Interactive effects of season and temperature on enzyme activities,tissue and whole animal respiration in roach,Rutilus rutilus

Synopsis This paper reviews investigations on the ecophysiology of a population of roach, Rutilus rutilus, from a subalpine oligotrophic lake in the Austrian Tirol. Metabolic responses to season and temperature were studied in whole animals, tissues and selected enzymes. The exponent of the relationship between body mass and three levels of the metabolic rate of acclimated fish was 0.82 ± 0.02, 0.60 ± 0.15, and 0.75 ± 0.01 at 4, 12, and 20° C respectively. Various combinations of long-term acclimation to constant or seasonally fluctuating temperatures and long-term (up to 14 days) monitoring of O₂ at the acclimation temperature led to the conclusion that the aerobic power of fish swimming in the routine mode does not show any sign of being temperature compensated. On the other hand, there are several indications that the energy expenditure of spontaneously swimming fish is adjusted to the seasonal pattern of environmental change and that these responses of metabolism and behaviour are controlled by both endogenous and exogenous factors. The rate of oxygen consumption of gill and muscle tissue brei from fish caught during a seasonal cycle and measured at 15° C appears to follow closely the reproductive and gonadal cycle of the living fish. The same holds for the activities of phosphofructokinase, acetoacetyl-CoA thiolase, and cytochrome oxidase. On the other hand, the Na⁺, K⁺-ATPase of the kidney shows near perfect temperature compensation when fish acclimated to 5 and 25° C are compared, whereas an equally pronounced case of inverse temperature acclimation has been reported for the activity of digestive enzymes in the gut. Summarizing these data it is pointed out that the temperature relationship of a poikilothermic organism is the sum of often very diverse temperature relationships of specific metabolic and behavioural functions. In the case of the roach, strong effects of acclimation temperature on the molecular level, sometimes in the opposite direction, combine with seasonal effects on enzyme activities and tissue respiration. However, on the whole animal level the fish behave as strictly non-compensating poikilotherms, the reproductive cycle being the only detectable influence capable of modulating the basic temperature relationship of energy expenditure.     

Growth of swimming muscles and its metabolic cost in larvae of whitefish at different temperatures

The temperature relationship of routine metabolic rate (R_r) of non-feeding, non-growing Coregonus lavaretus larvae between 2 and 15°C is characterized by Q₁₀-values ranging from l.8-2.45. The rate of growth, based on weight determinations, of first-feeding larvae amounted to 3.5, 7.6 and 9.4% day-¹ at 5, 10 and 12°C respectively, from which Q₁₀-values between 4.0 and 4.8 can be calculated. The rate of increase of muscle mass between 5 and 10°C, based on the determination of the cross-sectional area of inner muscle fibres, resulted in a Q₁₀-value of 4.5. Water temperature influenced the pattern of growth of the inner muscle fibres. At hatching, after 360 day degrees, total muscle mass of larvae reared at 4 and 8°C was independent of temperature, but at 4°C the rate of mass increase owed more to hyperplasia (increase in fibre number) than to hypertrophy (increase in fibre mass), whereas at 8°C the opposite was the case. The calculation of power budgets (including the metabolic cost of growth) of first-feeding larvae yielded net conversion efficiencies (K₂) increasing with temperature from 46.3% at 5°C to 54.7% at 12°C. Comparing our data with literature data two general conclusions can be drawn. (1) In first-feeding larvae the net, but not the gross, conversion efficiency of food energy increases with temperature. This is due to net energy input being characterized by a much higher Q₁₀-value than energy expenditures. (2) In embryos of freshwater fish so far investigated hyperplasia plays a greater role in the increase of fibre mass than hypertrophy at the lower temperature, whereas in embryos of marine fish hyperplasia prevails at the higher temperature. It is suggested that this discrepancy correlates with the high concentration of free amino acids in the eggs of marine species which provide an additional, easily available, source of metabolic energy absent in freshwater species.     

Identification of plasma membrane glycoproteins involved in the contact-dependent inhibition of growth of diploid human fibroblasts

Growth of normal, nontransformed cells is regulated by the interplay between growth stimulating compounds and growth inhibiting cell-cell contacts. We have previously shown that the growth of normal diploid human fibroblasts is mainly regulated by a specific class of plasma membrane glycoproteins (R. J. Wieser and F. Oesch (1986) J. Cell Biol. 103, 361-367). Because it was found that immobilization of the glycoproteins involved in contact-dependent inhibition of growth is an essential step in the recovery of the biological activity of the glycoproteins, we developed a technique for a first characterization of the active compounds. After SDS-PAGE separation of plasma membrane glycoproteins, they were transferred onto nitrocellulose. The nitrocellulose was cut along the separation track into circles which fit into wells of a 96-well microtiter plate. Culturing human diploid fibroblasts on the nitrocellulose circles resulted in characteristic growth patterns, which were dependent upon the source and the treatment of the plasma membrane proteins which had been separated. Five major inhibitory fractions with apparent molecular masses of 300, 170, 90, 50, and 25 kDa have been identified in plasma membranes from confluent fibroblast cultures.     

