Integration of natural selection across the life cycle stabilizes a marine mussel hybrid zone.

Hybridization among related species is now recognized as common but it remains unclear how hybrid zones persist for prolonged periods. Here, we test the hypothesis that selection in different components of the life cycle may stabilize a hybrid zone. A hybrid zone occurs in southwest England between the marine mussels and .

Glucose-6-phosphatase flux and the hepatic glucose balance model.

Mice were studied with the euglycemic hyperinsulinemic and the hyperglycemic clamp techniques after a 6-h fast: 1) euglycemic (6.7 +/- 0.2 mM) hyperinsulinemia (approximately 800 microU/ml); 2) hyperglycemic (15.

Dicarboxylic acid fluxes during gluconeogenesis. No channelling of mitochondrial oxalacetate.

A rather complete model of the gluconeogenic pathway was used, with the known separate pools of mitochondrial and cytosolic oxalacetate, malate and aspartate. The fumarase, malate dehydrogenase and glutamate oxalacetate transaminase reactions were assumed to be isotopically actively reversible, but none at isotopic equilibrium. Malate was assumed to exchange actively between the mitochondria and cytosol, while aspartate exchange was more limited, in agreement with the known electrogenic nature of aspartate export from the mitochondria.

Models of the liver pentose cycle.

Simple and complete models of the classical liver pentose cycle, and a model of Williams' proposed "L-type" pentose cycle, are compared. All extant experimental data on well-oxygenated whole cell systems can be fitted to the predicted output of the complete classical pentose cycle model; however, there are gross discrepancies between key experimental data and Williams' proposed scheme. The complete classical model allows isotopic reversibility in the non-oxidative segment of the cycle, but none of the reversible enzymes are extremely close to isotopic equilibrium.

On the estimation of alternative pathways of fatty acid oxidation in the liver in vivo.

The relative contributions of mitochondrial beta-oxidation and peroxisomal beta-oxidation and peroxisomal omega-oxidation to the oxidation of a given fatty acid in vivo can be quantitated by an isotopic method. The approach requires infusion of a fatty acid labelled on two specific carbon atoms (e.g.

Tests of the liver specificity of drug glucuronidation.

Hellerstein and Landau and their coworkers have developed the glucuronide conjugate approach to aid in the analysis of pathways of liver carbohydrate metabolism. This approach requires that the liver is essentially the sole site of glucuronidation of the given drug. Since UDPglucuronyl transferases are present also in other tissues, most notably the kidney and intestines, we need to test the liver specificity of this process.

Isotopic estimation of the hepatic glucose balance in vivo.

Conventional determination of "hepatic glucose production" in the refed state is based on the 30-year-old Steele model, which omits glucose<-->glucose-6P cycling. Using the more complete model, conventional hepatic glucose production is shown to be a complex ratio of fluxes, and not a simple physiological flux. Our new model allows some prospective isotopic approaches to the estimation of glucose-6-phosphatase and glucokinase fluxes, and thus real net hepatic glucose production or uptake.

Estimation of peroxisomal and mitochondrial fatty acid oxidation in rat hepatocytes using tritiated substrates.

The pathways of peroxisomal and mitochondrial fatty acid oxidation were monitored with the use of substrates which produce NAD3H. I used as marker substrates: D-[3-3H]3-hydroxybutyrate for mitochondrial NAD3H production, [2-3H]glycerol for cytosolic NAD3H production, and [2-3H]acetate to measure carbon-bound 3H which was also generated by the metabolism of the commercial 9,10-3H-labelled fatty acids. The assumption that peroxisomal NAD3H can be considered to be equivalent to cytosolic NAD3H was supported using a specific inhibitor of mitochondrial fatty acid oxidation.

Evidence against tight channelling of NADH in hepatocytes.

Tritiated substrates at tracer levels were incubated with rat hepatocytes plus 10 mM L-lactate, and the yields of tritium in glucose and water, as well as the tritium distribution on C-6 and C-4 of glucose, determined. Substrates of cytosolic type A NAD-linked dehydrogenases showed some preferential labeling of C-6 of glucose (the pathway involving type A malate dehydrogenase), whereas substrates of cytosolic type B NAD-linked dehydrogenases showed some preferential labeling of C-4 of glucose (the pathway involving type B glyceraldehyde-3P dehydrogenase). The results found are consistent with a classical diffusion model of NADH metabolism, and are at odds with the Srivastava hypothesis (based on isolated enzyme studies) which indicated that direct transfer of NADH can occur between many NAD-linked enzymes but only when they are of opposite (A or B) specificity.

Effects of salicylate on hepatocyte lactate metabolism.

