Acceleration of plasma bicarbonate conversion to carbon dioxide by pulmonary carbonic anhydrase.

In isolated rabbit lungs perfused with solutions containing little or no carbonic anhydrase activity, nearly complete equilibration between H14CO3- and 14CO2 occurs during a single circulation. This equilibration can be inhibited by blocking pulmonary carbonic anhydrase with acetazolamide.

Osmotic extraction of hypotonic fluid from the lungs.

After injections of sucrose, NaCl, and urea solutions, the flow of tissue fluid from the lungs amounted to 0.182, 0.216, and 0.152 x 10(-3) ml/s per mosmol/kg of concentration difference between plasma and tissues in each gram of wet tissue weight. The extracted fluid contained less than 20% of the Na(+), K(+) and urea concentrations of the plasma. It was concluded that this fluid was distinctly hypotonic in comparision with the fluids of the plasma and tissue compartments both before and after the injection of hypertonic solutions. The presence of low solute concentrations in the extracted fluid is attributed to the passage of this fluid across cellular membranes, which are relatively impermeable to small hydrophilic solutes. Movement of fluid out of the junctions appears to be less than that through the endothelial cells. It is suggested that the injected solutes rapidly leak into the junctions and consequently induce relatively little movement of water or tissue solutes out of the junctions. Concentrations of tritiated water and [(14)C]antipyrine in the extracted fluid are essentially the same as base-line plasma concentrations when the animals have been primed with these tracers. It is therefore likely that these tracers can readily traverse cellular membranes. Red cell transit through the lungs is impaired by hypertonic solutions of sucrose and NaCl with transient increases in pulmonary arterial hemoglobin concentrations of as much as 35% of base-line values.

The in vivo pH of the extravascular space of the lung.

The partition of 5,5-dimethyloxazolidine-2,4-dione (DMO) and of 11 amines between the vascular and extravascular spaces of the lung has been determined by the multiple indicator dilution technique. Four amines (nicotine, pentylamine, quinine, and benzylamine) were found to have pH-sensitive tissue to blood concentration ratios. Of these, tritiated nicotine appears to be the nost satisfactory indicator of tissue pH and values for the pH of the pulmonary extravascular space (pH(e)) have been calculated from the nicotine data. At an arterial pH (pH(art)) between 7.38 and 7.43 pH(e) averaged 6.69 +/-0.07. Changes in pH(e) usually paralleled but were consistently less than concomitant changes in pH(art). Alterations in P(CO2) at constant pH(art) regularly produced relatively small, parallel changes in extravascular hydrogen ion concentrations. Local alterations in tissue pH due to P(CO2) changes are apparently buffered quite rapidly and the pH(e) of the lung seems more closely linked to pH(art) than the cellular pH of other tissues.DMO, guanidine, methylamine, morphine, and atropine were confined to the vascular volume during the first circulation and could not be used to measure tissue pH. Histamine appeared to be bound to a pH-insensitive site. The extravascular distributions of antipyrine and aniline were unresponsive to alterations in arterial pH, presumably because they are essentially uncharged at pH levels found in the lung.

Deficiency of carbonic anhydrase in the vasculature of rabbit kidneys.

The transit of 14CO2 and H14CO3- through the renal vasculature was studied in rabbit kidneys perfused without erythrocytes and in an in vivo preparation in which erythrocytes were present. In the absence of erythrocytes, the transit of 14CO2 from the renal artery to renal vein was much more rapid than that of H14CO3-. This suggests that (a) there is insufficient carbonic anhydrase (c.a.) in the vasculature between the renal artery and the exchange vessels of the kidney to ensure equilibration between CO2 and HCO3- and (b) CO2 can diffuse directly between arterial and venous vessels in the kidney. Following infusions of carbonic anhydrase, the renal venous outflow patterns of 14CO2 and H14CO3- became the same in the perfused kidneys. Although the initial recovery of 14CO2 remained greater than that of H14CO3- after infusions of acetazolamide (a c.a. inhibitor), arteriovenous diffusion of 14CO2 was diminished by this agent. This is attributed to inhibition of renal tubular c.a. The outflow patterns of H14CO3- and 14CO2 were nearly the same in the presence of erythrocytes, indicating that erythrocyte c.a. is sufficiently accessible to permit virtual equilibration of these radionuclides during the interval required for transit between the renal artery and exchange vessels. However, addition of carbonic anhydrase to the plasma seemed to accelerate transit of both 14CO2 and H14CO3- through the kidneys, and a small disequilibrium between CO2 and HCO3- may therefore normally be present in the renal interstitium and capillaries.

