The redox state of free nicotinamide–adenine dinucleotide phosphate in the cytoplasm of rat liver

1. The concentrations of the oxidized and reduced substrates of the ;malic' enzyme (EC 1.1.1.40) and isocitrate dehydrogenase (EC 1.1.1.42) were measured in freeze-clamped rat livers. By assuming that the reactants of these dehydrogenase systems are at equilibrium in the cytoplasm the [free NADP(+)]/[free NADPH] ratio was calculated. The justification of the assumption is discussed. 2. The values of this ratio obtained under different nutritional conditions (well-fed, 48hr.-starved, fed with a low-carbohydrate diet, fed with a high-sucrose diet) were all of the same order of magnitude although characteristic changes occurred on varying the diet. The value of the ratio fell on starvation and on feeding with the low-carbohydrate diet and rose slightly on feeding with the high-sucrose diet. 3. The mean values of the ratio were calculated to be between 0.001 and 0.015, which is about 100000 times lower than the values of the cytoplasmic [free NAD(+)]/[free NADH] ratio. 4. The differences in the redox state of the two nicotinamide-adenine dinucleotide couples can be explained on a simple physicochemical basis. The differences are the result of equilibria that are determined by the equilibrium constants of a number of highly active readily reversible dehydrogenases and transaminases and the concentrations of the substrates and products of these enzymes. 5. The decisive feature is the fact that the NAD and NADP couples share substrates. This sharing provides a link between the redox states of the two couples. 6. The application of the method of calculation to data published by Kraupp, Adler-Kastner, Niessner & Plank (1967), Goldberg, Passonneau & Lowry (1966) and Kauffman, Brown, Passonneau & Lowry (1968) shows that the redox states of the NAD and NADP couples in cardiac-muscle cytoplasm and in mouse-brain cytoplasm are of the same order as those in rat liver. 7. The determination of the equilibrium constant at 38 degrees , pH7.0 and I 0.25 (required for the calculation of the [free NADP(+)]/[free NADPH] ratio), gave a value of 3.44x10(-2)m for the ;malic' enzyme (with CO(2) rather than HCO(3) (-) as the reactant) and a value of 1.98x10(-2)m(-1) for glutathione reductase.     

Size-specific social interactions and foraging styles in a shallow water population of mutton snapper,Lutjanus analis (Pisces: Lutjanidae), in the central Bahamas

Synopsis Field observations quantified the effects of fish size and time of day on activity patterns, intraspecific encounters, and foraging styles in mutton snapper,Lutjanus analis, during the spring and winter of 1991. Fish ranged in size from 15 to 65 cm fork length (FL), and were associated with an artificial patch reef system located on a shallow seagrass meadow in the Exuma Cays, Bahamas. The most common, non-resting diurnal activities were intraspecific chasing and displacing, and feeding. Intraspecific displacing was significantly higher during midday compared to morning or evening. The highest proportion of intraspecific encounters (combined chasing and displacing events) occurred among medium (25–35 cm FL) and large (> 35 cm FL) fish. The few large fish observed (13% of population) initiated the same proportion of encounters as the predominant (50%) medium fish. The remaining (37%) small fish (> 25 cm FL) were the least aggressive. Dark barred and dark nape color patterns were associated with displacing and chasing, respectively. Fish exhibited considerable variability in feeding behavior, Proportionally fewer fish fed during midday compared to morning or evening, although small fish fed proportionally more often than medium or large fish despite time of day. Picking was the primary feeding mode and was observed during all times of day. Winnowing was observed during midday and evening, whereas midwater strikes were confined to morning and evening. Small fish displayed proportionally higher picking and midwater strikes during morning and evening, respectively, compared to medium or large fish. However, large fish winnowed proportionally more often than small or medium fish during evening. Dark barred color patterns were associated with feeding on the substrate, whereas no color changes occurred during midwater strikes. Our results indicate thatL. analis forms dominance hierarchies and that high variability in foraging styles, according to fish size and time of day, may be a means to reduce intraspecific competition.     

