Conducting airway gas exchange: diffusion-related differences in inert gas elimination.

We studied CO2 and inert gas elimination in the isolated in situ trachea as a model of conducting airway gas exchange. Six inert gases with various solubilities and molecular weights (MW) were infused into the left atria of six pentobarbital-anesthetized dogs (group 1). The unidirectionally ventilated trachea behaved as a high ventilation-perfusion unit (ratio = 60) with no appreciable dead space. Excretion of higher-MW gases appeared to be depressed, suggesting a MW dependence to inert gas exchange. This was further explored in another six dogs (group 2) with three gases of nearly equal solubility but widely divergent MWs (acetylene, 26; Freon-22, 86.5; isoflurane, 184.5). Isoflurane and Freon-22 excretions were depressed 47 and 30%, respectively, relative to acetylene. In a theoretical model of airway gas exchange, neither a tissue nor a gas phase diffusion resistance predicted our results better than the standard equation for steady-state alveolar inert gas elimination. However, addition of a simple ln (MW) term reduced the remaining residual sum of squares by 40% in group 1 and by 83% in group 2. Despite this significant MW influence on tracheal gas exchange, we calculate that the quantitative gas exchange capacity of the conducting airways in total can account for less than or equal to 16% of any MW-dependent differences observed in pulmonary inert gas elimination.     

Alveolar slope and dead space of He and SF6 in dogs: comparison of airway and venous loading

Series (Fowler) dead space (VD) and slope of the alveolar plateau of two inert gases (He and SF6) with similar blood-gas partition coefficients (approximately 0.01) but different diffusivities were analyzed in 10 anesthetized paralyzed mechanically ventilated dogs (mean body wt 20 kg). Single-breath constant-flow expirograms were simultaneously recorded in two conditions: 1) after equilibration of lung gas with the inert gases at tracer concentrations [airway loading (AL)] and 2) during steady-state elimination of the inert gases continuously introduced into venous blood by a membrane oxygenator and partial arteriovenous bypass [venous loading (VL)]. VD was consistently larger for SF6 than for He, but there was no difference between AL and VL. The relative alveolar slope, defined as increment of partial pressure per increment of expired volume and normalized to mixed expired-inspired partial pressure difference, was larger by a factor of two in VL than in AL for both He and SF6. The He-to-SF6 ratio of relative alveolar slope was generally smaller than unity in both VL and AL. Whereas unequal ventilation-volume distribution combined with sequential emptying of parallel lung regions appears to be responsible for the sloping alveolar plateau during AL, the steeper slope during VL is attributed to the combined effects of continuing gas exchange and ventilation-perfusion inequality coupled with sequential emptying. The differences between He and SF6 point at the contributing role of diffusion-dependent mechanisms in intrapulmonary gas mixing.     

Ventilation-perfusion lines and gas exchange in liquid breathing: theory

Alveolar exchange of a gas is governed by the ventilation-perfusion ratio (VA/Q) and the Ostwald partition coefficient for that species. We altered the Ostwald coefficients for O2 and CO2 by considering an animal breathing water or a fluorocarbon (FC-80) and studied the effects on gas exchange. Among our conclusions are the following. 1) When the ratio of the CO2 to O2 solubility in the inspirate exceeds the ratio of the O2 to the CO2 slope of the blood dissociation curve, as in water breathing, the VA/Q line becomes concave upward, and elements having a low VA/Q differ from each other more in terms of CO2 than of O2. 2) As the ratio of the CO2 to O2 solubility in the inspired medium increases, CO2 elimination becomes more dependent on perfusion. 3) At times, the same R will prevail in areas having different VA/Q values. 4) The alveolar-to-arterial O2 and CO2 differences resulting from a given VA/Q distribution do not depend on the O2 and CO2 solubility coefficients of the inspired medium, but on the inspired and mixed venous concentrations necessary to maintain adequate arterial gas levels in the presence of different inspired media.     

Can the stereoselective effects of the anesthetic isoflurane be accounted for by lipid solubility?

