Uptake of materials by the intact liver. The exchange of glucose across the cell membranes.

D-Glucose equilibrates within liver cells. We have studied its process of entry into and exit from these cells with the multiple indicator dilution technique. Labeled red cells (a vascular indicator), labeled sucrose (an extracellular reference), and labeled D-glucose were rapidly injected into the portal vein, and from serially sampled hepatic venous blood, normalized outflow-time patterns were obtained. The labeled red cell curve rises to an early high peak, and decays rapidly; and that for sucrose reaches a later and lower peak and decays less rapidly, but generates an equivalent area. The curve for labeled D-glucose begins with that for labeled sucrose, gradually rises to a peak which is later and substantially lower than that for sucrose, and then decreases slowly. At high glucose levels this curve assumes a squared-off shape, rises fairly quickly to its highest level, at the time of the sucrose peak, and then slowly decreases. Phlorizin and galactose infusion result in the emergence of a pronounced early peak, under the sucrose peak; and the curve for tracer L-glucose approaches that for sucrose. We resolve from the D-glucose curves, by model analysis, two components: throughout material, which has not entered the cells; and exchanging material, which has entered and later returned to the circulation. The analysis provides estimates of the kinetic entrance and exist coefficients; and from these, saturation of both the entrance and exit processes was evident. The characteristic transport parameters were determined. For both entrance and exit, a common Km, 2,170 mg/100 ml, and transport maximum, 5.13 mg s-1 (ml intracellular fluid)-1, were found. Both these values are exceedingly large. Several other phenomena were defined which additionally characterize the transport process: phlorizin and galacose produced competitive inhibition; the transport process was found to be relatively stereospecific; and sudden infusion of hypertonic glucose produced counter-transport of labeled D-glucose.

On the uptake of materials by the intact liver. The concentrative transport of rubidium-86.

In this study we use the multiple indicator dilution technique to outline the kinetic mechanisms underlying the uptake of rubidium, a cation which, in the steady state, is concentrated by hepatic parenchymal cells. We inject a mixture of (51)Cr-labeled red blood cells (a vascular reference substance), (22)Na (which is confined to the extracellular space, the expected extravascular distribution space for rubidium, in the absence of cellular uptake), and (86)Rb into the portal vein and obtain normalized outflow patterns, expressed as outflowing fractions of each injected mass per milliliter vs. time. The labeled red cell curve rises to the highest and earliest peak and decays rapidly. That for labeled sodium rises to a later and lower peak, and decays less rapidly. Its extrapolated recovery is equal to that for the red cells. The observed (86)Rb curve consists of two parts: an early clearly defined peak of reduced area, related to the (22)Na peak in timing; and a later tailing, obscured by recirculation, so that total outflow recovery cannot be defined (even though it would be expected to be the same). We model the concentrative uptake of (86)Rb and find two corresponding outflow fractions: throughput material, which sweeps past the cell surface as a wave delayed with respect to the vascular reference (tracer which has not entered cells); and exchanging material (tracer which has entered cells and later returns to the circulation). We find that the outflow form of the rubidium curve, the presence of both a relatively clearly defined throughput component and a relatively prolonged low-in-magnitude tailing, is consequent to the concentrative character of the transport mechanism, to the presence of an influx rate constant many times the efflux rate constant. The modeling which we develop is general, and has potential application in situations where transport is nonconcentrative.

On the uptake of materials by the intact liver. The transport and net removal of galactose.

D-galactose, a monosaccharide rapidly phosphorylated within liver cells, is irreversibly removed from the portal circulation. We have studied the kinetic relations between the hepatic cell entry process and the metabolic sequestration process, by means of the multiple indicator dilution technique. Labeled red blood cells (a vascular indicator), labeled sucrose (an extracellular reference), and labeled galactose were rapidly injected into the portal vein, and from rapidly sampled hepatic venous blood, normalized outflow-time patterns were secured. The labeled red cell curve rises to the highest and earliest peak, and decays rapidly; and that for labeled sucrose rises to a later and lower peak. Its extrapolated recovery is equivalent to that of the labeled red cells. At low blood galactose concentrations, the labeled galactose appears at the outflow with labeled sucrose, but is much reduced in magnitude, and exhibits a long tailing. Its outflow recovery is much reduced. At high blood galactose concentrations, the initial part of the profile increases towards that for labeled sucrose, the tailing becomes much larger in magnitude, and the outflow recovery becomes virtually complete. We have modeled the uptake of labeled galactose, and find two parts to the predicted outflow pattern, corresponding to our experimental observations; throughput material, which sweeps past the cell surface in the extracellular space; and returning material, which has entered the cells but escaped the sequestration process. Analysis of the data by use of this model provides estimates of both transmembrane fluxes and rates of sequestration. The capacity of the process subserving cell entry is found to be 40 times that for phosphorylation; and, whereas the K(m) value for sequestration is less than 15 mg/100 ml, that for entry is approximately 500 mg/100 ml. Both processes are relatively stereospecific; the entry of the L-stereoisomer is very slow and it undergoes no significant amount of metabolic sequestration. The sequestration process produces a lobular intracellular concentration gradient; and this gradient, in turn, produces some uncertainty in the estimate of the true K(m) value for the sequestration process.