Germ-Line and Somatic Recombination Induced by in Vitro Modified P Elements in Drosophila Melanogaster

The P element insertion Δ2-3(99B) has previously been shown to activate incomplete P elements elsewhere in the genome. We show that this element, in conjunction with a second incomplete P element, P[CaSpeR], also induces recombination in the male germ line. The recombination is induced preferentially in the region of the P[CaSpeR] element. Recombinant chromosomes contain the P[CaSpeR] element in more than 50% of cases, and alternative models of transposon replication and preferential chromosome breakage are put forward to explain this finding. As is the case with male recombination induced by P-M dysgenic crosses, recombination appears to be premeiotic in a high proportion of cases. The Δ2-3(99B) element is known to act in somatic cells. Correspondingly, we show that the Δ2-3(99B)-P[CaSpeR] combination elevates the incidence of somatic recombination.     

Establishment and maintenance of nuclear position during zoospore formation in allomyces macrogynus: roles of the cytoskeleton

The involvement of the microtubule (MT) and actin microfilament (MF) cytoskeletons in establishing nuclear positions during zoosporogenesis in Allomyces macrogynus was assessed using selective cytoskeletal disrupting treatments and documented with light microscopy. These experiments were coupled with low-speed centrifugation studies to determine the degree to which cytoskeletal elements anchor nuclear position. At the onset of zoospore formation, nuclei were positioned only in cortical cytoplasmic regions of the zoosporangia (ZS). Immunofluorescence microscopy revealed that MTs primarily emanated from centrosomal regions into the surrounding cytoplasm at this stage. During delimitation of the cytoplasm into individual uninucleate zoospores, nuclei migrated from cortical regions to become distributed throughout the cytoplasm. Coincident with nuclear migrations, MTs were primarily organized at and emanated from nuclear surfaces, forming extensive perinuclear arrays. Nuclear migrations were suppressed in ZS induced to sporulate in the presence of cytochalasin D, an actin MF inhibiting compound. Disruption of MTs with nocodazole did not block nuclear migrations, although resultant nuclear spacing was irregular. Centrifugation treatments of control and drug-treated ZS demonstrated that nuclear positions were stabilized by perinuclear MT arrays. The results indicate that nuclear motility in ZS of A. macrogynus is the result of an actin-based system while perinuclear MTs arrays function to establish and fix nuclear position during zoospore formation. Copyright 1998 Academic Press.     

Quantitative determination of 5-aminolaevulinic acid and its esters in cell lysates by HPLC-fluorescence

The development of a reliable sensitive method for the HPLC determination of 5-aminolaevulinic acid (ALA) and ALA esters in cell lysates is described. The method relies on the quantification of a fluorescent derivative of ALA following its derivatisation with acetylacetone and formaldehyde. Following this procedure it is possible to quantify ALA in cell lysates with no need for pre-purification of the sample. The method has been validated in two ranges of concentration (0.6-65 microM, 0.1-10 microg/mL, and 30-600 microM, 5-100 microg/mL), and has also been extended and validated for the determination of ALA released from ALA prodrugs after acidic hydrolysis.     

Analysis of genetic variation in 28 dog breed populations with 100 microsatellite markers

Dog breeds were created by man choosing for select phenotypic traits such as size, shape, coat color, conformation, and behavior. Rigorous phenotypic selection likely resulted in a loss of genetic information. The present study extends previous dog population observations by assessing the genotypic variation within and across 28 breeds representing the seven recognized breed groups of the American Kennel Club (AKC). One hundred autosomal microsatellite markers distributed across the canine genome were used to examine variation within breeds. Resulting breed-specific allele frequencies were then used in an attempt to elucidate phylogeny and genetic distances between breeds. While the set of autosomal microsatellites was useful in describing genetic variation within breeds, establishing the genetic relatedness between breeds was less conclusive. A more accurate determination of breed phylogeny will likely require the use of single-nucleotide polymorphisms (SNPs).     

