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J R Hazel

Publications and source records attributed to J R Hazel.

At least 37 records · Page 2Linked to original sources
PubMed1988

Acylation of lysophosphatidylcholine in liver microsomes of thermally acclimated trout.

The specificity of acyl-coenzyme A (CoA): lysophosphatidylcholine acyltransferase (LPCAT; EC 2.3.1.23) was determined for a range for acyl-CoA substrates differing with respect to chain length and degree of unsaturation in liver microsomes of thermally acclimated (5 and 20 degrees C) rainbow trout, Salmo gairdneri. Absolute levels of oleate incorporation into phosphatidylcholine (PC) were determined at substrate concentrations in the physiological range (12 microM) and higher (64 microM). The specificity of LPCAT was determined by the extent to which competing substrates decreased the incorporation of oleate. LPCAT specificity was significantly influenced by both assay and acclimation temperature at total substrate concentrations of both 72 and 256 microM. A clear preference for 14- and 16-carbon monoenes was exhibited by LPCAT from 20 but not 5 degrees C-acclimated trout. Furthermore, LPCAT from 5 degrees C-acclimated trout preferentially incorporated long chain and polyunsaturated fatty acids (eicosadienoyl, arachidonoyl, erucoyl) and excluded 18-carbon unsaturates at an assay temperature of 5 degrees C compared with 20 degrees C; at 20 degrees C, 18-carbon unsaturates were incorporated more readily than 20-carbon species. Linolenic acid (18:3N3) was generally excluded from incorporation, reflecting a possible mechanism by which this precursor of docasahexaenoic acid (22:6N3, n - 3) remains available for modification. These results indicate that trout liver LPCAT preferentially incorporates fatty acids into PC on the basis of both chain length and degree of unsaturation in a manner consistent with the temperature-induced restructuring of membrane phospholipids.

Acclimatization↗
PubMed1987

A method for the determination of diffusion coefficients for small molecules in aqueous solution.

A method is described for determining the diffusion coefficients of small solutes in limited volumes (approximately equal to 4-9 ml) of fluid. Diffusion is measured in a three-chamber diffusion cell across a central unstirred compartment. Compartments are separated by nitrocellulose membranes. The instantaneous concentration gradient and the instantaneous flux of solute into the dilute end compartment are derived from changes in the concentration of solute in the two stirred end compartments through time. The diffusion coefficient is calculated from the slope of the least-squares regression line relating the magnitude of the instantaneous solute flux to that of the instantaneous concentration gradient. The apparatus is calibrated with a solute of known diffusivity (KCl). Diffusion coefficients thus determined in water at 25 degrees C for CaCl2 (7.54 X 10(-6) cm2.s-1), Na2-ATP (7.01 X 10(-6) cm2.s-1), 2-deoxyglucose (5.31 X 10(-6) cm2.s-1), and D-Na-lactate (5.62 X 10(-6) cm2.s-1) differed by an average of 3.7% from literature values. The method described results in accurate estimates of diffusion coefficients by a simple and relatively rapid procedure.

Calcium Chloride↗
PubMed1987

Temperature affects the diffusion of small molecules through cytosol of fish muscle.

Undiluted cytosolic extracts were prepared from fast glycolytic muscle tissue of white perch (Morone americanus). Diffusion coefficients (D) through the cytosol preparations were estimated in vitro for a series of selected low molecular weight compounds using an experimental diffusion chamber. Determinations were made at 5 degrees and 25 degrees C to assess thermal sensitivity of the process. Non-metabolizable analogues of naturally occurring compounds were employed to avoid chemical alteration of solutes by the catalytically competent preparations during diffusion experiments. Kinematic viscosity of cytosolic extracts, which is a major determinant of diffusive resistance, increases from 2.94 +/- 0.06 to 5.35 +/- 0.02 X 10(-2) cm2 s-1 between temperatures of 25 degrees and 5 degrees C (Q10 = 1.35 +/- 0.01). The diffusion coefficients (D) of D-lactic acid are 2.26 +/- 0.84 and 0.79 +/- 0.15 X 10(-6) cm2s-1 at 25 degrees and 5 degrees C, respectively (Q10 = 1.84 +/- 0.36). The D values of 2-deoxyglucose are 2.87 +/- 1.01 and 1.22 +/- 0.36 X 10(-6) cm2s-1 at 25 degrees and 5 degrees C (Q10 = 1.75 +/- 0.54). The D values of Ca2+ are 2.47 +/- 0.28 and 1.09 +/- 0.36 X 10(-6) cm2s-1 at 25 degrees and 5 degrees C (Q10 = 2.04 +/- 0.36). The D values for the ATP analogue, AMP-PNP, are 0.87 +/- 0.33 and 0.81 +/- 0.15 X 10(-6) cm2s-1 at 25 degrees and 5 degrees C (Q10 = 0.98 +/- 0.12). AMP-PNP is the only compound tested which did not show significant thermal sensitivity of diffusion. Recently reported changes in muscle cell ultrastructure induced by temperature acclimation of fishes may serve to counteract the effect of temperature change on diffusion of key small molecules through the aqueous cytoplasm, thus maintaining flux rates between cellular compartments. These mechanisms may be of considerable import in achieving relative temperature independence of cellular function that is characteristic of many eurythermal aquatic animals.