A nonocclusive cannula for sampling venous blood from the interscapular brown fat of conscious rats.

Lack of an adequate method for sampling venous blood from the brown adipose tissue (BAT) of conscious animals has impeded study of the in vivo metabolism of this tissue during physiological activation of its thermogenic function. This paper describes a technique for cannulating the main vein (Sulzer's) of the interscapular BAT (IBAT) of rats in a manner that does not impair blood flow and allows multiple venous sampling over several hours in conscious animals. The technique was tested over the widest possible range of IBAT blood flows by applying it to measurements of IBAT arteriovenous O2 differences in barbital-anesthetized, cold-acclimated rats infused with vehicle or with various doses of noradrenaline. Comparison was made with controls in which samples of IBAT venous blood were obtained by cutting Sulzer's vein. Blood flow was measured by the microsphere method. These tests showed that the presence of the special cannula in Sulzer's vein had no significant effect on the blood flow, arteriovenous O2 difference, or O2 consumption of the IBAT at any level of noradrenaline-induced thermogenesis. The new technique will permit examination of the functioning of BAT in nonshivering thermogenesis and diet-induced thermogenesis under much more physiological conditions than hitherto possible. It should also significantly reduce the number of animals required for such studies.

Red cell carriage of label: its limiting effect on the exchange of materials in the liver.

The red cell membrane is a permeability barrier that limits the equilibration of a variety of solutes between red cell and plasma water. We utilized the multiple indicator dilution technique to investigate the effect of this barrier on the exchange in the liver of a group of tracer substances that are not removed in net fashion from the hepatic circulation: thiourea, urea, and chloride. We demonstrated that, after preequilibration of the label with red cells, a red cell carriage effect appeared (the trapping and translocation of label in the red cells), that this effect was most marked when the permeability of the red cell was relatively low for the substance under consideration (thiourea), and that the effect became small when the permeability of the red cells was large for the exchanging substance (urea and chloride). We developed a theoretical description of the retarding effect of the red cell permeability barrier on the extravascular exchange of label and were able to use this description to obtain estimates of the red cell permeability from the in vivo dilution curves. We examined the effect of plasma injection, of changing the input in such a fashion that the label was not preequilibrated with red cells, and found both experimentally and theoretically, that for substances of low permeability the transit time from these experiments, if multiplied by the total water flow or solute flux, gave an overestimate of both the apparent total volume of distribution and the mass of traced material in the system. This last effect is of great importance for the practical design of many biological experiments. Reliable volume and mass estimates can be made only when the labeled material has been preequilibrated with red cells.

Evidence for a contribution by brown adipose tissue to the development of fever in the young rabbit.

This study was undertaken to determine if brown adipose tissue was involved in heat production during fever produced by S. abortus equi (1 micrograms) in unanesthetized rabbits aged 19-26 days. The fever (0.9-1.6 degrees C) occurred after a delay of 20-30 min and was frequently biphasic. Radiolabelled microspheres for measuring tissue blood flow were injected intraventricularly into three groups of animals: rabbits not given pyrogen, rabbits in which the febrile response to pyrogen was developing, and rabbits in which the febrile response had peaked. Blood flow to brown fat deposits and other organs was calculated from the fractional distribution of the microspheres and the recovery of microspheres in a reference arterial blood sample. At the fever peak, blood flow to brown fat was not significantly different (p greater than 0.05) from the control value (0.9 +/- 0.2), but during the rising phase of the fever the flow increased significantly (p less than 0.01) to 2.6 +/- 0.4 mL min-1 g-1. The blood flow to muscles of the forelimbs and hind limbs was also increased significantly (p less than 0.05) during the rising phase of the fever. No significant change in blood flow to other organs or tissues was found during the rising phase of the fever. These results indicate that both nonshivering as well as shivering thermogenesis contribute to heat production during development of fever in the young rabbit. However, nonshivering thermogenesis was not involved in the maintenance of the elevated body temperature after the fever had peaked.

The relationship between the thermogenic response of brown adipose tissue and age during noradrenaline infusion in the young rabbit.

This work was undertaken to determine if the thermogenic activity of brown fat decreased with age in young rabbits despite the morphological evidence indicating persistence of the brown adipocytes at 4 weeks of age. Data obtained by infusing five doses of noradrenaline and measuring oxygen consumption were used to construct cumulative dose-response curves for five age groups between 3 and 32 days of age. Blood flow to brown fat and other tissues was measured by the microsphere method at 1 and 3 weeks of age. The noradrenaline-induced increase in oxygen consumption when expressed as a percentage of resting oxygen consumption in millilitres per 100 g of body weight decreased (p less than 0.05) with age. However, the absolute noradrenaline-induced increase in metabolic rate (millilitres per minute) increased with age. Total blood flow to brown fat (millilitres per minute) during noradrenaline infusion was unchanged between 1 and 3 weeks of age, but when the blood flow was expressed in millilitres per minute per gram of tissue flow decreased significantly (p less than 0.05) probably because of infiltration of brown fat with white fat. These data suggest that the amount of brown fat and its thermogenic capacity remain relatively constant between 1 and 3 weeks of age, but as a thermogenic organ, brown fat becomes proportionally less effective with age because of the large increase in body mass.

