Eugene M. Renkin

Microvascular supply in relation to fiber metabolic type in mixed skeletal muscles of rabbits

Abstract Medial gastrocnemius (MG), lateral gastrocnemius (LG), tibialis anterior (TA), and soleus muscles (SO) of New Zealand White female rabbits were frozen, and serial sections were cut for histochemical demonstration of fiber types (succinic dehydrogenase and myofibrillar ATPase) and capillaries (alkaline phosphatase). Fibers were identified as fast white (αW), fast red (αR), and slow red (β). MG, LG, and TA contained all three types. Most SO had only β fibers, a few had a small proportion of αR. Capillary/fiber ratios for whole muscles or for individual fields of a single muscle increased with the number of red fibers (αR + β) present. Capillary fiber ratios for αR and β fibers in LG and TA were greater than for αW fibers in the same muscle, but in MG ratios were similar for all fiber types. In MG and LG the number of capillaries per αR or β fiber was less than for the all-β soleus; only in TA were comparable ratios observed. However, in all three mixed muscles, the αR fibers were smaller than the αW, and in MG and LG the β fibers were also smaller, and these size differences masked uniformly higher capillary densities (capillaries/mm 2 ) for αR and β fibers in mixed muscles. On the average, capillary densities for red fibers were twice those for white in the same muscles, and were comparable to densities in the all-red soleus. Estimated maximal oxygen uptakes for red and white fibers (based on data in the literature) are in the ratio of approximately five to one. If these estimates are valid, capillary supply to the white fibers in mixed muscles is excessive, or the supply to red fibers is deficient, in relation to their capacities for oxidative metabolism.

Transport pathways for fluid and large molecules in microvascular endothelium of the dog's paw

Abstract Solvent drag coefficients ( σ f ) and permeability surface area products ( PS ) for six plasma proteins and Dextran 110, determined in the preceding publication, were analyzed according to irreversible thermodynamic and hydrodynamic principles. The relation of σ f to molecular size indicates that endothelial pathways admitting these molecules are not the only hydraulically conductive pathways present. At least 81.5% of the volume flow must be assigned to routes essentially impermeable to serum albumin and larger molecules. Presumably, these are the cell membranes and small pore system. The convective pathway for large molecules may represent openings between endothelial cells or fused chains of cytoplasmic vesicles forming transitory channels. Calculated PS values for large molecules in these channels are no larger than one-fourth the measured values. The residual PS must be attributed to nonhydraulically conductive pathways. If these are supposed to be nonfused endothelial vesicles, their turnover rate is estimated to be 0.4 × 10 −4 ml/sec per 24-g paw, accounting for three-fourths to five-sixths of the total dissipative solute flux. Due to the composite (parallel pathway) structure of the microvascular endothelium, σ f for the whole membrane does not appear to be the same as the osmotic reflection coefficient, ( σ d ), even though the two may be equal for individual membrane pathways (Onsager reciprocity). Over the range of experimental pressures and flows, σ f for total plasma proteins is 0.83, while σ d is probably 0.89 or higher. The relation between the two σs is sensitive to the microscopic disposition of osmotic and hydrostatic forces in relation to individual transport paths.

B. W. Zweifach award lecture: Regulation of the microcirculation☆

Microcirculatory blood flow and transport are controlled to meet local and systemic demands for material exchange and body fluid balance. Control mechanisms act through effectors (smooth muscle cells) at many sites within the microvascular bed. Responses at different sites are not uniform, resulting in a broadly heterogeneous distribution of pressures and flows which is constantly changing. Simplified, uniform models of microvascular networks have made it possible to identify the principles governing blood circulation and blood-tissue transport. However, knowledge of how these principles are integrated at the microcirculatory level requires the variability and heterogeneity to be taken into account. Indeed, there is much reason to believe that heterogeneity is an important part of microcirculatory control.

Measurement of microvascular transport parameters of macromolecules in tissues and organs of intact animals.

A critical analysis is presented of two widely used approaches to measurement of microvascular transport of large molecules in intact animals: (1) measurement of lymph flow and macromolecular solute concentration relative to plasma, and (2) tissue accumulation of tracer macromolecules. To demonstrate the advantages and limitations of each method, experimental results which permit direct comparison of the two methodologies are reviewed and analyzed, and sources or error in each pointed out. It is concluded that both approaches are valid under the appropriate conditions: steady state for lymph, initial transient state for tissue uptake. These conditions are mutually exclusive, but complementary. When the requirements for neither lymph collection or tissue accumulation alone can be satisfied experimentally, a combination of the two approaches can yield valid results.

Wick sampling of interstitial fluid in rat skin: Further analysis and modifications of the method ☆

UNLABELLED We compared modifications of the wick technique for analysis of interstitial fluid in rat subcutis. Nylon wicks were implanted for 60 min in back skin of rats after anesthesia with pentobarbital or after sacrifice by potassium chloride injection. Wicks were implanted dry or loaded with saline or varied dilutions of rat serum. Implantation of dry wicks and wicks loaded with diluted serum in living, anesthetized animals produced similar results; the protein concentration of wick fluid averaged about 60% that of the plasma protein concentration. The saline loaded wicks produced wick fluid with a lower protein concentration, average about 45% that of plasma protein concentration. The lower concentrations apparently resulted from simple dilution. Wick fluid sampled from dead animals had similar total protein concentrations, but in the dead animals there was a lower concentration of the large plasma proteins and a relatively higher concentration of the smaller proteins. CONCLUSIONS Wick implantation in living animals causes a transitory inflammatory reaction and a decrease in the size selectivity of macromolecular sieving, but local osmotic forces bring about a concentration equilibrium with undisturbed interstitium. Implantation of dry wicks in subcutis either in vivo or post mortem provides a simple, direct method for sampling the total protein concentration and colloid osmotic pressure of interstitial fluid. Implantation of dry wicks postmortem permits measurement of individual component protein concentrations and evaluation of molecular selectivity between plasma and interstitium.