Anaerobic and aerobic energy production of young rainbow trout (Salmo gairdneri) during and after bursts of activity

Journal of Comparative Physiology B -     

Plasma membrane glycoproteins covalently bound to silica beads as a model for molecular studies of cell-cell interactions in culture

In previous studies, we have shown that plasma membrane glycoproteins are of major importance in the density-dependent regulation of growth of normal diploid fibroblasts. Due to the hydrophobic portions of these molecules, functional studies in cell culture are often difficult to perform and to interpret. Specifically, the addition of these molecules in soluble form to cell culture, after depletion of detergents needed for their solubilization, leads to aggregation and internalization. Therefore, we developed a method for the covalent immobilization of the solubilized plasma membrane proteins to derivatized silica beads for further investigations on the molecular nature of the active molecules. The addition of immobilized plasma membrane glycoproteins to sparsely seeded human fibroblasts resulted in cellular reactions similar to those found in confluent cell cultures (strongly reduced cell proliferation; high collagen type III synthesis). The method consists in the derivatization of silica beads (Lichrosphere Si 500, 10 microns) with isothiocyanatopropyltriethoxysilane. Amino-groups react with the SCN group under physiological conditions, resulting in a stable linkage of amino-group bearing molecules with the silica beads. Due to the easy handling of the silica beads (e.g. washing by short centrifugation steps), the mild coupling conditions, and the stable bondings this system is highly suited for functional studies of molecules involved in cell-cell interactions.     

Contact inhibition of growth of human diploid fibroblasts by immobilized plasma membrane glycoproteins

The human embryonic fibroblasts used in this study show pronounced inhibition of growth when reaching a critical cell density. High cell density and growth inhibition has previously been mimicked by the addition of glutaraldehyde-fixed cells or of isolated plasma membranes to sparsely seeded proliferating fibroblasts (Wieser, R. J., R. Heck, and F. Oesch, 1985, Exp. Cell Res., 158:493-499). In this report, we describe the successful solubilization of the growth-inhibiting glycoproteins and their covalent coupling to silicabeads (10 microns), which had been derivatized with 3-isothiocyanatopropyltriethoxysilane. The beads, bearing the plasma membrane proteins, were added to sparsely seeded, actively proliferating fibroblasts, and growth was measured by the determination of cell number or of incorporation of [3H]thymidine into DNA. The growth was inhibited in a concentration-dependent manner, whereby 50% inhibition was achieved with 0.3 micrograms of immobilized protein added to 5 X 10(3) cells. Terminal galactose residues of plasma membrane glycoproteins with N-glycosydically bound carbohydrates were responsible for the inhibition of growth. Dense cultures of human fibroblasts are characterized by an accelerated synthesis of procollagen type III. We have found that this cellular response can also be induced by the addition of immobilized plasma membrane glycoproteins to sparsely seeded cells. These observations support the conclusion that the addition of immobilized plasma membrane glycoproteins to sparsely seeded fibroblasts mimics the situation occurring at high cell density. These results show that cell-cell contacts via plasma membrane glycoproteins carrying terminal galactose residues are important for the regulation of the proliferation of cultured human fibroblasts and presumably of the accelerated synthesis of collagen type III.     

Sap flow measurements as a basis for assessing trace-gas exchange of trees

The assessment of trace-gas uptake by plants is of basic interest in plant ecophysiology and atmospheric chemistry. For tall vegetation and extensive canopies micrometeorological methods and modelling of deposition or combinations of both are usually the methods of choice. However, distinguishing between the aerodynamically driven components of deposition and stomatal uptake is difficult and estimates of plant uptake remain uncertain. Canopy conductance derived from sapflow measurements of trees represents an important and highly variable component to determine the uptake of trace gases by trees under free atmospheric conditions. The theory of the assessment of trace-gas uptake by the sapflow-based approach is reviewed and exemplified for the uptake of ozone, nitrogen oxides, and ammonia into coniferous and deciduous tree species. First results on the stomatal ammonia compensation point (χ_s=0.11–0.27 nmol mol⁻¹) of the coniferous tree species Picea abies determined by a bio-assay are reported and compared with published values on herbaceous plants and gas-exchange approaches for P. abies. For a summer period in 2003, the ground-area scaled uptake rate of gaseous NH₃ by P. abies was more than twice as high (1.38 nmol m⁻² s⁻¹) than the uptake of NO_x (0.53 nmol m⁻² s⁻¹). Estimates of ground-area scaled O₃ uptake and phytomedically relevant O₃ doses of Fagus sylvatica were found to be significantly less under dry conditions in August 2003 (cumulative uptake 2.3 mmol m⁻²) than in years with sufficient soil water supply despite higher atmospheric O₃ concentrations in 2003. Cumulative ground-area scaled O₃ uptake of Pinus cembra reached 150 mmol m⁻² during the growing season at an alpine site. Preliminary results and future perspectives are discussed for the transfer of the approach to the uptake of carbon dioxide and hence to determine total net primary production of trees. This novel approach has the potential to reduce uncertainties of C fluxes measured by the eddy-covariance technique and biogeochemical plot studies. It also allows to determine flux components like heterotrophic and autotrophic respiration separately as residuals from budget equations. Overall, it is concluded that the sapflow-based methodology contributes a new quality of flux data significantly improving our current understanding of biospheric aspects of trace-gas fluxes into tall vegetation.