Steady-state 125I-labeled rat serum albumin (125I-labeled RSA) concentration in plasma was maintained by intravenous infusion of tracer for 72-168 h with an implanted osmotic pump. At the end of the infusion period, the rat was anesthetized and nephrectomized, and extracellular fluid was equilibrated with intravenous 51Cr-labeled EDTA for 4 h. Five minutes before final plasma and tissue sampling, 131I-labeled bovine serum albumin (131I-labeled BSA) was injected intravenously as a plasma volume marker. Samples of skin, muscle, tendon, and intestine were assayed for all three tracers. Apparent distribution volumes were calculated as tissue tracer content/plasma tracer concentration. Interstitial fluid volume (Vi) was calculated as V51Cr-EDTA-V131I-BSA. Steady-state extravascular distribution of 125I-labeled RSA as plasma equivalent volume (Va,p) was calculated as V125I-RSA-V131I-BSA. Steady-state interstitial fluid concentrations of 125I-labeled RSA in skin, muscles, and tendon were measured with nylon wicks implanted postmortem, and steady-state interstitial albumin distribution volumes were recalculated as wick-fluid equivalent volumes (Va,w). Relative albumin exclusion fraction (Ve/Vi) was calculated as 1-Va,w/Vi. For skin and muscle, steady-state 125I-labeled RSA tissue concentrations were reached at 72 h. Ve/Vi for albumin averaged 26% in hindlimb muscle, 41% in hindlimb skin, 30% in back skin, 39% in tail skin, and 54% in tail tendon. For muscle, Ve/Vi corresponds to expectation if all tissue collagen and hyaluronan is dispersed in the interstitium. However, for skin and tendon, albumin exclusion is considerably lower than expected on this basis, suggesting that much of their collagen is organized into dense bundles of fibers containing no fluid accessible to 51Cr-labeled EDTA or 125I-labeled RSA.
A modification of the implanted wick method (K. Aukland and H. O. Fadnes. Acta Physiol. Scand. 88: 350-358, 1973) was devised to sample interstitial fluid from rat muscles. Dry nylon wicks were inserted postmortem into intermuscular spaces between leg muscles by means of a plastic catheter, which was subsequently withdrawn. Inserting the wicks postmortem avoids contaminating wick fluid with proteins extravasated as a result of local inflammatory reactions; placing them intermuscularly avoids contamination by fluid and proteins from damaged muscle cells. Wick fluid protein concentrations (mg/ml) averaged 24.1 +/- 1.1 and 28.5 +/- 1.5 (means +/- SE) in medial and lateral hindlimbs muscles, respectively. The corresponding albumin concentrations were 13.0 +/- 0.7 and 13.9 +/- 0.7 mg/ml. Total protein and albumin concentrations in plasma were 54.1 +/- 0.8 and 22.5 +/- 0.3 mg/ml. Electrophoresis of wick fluid showed a pattern of peaks similar to that of plasma, with albumin relatively high and larger molecules relatively low. Proteins from muscle cells were not detected. Isotope studies (125I-labeled albumin, 51Cr-EDTA) showed that less than 2% of the albumin in wick fluid came directly from plasma and that wick fluid was not concentrated by cell swelling postmortem. Wick fluid from intermuscular wicks implanted in anesthetized rats in vivo had nearly the same total protein concentration as fluid from postmortem wicks, but albumin-to-globulin (A/G) ratios were slightly lower (1.22 +/- 0.07 vs. 1.53 +/- 0.21 measured by gel electrophoresis), and more significantly, nearly 50% of the albumin leaked to wick fluid from plasma as a result of wick implantation.(ABSTRACT TRUNCATED AT 250 WORDS)
Bovine serum albumin (BSA) labeled with 131I or 125I was injected intravenously in pentobarbital sodium-anesthetized rats, and tracer clearances into leg skin and muscles were measured over 30, 60, and 120 min. BSA labeled with the alternate tracer was used as vascular volume reference. Two minutes before injection of the tracer, a ligature was tied around one femoral vein to occlude outflow partially and raise capillary pressure in that leg. The unoccluded leg served as control. Skin and muscles of the occluded leg had variably and substantially higher water contents (delta W) than paired control tissues and slightly but consistently increased albumin clearances (CA). The delta CA/delta W, equivalent to the albumin concentration of capillary filtrate relative to plasma determined by linear regression, were as follows: leg skin 0.004 (95% confidence limits -0.001 to +0.009), muscle biceps femoris 0.005 (0.001-0.010), muscle gastrocnemius 0.011 (0.004-0.019), muscle tibialis anterior 0.016 (0.012-0.021). All these values are significantly less than 0.10, which corresponds to a reflection coefficient for serum albumin (sigma A) of 0.90. Convective coupling of albumin flux to volume flux in skin and muscles of intact, anesthetized rats is low, with sigma AS in the range 0.98 to greater than 0.99.
