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Biomedical subjects

E D Crandall

Publications and source records attributed to E D Crandall.

At least 19 recordsLinked to original sources
PubMed1979

HCO3-/Cl- exchange across the human erythrocyte membrane: effects of pH and temperature.

Changes in extracellular pH (PH0) in red cell suspensions were monitored in a stopped-flow rapid reaction apparatus under conditions where dpH0/dt was determined by the rate of HCO3-/Cl- exchange across the membrane. Experiments were performed at 5 degrees C less than T less than 40 degrees C using either untreated cells or cells exposed to 0.11 mM SITS (4-acetamido-4'-isothiocyanostilbene-2,2'-disulfonic acid). Although SITS exposure reduced the rate of exchange by 90%, both untreated and SITS-treated cells are similarly affected by changes in pH0 and temperature. The rate of HCO3-/Cl- exchange exhibits a minimum at about pH0 5 and a maximum at about pH0 7.4 at all temperatures. A transition temperature of 17 degrees C was observed in the Arrhenius relationship for all pH0. The activation energies (Ea) in kcal/mol are 19.6 below and 11.7 above 17 degrees C for 5 less than pH0 less than 8. These findings, similar to those reported for Cl- self-exchange, suggest that: (i) a change in the rate-limiting step for HCO3-/Cl- exchange occurs at 17 degrees C, possibly due to an altered interaction between the transport pathway and membrane lipids; (ii) the carrier system can be titrated by either H+ or SITS from the outside of the membrane, but the untitrated sites continue to transport normally; (iii) the pH0 dependence of the rate of exchange is consistent with the titratable carrier having its most alkaline pK in the range expected for amino groups; and (iv) below pH0 5, the nature of the exchange is markedly altered.

4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfo↗
PubMed1979

Kinetics of bicarbonate/chloride exchange in dogfish erythrocytes.

A stopped-flow rapid reaction apparatus was used to monitor changes in extracellular pH in dogfish (Mustelus canis) erythrocyte suspensions under conditions where dpH/dt was determined by the rate of HCO3-/Cl- exchange across the red cell membrane. Experiments were performed on erythrocytes suspended either in their own plasma or in elasmobranch Ringer solution over a range of temperatures from 5 to 35 degrees C. The exchange fluxes at 25 degrees C for red blood cells suspended in their own plasma (2.03 nmol/cm2-s) or in Ringer solution (2.00 nmol/cm2-s) are not significantly different and can be compared to those obtained under similar conditions for human red cell suspensions (0.910 nmol/cm2-s). The flux for dogfish erythrocytes suspended in Ringer solution was reduced by 80% after exposure of the cells to SITS. An Arrhenius plot of the exchange rate constant yielded an activation energy of about 13.2 kcal/mol. We conclude that 1) plasma has no inhibitory effect of HCO3-/Cl- exchange across the dogfish erythrocyte membrane or on activity of intraerythrocyte carbonic anhydrase, 2) HCO3-/Cl- exchange probably occurs via the same mechanism in fish and mammalian erythrocytes, and 3) the conversion of plasms HCO3- to CO2 in dogfish can be catalyzed by intraerythrocyte carbonic anhydrase.

Animals↗
PubMed1979

Postcapillary pH disequilibrium after gas exchange in isolated perfused liver.

pH equilibration after gas exchange in a systemic vascular bed was investigated using an isolated guinea pig liver preparation perfused with a blood-free Krebs-Ringer-bicarbonate solution. Effluent perfusate was withdrawn into a stopped-flow apparatus in which pH and temperature were continuously monitored. A decrease in pH of the perfusate after exit from the liver was observed. Addition of acetazolamide to inflowing perfusate had little effect on this fall in effluent fluid pH, while the addition of carbonic anhydrase completely abolished the decrease in pH. These results suggest that: 1) after the addition of metabolically produced CO2, the reaction CO2 leads to H2CO3 leads to H+ + HCO3- in the perfusate did not reach equilibrium during passage through the liver; 2) acetazolamide had little effect on the magnitude of the pH disequilibrium; and 3) carbonic anhydrase accelerated the reaction to equilibrium. We conclude that little or no catalysis of the conversion of perfusate CO2 to H2CO3 takes place within the hepatic circulation and that the presence of slow postcapillary blood pH changes in vivo may be dependent on the specific species and organ vascular bed from which the blood exited after participating in gas exchange.

Acetazolamide↗
PubMed1979

Transport of water and solutes across sheep visceral pleura.