We have examined the effects of salicylate on fluxes of lactate metabolism in rat hepatocytes using a steady state model. Salicylate produces an uncoupling effect, an inhibition of gluconeogenesis, a marked activation of pyruvate dehydrogenase flux, and an inhibition of endogenous fatty acid oxidation. Agents with known functions such as dinitrophenol and dichloroacetate were also compared in this system.

Errors in the isotopic estimations of hepatic glycogen synthesis and glucose output.

Ignoring the reality of glucose in equilibrium glucose-6P cycling in the liver in vivo produces large potential errors in the isotopic quantitation of percentage contributions of direct and indirect pathways of glycogen synthesis and of hepatic glucose output.

Possible role for carbamyl phosphate in the control of liver glycogen synthesis.

Isotopic data suggested a possible correlation between the activation of liver glycogen synthesis and some unstable intermediate. Carbamyl phosphate was one candidate considered. L-Norvaline, an inhibitor of ornithine transcarbamylase, is known to increase intracellular carbamyl phosphate levels.

Inhibition of glycogen synthesis in rat hepatocytes by medium Zn2+.

In hepatocytes from fasted rats, Zn2+ in the range from 0 to 500 microM has relatively minor effects on gluconeogenesis from most substrates, or on ureagenesis from NH3. In hepatocytes from fed rats, Zn2+ does not affect glycogenolysis. In hepatocytes from fasted rats, in which glycogen is being actively synthesized using the substrate combination (Katz et al.

Gluconeogenesis in rat hepatocytes from monomethyl succinate and other esters.

Although little glucose is formed from succinate in rat hepatocytes, the rate of gluconeogenesis from monomethyl succinate approaches that from L-lactate. Dimethyl succinate is as good as monomethyl succinate at 5 mM, but not at 20 mM. Monoethyl fumarate and 4-methyl malate are only fair glucogenic substrates, but 1-methyl malate is another good substrate at high concentrations.

The role of mitochondrial pyruvate transport in the control of lactate gluconeogenesis.

At low L-lactate concentrations, plots of the reciprocal of the gluconeogenic rate vs hydroxycyanocinnamate concentration were linear up to high inhibitor concentrations, indicating that the pyruvate transporter was a rate-limiting step and alternate pathways were at best minor. At 10 mM L-lactate and in the absence of added acids, the 1/V vs I plots became sigmoidal, indicating both some excess capacity of the transporter, and a significant alternative pathway. Use of transaminase inhibitors suggests that the alternate pathway does not primarily involve a dual glutamate-pyruvate transaminase mechanism.

Further evidence for the classical pentose phosphate cycle in the liver.

Isolated rat hepatocytes were incubated with [3-(14)C]xylitol or d-[3-(14)C]xylulose plus xylitol or glucose at substrate concentrations. The glucose formed was isolated and degraded to give the relative specific radioactivities in each carbon atom. C-4 of glucose had the highest specific radioactivity, followed by C-3, with half to one-fifth that of C-4.

14CO2 fixation by phosphoenolpyruvate carboxykinase during gluconeogenesis in the intact rat liver cell.

During gluconeogenesis from L-glutamine, 14CO2 is fixed into glucose. Inhibitors of pyruvate transport or pyruvate carboxylase only slightly decrease the 14CO2 incorporation, indicating that a pathway of formation of pyruvate, followed by pyruvate carboxylation, is not primarily involved. These results suggest that 14CO2 fixation is effected by a reverse (exchange) reaction of P-enolpyruvate carboxykinase.

Effects of alterations in energy supply on gluconeogenesis from L-lactate.

1. We have examined effects, on gluconeogenesis from lactate, of altering energy metabolism in two ways: (a) by primarily lowering cytosolic ATP levels with the use of atractyloside or 2,5 anhydromannose; and (b) by decreasing mitochondrial energy generation with the use of the classical uncoupler, dinitrophenol. 2.

Effects of glucose and of lactate on phosphofructokinase flux during gluconeogenesis.

We have examined the effects of glucose and lactate, the products of the gluconeogenic-glycolytic pathways, on phosphofructokinase flux during gluconeogenesis in hepatocytes from fasted rats. With dihydroxyacetone as substrate, phosphofructokinase flux is rather active. Addition of lactate, at concentrations of 5-10 mM, causes a lowering of this flux to the levels found when lactate alone is the substrate.

Control of ethanol utilization by rat hepatocytes.

Ethanol oxidation by hepatocytes from fasted rats was determined in the presence and absence of 0.2 mM ethyl hydrazinoacetate, a transaminase inhibitor which blocks the malate-aspartate cycle. 20 muM phenazine methosulfate caused the largest increase (nearly 150%) in ethanol utilization.