Reversible inhibition of urea exchange in rat hepatocytes.

Urea exchange is enhanced in renal collecting duct cells and erythrocytes by transporters which can be inhibited by phloretin and urea analogs such as thiourea. In this study, evidence for a comparable transporter was found in rat livers perfused with solutions which contained no red cells and in suspensions of hepatocytes. Bolus injections containing 125I-albumin (intravascular indicator), 99mTc-DTPA (extracellular indicator), 3HOH (water indicator), and [14C]urea were administered into the portal vein and fluid was collected from the hepatic vein. Under control conditions, [14C]urea and 3HOH emerged from the hepatic vein at nearly the same rate. However when the perfusate contained 2.5 mM phloretin (equivalent to 0.058 mM phloretin not bound to albumin), the amount of [14C]urea which had been recovered in the hepatic venous outflow by the time of peak 125I-albumin concentrations exceeded 3HOH recovery by a factor of 2.31 +/- 0.23 (n = 7). When the perfusate contained 200 mM thiourea, the comparable recovery of [14C]urea from the hepatic veins exceeded that of 3HOH by a factor of 3.48 +/- 0.44 (n = 7). These effects were at least partially reversible and suggested inhibition of urea transporters in hepatocytes. This conclusion was supported by studies of unloading of [14C]urea from hepatocytes which were exposed to unlabeled solutions: in the presence of phloretin, the amount of [14C]urea remaining within hepatocytes at 4 s was approximately twice that remaining in hepatocytes which had not been exposed to phloretin. Rapid transport of urea out of hepatocytes may increase urea synthesis and minimize cellular swelling due to urea accumulation.

In vivo myocardial cell pH in the dog. Response to ischemia and infusion of alkali.

Myocardial cell pH has been measured with 5,5-dimethyl-2,4-oxazolidinedione (DMO) in intact anesthetized dogs by a transient indicator dilution technique. Bolus injections of labeled DMO, vascular, extracellular, and water indicators were made into the anterior descending coronary artery, and blood samples were collected from the great cardiac vein. The steady-state distribution of DMO between cells and plasma was calculated from the indicator mean transit times, and the plasma pH. Myocardial cell pH was determined from the distribution value and plasma pH. Normal myocardial cell pH averaged 6.94. Changes in myocardial cell pH after infusions of acid or alkali. Myocardial ischemia induced by inflation of a coronary artery balloon resulted in progressive decreases in cellular pH to average values of 6.83 within the initial 15 min and to 6.59 within the interval between 20 and 70 min. Infusions of Na2CO3 tended to diminish intracellular acidosis although these infusions had little effect on the difference in pH between the myocardial cell and extracellular fluid.

Response of the lungs to aspiration.

Aspiration of acid from the stomach and water from the mouth can cause significant lung injury. Animal experiments suggest that acid entering the lungs is normally neutralized by bicarbonate derived from the plasma. It is hypothesized that this process may be impaired in patients with cystic fibrosis and that some of the airway injury that they experience may be related to this defect. This disease is characterized by abnormalities in the cystic fibrosis transmembrane conductance regulator, which normally conducts bicarbonate and chloride exchange. Evidence is discussed regarding the role of water channels (aquaporins) in transporting water from the airspaces into the vasculature.

Protection of the lungs from acid during aspiration.