Interstitial exclusion, defined as the fraction of interstitial fluid volume inaccessible to a solute, was evaluated for immunoglobulin G (IgG) in selected tissues of rats by a method previously applied to serum albumin (29). IgG distribution volumes were also measured for intestine. 125I-labeled rat IgG was infused for 5 or 7 days (n = 4 rats each) with an implanted osmotic pump (Alzet). At the termination of infusion, the rat was anesthetized, nephrectomized, and injected with 51Cr-labeled EDTA (4 h) to label total extracellular fluid volume and 131I-labeled bovine IgG (5 min) to label plasma volume. Samples of skin, muscle, and tendon were assayed for total and extractable tracer activity. Interstitial fluid from these tissues was sampled postmortem with nylon wicks for assay of 125I-labeled IgG and endogenous albumin and IgG. Exclusion of IgG was calculated from the difference between extravascular 125I-labeled IgG and 51Cr-labeled EDTA distribution volumes. In contrast to our previous experience with tracer albumin, 125I-labeled IgG was not fully extractable from minced skin, muscle, or tendon by isotonic saline; only 71-83% was recovered under conditions that eluted 92-96% of tracer albumin and 94-99% of tracer EDTA. We conclude that approximately 20% of extravascular 125I-labeled IgG in these tissues is sequestered or bound in the interstitium. Calculation of IgG fractional exclusion from extractable tracer yielded the following values (means +/- SE, n = 8 rats): leg muscles 0.37 +/- 0.09, leg skin 0.44 +/- 0.03, back skin 0.36 +/- 0.04, tail skin 0.40 +/- 0.08, and tail tendon 0.55 +/- 0.04.(ABSTRACT TRUNCATED AT 250 WORDS)
Anesthetized rats were infused with lactated Ringer solution (LR) at constant rate for 30 or 60 min; delivered volume loads ranged from 0.03 to 0.08 ml/g body wt. Controls were given only a sustaining infusion of saline at 0.002 ml.g-1.h-1. Only 7-14% of the LR remained in the plasma at the end of the infusion; 76-88% entered the interstitial compartment, and 7-17% was excreted. The amount of plasma protein lost from the circulation with the extravasated fluid was studied simultaneously by two methods: 1) material balance in the whole animal and 2) changes in 131I-labeled albumin uptake (VA) and water content (VW) in individual tissues. The extravasation of 0.03-0.06 ml fluid/g body wt (75-160% initial plasma volume) did not significantly increase plasma protein extravasation in the whole rat. Nearly all of the sampled tissues of LR-infused rats had higher VW than controls. Tissue VA tended to increase with VW, but the regression slopes (delta VA/delta VW), a measure of the tracer albumin concentration of capillary filtrate relative to plasma, were low; skin, 0.006; paw, 0.018; skeletal muscles, 0.007; heart, 0.057; jejunum, 0.095; ileum, 0.045; cecum, 0.026; and colon, 0.027. These ratios are consistent with the very small loss of total plasma protein observed and attest to high solvent-drag reflection coefficients (sigma approximately equal to 1 - delta VA/delta VW): greater than 0.98 in capillaries of skeletal muscles, skin, and paw and 0.91-0.97 in heart and intestine.
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