Isoflurane is an inhalational general anesthetic widely used in surgical operations as a racemic mixture of its two optical isomers. The recent availability of pure enantiomers of isoflurane has encouraged their use in experimental studies, and stereoselective effects have now been observed on anesthetic-sensitive neuronal ion channels. Although it has been assumed that such chiral effects demonstrate direct interactions with proteins, it is possible that they could be due to stereoselective interactions with chiral membrane lipids. We have determined the partition coefficients of the two optical isomers of isoflurane between lipid bilayers and water, using racemic isoflurane and gas chromatography with a chiral column. For lipid bilayers of phosphatidylcholine (PC) and 4 mol% phosphatidic acid (PA), both with and without cholesterol (CHOL), we found equal partitioning of the isoflurane optical isomers. The ratios of the S(+) to R(-) isoflurane partition coefficients were (mean +/- SEM): 1.018 +/- 0.010 for bilayers of PC/CHOL/PA (mole ratios 56:40:4) and 1.011 +/- 0.002 for bilayers of PC/PA (mole ratio 96:4). Molar partition coefficients for racemic isoflurane were 49 +/- 4 and 165 +/- 10, respectively. These findings support the view that the stereoselective effects on ion channels observed with isoflurane are due to direct actions on proteins rather than lipids.     

19F-NMR spin-spin relaxation (T2) method for characterizing volatile anesthetic binding to proteins. Analysis of isoflurane binding to serum albumin.

This paper characterizes the low-affinity ligand binding interactions of a fluorinated volatile anesthetic, isoflurane (CHF2OCHClCF3), with bovine serum albumin (BSA) using 19F-NMR transverse relaxation (T2). 19F-NMR spectra of isoflurane in aqueous BSA reveal a single isoflurane trifluoromethyl resonance, indicative of rapid exchange of isoflurane between protein-bound and aqueous (free) environments. The exchange is slow enough, however, that the chemical shift difference between bound and free isoflurane (delta omega = 0.545 ppm) contributes to the observed isoflurane T2. The contribution of delta omega to T2 can be minimized by shortening the interval between 180 degrees refocusing pulses in the Carr-Purcell-Meiboom-Gill pulse sequence used to monitor T2. Analysis of the dependence of T2 on interpulse interval additionally allows determination of the T2 (6.2 ms) and the average lifetime (tau b = 187 microseconds) of bound isoflurane molecules. By use of a short interpulse interval (less than 100 microseconds), T2 measurements can readily be used to analyze equilibrium binding of isoflurane to BSA. This analysis revealed a discrete saturable binding component with a KD = 1.4 mM that was eliminated either by coincubation with oleic acid (6 mol/mol of BSA) or by conversion of BSA to its "expanded" form by titration to pH 2.5. The binding was independently characterized using a gas chromatographic partition analysis (KD = 1.4 mM, Bmax = 3-4 sites). In summary, this paper describes a method whereby T2 measurements can be used to characterize equilibrium binding of low-affinity ligands to proteins without the confounding contributions of chemical shift.(ABSTRACT TRUNCATED AT 250 WORDS)     

Alaskan malamute chondrodysplasia--VII. Isolation and characterization of copper, zinc and iron binding proteins in Canis familiaris.

1. Three low molecular weight (12,000, 10,000 and 7,000) metal binding proteins have been isolated from the livers of normal and chondrodysplastic Alaskan Malamutes. 2. Comparison studies between kennel (mixed breed) dogs and both adult and immature Alaskan Malamutes suggested that the disturbance in trace mineral metabolism found in the Malamutes is almost entirely reflected in the 12,000 mol. wt species. 3. The major copper-inducible protein (10,000 mol. wt) observed in kennel dogs was not found to be inducible in Malamutes and contained constant ratios of both copper and zinc to protein in metal binding proteins isolated from the livers of both normal and dwarf Malamutes. 4. The copper and zinc found in the UM2 concentrates (mol. wt greater than 2000) of immature Malamutes showed very little affinity to the proteins and these metals were found chiefly in a peptide fraction which apparently serves as a reservoir from which the storage proteins obtain the metals that they bind. 5. Regression analysis indicated a statistically significant correlation between both copper and zinc concentrations and the carbohydrate concentration in the proteins investigated.     