Unlike the thick mucosa that normally covers the upper gastrointestinal tract, the membranes that cover the distal surfaces of the lungs are remarkably attenuated. This permits rapid exchange of gases between the airspaces and pulmonary vasculature, and may make the lungs more susceptible to acid challenges associated with acid reflux and aspiration. Any injury to the alveolar epithelium could result in the movement of solute and water into the airspaces (chemical pneumonia) and impair gas exchange. In this study, we used a fluorescent approach to compare the relative permeability of the apical basolateral surfaces of the lungs to the exchange of the ionic forms of acids and bases. The apical membranes proved to be much less permeable to NH(4)(+) and HCO(3)(+) than the basolateral membranes. This asymmetry in permeability should enhance resistance of the epithelium to inspired acidic challenges by slowing entry of acid into the cells and by linking the intracellular pH of the alveolar cells to that of the plasma, which is a relatively large, well-buffered compartment. Evidence also was obtained that the acid is secreted by the membranes covering the lungs.

Myocardial ischemia and cell acidosis: Modification by alkali and the effects on ventricular function and cation composition.

Myocardial cell pH was measured with 5, 5 dimethyl-2, 4-oxazolidinedione (DMO) in intact anesthetized dogs by a transient indicator dilution technique. Bolus injections of labeled DMO, vascular, extracellular and water indicators were made into the left anterior descending coronary artery, and blood samples were collected from the great cardiac vein. The steady state distribution of DMO between cells and plasma was calculated from the mean transit times of the indicator. Normal myocardial cell pH averaged 6.94 and changed by 58% of the concomitant alterations in plasma pH after infusions of acid or alkali. Myocardial ischemia induced by inflation of a balloon tip catheter in the left anterior descending coronary artery resulted in progressive decreases in cell pH to 6.59 by 1 hour. Infusions of sodium carbonate diminished intracellular acidosis. Hemodynamic studies during 4 hours of ischemia with blood pH at 7.55 to 7.60 indicated a significantly reduced left ventricular end-diastolic pressure and increased stroke volume by comparison with findings in animals given infusions of saline solution. Ventriculograms revealed improved wall motion in the ischemic segment after infusion of alkali. Precordial mapping showed a significant reduction in the number of leads with S-T segment elevation as well as in the sum of S-T segment elevations, but R wave amplitudes did not differ from those in control studies. Calculations of extracellular space, tissue water and cation content revealed a reduced gain of cell sodium ion and loss of cell potassium ion during ischemia after alkali treatment. The latter may account for the S-T segment responses, whereas enhanced ventricular performance may be related to reduced competition of hydrogen ion with calcium ion for binding sites on contractile protein.

Small solute clearance from the lungs of patients with cardiogenic and noncardiogenic pulmonary edema.

The regional clearance of 99mTc-diethylenetriamine penta-acetate (99mTc-DTPA) from the lungs was measured in 14 patients with noncardiogenic pulmonary edema, six patients with acute pulmonary edema secondary to heart failure, and 29 normal subjects. The radionuclide was delivered in an aerosol which was inhaled for 120 seconds, and the subsequent decline of radioactivity from the lungs was monitored for seven minutes over each of six peripheral regions of interest with a computerized scintillation camera. The average 99mTc-DTPA clearance of these regions was accelerated above the 98 percent confidence limits in all but three of the patients with noncardiogenic edema. The mean clearance value in this group of patients was significantly greater than those in normal subjects or patients with cardiogenic pulmonary edema. Clearances returned toward normal in each of seven subjects who improved clinically. Only one of the patients with cardiogenic pulmonary edema had an elevated average clearance rate, and the mean clearance for this population was not statistically greater than normal. This procedure appears to detect increased epithelial permeability caused by lung injury and may help distinguish between cardiogenic and noncardiogenic pulmonary edema.

Rapidly reversible alterations of pulmonary epithelial permeability induced by smoking.

A radioaerosol procedure using 99mcTc-DTPA (diethylene triamine penta acetate) was used to evaluate the permeability of the pulmonary epithelium in smokers and nonsmokers. The average clearance of this indicator from the lungs of smokers without significant airway obstruction exceeded that found in normal subjects by an average factor of more than five. This abnormality was observed throughout all lung regions. 99mTc-DTPA clearance decreased rapidly during the week after smoking was discontinued. It is concluded that smoking results in a rapidly reversible increase in pulmonary epithelial permeability.