Adenosine Triphosphate↗
PubMed1985

Partial purification and kinetic characterization of the microsomal phospholipase A2 from thermally acclimated rainbow trout (Salmo gairdneri).

Phospholipase A2 (PLA2) was extracted from liver microsomal membranes of both 5 and 20 degrees C-acclimated rainbow trout (Salmo gairdneri), using the non-ionic detergent, Triton X-100. Further purification was achieved by precipitation with 35-65% ammonium sulfate followed by gel filtration chromatography in the presence of 0.1% Triton X-100 on Sephadex G-200. These procedures resulted in a 30-fold purification and the removal of all traces of phospholipid from the enzyme of both warm- and cold-acclimated trout. Column elution profiles were similar for both acclimation groups, yielding a molecular weight estimate for the trout liver enzyme of 73,000. Comparisons of activity levels and kinetic parameters of PLA2 from warm- and cold-acclimated fish indicated that compensation for temperature at non-saturating substrate concentrations was an attribute of both the particulate (microsomal) enzyme and the lipid-free protein. Cold acclimation resulted in higher activity below Vmax due primarily to decreased apparent Km values. These adaptations to temperature could not be attributed to the interaction of the enzyme with the membrane lipids, but were due to qualitative changes in the enzyme that resulted from acclimation. Other adaptive qualities of PLA2, such as reduced Km in response to acute decreases in temperature in warm-acclimated fish, were only apparent in particulate preparations, and thus were a function of the protein-lipid complex. These data suggest that an acclimation-induced increase in the activity of PLA2 may result in the activation of a deacylation-reacylation cycle at cold temperatures.

Acclimatization↗
PubMed1985

Rapid changes in the phospholipid composition of gill membranes during thermal acclimation of the rainbow trout, Salmo gairdneri.

The phospholipid composition of gill tissue was determined in rainbow trout (Salmo gairdneri) undergoing thermal acclimation between 5 degrees C and 20 degrees C for a period of up to 28 days. Proportions of phosphatidylethanolamine (PE) and cardiolipin (CL) increased during cold acclimation and decreased during warm acclimation; proportions of phosphatidylcholine (PC) changed in the opposite direction (i.e., decreased during cold acclimation). In contrast, levels of phosphatidylserine, -inositol, and sphingomyelin did not vary significantly. Thermal modulation of headgroup composition occurred rapidly as reflected by changes in the ratio of PC-to-PE, which rose significantly from 2.40 +/- 0.09 to 2.92 +/- 0.09 within 72 h of transfer from 5 to 20 degrees C; adaptation to 5 degrees C was equally rapid. Proportions of PE changed more rapidly than those of PC during cold adaptation, whereas the opposite was true during warm acclimation. Both the time course and the direction of the observed changes in phospholipid composition suggest that such adjustments may contribute to the homeoviscous regulation of membrane properties, particularly during the initial stages of thermal adaptation.

Acclimatization↗
PubMed1985

Determination of the phospholipid composition of trout gill by Iatroscan TLC/FID: effect of thermal acclimation.