Carbon dioxide-oxygen relationships in gas exchange of animals. In memory of Hermann Rahn.

In external gas exchange of vertebrates, behavior of the respiratory gases CO2 and O2 can in many cases adequately be explained by the different physico-chemical properties of the gases, including solubility, chemical combination in blood and tissue, and diffusivity. In particular, the differences in behavior between CO2 and O2 are often of particular relevance. This is demonstrated on a number of examples of gas exchange mechanisms in vertebrates, including (1) exchange ratio after changes in ventilation, (2) local variations of pulmonary ventilation/perfusion ratio, (3) absorption of gas from gas pockets, (4) water vs. air breathing, (5) multimodal breathing, (6) skin breathing, (7) gas exchange of avian eggs, (8) anomalous gas/blood CO2 equilibration, (9) blood/gas CO2 equilibration in avian lungs, (10) pulmonary diffusing capacity, (11) blood/water CO2 equilibration in fish gills, (12) deposition of gas into fish swim bladder.     

Lobar contribution to VA/Q inequality during constant-flow ventilation

Previous studies have shown that normal arterial PCO2 can be maintained during apnea in anesthetized dogs by delivering a continuous stream of inspired ventilation through cannulas aimed down the main stem bronchi, although this constant-flow ventilation (CFV) was also associated with a significant increase in ventilation-perfusion (VA/Q) inequality, compared with conventional mechanical ventilation (IPPV). Conceivably, this VA/Q inequality might result from differences in VA/Q ratios among lobes caused by nonuniform distribution of ventilation, even though individual lobes are relatively homogeneous. Alternatively, the VA/Q inequality may occur at a lobar level if those factors causing the VA/Q mismatch also existed within lobes. We compared the efficiency of gas exchange simultaneously in whole lung and left lower lobe by use of the multiple inert gas elimination technique in nine anesthetized open-chest dogs. Measurements of whole lung and left lower lobe gas exchange allowed comparison of the degree of VA/Q inequality within vs. among lobes. During IPPV with positive end-expiratory pressure, arterial PO2 and PCO2 (183 +/- 41 and 34.3 +/- 3.1 Torr, respectively) were similar to lobar venous PO2 and PCO2 (172 +/- 64 and 35.7 +/- 4.1 Torr, respectively; inspired O2 fraction = 0.44 +/- 0.02). Switching to CFV (3 l.kg-1.min-1) decreased arterial PO2 (112 +/- 26 Torr, P less than 0.001) and lobar venous PO2 (120 +/- 27 Torr, P less than 0.01) but did not change the shunt measured with inert gases (P greater than 0.5).(ABSTRACT TRUNCATED AT 250 WORDS)     

Validation of measurements of ventilation-to-perfusion ratio inequality in the lung from expired gas.

The analysis of the gas in a single expirate has long been used to estimate the degree of ventilation-perfusion (Va/Q) inequality in the lung. To further validate this estimate, we examined three measures of Va/Q inhomogeneity calculated from a single full exhalation in nine anesthetized mongrel dogs under control conditions and after exposure to aerosolized methacholine. These measurements were then compared with arterial blood gases and with measurements of Va/Q inhomogeneity obtained using the multiple inert gas elimination technique. The slope of the instantaneous respiratory exchange ratio (R slope) vs. expired volume was poorly correlated with independent measures, probably because of the curvilinear nature of the relationship due to continuing gas exchange. When R was converted to the intrabreath Va/Q (iV/Q), the best index was the slope of iV/Q vs. volume over phase III (iV/Q slope). This was strongly correlated with independent measures, especially those relating to inhomogeneity of perfusion. The correlations for iV/Q slope and R slope considerably improved when only the first half of phase III was considered. We conclude that a useful noninvasive measurement of Va/Q inhomogeneity can be derived from the intrabreath respiratory exchange ratio.     