Lung water measurements with the mean transit time approach.

The potential usefulness and limitations of the double-indicator mean transit time approach for measuring lung water are evaluated from both theoretical and empirical points of view. It is concluded that poor tissue perfusion is the most serious factor that can compromise the reliability of this approach. Replacement of the conventional water isotopes with a thermal signal enhances indicator delivery to ischemic areas but the diffusion of heat is not sufficiently rapid to permit measurements of water in macroscopic collections of fluid which remain unperfused. The frequency of pulmonary vascular obstruction in patients with pulmonary edema related to lung injury suggests that interpretation of transit time data will be complicated by uncertainties concerning perfusion. Thermal-dye measurements of lung water may prove more helpful in situations where pulmonary blood flow remains relatively uniform.

CO2 and bicarbonate exchange in the rat liver.

Several forms of carbonic anhydrase (CA) have been detected in hepatocytes. The distribution of these enzymes appears to be heterogeneous in the hepatic lobule, and the specific isoenzyme that predominates is influenced by sex steroid levels in the animal. In the present study, experiments were conducted in isolated male rat livers perfused with erythrocyte-free solutions, which were devoid of CA to see if there were sufficient tissue CA activity accessible to the plasma to ensure equilibration between labeled HCO3- and CO2 during a single passage from the portal vein to the hepatic vein. After injection of H14CO3- into the portal vein, emergence of the 14C label from the hepatic vein was slightly more rapid than after injections of 14CO2. After infusion of 5-250 microM of acetazolamide, an inhibitor of CA, H14CO3- was virtually confined to the extracellular space during a single transit through the organ, whereas the outflow of 14CO2 was very prolonged, suggesting that some of the 14C had been "trapped" within the hepatic cells as H14CO3-. Inhibition of CA activity in the intact organ with low doses of acetazolamide suggests the presence of a readily inhibitable isoenzyme of CA on the surface of the hepatocytes, which is directly accessible to both HCO3- and acetazolamide. The outflow patterns of 14CO2 and H14CO3- became the same after infusion of erythrocyte CA into the portal vein. On the basis of the pH of the perfusate and the cellular distribution of 14CO2 and H14CO3- in the presence of CA, an intracellular pH value of 7.26 was calculated.(ABSTRACT TRUNCATED AT 250 WORDS)

Solute exchange between the plasma and epithelial lining fluid of rat lungs.

Although the transport of solutes from air spaces to plasma has been extensively studied, comparatively little information is available concerning solute equilibration between the plasma and the epithelial lining fluid (ELF) of air-filled lungs. In the present study, 11 lipophobic indicators varying in molecular mass between 22 and 80,000 Da were injected intravenously and/or intramuscularly into anesthetized rats in a manner designed to keep blood concentrations constant. The animals were killed by rapid lavage of their lungs at various intervals up to 120 min after the injections had been made. Indicator concentrations in the bronchoalveolar lavage (BAL) fluid and plasma were determined, and BAL-to-plasma concentration ratios were calculated for indicators that were injected (exogenous: [14C]urea, 22Na+, [3H]mannitol, 99mTc-diethylenetriaminepentaacetate (a chelate), 51Cr-(ethylene dinitrilo)tetraacetate (a chelate), 113mIn-transferrin, human albumin, and Evans blue-labeled rat albumin) and those that were already present from the plasma and ELF (unlabeled urea, rat albumin, and rat transferrin). Leakage of exogenous indicators in the blood into the BAL fluid was observed during the lavage procedure. Leakage of [14C]urea, 22Na+, and [3H]mannitol exceeded that of the heavier solute molecules. Diffusion of proteins and the labeled chelates into the ELF before lavage occurred at similar rates, suggesting vesicular transport. Use of rapidly diffusible solutes such as urea for determining dilution of ELF by BAL should be accompanied by intravascular injections of labeled solutes to correct for diffusion from the blood during lavage. Alternatively, labeled chelates or serum proteins can be used to estimate dilution of ELF by BAL. Interstitial sampling may be inevitable if the epithelium has been injured before lavage.