The phospholipid composition of gill tissue from thermally acclimated rainbow trout, Salmo gairdneri, was determined by Iatroscan analysis following an initial development of the chromarods in a non-polar solvent to remove neutral lipids. Standard curves for all phospholipids, although linear through most of the concentration range tested (1-40 micrograms), extrapolated to negative intercepts on the ordinate, indicating a decline in sensitivity at low phospholipid levels. In addition, the concentration dependence of the Iatroscan response varied by nearly 6-fold among phospholipids. Of the major phospholipids, only lysophosphatidylcholine could not be quantitated accurately because of the presence of an interfering peak. Quantitation by Iatroscan yielded results which, in general, agreed well (within 5%) with results obtained by an independent phosphate analysis. Only in the case of phosphatidylinositol (PI) did the two analytical methods differ significantly; proportions of PI were 55% higher when determined by Iatroscan as opposed to phosphate analysis. Gill tissue from 5 C-acclimated trout possessed higher proportions of phosphatidylethanolamine than tissue from 20 C-acclimated trout. The Iatroscan provided a rapid and reliable means of quantitating the proportions of all the major phospholipids of trout gill, although there are some limitations to the general applicability of the technique.

Acclimatization↗
PubMed1984

Temperature-dependent deacylation of molecular species of phosphatidylcholine by microsomal phospholipase A2 of thermally acclimated rainbow trout, Salmo gairdneri.

Using the ratios of kinetic parameters, V/Km, the deacylation of different molecular species of 1-palmitoyl,2-acyl phosphatidylcholine via microsomal phospholipase A2 (PLA2) was studied in liver tissue of thermally acclimated rainbow trout (Salmo gairdneri). In general, PLA2 from fish acclimated to cold temperatures showed an order of preference for the acyl moieties of 18:1 greater than 18:2 greater than 18:0. Trout acclimated to warm temperatures generally preferred 18:0 PC, but the actual order of preference depended on the temperature of the assays and the presence of endogenous lipids in the enzyme preparation. At 5 C, the particulate (microsomal) enzyme preferred 18:0 greater than 18:2 greater than 18:1, but a lipid-free preparation of the enzyme preferred 18:2 greater than 18:0 greater than 18:1. At 20 C, particulate enzyme preferred 18:1 greater than 18:0 greater than 18:2 but purified enzyme preferred 18:0 greater than 18:2 greater than 18:1. Thus, assay temperature and the presence of microsomal lipids had a greater effect on PLA2 from fish acclimated to warm temperatures than fish acclimated to cold temperatures. The substrate preference of PLA2 is discussed with reference to the previously observed changes in membrane fatty acid composition that occur with thermal acclimation in rainbow trout.

Acclimatization↗
PubMed1984

Effects of temperature on the structure and metabolism of cell membranes in fish.

The metabolic adjustments responsible for the "homeoviscous adaptation" of membrane lipid composition in fish are examined with special reference to the rainbow trout, Salmo gairdneri. The percentage of fatty acid lipogenesis attributable to unsaturates was elevated after an acute drop in temperature but declined with continued cold exposure (i.e., cold acclimation). In contrast, selected desaturation reactions [particularly those involved in the production of polyunsaturated fatty acids (PUFA) of the n-3 and/or n-6 families] proceeded more rapidly in cold-than in warm-acclimated trout. Different time courses for the change in monoene and PUFA levels of hepatic microsomal membranes during thermal acclimation suggest that the various desaturase enzymes contribute to the acclimatory response at different times. Certain fatty acids, particularly the delta 5-desaturation products of the n-3 (20:5 delta 5,8,11,14,17) and n-6 (20:4 delta 5,8,11,14) series, were preferentially incorporated into phospholipids at cold temperatures and by cold-acclimated trout, due in part to the direct effect of temperature on the substrate preferences of the phospho- and acyltransferase enzymes of de novo phospholipid biosynthesis; however, chain length rather than degree of unsaturation per se may determine the temperature-dependent pattern of fatty acid incorporation. Both acute and chronic cold exposure elevated the incorporation of PUFA into phosphatidylserine (PS), suggesting that the conversion of PS to phosphatidylethanolamine (PE) may be activated at cold temperatures. The rate of homeoviscous adaptation appears to be limited by the rate of membrane lipid turnover, which although generally positively correlated with acclimation temperature, did vary depending on the phospholipid moiety and tissue considered. Finally the direct acylation of lysophospholipids formed during the process of membrane turnover may contribute to both rapid and acclimatory adjustments in membrane lipid composition.