Phosphodiesterase 4 inhibition attenuates atrial natriuretic peptide-induced vascular hyperpermeability and loss of plasma volume

Natriuretic peptides (such as atrial natriuretic peptide, ANP) are normally present at very low levels in the blood and are part of physiological systems that control blood volume. During diseases such as heart failure and sepsis, circulating levels of ANP increase, leading to an increase in blood vessel permeability and loss of blood fluid volume to the tissues. Other studies show that some inflammatory responses are strongly blocked by increased intracellular cAMP. Here we tested whether rolipram, an inhibitor of the degradation of cAMP, could counteract the movement of protein and fluid out of the blood that is induced by ANP. We found that rolipram almost completely blocked the ANP‐induced loss of blood volume. Stabilizing the endothelial barrier by controlling cAMP levels to offset ANP‐induced increases in vascular permeability may be part of a strategy to maintain plasma volume in disease states with elevated natriuretic peptides.

Interstitial exclusion of macromolecules studied by graded centrifugation of rat tail tendon.

Mechanical compression of cartilage and tendon has been shown to expel fluid both from collagen fibrils and from the extrafibrillar space. As reported previously, albumin (Alb) concentration and colloid osmotic pressure in tendon fluid (TF) expelled by repeated centrifugations fell progressively at increasing centrifugation force (G = 600, 2,400, and 13,100), suggesting either molecular sieving in compressed tendon or mobilization of protein-free (excluded) fluid. The present experiments, including analysis of 51Cr-EDTA, aprotinin (Ap), Alb, immunoglobulin G (IgG), and hyaluronan (hyaluronic acid; HA) with molecular weight (MW) ranging from 341 to 5 × 106, strongly favored the exclusion hypothesis; the fraction of Alb, IgG, and HA-free fluid (excluded) was already 0.23-0.36 in the first centrifugate, increasing to 0.73-0.82 in the third. The corresponding numbers were, respectively, 0.11 and 0.43 for Ap (MW 6,500), and 0 and 0.08 for51Cr-EDTA. These data, combined with calculated exclusion by collagen fibrils, proteoglycans, and HA, indicated that the first centrifugate was mainly derived from the extrafibrillar space, with increasing addition of macromolecular free intrafibrillar fluid in the second and third centrifugates, with each space contributing about equally to the total centrifugate volume. The calculations also indicated that Alb-, IgG-, and Ap-free fluid was mobilized from extrafibrillar space by increasing overlap of excluded territories. An excess of HA in tendon compared with that estimated from centrifugate concentrations suggests a large bound or immobilized HA fraction.

Cellular Aspects of Transvascular Exchange: A 40‐Year Perspective

Study of the microcirculation began with microscopic anatomy in the 17th century, but it was not until the mid‐19th century that the concept of microcirculatory exchange was developed, in association with the great expansion in knowledge of physical chemistry and cell biology. In the first half of the 20th century, the roles of diffusion and osmosis (ultrafiltration) were put on a quantitative foundation and the physiological basis for regulation of the microcirculation was established. During the past 40 years, our understanding of transvascular exchange has been deepened and widened through two major developments: 1) the application of electron microscopy to the study of microvascular structure and 2) the use of sophisticated physical and mathematical models of passive transport processes to analyze and interpret experimental data. In the past few years, the focus of microvascular research has been returning to cell biology and the role of endothelial cells in controlling transvascular exchange. The 21st century holds promise of many exciting new developments.

Flow and distribution of India ink in microvessels of the frog.

Velocities of an India ink front and of RBCs moving ahead of it were studied by intravital microscopy in capillaries of frog mesentery and skeletal muscle. Measurements were made during microperfusion of single vessels or groups of connected vessels in mesentery, and following intravenous ink injection in both tissues. The presence of ink did not appear to interfere with microvascular flow or vasomotion during the period of observation. On the average, the ink spearhead moved only slightly faster than the RBCs. There was substantial variation in relative velocities of RBCs in the same vessel and in the relative velocities of ink front and RBCs. The time course of ink filling showed substantial heterogeneity of flow in mesentery, more nearly uniform flow in skeletal muscle. Comparison of the measured velocity ratios of ink to RBCs with published observations on relative velocities of RBC to blood suggest that the advancing, apparently parabolic front of ink moves at less than twice the mean blood velocity. This is due in small part to diffusive dispersion of the ink particles in the laminar flow gradient, but more largely to stochastic dispersion of the front by interaction with RBCs and by displacement at branches.

DETERMINANTS OF LYMPH FLOW AND COMPOSITION

Publisher Summary According to Starlings hypothesis, the rate of fluid movement across capillary walls depends on the balance of hydrostatic (P) and colloid osmotic (Π) forces. Under normal conditions, a small net loss of fluid from the capillaries is compensated for by lymphatic drainage of leaked fluid and solutes from the interstitium. Filtration rate and lymph flow can be increased by raising capillary hydrostatic pressure or by lowering plasma colloid osmotic pressure or by combinations of these procedures. The results of a study described in the chapter show that the increase in lymph flow for a given decrease in transcapillary colloid osmotic pressure difference is 1.4 to 1.8 times as great as that produced by an equivalent increase in hydrostatic pressure. For a given increase in lymph flow, decreasing colloid osmotic pressure produces a larger increase in blood–lymph protein clearance. Both these observations are contrary to expectation based on fluid and protein transport through large pores or channels, with low reflection coefficients for plasma proteins.