New evidence for active sodium transport from fluid-filled rat lungs.

The hypothesis that fluid reabsorption from the air spaces is mediated at least in part by active transport of Na+ was investigated in six sets of experiments conducted in isolated fluid-filled rat lungs. Fluid reabsorption was monitored by following the changes in the air space concentration of labeled albumin. We found that incorporation of bicarbonate rather than a nonvolatile buffer (N-2-hydroxy-ethylpiperazine-N'-2-ethanesulfonic acid) in the air space solution more than doubled the rate of fluid reabsorption. Addition of 10(-4) M amiloride to the air space solution reduced the rate of fluid reabsorption over a 2-h experiment from 1.2 +/- 0.1 to 0.7 +/- 0.1 ml and decreased reabsorption of both labeled and unlabeled Na+ from the air spaces. To show that Na+ could be reabsorbed from the air spaces even if the concentrations of Na+ in the perfusate increased above those in the air space, mannitol (150 mM) was added to the perfusate and air space solutions and the concentrations of Na+ and Cl- were reduced to 90 and 60 mM, respectively. Mannitol diffuses across the pulmonary epithelium very slowly, and it osmotically restrained the movement of water out of the air spaces. Na+ concentrations in the perfusate increased by 10 +/- 2 mM, but concentrations in the air space remained unchanged. Despite an increasingly unfavorable concentration gradient for Na+, 0.2 mmol Na+ and 0.6 ml water were reabsorbed from the air spaces in 2 h. Ouabain (10(-4) M) did not appear to slow fluid reabsorption in the presence of mannitol, but it reduced K+ secretion into the air spaces and increased K+ appearance in the perfusate in a manner consistent with inhibition of Na+-K+-adenosinetriphosphatase at the basolateral surface of the epithelial cells. Fluid reabsorption was not altered when the lungs were exposed to a hypotonic solution (185 mM), but secretion of K+ into the air spaces was accelerated and K+ was lost from the perfusate. These experiments are consistent with active Na+ transport from the air spaces.

Continuous measurements of changes in pulmonary capillary surface area with 201Tl infusions.

The impact of physiological and pathological processes on metabolism and transport of a variety of substances traversing the pulmonary vasculature depends in part on the capillary surface area available for exchange, and a reliable method for detecting changes in this parameter is needed. In this study, a continuous-infusion approach was used to investigate the response of the pulmonary capillary surface area to increases in flow and left atrial pressure. Isolated rat lungs were perfused with an acellular perfusion solution containing 125I-labeled albumin (an intravascular indicator) and 201Tl, a K+ analogue which is concentrated within lung cells. The extraction of 201Tl from the perfusate was 61% greater at low flow (8.5 ml/min) than at high flow (26 ml/min), and rapid changes in extraction were observed when flow was altered. In contrast, the permeability-surface area product was 76% greater when lungs were perfused at high flow than at low flow, suggesting comparable increases in pulmonary capillary surface area in these zone 2 lungs (airway pressure = 5 cmH2O, left atrial pressure < 0 cmH2O). In a second group of experiments, increases in left atrial pressure to 14 cmH2O (zone 3 lungs) at a constant flow of 8.5 ml/min increased the permeability-surface area product by only 18% despite increases in average intravascular pressure that were at least as high as those associated with high perfusion rates. 201Tl infusions provide a useful method for detecting and quantifying changes in pulmonary capillary surface area.

Abrupt changes in mixed venous blood gas composition after the onset of exercise.