Acyl Coenzyme A↗
PubMed1983

The incorporation of unsaturated fatty acids of the n-9, n-6, and n-3 families into individual phospholipids by isolated hepatocytes of thermally-acclimated rainbow trout, Salmo gairdneri.

Rates of incorporation of 1-14C-oleic (18:1n9), -linoleic (18:2n6), and -linolenic (18:3n3) acids into individual phosphatides were determined in isolated hepatocytes from cold (5 degrees C)- and warm (20 degrees C)-acclimated rainbow trout, Salmo gairdneri. Fatty acid incorporation into phosphatidylcholine (PC) exceeded that into all other phospholipids, but at assay and acclimation temperatures of 5 degrees C, incorporation into phosphatidylethanolamine (PE) was generally intermediate between that of PC and the remaining phosphatides. Specific radioactivities (ratios of percentage isotope incorporation-to-mole percentage of phosphatide) were consistently less than one for both PC and PE, and greater than one for phosphatidic acid (PA), lysophosphatidylcholine (LPC), phosphatidylserine (PS), and cardiolipin (CL). For PS, specific radioactivities were greater in cold- than warm-acclimated trout, and greater at 5 degrees C than 20 degrees C. Rates of oleate incorporation were generally higher, and rates of incorporation of 18:2 and 18:3 lower in cold- than warm-acclimated trout. Most phospholipids demonstrated a clear preference for the incorporation of 18:2 when assayed at 20 degrees C; however, at 5 degrees C the incorporation of 18:2 was reduced and 18:3 was generally the preferred substrate. A reduction in assay temperature from 20 degrees C to 5 degrees C also shifted the incorporation of 18:2 away from PC into PS and PA. These data were interpreted to indicate 1) a cold-induced activation of PS metabolism, possibly resulting in elevated levels of PE; 2) lower rates of general acyl group turnover in animals acclimated to 5 degrees C than 20 degrees C; 3) a specificity to the acclimation response that favors the incorporation at cold temperatures of polyunsaturated fatty acids, but not the parent acids from which they are derived; and 4) the participation of a deacylation-reacylation cycle in the metabolism of phospholipids, particularly at cold temperatures.

Adaptation, Physiological↗
PubMed1982

Incorporation of polyunsaturated fatty acids into lipids of rainbow trout hepatocytes.

Hepatocytes from 5- or 20 degrees C-acclimated rainbow trout (Salmo gairdneri) were incubated with [1-14C]oleic, -linoleic, or -linolenic acid. Both acclimation groups demonstrated greater incorporation of derivatives from linolenic and linoleic acids into phospholipids when assayed at 5 and 20 degrees C; few derivatives of oleic acid were formed. Cells from cold-acclimated trout, when assayed at 5 degrees C with linolenic acid, incorporated a large proportion of radioactivity into free fatty acids. Analysis of each lipid fraction revealed a relatively specific incorporation of certain fatty acids. For example, "dead end" elongation products of the three substrates were preferentially incorporated into neutral lipids, while delta 6 desaturation products of the three acids were retained in the free fatty acid fraction. Twenty-carbon acid derivatives of linoleic and linolenic acids were directed into the phospholipid fraction. Incorporation of the delta 5 desaturation products was temperature sensitive in cells from cold-acclimated but not warm-acclimated trout. The results suggest that selectivity of incorporation of specific fatty acids into phospholipids may be of importance in restructuring membranes of poikilotherms during thermal adaptation.

Animals↗
PubMed1979

Fatty acid and sterol synthesis by hepatocytes of thermally acclimated rainbow trout (Salmo gairdneri).