Use of holographic laser interferometry to study the diffusion of polymers in gels

The aim of this study was to demonstrate the potential for holographic interferometry to be used for diffusion studies of large molecules in gels. The diffusion and partitioning of BSA (67,000 g/mol) and pullulans (5,900-112,000 g/mol) in agarose gel were investigated. The gel diffusion coefficients obtained for BSA were higher when distilled water was used as a solvent compared to those obtained with 0.1 M NaCl as the solvent. Furthermore, the gel diffusion coefficient increased with increasing BSA concentration. The same trend was found for liquid BSA diffusion coefficients obtained by DLS. BSA partition coefficients obtained at different agarose gel concentrations (2-6%, w/w) decreased slightly with increasing gel concentration. However, all BSA gel diffusion coefficients measured were significantly lower than those in pure solvent and they decreased with increasing agarose concentration. The gel diffusion coefficients obtained for pullulans decreased with increasing pullulan molecular weight. The same effect from increased molecular weight was seen in the liquid diffusion coefficients measured by DLS. The pullulan partition coefficients obtained decreased with increasing molecular weight. However, pullulans with a larger Stokes' radius than BSA had partition coefficients that were higher or approximately the same as BSA. This implied that the pullulan molecules were more flexible than the BSA molecules. The results obtained for BSA in this study agreed well with other experimental studies. In addition, the magnitude of the relative standard deviation was acceptable and in the same range as for many other methods. The results thereby obtained showed that holographic interferometry is a suitable method for studying diffusion of macromolecules in gels.     

Blood viscosity and cardiac output in acute experimental anemia.

The significance of blood viscosity alterations during anemia was evaluated in dogs under morphine-chloralose anesthesia. In group I, anemia (mean hematocrit 18.1 +/- 1.3 vol %) was produced by exchange transfusion with clinical dextran (avg mol wt 70,000). In group II, anemia was produced (mean hematocrit 19.9 +/- 0.88 vol %) with 500,000 molecular weight dextran, thus preventing the decrease in blood viscosity in group I. The cardiac output increase in group I (93.4%) with low-viscosity anemia was significantly greater than in group II (43.3%) with unchanged blood viscosity. Group III animals were transfused with a clinical dextran-red cell mixture, and group IV animals received a 500,000 mol wt dextran-red cell mixture. In group III, blood viscosity and cardiac output did not change. In group IV, blood viscosity rose and cardiac output fell significantly. The results suggest that a change in blood viscosity exerts a significant effect upon cardiac output, especially during acute dextran-exchange anemia.     

Determination of Natural In Vivo Noble-Gas Concentrations in Human Blood

Although the naturally occurring atmospheric noble gases He, Ne, Ar, Kr, and Xe possess great potential as tracers for studying gas exchange in living beings, no direct analytical technique exists for simultaneously determining the absolute concentrations of these noble gases in body fluids in vivo. In this study, using human blood as an example, the absolute concentrations of all stable atmospheric noble gases were measured simultaneously by combining and adapting two analytical methods recently developed for geochemical research purposes. The partition coefficients determined between blood and air, and between blood plasma and red blood cells, agree with values from the literature. While the noble-gas concentrations in the plasma agree rather well with the expected solubility equilibrium concentrations for air-saturated water, the red blood cells are characterized by a distinct supersaturation pattern, in which the gas excess increases in proportion to the atomic mass of the noble-gas species, indicating adsorption on to the red blood cells. This study shows that the absolute concentrations of noble gases in body fluids can be easily measured using geochemical techniques that rely only on standard materials and equipment, and for which the underlying concepts are already well established in the field of noble-gas geochemistry.     