PCR multiplexed microsatellite panels to expedite canine genetic disease linkage analysis

Modern dog breeds possess large numbers of genetic diseases for which there are currently few candidate genes or diagnostic tests. Linkage of a microsatellite marker to a disease phenotype is often the only available tool to aid in the development of screening tests for disease carriers. Detection of linkage to a specific disease phenotype requires screening of large numbers of markers across known affected and unaffected animals. To establish high throughput genome scanning this study placed 100 canine microsatellite markers, arranged by fragment size and fluorescent dye label, into 12 PCR multiplexed panels. The highest degree of multiplexing was 11 markers per panel while the lowest was five markers per panel; each panel was run in one gel lane on automated DNA sequencers. Selection of the markers was based upon chromosomal or linkage group locations, degree of polymorphism, PCR multiplex compatibility and ease of interpretation. The marker set has an average spacing of 22.25 centiMorgan (cM). Marker polymorphism was evaluated across 28 American Kennel Club (AKC) recognized breeds. The utility of buccal swab vs. blood samples was also validated in this study as all template DNA was derived from swabs obtained and submitted by participating dog breeders and owners. The PCR multiplexed microsatellite panels and sampling method described in this report will provide investigators with a cost effective and expedient means of pursuing linkage studies of specific canine genetic diseases.     

A genomewide scan for early-onset coronary artery disease in 438 families: the GENECARD Study

A family history of coronary artery disease (CAD), especially when the disease occurs at a young age, is a potent risk factor for CAD. DNA collection in families in which two or more siblings are affected at an early age allows identification of genetic factors for CAD by linkage analysis. We performed a genomewide scan in 1,168 individuals from 438 families, including 493 affected sibling pairs with documented onset of CAD before 51 years of age in men and before 56 years of age in women. We prospectively defined three phenotypic subsets of families: (1) acute coronary syndrome in two or more siblings; (2) absence of type 2 diabetes in all affected siblings; and (3) atherogenic dyslipidemia in any one sibling. Genotypes were analyzed for 395 microsatellite markers. Regions were defined as providing evidence for linkage if they provided parametric two-point LOD scores >1.5, together with nonparametric multipoint LOD scores >1.0. Regions on chromosomes 3q13 (multipoint LOD = 3.3; empirical P value <.001) and 5q31 (multipoint LOD = 1.4; empirical P value <.081) met these criteria in the entire data set, and regions on chromosomes 1q25, 3q13, 7p14, and 19p13 met these criteria in one or more of the subsets. Two regions, 3q13 and 1q25, met the criteria for genomewide significance. We have identified a region on chromosome 3q13 that is linked to early-onset CAD, as well as additional regions of interest that will require further analysis. These data provide initial areas of the human genome where further investigation may reveal susceptibility genes for early-onset CAD.     

Unintended pregnancy and women's psychological well-being in Indonesia

Few studies have examined the impact of unintended pregnancy on women in developing countries. This paper examines the impact of unintended pregnancy on Indonesian women's psychological well-being. It is hypothesized that experiencing unintended pregnancy is associated with lower psychological well-being and that use of family planning and small family size are associated with higher levels of psychological well-being. Data are drawn from a 1996 survey of 796 women aged 15-49 from two Indonesian provinces, Lampung and South Sumatra. This article focuses on the 71% of women (n=562) who answered all 41 survey items related to psychological well-being. In cluster analysis, women grouped into three clusters, differentiated by their scores on four scales of well-being established through factor analysis (general negative feelings, satisfaction with relationships, satisfaction with economic/family/personal conditions, and negative feelings regarding domestic issues). Women in cluster 3 were characterized mainly by their high level of psychological well-being. Women in cluster 1 had the lowest level of well-being, and women in cluster 2 were in the middle. Multinomial logistic regression was used to assess jointly the effect of unintended pregnancy, contraceptive use, number of children and other factors on a woman's level of psychological well-being. Unintended pregnancy was associated with lower levels of psychological well-being and contraceptive use was associated with higher levels of psychological well-being, while number of children was not associated with level of well-being. Women who had experienced an unintended pregnancy were less likely to be in the high psychosocial well-being cluster versus both the medium and low clusters. In addition, women using contraception were more likely to be classified in the high than in the low or medium well-being clusters.