Incorporation of tritium from tritiated water into lipid fractions was measured in isolated hepatocytes from rainbow trout (Salmo gairdneri) acclimated to 5 degrees C and 20 degrees C. Hepatocytes from cold-acclimated trout exhibited significantly higher rates of tritium incorporation into both fatty acid and sterol fractions at assay temperatures of 15 degrees C and 20 degrees C than did hepatocytes from warm-acclimated trout. Tritium incorporation into the fatty acid fraction was nearly temperature independent in hepatocytes from warm-acclimated trout (Q10 = 1.39) but markedly temperature dependent (Q10 = 2.63) in hepatocytes from cold-acclimated trout; in contrast, rates of sterol synthesis were more temperature dependent in warm-acclimated trout. At 5 degrees C, fatty acid lipogenesis comprised a significantly greater percentage of the total tritium incorporation in hepatocytes from warm-acclimated trout and the percentage of total lipogenesis attributable to fatty acids decreased significantly in warm-acclimated trout as the assay temperature increased; the opposite trends were observed in cold-acclimated trout.

Adaptation, Physiological↗
PubMed1979

Influence of thermal acclimation on membrane lipid composition of rainbow trout liver.

Rainbow trout (Salmo gairdneri) acclimated to 5 degrees C possessed larger livers and less neutral lipid per gram of liver than 20 degrees C-acclimated animals; quantities of liver glycolipid, phospholipid, and cholesterol did not vary significantly with acclimation temperature. The relative proportions of phosphatidylethanolamine increased significantly following cold exposure, whereas the quantities of sphingomyelin and cardiolipin declined. For all phosphatides examined (phosphatidylethanolamine, phosphatidylcholine, phosphatidylserine, phosphatidylinositol, lysolecithin, cardiolipin, sphingomyelin) cold acclimation resulted in 1) an increase in the quantity of polyunsaturated fatty acids, 2) a reduction in the level of saturated fatty acids, and 3) little change in the total content of monoenes and dienes. The increased content of polyunsaturated fatty acids in choline and ethanolamine phosphatides following cold acclimation was confined to the 2-position and occurred at the expense of monoenes and dienes. The relative proportions of n - 3 fatty acids, and less frequently n - 6 fatty acids, increased in phosphatides of cold-acclimated trout, whereas the relative proportions of n - 9 fatty acids declined. These data suggest a preferential incorporation of fatty acids belonging to the linolenic acid family at reduced temperatures. Temperature-induced changes in the chemical composition of trout liver phospholipids counteracted the effects of acute temperature change on nonelectrolyte permeability of isolated liposomes.

Adaptation, Physiological↗
PubMed1976

The effects of temperature and thermal acclimation upon the osmotic properties and nonelectrolyte permeability of liver and gill mitochondria from rainbow trout (Salmo gairdneri).

Thermal acclimation of rainbow trout (Salmo gairdneri) taken from 20 degrees C to 7 degrees C resulted in adaptation of mitochondrial function, as evidenced by increases in the specific activities of NADH- and succinate-cytochrome c reductase of 1.93- and 2.7-fold respectively. Mitochondria from both gill and liver obeyed the Boyle-van't Hoff relationship in the range from 400 to 60 mosM. Thermal acclimation had no effect on the osmotic properties of liver mitochondria, whereas gill mitochondria from cold-acclimated trout were more sensitive to osmotic swelling than mitochondria from warm-acclimated individuals. The non-electrolyte permeability of liver mitochondria was assessed by optically monitoring mitochondrial swelling rates in isosmotic solutions of urea, glycerol, mannitol and glucose. Two parameters of mitochondrial swelling were determined: (a) initial swelling rates, d(1/A)dt, and (b) swelling constants, ks, derived from the time required to swell a fixed volume. Regardless of the assay temperature or the permeant employed, liver mitochondria from cold-acclimated trout exhibited greater initial swelling rates than mitochondria from warm-acclimated trout, indicating properties of temperature-compensated permeability. The apparent ranking of nonelectrolyte permeabilities was urea greater than glycerol greater than mannitol greater than glucose. ks values for urea and glycerol from cold-acclimated trout were greater than values typical of warm-acclimated populations; however ks values for glucose and mannitol were not influenced by thermal acclimation. Regardless of the permeant considered, activation energies for ks values were 3- to 5-fold greater than those for initial swelling rates. The time course of mitochondrial swelling consists of two components, an initial rapid swelling phase characterized by a half-life of 3-12 seconds, and a slower swelling phase characterized by a half life of 1-6 minutes. Initial swelling rates, which approximate the rapid swelling component, are considered to be the least ambiguous index of permeability, whereas ks values are more complex and strongly influenced by the slower swelling component.

Acclimatization↗