Evidence for liver as the major site of the diet-induced thermogenesis of rats fed a "cafeteria" diet.

The resting metabolic rates (VO2) of rats fed chow (CH) or a "cafeteria" (CAF) diet of highly palatable human foods were measured at thermoneutrality (28 degrees C) before and shortly after two-thirds hepatectomy or sham operation, and again after administration of propranolol (5 mg/kg). CAF rats initially had a 17% and 1.2 mL/min higher mean resting VO2 than CH rats, a difference usually considered to represent the diet-induced thermogenesis (DIT) that CAF rats develop during overconsumption of the diet. Sham operation did not significantly affect resting VO2 in either diet group. Two-thirds hepatectomy decreased VO2 by about 1.0 mL/min more (125% more) in CAF rats than in CH rats, from which it may be estimated that the CAF rats initially had a liver VO2 about 1.6 mL/min higher than that of the CH rats, a difference more than sufficient to fully account for their apparent DIT. Propranolol did not significantly affect the VO2 of CH rats. It reduced the VO2 of sham-operated CAF rats by 0.94 +/- 0.08 mL/min (12%), but had a significantly smaller effect (delta VO2 = -0.50 +/- 0.05 mL/min) in partially hepatectomized CAF rats. This difference suggests that about 70% of the propranolol-inhibitable fraction of the elevated VO2 of the CAF rats, presumably a measure of sympathetically mediated DIT, resided in the liver. This study thus points to the liver as the major (70-100%) effector of the DIT of CAF rats.

Effect of exercise on dilution estimates of extravascular lung water and on the carbon monoxide diffusing capacity in normal adults.

Previous studies in exercising animals have demonstrated that the extravascular lung water accessible to measurement by dilution methodology increases in the transition from rest to low-level exercise and thereafter does not change with progress to high-level exercise. In normal humans, similar systematic examination is essential to provide a background for the interpretation of changes in measured extravascular lung water in pathophysiological states. Moreover, such an examination might provide new insight into the mechanisms underlying the change in the pulmonary diffusing capacity with exercise. We therefore measured both the pulmonary extravascular lung water (by use of the triple indicator-dilution technique) and the diffusing capacity for carbon monoxide in 11 subjects, seated on an exercise bicycle, at rest and usually during two levels of exercise. The central blood volume increased by 50% with a tripling of the cardiac output. The accessible lung water increased from an average of 2.16 g/kg to 2.55 g/kg in the transition from rest to low-level exercise, but it did not increase further at the higher level of exercise. The simultaneously measured diffusing capacity for carbon monoxide (single breath and steady state) continued to increase over the whole range of cardiac outputs. We infer that the proportion of the pulmonary parenchyma perfused by blood flow increases slightly in the transition from rest to low-level exercise but increases no further at the higher level of exercise. The continued increase in the pulmonary diffusing capacity over the range in which the estimated lung water values do not change appears to imply that part of this increase may be blood flow dependent rather than dependent on the recruitment of additional surface for exchange.

Absence of increased oxygen consumption in brown adipose tissue of rats exhibiting "cafeteria" diet-induced thermogenesis.

Young male Sprague-Dawley rats were induced to overeat (approximately 45%) by provision of a "cafeteria" (CAF) diet of palatable human foods. Normophagic rats fed a commercial chow or a semisynthetic diet served as controls. The CAF rats exhibited (a) the reduced food efficiency and the propranolol-inhibitable elevation in resting metabolic rate (resting VO2) that are indicative of a facultative diet-induced thermogenesis (DIT) by which excess energy gain is resisted, and (b) certain changes in brown adipose tissue (BAT) that are among those taken as evidence for BAT as the effector of DIT, e.g., increased protein content and increased mitochondrial binding of GDP. To assess directly and quantitatively the contribution by BAT to the elevation in VO2 (apparent DIT) of the CAF rats, BAT O2 consumption was determined (Fick principle) from measurements of tissue blood flow (microsphere method) and the arteriovenous difference in blood O2 across interscapular BAT (IBAT). To obtain the measurements, the animals were fitted under halothane anesthesia with vascular cannulas for intraventricular injection of microspheres and sampling of arterial blood and the venous effluent of IBAT. After recovery from anesthesia and rewarming to normal body temperature the animals were placed singly in a temperature-controlled metabolic chamber and the measurements, which also included determination of resting VO2, were made 1.5-2 h later about 11:30 h. As determined from measurements made at 28 degrees C (thermoneutrality) mean values of resting VO2 for the cannulated rats were unchanged from those of intact (unoperated) CAF or control rats.(ABSTRACT TRUNCATED AT 250 WORDS)