Crucial role of intracellular effectors on glycogenolysis in the isolated rat heart: Potential consequences on the myocardial tolerance to ischemia

The role played by glycogenolysis in the ischemic heart has been recently put into question because it is suspected that a slowing down of this process could be beneficial for the tolerance of the myocardium to ischemia. The role of the intracellular effectors that control the rate of glycogenolysis has therefore regained interest. We aimed to understand the role played by those intracellular effectors which are directly related to the energy balance of the heart. To this end, we review some of the previously published data on this subject and we present new data obtained from P-31 and C-13 NMR spectroscopic measurement on isolated rat heart. Two conditions of ischemia were studied: 15 min global no-flow and 25 min low-flow ischemia. The hearts were isolated either from control animals or from rats pre-treated with isoproterenol (5 mg.kg^–1 b.w. i.p.) 1 h before the perfusion in order to C-13 label glycogen stores. Our main results are as follows: (1) the biochemically determined glycogenolysis rate during the early phase of ischemia (up to 10–15 min) was larger in no-flow ischemia than in low-flow conditions for both groups, (2) direct measurement of the glycogenolysis rate, as determined by C-13 NMR, after labelling of the glycogen pool in the hearts from isoproterenol-treated rats, confirms the estimations from the biochemical data, (3) glycogenolysis was slower in the hearts from pre-treated animals than in control hearts for both conditions of ischemia, (4) the total activity of glycogen phosphorylase (a + b) increased, by 50%, after 5 min no-flow ischemia, whereas it decreased by 42% after the same time of low-flow ischemia. However, the ratio phosphorylase a/a + b was not altered, whatever the conditions, (5) the concentration of inorganic phosphate (Pi) increased sharply during the first minutes of ischemia, to values above 8–10 mM, under all conditions studied. The rate of increase was larger during no-flow ischemia than during low-flow ischemia. The concentration of Pi was thereafter higher in controls than in the hearts from isoproterenol-treated animals.The calculated cytosolic concentration of free 5 AMP increased sharply at the onset of ischemia, reaching in a few minutes values above 30 M in controls and significantly lower values, around 15 M, in the hearts from isoproterenol-treated rats. (6) The hearts from isoproterenol-treated rats displayed a reduced intracellular acidosis, when compared to controls, under both conditions of ischemia.We conclude that the intracellular effectors, mainly free AMP, play an essential role in the control of glycogenolysis via allosteric control of phosphorylase b activity. The alteration in the concentration of free Pi, the substrate of both forms of phosphorylase, can also be considered as determinant in the control of the rate of glycogenolysis.The attenuation of ischemia-induced intracellular acidosis in the hearts from isoproterenol-treated rats could be a consequence of a reduced glycogenolytic rate and is likely to be related to a better resumption of the mechanical function on reperfusion.     

Selenium,zinc, and thyroid hormones in healthy subjects

Iodothyronine 5′ deiodinase, which is mainly responsible for peripheral T₃ production, has recently been demonstrated to be a selenium (Se)-containing enzyme. The structure of nuclear thyroid hormone receptors contains Zinc (Zn) ions, crucial for the functional properties of the protein. In the elderly, reduced peripheral conversion of T₄ to T₃ with a lower T₃/T₄ ratio and overt hypothyroidism are frequently observed. We measured serum Se and RBC GSH-Px (as indices of Se status), circulating and RBC Zinc (as indices of Zn status), thyroid hormones and TSH in 109 healthy euthyroid subjects (52 women, 57 men), carefully selected to avoid abnormally low thyroid hormone levels induced by acute or chronic diseases or calorie restriction. The subjects were subdivided into three age groups. To avoid under- or malnutrition conditions, dietary records were obtained for a sample of 24 subjects, randomly selected and representative of the whole population for age and sex. Low T₃/T₄ ratios and reduced Se and RBC GSH-Px activity were observed only in the older group. A highly significant linear correlation between the T₃/T₄ ratio and indices of Se status was observed in the older group of subjects (r=0.54;p<0.002, for Se;r=0.50;p<0.002, for RBC GSH-Px). Indices of Zn status did not correlate with thyroid hormones, but RBC Zn was decreased in older as compared with younger subjects. We concluded that reduced peripheral T₄ conversion is related to impaired Se status in the elderly.     

