Andreas J. Schwab

Tracer Oxygen Distribution Is Barrier-Limited in the Cerebral Microcirculation

The kinetics of tracer oxygen distribution in the brain microcirculation of the awake dog were investigated with the multiple indicator dilution technique. A bolus containing 51Cr-labeled red blood cells, previously totally desaturated and then resaturated with [18O]2 (oxygen), 125I-albumin, 22Na, and [3H]water, was injected into the carotid artery, and serial anaerobic blood samples were collected from the sagittal sinus over the next 30 seconds. The outflow recovery curves were analyzed with a distributed-in-space two-barrier model for water and a one-barrier model for oxygen. The analysis provided an estimate of flow per gram brain weight as well as estimates for the tracer water and oxygen rate constants for blood-to-brain exchange and tracer oxygen parenchymal sequestration. Flow to tissue was found to vary between different animals, in concert with parallel changes in oxygen consumption. The 18O2 outflow curves showed an early peak, coincident with and more than half the magnitude of its vascular reference curve (labeled red blood cells), whereas the [3H]water curve increased abruptly to a low-in-magnitude curve at low flow values and to a small early peak at high flow values. Analysis indicates that the transfers of both 18O2 and [3H]water indicators from blood to brain are barrier-limited, with the former highly so because of the large red blood cell capacity for oxygen, and that the proportion of the tracer oxygen returning to the circulation from tissue is a small fraction of the total tracer emerging at the outflow.

The capillary transport system for free fatty acids in the heart.

The nature of the process by which free fatty acids, which are tightly bound to albumin, traverse the endothelium of cardiac capillaries to reach the cardiac muscle cells, so that they are extracted to a net extent of approximately 40%, needs clarification. Previous studies have indicated that a membrane fatty acid-binding protein provides for carrier-mediated uptake of free fatty acids by isolated hepatocytes, cardiomyocytes, and jejunal mucosal cells. A monoclonal monospecific antibody was prepared against purified membrane fatty acid-binding protein from rat liver. Multiple-indicator dilution experiments were carried out in the isolated rat heart with labeled albumin, sucrose, and palmitate in the presence of control perfusate or perfusate containing either specific antibody or comparable nonspecific myeloma cell supernatant (each of the latter containing additional albumin, in identical concentrations). Analysis of the labeled-sucrose curves provided a permeability-surface area product for sucrose to which that for palmitate could be compared. In comparison with control supernatants, myeloma supernatant produced a minor inhibition of palmitate uptake, as a result of the increase in albumin concentration. The specific antibody, which contained identical albumin concentrations, produced a major inhibition of palmitate uptake, significantly greater than with the myeloma supernatant. The data indicate that the membrane fatty acid-binding protein mediates the transfer of free fatty acid across the endothelial cells of cardiac capillaries for presentation to heart muscle. Passive intramembrane lateral diffusion of palmitate could not provide an explanation for the findings.

Transport, binding, and metabolism of sulfate conjugates in the liver

Sulfate conjugates are a heterogeneous class of polar, anionic metabolites that result from the conjugation of endogenous and exogenous compounds. Sulfate conjugates exhibit a high degree of binding to albumin, the extent of which usually exceeds those of their parent compounds. Preponderant direct and indirect evidence suggests that sulfation activity is slightly higher in the periportal than in the perivenous (centrilobular) region of the liver, but recent immunohistochemical studies imply that specific isoforms of the sulfotransferases may also be preferentially localized in the perivenous region. Entry of sulfate conjugates into the liver cell is poor unless discrete carriers are present. Although known transport carriers exist for the sulfated bile acids, the specificity of the carriers for drug sulfate conjugates is presently unknown. The removal of sulfates is usually by way of biliary excretion while, on occasion, sulfates can be desulfated and participate in futile cycling with their parent compounds. The binding, transport, and hepatic elimination of various drug sulfate conjugates are examined. Non-recirculating studies carried out in the perfused rat liver with the multiple indicator dilution technique under varying input sulfate conjugate concentrations have provided essential information on the effects of vascular (red blood cells and plasma protein) binding on transport and removal of the conjugates. These studies clearly demonstrate the need to study protein binding, transmembrane transfer characteristics across the liver basolateral (sinusoidal) and canalicular membranes, and enzyme zonation in a distributed-in-space fashion in order to properly define the handling of sulfate conjugates in the liver.

Xenon handling in the liver: red cell capacity effect.