Mechanisms of gas exchange with different gases during constant-flow ventilation

To investigate the mechanisms responsible for the difference in gas exchange during constant-flow ventilation (CFV) when using gases with different physical properties, we used mixtures of 70% N2-30% O2 (N2-O2) and 70% He-30% O2 (He-O2) as the insufflating gases in 12 dogs. All dogs but one had higher arterial PCO2 (PaCO2) with He-O2 compared with N2-O2. At a flow of 0.37 +/- 0.12 l/s, the mean PaCO2's with N2-O2 and He-O2 were 41.3 +/- 13.9 and 53.7 +/- 20.3 Torr, respectively (P less than 0.01); at a flow rate of 0.84 +/- 0.17 l/s, the mean PaCO2's were 29.1 +/- 11.3 and 35.3 +/- 13.6 Torr, respectively (P less than 0.01). The chest was then opened to alter the apposition between heart and the lungs, thereby reducing the extent of cardiogenic oscillations by 58.4 +/- 18.4%. This intervention did not significantly alter the difference in PaCO2 between N2-O2 and He-O2 from that observed in the intact animals, although the individual PaCO2 values for each gas mixture did increase. When the PaCO2 was plotted against stagnation pressure (rho V2), the difference in PaCO2 between N2-O2 and He-O2 was nearly abolished in both the closed- and open-chest animals. These findings suggest that the different PaCO2's obtained by insufflating gases with different physical properties at a fixed flow rate, catheter position, and lung volume result mainly from a difference in the properties of the jet.     

Nucleocytoplasmic exchange of macromolecules

Quantitative measurements of the cytoplasm-to-nucleus exchange of specific protein tracers were correlated with known physical properties (size and electrical charge) of the proteins. Tracers differing in their molecular parameters were produced by fluorescence labelling of wellcharacterized proteins (bovine serum albumin, mol. wt 67 500; ovalbumin, mol. wt 45 000; myoglobin, mol. wt 17 500; lysozyme, mol. wt 14 500; and cytochrome c, mol. wt 13 000) with fluorescein isothiocyanate. The labelled proteins were microinjected into the cytoplasm of living cells, and their uptake into the nucleus was followed by quantitative fluorescence microscopy. In addition, the distribution of cytoplasmically injected ferritin (mol. wt 465 000) was observed with the electron microscope.     

Isolation and characterization of a dog serum lipoprotein having apolipoprotein A-I as its predominant protein constituent.

The serum high density lipoproteins (HDL) of normolipemic dogs (beagles) were isolated in the density range of p 1.063 to 1.21 g/ml, and characterized in terms of composition and physical properties (flotation and diffusion coefficients, partial specific volume, molecular weight, electrophoretic mobility, ultraviolet absorption, and circular dichroism). The results indicated that canine HDL is a relatively homogeneous class with a molecular weight of about 230 000 and general properties similar to those reported for human HDL. After delipidation, the resulting apolipoprotein, apo-HDL, was fractionated by Sephadex G-200 column chromatography in urea or guanidine hydrochloride solutions. About 90% of the apo-HDL consisted of a protein with a molecular weight of about 28 000, similar in amino acid composition to human apolipoprotein A-I and having the same NH2 terminus (aspartic acid) and COOH terminus (glutamine) and no carbohydrates. Two other proteins were isolated, one having an apparent mol wt of 55 000 and representing, at least in part, an aggregate of apolipoprotein A-I and the other component with a mol wt of 8000, not yet characterized. The results indicate that canine HDL, as an intact complex, has general physical properties that lie between those reported for human HDL2 and HDL3, and that it differs compositionally from the human products mainly in its predominant content of apo-A-I. These findings together with evidence for the relatively homogeneous nature of the canine HDL provide new prospects for unraveling the relationship between polypeptide composition and HDL structure.     

Estimation of cardiac output by analysis of respiratory gas exchange.

Cardiac output is estimated by least squares fitting of a model of pulmonary gas exchange to measurements of respiratory gas composition obtained with a mass spectrometer during a rebreathing maneuver. This new technique estimates cardiac output on spontaneously breathing subjects at rest and requires neither central venous nor arterial blood samples. Principal features of the technique are the use of multiple gases simultaneously in the analysis, the use of a mathematical model for breath-to-breath evaluation of gas exchange, and simultaneous estimation of gas exchange and alveolar gas tensions with the same instrumentation. The technique is compared with thermal dilution estimates in dogs before and during hemorrhagic shock. Two-thirds of these estimates were within 20% of one another. The standard deviation of replication was 15%. Shortcomings, possibilities for improvement, and possible applications are discussed.     

