the conformational change poised far (K _c1000) in the direction of E₂ · (K). It is proposed in addition that at least two binding sites for K⁺ exist at the cytoplasmic surface of isolated (Na,K) ATPase in state E₁ but a large difference in affinities precludes detection in fluorescence titrations of more than one site. A variety of ligands in addition to K⁺ produce fluorescence-quenched or E₂ forms of the labeled (Na,K) ATPase. These include Mg²⁺ plus inorganic phosphate, without or with K⁺ ions (E₂P or E₂P · K) or with ouabain (E₂-ouabain or E₂P · ouabain). Na⁺ ions antagonize these effects. The collected data support the notion that there may be many subspecies of the E₁ and E₂ forms (either phosphorylated or nonphosphorylated) with different numbers of Na⁺ and/or K⁺ ions bound or occluded, each subspecies having a characteristic ability to catalyze reactions and/or transport cations. The relationship between the conformational changes in fluorescein-labeled enzyme and the subunit structure of the (Na,K) ATPase is discussed with particular reference to half of the site models for ATP hydrolysis.     

Fractional contribution of major ions to the membrane potential of Drosophila melanogaster oocytes

In ovarian follicles of Drosophila melanogaster, ion substitution experiments revealed that K⁺ is the greatest contributor (68%) in setting oocyte steady‐state potential (E_m), while Mg²⁺ and a metabolic component account for the rest. Because of the intense use made of Drosophila ovarian follicles in many lines of research, it is important to know how changes in the surrounding medium, particularly in major diffusible ions, may affect the physiology of the cells. The contributions made to the Drosophila oocyte membrane potential (E_m) by [Na⁺]_o, [K⁺]_o, [Mg²⁺]_o, [Ca²⁺]_o, [Cl^?]_o, and pH (protons) were determined by substitutions made to the composition of the incubation medium. Only K⁺ and Mg²⁺ were found to participate in setting the level of E_m. In follicles subjected to changes in external pH from the normal 7.3 to either pH 6 or pH 8, E_m changed rapidly by about 6 mV, but within 8 min had returned to the original E_m. Approximately half of all follicles exposed to reduced [Cl^?]_o showed no change in E_m, and these all had input resistances of 330 kΩ or greater. The remaining follicles had smaller input resistances, and these first depolarized by about 5 mV. Over several minutes, their input resistances increased and they repolarized to a value more electronegative than their value prior to reduction in [Cl^?]_o. Together, K⁺ and Mg²⁺ accounted for up to 87% of measured steady‐state potential. Treatment with sodium azide, ammonium vanadate, or chilling revealed a metabolically driven component that could account for the remaining 13%. © 2009 Wiley Periodicals, Inc.     

Beta-adrenergic receptors and adenylate cyclase activity in heart, muscle and adipose tissue of German Landrace pigs selectively bred for differences in backfat deposition

The density of beta-adrenergic receptors was examined in heart, muscle (m. longissimus dorsi) and backfat tissue of German Landrace pigs using (-)-[3H]-dihydroalprenolol. The animals were selected for high (E(+)-line) or low (E(-)-line) activity of lipogenic enzymes in backfat and for low backfat thickness (U(-)-line) based on ultrasonic measurement. An unselected control (K0-line) also exists. The correlated selection response on stress susceptibility (measured by halothane reaction and by meat quality parameters) showed the following sequence of selection lines: U- greater than E- greater than E+ greater than K0. The strains used for examination descended from the 9th generation (heart and muscle tissue) and the 10th generation (heart, muscle and adipose tissue) of this selection experiment. No further selection took place between the 9th and 10th generation. Basal adenylate cyclase activity was determined as well as (-)-Isoproterenol + GTP or sodium fluoride (NaF) stimulated activity in heart and muscle of the 10th generation measuring [3H]-cAMP formation from [3H]-ATP. A good correlation (r greater than or equal to 0.59, n = 9) between beta-adrenergic receptor density and adenylate cyclase activity was found which pointed to an efficient receptor-effector system. The number of beta-adrenergic receptors and adenylate cyclase activity was higher in the U(-)-line in all investigated tissues compared to the other lines in the 9th and 10th generation. Most differences between these lines were also significant.(ABSTRACT TRUNCATED AT 250 WORDS)     

Heat-induced hormesis in longevity as correlated response to thermal-stress selection in Drosophila buzzatii