The fluid and solute transport properties of pleural tissue were studied using specimens of intact visceral pleura from adult sheep lungs. After thoracotomy, a shallow incision through the pleural surface permitted 10-cm2 by 10-micrometer pieces of visceral pleura free of lung parenchyma to be peeled off the lung surface. The pleura was then mounted as a planar sheet separating 2 reservoirs of Krebs-Ringer solution. Electrical potential and resistance, hydraulic water permeability, and diffusional permeability to water and several hydrophilic solutes were measured. The results showed that (1) no spontaneous voltage difference was present across the pleura; (2) electrical resistance (27.1 omega/cm2) was very low; (3) hydraulic water permeability was extremely high (1.64 X 10(-8) ml/dyne-s); and (4) diffusional permeability was high, varying from 5.24 X 10(-4) cm/s for water to 4 X 10(-5) cm/s for hemoglobin. Blue dextran (molecular weight, 2 X 10(6) daltons) did not cross the pleura in measurable quantities. We concluded that the isolated visceral pleura of the adult sheep is an extremely "leaky" tissue that probably does not actively transport salt and water. These findings are consistent with a passive model of pleural fluid formation and reabsorption, and suggest that the transport properties of normal pleural tissue are unlikely to be responsible for any differences in composition between interstitial and pleural fluids.

Animals↗
PubMed1978

Analysis of the effects of pulsatile capillary blood flow and volume on gas exchange.

Blood flow into the pulmonary capillaries and the volume of blood within the capillary bed are both pulsatile with the cardiac cycle. We have developed a quantitative model of diffusional gas exchange in the lung to investigate the effects of coupling between these two time-varying parameters on lung O2 and CO2 exchange. For normal man breathing room air at rest, the computed results agree well with previous predictions for the constant flow and volume case, and for the case of pulsatile flow alone. When coupled time-varying pulmonary capillary blood flow and volume are included, using the best data available in the literature to define these parameters, diffusional O2 exchange is improved over the cases of pulsatile flow or volume alone, and closely approximates that obtained for the hypothetical constant flow and volume case. CO2 exchanges, and O2 exchange during hypoxia, are not affected by pulsatile flow and/or volume. These results suggest that O2 exchange is efficient in the presence of coupled blood flow and blood volume pulsations as they exist in the lung capillaries, and that these conditions may be optimal for gas exchange under certain physiological (or pathological) conditions.

Blood Volume↗
PubMed1978

Bicarbonate-chloride exchange in erythrocyte suspensions. Stopped-flow pH electrode measurements.

A pH-sensitive glass electrode was used in a temperature-controlled stopped-flow rapid reaction apparatus to determine rates of pH equilibration in red cell suspensions. The apparatus requires less than 2 ml of reactants. The electrode is insensitive to pressure and flow variations, and has a response time of < 5 ms. A 20% suspension of washed fresh human erythrocytes in saline at pH 7.7 containing NaHCO(3) and extracellular carbonic anhydrase is mixed with an equal volume of 30 mM phosphate buffer at pH 6.7. Within a few milliseconds after mixing, extracellular HCO(3) (-) reacts with H(+) to form CO(2), which enters the red cells and rehydrates to form HCO(3) (-), producing an electrochemical potential gradient for HCO(3) (-) from inside to outside the cells. HCO(3) (-) then leaves the cells in exchange for Cl(-), and extracellular pH increases as the HCO(3) (-) flowing out of the cells reacts with H(+). Flux of HCO(3) (-) is calculated from the dpH/dt during HCO(3) (-)-Cl(-) exchange, and a velocity constant is computed from the flux and the calculated intracellular and extracellular [HCO(3) (-)]. The activation energy for the exchange process is 18.6 kcal/mol between 5 degrees C and 17 degrees C (transition temperature), and 11.4 kcal/mol from 17 degrees C to 40 degrees C. The activation energies and transition temperature are not significantly altered in the presence of a potent anion exchange inhibitor (SITS), although the fluxes are markedly decreased. These findings suggest that the rate-limiting step in red cell anion exchange changes at 17 degrees C, either because of an alteration in the nature of the transport site or because of a transition in the physical state of membrane lipids affecting protein-lipid interactions.

Bicarbonates↗
PubMed1978

Influence of pH on elastic deformability of the human erythrocyte membrane.

Fresh human blood was diluted 1:5000 in buffered saline-sucrose solution and titrated to a pH varying from 4.5 to 10.5 with 0.1 N HCl or 0.1 N NaOH. Circular regions of the membrane of individual cells were then deformed at 25 degrees C by aspiration into a micropipette having an internal tip diameter of 0.9-1.4 micron. A membrane surface elasticity modulus, mu (dyn/cm), was computed from the relationship between length of the aspirated membrane and the deforming pressure according to a two-dimensional membrane model. Surface elasticity increases with decreasing pH and with time after the cell suspension is acidified, rising several orders of magnitude with a t1/2 of 1--5 h as pH is lowered from 7.2 to 4.6. This increase in mu is only partially reversible. pH greater than 7.2 had little effect on mu. Membrane surface elasticity is not affected by variations in external [Ca2+] over the range of 0--50 mM, tonicity of the suspension medium from 275--400 mosM, or age of 0--50 h. Addition of 50 mM NaHCO3 to the medium increases the rate of change of mu at a given pH. These results suggest that the elastic properties of the red cell membrane are largely determined by interactions among structural proteins located on the cytoplasmic surface of the membrane and that these interactions are initiated by changes in intracellular pH.