Does observed fertility maximize fitness among New Mexican men?

Our objective is to test an optimality model of human fertility that specifies the behavioral requirements for fitness maximization in order (a) to determine whether current behavior does maximize fitness and, if not, (b) to use the specific nature of the behavioral deviations from fitness maximization towards the development of models of evolved proximate mechanisms that may have maximized fitness in the past but lead to deviations under present conditions. To test the model we use data from a representative sample of 7,107 men living in Albuquerque, New Mexico, between 1990 and 1993. The model we test proposes that low fertility in modern settings maximizes number of grandchildren as a result of a trade-off between parental fertility and next generation fertility. Results do not show the optimization, although the data do reveal a trade-off between parental fertility and offspring education and income. We propose that two characteristics of modern economies have led to a period of sustained fertility reduction and to a corresponding lack of association between income and fertility. The first is the direct link between costs of investment and wage rates due to the forces of supply and demand for labor in competitive economies. The second is the increasing emphasis on cumulative knowledge, skills, and technologies in the production of resources. Together they produce historically novel conditions. These two features of modern economies may interact with evolved psychological and physiological mechanisms governing fertility and parental investment to produce behavior that maximizes the economic productivity of lineages at the expense of fitness. If cognitive processes evolved to track diminishing returns to parental investment and if physiological processes evolved to regulate fertility in response to nutritional state and patterns of breast feeding, we might expect non-adaptive responses when returns from parental investment do not diminish until extremely high levels are reached. With high economic payoffs from parental investment, people have begun to exercise cognitive regulation of fertility through contraception and family planning practices. Those cognitive processes maynot have evolved to handle fitness trade-offs between fertility and parental investment. A preliminary presentation of this data was published in R. I. M. Dunbar, ed.,Human Reproduction Decisions: Biological and Social Perspectives. New York: St. Martin’s Press, 1995. Support for the research project, “Male Fertility and Parenting in New Mexico,” began with two seed grants from the University of New Mexico’s Biomedical Research Grants Program, 1988 and 1989, and one from the University of New Mexico Research Allocations Committee, 1988. Further seed money as well as interim funding came from the William T. Grant Foundation (#89130589 and #91130501). The major support for the project came from the National Science Foundation from 1990 to 1993 (#BNS-9011723 and #DBS-911552). Both National Science Foundation grants included Research Experience for Undergraduates supplements. Hillard S. Kaplan is an Associate Professor of Anthropology at the University of New Mexico. His earlier research and publications focused on food sharing, time allocation, parental investment, and reproductive strategies among Ache hunter-gatherers in Paraguay, Machiguenga and Piro forager-horticulturalists in Peru, and villagers of several ethnicities in Botswana. New research and theory concern fertility, parental investment, and mating strategies in developed and developing nations. This research formulates a new theory of reproductive decision-making and the demographic transition, integrating human capital and parental investment theory in a synthesis of economic and evolutionary approaches. Jane B. Lancaster is a Professor of Anthropology at the University of New Mexico. Her research and publications are on human reproductive biology and behavior, especially human parental investment; women’s reproductive biology of pregnancy, lactation, and child-spacing; and male fertility and investment in children. Current research with Hillard S. Kaplan is on male life history strategies among a large sample of men in New Mexico. She has coedited three books on human parental investment:School-Age Pregnancy and Parenthood (with B. Hamburg),Parenting across the Life Span (with J. Altmann, A. Rossi, and L. Sherrod), andOffspring Abuse and Neglect (with R. Gelles). She is scientific editor of a quarterly journal,Human Nature: An Interdisciplinary, Biosocial Perspective published by Aldine de Gruyter. She is also a council member of the newly formed Human Behavior and Evolution Society. John A. Bock is Andrew W. Mellon Post-Doctoral Fellow in Epidemiology and Population Health at the National Centre for Epidemiology and Population Health, The Australian National University. His research focuses on the allocation of parental investment and the determinants of children’s activities, integrating aspects of economic and evolutionary theory. He has ongoing field research with Bantu and Bushmen agro-pastoralists and forager-horticulturalists in the Okavango Delta, Botswana. He is also collaborating with Lancaster and Kaplan on the determinants of progeny distribution and homosexuality among New Mexican men. Sara E. Johnson is a Ph.D. candidate at the University of New Mexico. Her major research trajectory focuses on trade-offs in life history characters. Her research experience includes participation in a study of variation in growth and development among children in a multi-ethnic community in the Okavango Delta, Botswana, in addition to her dissertation work on individual variation in growth and mortality among juvenile baboons. She is collaborating with Lancaster and Kaplan on the association between survival and fertility among Albuquerque men.     