It has been assumed that increases in both O2 uptake and ventilation occurring within the first few seconds after the onset of exercise cannot be the result of changes in blood gas composition reaching the central circulation because of the circulatory delay from the exercising limbs (A. Krogh and J. Lindhard, J. Physiol. Lond. 42: 112-136, 1913). We sought to validate this assumption by measuring the time course of pulmonary arterial blood gases during the transition from rest to exercise. Six healthy men underwent pulmonary arterial catheterization and then performed transitions from rest to moderate cycle ergometer exercise. An anaerobic sampling manifold withdrew 19 samples of blood during the rest-to-exercise transition; sampling interval was usually 4 s. Blood gas analysis showed that, on average, from rest-to-steady-state exercise, O2 saturation (Svo2) fell from 71 to 41% and mixed venous PCO2 (PvCO2) rose from 42 to 59 Torr. Contrary to our expectations, Svo2 decreased and PvCO2 increased with no discernible latency after exercise onset (by 10% and 2 Torr, respectively, within 6 s). The half time for the Svo2 decrease was 32 s, whereas for the PvCO2 increase it was 80 s. The time course of superior vena cava blood gas composition was determined in several experiments; no rapid changes after exercise onset were found. We conclude that at exercise onset there is a rapid fall in Svo2 and rise in PvCO2 well in advance of arrival of blood produced by exercising legs.(ABSTRACT TRUNCATED AT 250 WORDS)

Protein concentrations have little effect on reabsorption of fluid from isolated rat lungs.

A study was conducted to determine whether differences in the concentrations of large molecules between the air space and perfusate solutions altered the rates at which fluid was reabsorbed from isolated fluid-filled perfused rat lungs. Four groups of experiments were conducted: 1) 5 g/dl albumin in the air spaces and perfusate, 2) 15 g/dl albumin in the air space and 5 g/dl albumin in the perfusate, 3) 5 g/dl albumin in the air space and 15 g/dl albumin in the perfusate, and 4) a mixture of 5 g/dl albumin and 7 g/dl Dextran 70 in the air spaces and 5 g/dl albumin in the perfusate. Fluid reabsorption was determined by following the concentration of albumin labeled with Evans blue (T-1824) in the air space and perfusate compartments. Because leakage of protein between the air space and perfusate compartments is very slow, increases in T-1824 concentrations in the air spaces indicated loss of fluid from this compartment, whereas decreases in these concentrations in the perfusate compartment provided evidence of fluid transport into the vasculature. Approximately 30% of the air space fluid was reabsorbed in a 2-h period, and virtually all of this fluid reached the perfusate compartment. Despite oncotic differences that ranged from -65 to 65 Torr, variations in air space or perfusate albumin concentrations did not have a significant effect on this process. A 30% decrease in fluid reabsorption was observed when dextran was in the air space solution, but this decrease did not appear to be due to the oncotic properties of this solution because albumin did not have a measurable effect on reabsorption.(ABSTRACT TRUNCATED AT 250 WORDS)

Pulmonary epithelial sieving of small solutes in rat lungs.

Transport and consumption of glucose from the air spaces of isolated, fluid-filled lungs can result in significantly lower glucose concentrations in the air spaces than in the perfusate compartment (11). This concentration difference could promote the osmotic movement of water from the air spaces to the perfusate, but the rate of fluid extraction from the air spaces would then be limited by the rates of electrolyte transport through the epithelium. In the present study, measurements were made of solute and water losses from the air spaces of fluid-filled rat lungs and the transport of these solutes and water into the vasculature after addition of hypertonic glucose or sucrose to the perfusate. Increases in the concentrations of Na+, Cl-, K+, and labeled mannitol in the air space were initially comparable to those of albumin labeled with Evans blue. Similarly, decreases in electrolyte concentrations in the perfusate were comparable to those of labeled albumin, indicating that very little solute accompanied the movement of water out of the lungs. Nor was evidence found that exposure of the vasculature to hypertonic glucose resulted in an increase in the rate at which fluid was reabsorbed from the air spaces over a 1-h interval, aside from an initial, abrupt loss of solute-free water from the lungs. These observations suggest that perfusion of fluid-filled lungs with hypertonic solutions of small solutes results in the extraction of water from the air spaces and pulmonary parenchyma across membranes that resist the movement of electrolytes and other lipophobic solutes.