Elasticity↗
PubMed1978

Analysis of postcapillary pH changes in blood in vivo after gas exchange.

A quantitative description of the reaction and transport processes that take place in blood during and after gas exchange in capillaries is developed and used to interpret recently reported experimental results. Included in the computation are 1) CO2-H2CO3 hydration-dehydration reactions in plasma and erythrocytes, 2) CO2 reactions with hemoglobin, 3) O2 binding to hemoglobin, 4) buffering of H+ intra- and extracellularly, 5) HCO3- Cl- exchange across the red cell membrane, 6) diffusion of gases between alveolar gas and blood, and 7) transcellular movement of water. Ion and water fluxes are described assuming passive diffusion down their electrochemical potential gradients. Recent data on the magnitude of the Bohr and Haldane shifts and on carbamate formation in the presence of 2,3-diphosphoglycerate are used. The analysis is used to examine the direction, magnitude, and time course of plasma pH changes in blood leaving the pulmonary capillaries and is shown to preduct results that agree very closely with recently reported experimental measurements in vivo. The time computed for plasma pH equilibration after gas exchange when carbonic anhydrase activity is absent from plasma is so great that blood may never be in complete electrochemical equilibrium as it travels around the circulation in normal man.

Bicarbonates↗
PubMed1978

Slow postcapillary changes in blood pH in vivo: titration with acetazolamide.

A stopped-flow pH electrode apparatus was used to investigate the mechanisms underlying slow changes in plasma pH (pHO) after blood leaves the pulmonary capillaries in carbonic anhydrase-inhibited animals. After acetazolamide was administered to an anesthetized dog or cat, arterial blood was withdrawn through the electrode apparatus into a syringe. Syringe movement was then suddenly stopped. Temperature and pHO of the blood in the electrode chamber were monitored both before and after blood withdrawal ceased. After stopping flow, pHO of the blood in the electrode chamber a) rose 0.02 after a dose of about 1 mg/kg acetazolamide; b) did not change after a dose of about 2 mg/kg acetazolamide; and c) fell 0.10 after a dose greater than about 5 mg/kg acetazolamide. With reasonable red cell and plasma carbonic anhydrase activities assumed for each dose level of acetazolamide, a computer model of the reaction and transport processes occurring in blood after gas exchange in the lung yielded predicted time courses of pHo that were in good agreement with the experimental results. The observed slow pHo changes are largely a result of disequilibrium of [H+] between red blood cells and plasma as blood leaves the pulmonary capillaries.

Acetazolamide↗
PubMed1978

Analysis of PCO2 differences during rebreathing due to slow pH equilibration in blood.

A quantitative analysis of the reaction and transport processes that occur in blood during and after gas exchange has been used to investigate mechanisms that might account for positive alveolar-mixed venous (A-V) and alveolar-arterial (Aa) PCO2 differences during rebreathing. The analysis was used to determine PCO2 changes that take place in blood as it travels from veins to arteries under conditions in which no CO2 is exchanged in the lung. The predicted A-V and Aa PCO2 differences are all positive and lie within the range of reported measured values. The differences are due to disequilibrium of [H+] between plasma and red blood cells, and to disequilibrium of the reactions CO2 in equilibrium HCO3- + H+ in plasma, as blood leaves the tissue and/or lung capillaries. The differences are increased with exercise and with continued O2 uptake in the lung, the latter due to the Haldane shift. We conclude that the two disequilibria and the Haldane shift contribute to the reported PCO2 differences in rebreathing animals but may not fully account for them. These mechanisms cannot explain any PCO2 differences that might exist during net CO2 elimination from blood in the lung.

Acid-Base Equilibrium↗
PubMed1978

Slow postcapillary pH changes in blood in anesthetized animals.

To investigate the hypothesis that blood pH and PCO2 continue to change after the blood leaves an exchange capillary, we used a rapidly responding, pressure-insensitive, stopped-flow pH electrode apparatus. Arterial blood from an anesthetized dog or cat is drawn through the apparatus into a syringe. Syringe movement is then suddenly stopped. Temperature and pH of the blood in the electrode assembly are continuously monitored, both before and after blood withdrawal ceases. Hemolysis was reduced by coating all blood contact surfaces with silicone and fasting the animal overnight, anesthetizing it with crystalline pentobarbital sodium, and allowing it to ventilate spontaneously. After stopping withdrawal, pH of blood in the electrode chamber continued to change, rising 0.01 unit with t1/2 of 4.4 s. After lysed blood was returned to the animal to provide carbonic anhydrase to the plasma, no pH change was seen after stopping the flow. The small pH rise occurring in arterial blood in vivo is probably due in large part to disequilibrium of [H+] between red blood cells and plasma at the end of the pulmonary capillary, the equilibration process being rate-limited by the extracellular CO2 hydration-dehydration reaction.