BET3 encodes a novel hydrophilic protein that acts in conjunction with yeast SNAREs.

Here we report the identification of BET3, a new member of a group of interacting genes whose products have been implicated in the targeting and fusion of endoplasmic reticulum (ER) to Golgi transport vesicles with their acceptor compartment. A temperature-sensitive mutant in bet3-1 was isolated in a synthetic lethal screen designed to identify new genes whose products may interact with BET1, a type II integral membrane protein that is required for ER to Golgi transport. At 37 degrees C, bet3-1 fails to transport invertase, alpha-factor, and carboxypeptidase Y from the ER to the Golgi complex. As a consequence, this mutant accumulates dilated ER and small vesicles. The SNARE complex, a docking/fusion complex, fails to form in this mutant. Furthermore, BET3 encodes an essential 22-kDa hydrophilic protein that is conserved in evolution, which is not a component of this complex. These findings support the hypothesis that Bet3p may act before the assembly of the SNARE complex.     

Phytochrome-mediated effects on extracellular peroxidase activity, lignin content and bending resistance in etiolated Vicia faba epicotyls

Etiolated Vicia faba seedlings were exposed to continuous red light to investigate whether changes in extracellular peroxidase activity were correlated in time and localization with changes in extension growth and/or lignin content in the subapical region of the epicotyl. Continuous red light: (a) increased extracellular peroxidase activity after a lag of ca 0.5 h, followed by a maximum peak after 2.5 h due to slightly acidic isoforms (pI = 6–6.5, according to isoelectrofocusing gels), a minimum after 4 h and a second maximum after 8 h due to acidic isoforms (pI=4–5), (b) increased lignin content and epicotyl resistance to bending after a lag of ca 4 h, i.e. simultaneously with changes in acidic extracellular peroxidase activity, and (c) reduced extension growth to a stable rate after a lag of ca 1 h, not coinciding with the kinetics of any of the extracellular peroxidase isoforms. These effects of continuous red light were at least partially mediated by phytochrome. Tissue printing and anatomical studies revealed red light effects on extracellular peroxidase activity and lignin content mainly in the outer cortical parenchyma. The results are consistent with the involvement of phyto-chrome-mediated effects on extracellular peroxidases (acidic isoforms) in the transduction chain leading to lignin responses to red light.     

Identification of Tuber magnatum Pico DNA markers by RAPD analysis

Summary Different species of truffle were studied in order to identify species-specific markers. The isolation of two Tuber magnatum Pico markers is reported. One of these could be used as a probe in dot blot hybridization, allowing the development of a rapid test able to identify Tuber magnatum species.     

The making of the minibody: An engineered β-protein for the display of conformationally constrained peptides

Conformationally constraining selectable peptides onto a suitable scaffold that enables their conformation to be predicted or readily determined by experimental techniques would considerably boost drug discovery process by reducing the gap between the discovery of a peptide lead and the design of a peptidomimetic with a more desirable pharmacological profile. With this in mind, we designed the minibody, a 61-residue β-protein aimed at retaining some desirable features of immunogloblin variable domains, such as tolerance to sequence variability in selected regions of the protein and predictability of main chain conformation of the same regions, based on the ‘canonical structures’ model. To test the ability of the minibody scaffold to support functional sites we also designed a metal binding version of the protein by suitably choosing the sequences of its loops. The minibody was produced both by chemical syntyhesis and expression in E. coli and charactgerized by size exclusion chromatography, UV CD (circular dichroism) spectroscopy and metal binding activity. All our data supported the model, but a more detailed structural characterization of the molecule was impaired by its low soubility. We were able to overcome this problem both by further; mutagenesis of the framework and by addition of a solublizing motif. The minibody is being used to select constrained human IL-6 peptidic ligands from a library displayed on the surface of the f1 bacteriophage.