Hormesis in longevity is a widespread phenomenon across the animal kingdom. It takes place when longevity is improved as an indirect effect of mild stress. We explored some possible evolutionary trajectories of hormesis in longevity in artificially selected lines of Drosophila buzzatii. The lines were bi-directionally selected for either knockdown resistance to heat stress (K⁻, K⁺) or chill-coma recovery (CCR⁻, CCR⁺, with the + and – signs indicating selection for decreased and increased tolerance, respectively). All K and CCR lines successfully diverged due to thermal-stress selection. The heat-inducible hormesis in longevity was substantial in both K⁻ and CCR⁻ females, whereas no hormesis was apparent for females in CCR⁺, K⁺ and control lines. Among-line differences in longevity of non-heat-treated females disappeared after a heat-hardening treatment. Hormesis effects on the demographic senescence rate were sex-specific and consistently higher in the shorter-lived than in the longer-lived lines. Hormesis is an adaptive response, as its magnitude can evolutionary increase with stress-sensitivity.     

Differences between CTP and ATP as substrates for the (Na + K)-ATPase

CTP was a poorer substrate than ATP when substituted in the (Na + K)-ATPase reaction assay, not only in terms of K_m but also of V. CDP was a poorer inhibitor than ADP, so product inhibition cannot account for CTP being a poorer substrate. In the Na-ATPase reaction, which the enzyme also catalyzes, substituting CTP for ATP resulted in greater activity, arguing against CTP being less effective than ATP in forming the enzyme-phosphate intermediate common to both reactions. Ligands that favor the E₂ conformational state of the enzyme, K⁺, Mg²⁺, and Mn₂⁺, inhibited the (Na + K)-CTPase reaction more than the (Na + K)-ATPase. Conversely, Triton X-100, which favors the E₁ conformational state of the enzyme, K⁺, Mg²⁺, and Mn²⁺, inhibited the (Na + K)-CTPase ATPase reaction but stimulated the (Na + K)-CTPase. Although the (Na + K)-ATPase reaction sequence probably involves cyclical interconversion between E₁ and E₂ conformational states (and is thus inhibitable by ligands favoring either state), the K-phosphatase reaction catalyzed by the enzyme apparently functions entirely in the E₂ state. This reaction is better stimulated by CTP plus Na⁺ than by ATP plus Na⁺; moreover, CTP lessens inhibition by Triton X-100, and ATP lessens inhibition by inorganic phosphate (which reacts with the E₂ state). These observations indicate that CTP is a poorer substrate than ATP because it is less effective in promoting conversion of E₂ to E₁, essential for the (Na + K)-dependent reaction mechanism. However, contrary to this rationale, dimethyl sulfoxide stimulated the (Na + K)-CTPase reaction although by other criteria, including inhibition of the (Na + K)-ATPase, the reagent appears to favor the E₂ over the E₁ conformational state.     

Theoretical studies and vibrational spectra of <Emphasis Type="Italic">1H</Emphasis>-indole-3-acetic acid. Exploratory conformational analysis of dimeric species

Theoretical studies on 1H-indole-3-acetic acid (IAA) were performed to investigate the conformational properties of dimeric species and vibrational spectra. Experimental infrared spectra at 100 K and 297 K and Raman spectrum at 297 K were analyzed and compared against calculations performed at B3LYP/6-31G** level. A exploratory study of the conformational space of dimeric species was performed. Our analysis showed that dimeric forms predicted theoretically contribute distinctively to the assignments of experimental results. These structures are defined by the orientation of the acetyl moieties with respect to the plane of indole ring. The dimers are formed by two symmetrical IAA monomers (one of them with the acetyl moiety upward oriented, Re-face, and the other isomer having the acetyl moiety downward oriented, Si-face) in tail-to-tail way. The X-ray geometry and FTIR vibrational frequencies were compared with the results of DFT calculations. A conformational equilibrium involving the non-equivalent IAA dimers: CCT-CCT, A⁺A⁺T-A^-A^-T, A⁺A^-T-A^-A⁺T, and A⁺CT-A^-CT was found. The relation of the conformational properties of the IAA molecule with the features of the vibrational spectra was described in detail. The band assignments were discussed as related to the conformations properties. Our analysis shows the significance of the theoretical study of the conformational space of the monomeric molecule in the rationalization of experimental results.