Xenon, despite its lack of chemical reactivity, associates preferentially with red cells in blood. To characterize the effect of this and the nature of xenon-tissue interaction in the liver, multiple indicator dilution studies were performed in the anesthetized normal dog through portal vein injection and hepatic vein collection of anaerobic blood samples. Two experimental runs were carried out in each animal, one at the prevailing hematocrit and the other at reduced hematocrit after bleeding and replacement with dextran. For comparison, the injection mixtures contained labeled red blood cells (a vascular reference), sucrose (an interstitial space reference), and labeled water (which freely enters liver cells), as well as labeled xenon. At the higher hematocrit, the labeled xenon curves generally rose earlier, peaked higher, and decayed more quickly than the labeled water curve; at the lower hematocrit, the xenon curve was delayed and diminished in magnitude in relation to the Libeled water curves. Analysis of the curve shapes indicated that xenon, like labeled sucrose and water, underwent delayed wave flow-limited distribution. With knowledge of the red cell plasma partition coefficient (2.89 ml/ml), it was possible to both account for the change in form of the xenon curves with hematocrit and to use the data to estimate the liver cell tissue plasma xenon partition coefficient. Values averaged 1.93 ml/ml liver space, or 1.79 ml/g, and did not change significantly from first to second runs. Theoretical analysis indicated that flow cannot be estimated from xenon downslopes.

Kinetics of endothelin-1 binding in the dog liver microcirculation in vivo

Endothelin-1 (ET-1) is a 21-amino acid peptide produced by vascular endothelial cells that acts as a potent constrictor of hepatic sinusoids. Hepatic binding of tracer 125I-labeled ET-1 was investigated in anesthetized dogs with the multiple-indicator dilution technique with simultaneous measurements of unlabeled immunoreactive ET-1 plasma levels. Despite 80% binding to albumin, tracer 125I-ET-1 was avidly extracted by the liver, with only 15 ± 6% of the peptide surviving passage through the organ. Exchange of ET-1 between plasma and binding sites, probably located on the surface of liver cells, was quantitatively described by a barrier-limited, space-distributed variable transit time model. Reversible and irreversible parallel binding sites were found. Reversible and irreversible plasma clearances of unbound 125I-ET-1 were 0.084 ± 0.033 ml ⋅ s-1 ⋅ g liver-1 and 0.17 ± 0.09 ml ⋅ s-1 ⋅ g liver-1, respectively, and the dissociation rate constant for reversible binding was 0.24 ± 0.12 s-1. The specific ETA receptor antagonist BMS-182874 did not modify binding to either site. The nonspecific ETA/ETBantagonist LU-224332 dose-dependently reduced irreversible binding only. ET-1 levels in the hepatic vein were significantly lower than in the portal vein but were not different from those in the hepatic artery. The ratio between hepatic vein and portal vein levels (0.64 ± 0.31) was considerably higher than survival fractions, suggesting a substantial simultaneous release of newly synthesized or stored ET-1 by the liver. These results demonstrate both substantial clearance and production of ET-1 by the intact liver. Hepatic ET-1 clearance is mediated by the ETB receptor, with the presence of reversible, nonspecific ET-1 binding at the liver surface.

Application of the dispersion model for description of the outflow dilution profiles of noneliminated reference indicators in rat liver perfusion studies.

AbstractThe dispersion model (DM) is a stochastic model describing the distribution of blood-borne substances within organ vascular beds. It is based on assumptions of concurrent convective and random-walk (pseudodiffusive) movements in the direction of flow, and is characterized by the mean transit time

Cardiac microcirculatory effects of beta-adrenergic blockade during sympathetic stimulation.

\left( {\overline {\text{t}} } \right)

Liver Cell Entry In Vivo and Enzymic Conversion

and the dispersion number (inverse Peclet number), DN. The model is used with either closed (reflective) boundary conditions at the inflow and the outflow point (Danckwerts conditions) or a closed condition at the inflow and an open (transparent) condition at the outflow (mixed conditions). The appropriateness of DM was assessed with outflow data from single-pass perfused rat liver multiple indicator dilution (MID) experiments, with varying lengths of the inflow and outflow catheters. The studies were performed by injection of bolus doses of 51Cr-labeled red blood cells (vascular indicator),125I-labeled albumin and [14C] sucrose (interstitual indicators), and [3H]2O (whole tissue indicator) into the portal vein at a perfusion rate of 12 ml/min. The outflow profiles based on the DM were convolved with the transport function of the catheters, then fitted to the data. A fairly good fit was obtained for most of the MID curve, with the exception of the late-in-time data (prolonged tail) beyond