Acid-Base Equilibrium↗
PubMed1978

Direct evidence of participation of rat lung carbonic anhydrase in CO2 reactions.

Isolated rat lungs were ventilated with air and perfused with a blood-free Krebs-Ringer bicarbonate solution under conditions of net CO2 elimination in the lung. Some of the effluent perfusate was drawn through a stop-flow pH electrode apparatus, arriving at the electrode within 4 s after passing through the pulmonary capillaries. pH and temperature of the fluid in the electrode chamber were continuously monitored both before and after withdrawal was suddenly stopped. Little or no change was observed in the pH of the perfusate after flow was stopped, despite the fact that CO2 was eliminated in the lung, suggesting that the conversion of H2CO3 to CO2 in the blood-free perfusion fluid was markedly accelerated and the rise in pH was complete by the time the perfusate reached the electrode. Because the effluent perfusate was shown to be free of carbonic anhydrase activity, the catalysis must have occurred during transit through the isolated lung. When acetazolamide was added to the perfusate, a rise in the pH of the perfusate after stopping flow was consistently seen. These results suggest that the carbonic anhydrase of isolated lungs accelerates the conversion of H2CO3 to CO2 and enhances COW elimination as perfusate passes through the pulmonary capillaries, and that the enzyme may be present on the capillary endothelial surface.

Acetazolamide↗
PubMed1977

Postcapillary changes in blood pH in vivo during carbonic anhydrase inhibition.

A rapidly responding stopped-flow glass pH electrode apparatus was used to investigate pH changes in blood in vivo after it exits from an exchange capillary. Arterial blood was drawn from anesthetized animals through the apparatus. Temperature and pH of the blood in the electrode chamber were continuously recorded, both during withdrawal and after flow was stopped. Blood pH did not change after stopping flow in control experiments. When benzolamide (2 mg/kg) was given to inhibit carbonic anhydrase activity available to plasma (e.g., due to lysis) while having less effect on intracellular activity, pH increased 0.02-0.04 (t1/2 approximately 8 s) after stopping flow. Administration of acetazolamide (50 mg/kg) resulted in pH decreasing 0.07-0.10 (t1/2 approximately 15 s) after stopping flow. Ventilation for 1 min with N2 resulted in an increased rise in pH for the benzolamide-treated animals but a decreased fall in pH for the acetazolamide-treated animals. These shifts in arterial blood pH after gas exchange are largely due to disequilibrium of [H+] between red cells and plasma at the end of the pulmonary capillary.

Acetazolamide↗
PubMed1976

Kinetics of bicarbonate-chloride exchange across the human red blood cell membrane.

The kinetics of bicarbonate-chloride exchange across the human red cell membrane was studied by following the time course of extracellular pH in a stopped-flow rapid-reaction apparatus during transfer of H+ into the cell by the CO2 hydration-dehydration cycle, under conditions where the rate of the process was determined by HCO3--Cl- exchange flux across the membrane. The flux of bicarbonate increased linearly with [HCO3-] gradient from 0.6 to 20 mM across the red cell membrane at both 37 degrees C and 2 degrees C, and decreased as transmembrane potential was increased by decreasing extracellular [Cl-]. An Arrhenius plot of the rate constants for the exchange indicates that the Q10 is strongly dependent on temperature, being about 1.7 between 24 degrees C and 42 degrees C and about 7 between 2 degrees C and 12 degrees C. These data agree well with the published values for Q10 of 1.2 between 24 degrees C and 40 degrees C and of 8 between 0 degrees C and 10 degrees C. The results suggest that different processes may determine the rate of HCO3--Cl- exchange at low vs. physiological temperatures, and that the functional (and/or structural) properties of the red cell membrane vary markedly with temperature.

Bicarbonates↗
PubMed1976

A pressure- and flow-insensitive reference electrode liquid junction.

The design and construction of a pressure- and flow-insensitive reference liquid junction for use in ion concentration electrode measuring systems is described. The junction is inexpensive, is very easily and rapidly constructed, is rugged, and is adaptable to various applications. When used in a pH-measuring system, drift, pressure artifacts, and flow artifacts are negligible. The response time of the system appears to be less than 10 ms. Using the pH electrode device as described, the dissociation reaction rate constant of H2CO3 at 24 degrees C was determined to be 22 s-1.

Electrodes↗