Constant-flow ventilation in pigs

Constant-flow ventilation (CFV) is a ventilatory technique in which physiological blood gases can be maintained in dogs by a constant flow of fresh gas introduced via two catheters placed in the main-stem bronchi (J. Appl. Physiol. 53: 483-489, 1982). High-velocity gas exiting from the catheters can create uneven pressure differences in adjacent lung segments, and these pressure differences could lead to gas flow through collateral channels. To examine this hypothesis, we studied CFV in pigs, animals known to have a high resistance to collateral ventilation. In three pigs we examined steady-state gas exchange, and in six others we studied unsteady gas exchange at three flow rates (20, 35, and 50 l/min) and three catheter positions (0.5, 1.5, and 2.5 cm distal to the tracheal carina). During steady-state runs we were unable to attain normocapnia; the arterial CO2 partial pressure (PaCO2) was approximately 300 Torr at all flow rates and all catheter positions, compared with 20-50 Torr at similar flows and positions in dogs studied previously. The initial unsteady gas-exchange experiments indicated no consistent effect of catheter position or flow rate on the rate of rise of PaCO2. In three other pigs, the rates of rise of PaCO2 were compared with the rates observed with apneic oxygenation (AO). At the maximum flow and deepest position, the rate of rise of PaCO2 was lower during CFV than during AO. These data suggest that flow through collateral channels might be important in producing adequate gas transport during CFV; however, other factors such as airway morphometry and the effects of cardiogenic oscillations may explain the differences between the results in pigs and dogs.     

Information content of multiple inert gas elimination measurements

The multiple inert gas elimination technique provides a fundamental assessment of the distribution of ventilation-perfusion (VA/Q) ratios in the lung. The resolution of the finer structure of this distribution is limited however. This study examines the theoretical basis of this limitation and presents an objective method for evaluating the independence of inert gas measurements. It demonstrates the linear dependence of the inert gas kernels and their filtering characteristics to be the factors most limiting information content. The limited number of gases available for measurement and experimental error are lesser limitations. At usual levels of experimental error, no more than seven different inert gases having partition coefficients between those of SF6 and acetone will provide independent information, and information content will be maximized by choosing gases with partition coefficients spaced equally on a logarithmic scale. A fivefold reduction in experimental error will not significantly alter the information content of the measurements. The analysis applies equally to other methods of multiple inert gas elimination data interpretation.     

Direct Test for Altered Gas Exchange Rates in Water stressed Soybean Nodules

Water stress usually lowers the nitrogenase activity of soybeanroot nodules. This loss in activity might result from an increasedbarrier to nodular gas exchange, from a general reduction inbiochemical function, or both. To test for the possibility ofan increased barrier to gas diffusion, we measured the apparentlag time for initiation of acetylene reduction by intact soybeanplants, both before and after water stress. Mild nodular waterloss (i.e. 10% of fresh weight or less) seldom altered the apparentlag time, whereas severe water stress (20–40% f. wt loss)frequently produced a small increase in apparent lag time. Severewater stress also produced a large decrease (24%) in the externaldiameter of the nodules and a loss of the white lenticel traces.Water stress usually caused a decrease in the apparent K_m foracetylene. The data do not suggest a large change in the diffusiveresistance to acetylene of nodules subjected to water stress.Thus, the observed decrease in nitrogenase activity may resultprimarily from biochemical, rather than physical, changes. However,because of the relatively greater importance of gas-phase diffusionfor oxygen entry, we cannot exclude the possibility of a largechange in a small gas pathway that affects oxygen influx morethan acetylene influx. Diffusion, Glycine max, nitrogen fixation, water stress     

A Simple Open Flow System Used to Measure Acetylene Reduction Activity of Sesbania rostrata Stem and Root Nodules

A simple method for mixing gases for the assay of acetylenereduction using an open flow system is described. Gas mixturesare obtained using simple regulator valves (described) to obtainequal gas pressures and then providing a resistance to gas flowproportional to the flow of gas required. The system describedallows any number of gases to be mixed over a wide range ofratios, at a wide range of flow rates. Using this system, theactivity of Sesbania rostrata stem and root nodules was examinedin undisturbed plants. Nodules in either location had high ratesof acetylene reduction (270 µmol C₂H₄ g