Unchanged in vivo P50 at high altitude despite decreased erythrocyte age and elevated 2,3-diphosphoglycerate

We measured hematological and erythrocyte O2 transport parameters in whole blood and density-separated erythrocytes in 11 mountaineers before and during 5 days of exposure to high altitude (4,559 m). We determined the in vivo (arterial pHblood and PCO2) and standard (pHblood = 7.4, PCO2 = 40 Torr) O2 tension at 50% O2 saturation of hemoglobin and (P50,vv and P50,st) and Bohr coefficients (BC) for fixed acid (H+) and CO2 and examined the contribution of the altered average age of circulating erythrocytes due to the stimulation of erythropoiesis on whole blood 2,3-diphosphoglycerate (2,3-DPG) and P50,st. At altitude, whole blood P50,vv remained almost unchanged, whereas P50,st and 2,3-DPG increased significantly (+4 Torr; 3.5 mumol/g hemoglobin). BCCO2 was elevated significantly at altitude. Serum erythropoietin increased transiently fourfold, iron utilization increased, and serum iron decreased by 66%. Reticulocyte counts increased, but other hematological parameters were unchanged. In density-separated erythrocytes, P50,st and 2,3-DPG increased with decreasing cell density but were higher in fractions with comparable reticulocyte counts in cells prepared at altitude than in those from control studies. Our data show that, despite the increase in 2,3-DPG and the decrease in average erythrocyte age, the in vivo hemoglobin-O2 affinity remains unchanged. P50,st values reflect the elevation of 2,3-DPG, and approximately 50% of the increase in both parameters can be ascribed to the increase in the number of reticulocytes and young erythrocytes.     

The 1988 Stevenson Memorial lecture. Physiological responses to severe hypoxia in man

Recent measurements at extreme altitude and in low pressure chamber simulations have clarified the human responses to extreme hypoxia. Man can only tolerate the severe oxygen deprivation of great altitudes by an enormous increase in ventilation which has the advantage of defending the alveolar PO2 against the reduced inspired PO2. Nevertheless the arterial PO2 on the Everest summit is less than 30 Torr (1 Torr = 133.3 Pa). An interesting consequence of the hyperventilation is that the respiratory alkalosis greatly increases the oxygen affinity of the hemoglobin and assists in oxygen loading by the pulmonary capillary. The severe hypoxemia impairs the function of many organ systems including the central nervous system, and there is evidence of residual impairment of memory and manipulative skill in climbers returning from great altitudes. At the altitude of Mt. Everest, maximal oxygen uptake is reduced to 20-25% of its sea level value, and it is exquisitely sensitive to barometric pressure. It is likely that the seasonal variation of barometric pressure affects the ability of man to reach the summit without supplementary oxygen.     

Nocturnal periodic breathing at altitudes of 6,300 and 8,050 m

Nocturnal periodic breathing was studied in eight well-acclimatized subjects living at an altitude of 6,300 m [barometric pressure (PB) 350-352 Torr] for 3-5 wk and in four subjects during one night at 8,050 m altitude (PB 281-285 Torr). The measurements at 6,300 m included tidal volume by inductance plethysmography, arterial O2 saturation by ear oximetry (calibrated by arterial blood samples), electrocardiogram (ECG), and electrooculogram. At 8,050 m, periodic breathing was inferred from the cyclical variation in heart rate obtained from a night-long ECG record. All subjects at 6,300 m altitude showed well-marked periodic breathing with apneic periods. Cycle length averaged 20.5 s with 7.9 s apnea. Minimal arterial O2 saturation averaged 63.4% corresponding to a PO2 of approximately 33 Torr, i.e., approximately 6 Torr lower than the normal value at rest during daytime. This was probably the most severe hypoxemia of the 24-h period. At 8,050 m altitude, the cycle length averaged 15.4 s, much longer than predicted by a theoretical model. Cyclical variations in heart rate caused by periodic breathing occurred in all subjects, but abnormal cardiac rhythms such as ventricular premature contractions were uncommon. The severe arterial hypoxemia caused by periodic breathing may be an important determinant of tolerance to these great altitudes.     

Human responses to extreme altitudes

It is a strange coincidence that the highest point on Earthis very close to the limit of human tolerance to hypoxia. Thephysiological changes that allow humans to reach these extremealtitudes involve enormous alterations of their normal state.It is useful to contrast this response with two others to highaltitude. One is acclimatization that allows lowlanders to ascendto altitudes of up to 5000 m and remain there for an indefiniteperiod. The other is evolutionary adaptation which allows highlandersto live continuously over generations at altitudes up to 5000m. These two responses enable humans to survive for an indefiniteperiod at high altitude. By contrast, the changes that allowascent to extreme altitudes are not compatible with an extendedstay because of a poorly-understood process called high-altitudedeterioration. The most important physiological response toextreme altitude is extreme hyperventilation which, on the summitof Mt. Everest, drives the alveolar P_CO2 down to 7–8 mmHg.This is associated with a marked respiratory alkalosis withan arterial pH exceeding 7.7. Interestingly this alkalosis increasesthe oxygen affinity of hemoglobin, a response which the successfulclimber shares with many other animals in oxygen-deprived environments.The arterial P_O2 on the Everest summit is only about 30 mmHgand falls on exercise because of diffusion limitation of oxygenacross the blood-gas barrier. Maximal oxygen consumption onthe summit is just over 1 liter.min^–1. Anaerobic metabolismas measured by blood lactate levels is paradoxically reducedat extreme altitudes.     

Operation Everest II: preservation of cardiac function at extreme altitude

Hypoxia at high altitude could depress cardiac function and decrease exercise capacity. If so, impaired cardiac function should occur with the extreme, chronic hypoxemia of the 40-day simulated climb of Mt. Everest (8,840 m, barometric pressure of 240 Torr, inspiratory O2 pressure of 43 Torr). In the five of eight subjects having resting and exercise measurements at the barometric pressures of 760 Torr (sea level), 347 Torr (6,100 m), 282 Torr (7,620 m), and 240 Torr, heart rate for a given O2 uptake was higher with more severe hypoxia. Slight (6 beats/min) slowing of the heart rate occurred only during exercise at the lowest barometric pressure when arterial blood O2 saturations were less than 50%. O2 breathing reversed hypoxemia but never increased heart rate, suggesting that hypoxic depression of rate, if present, was slight. For a given O2 uptake, cardiac output was maintained. The decrease in stroke volume appeared to reflect decreased ventricular filling (i.e., decreased right atrial and wedge pressures). O2 breathing did not increase stroke volume for a given filling pressure. We concluded that extreme, chronic hypoxemia caused little or no impairment of cardiac rate and pump functions.     

Operation Everest II: oxygen transport during exercise at extreme simulated altitude

A decrease in maximal O2 uptake has been demonstrated with increasing altitude. However, direct measurements of individual links in the O2 transport chain at extreme altitude have not been obtained previously. In this study we examined eight healthy males, aged 21-31 yr, at rest and during steady-state exercise at sea level and the following inspired O2 pressures (PIO2): 80, 63, 49, and 43 Torr, during a 40-day simulated ascent of Mt. Everest. The subjects exercised on a cycle ergometer, and heart rate was recorded by an electrocardiograph; ventilation, O2 uptake, and CO2 output were measured by open circuit. Arterial and mixed venous blood samples were collected from indwelling radial or brachial and pulmonary arterial catheters for analysis of blood gases, O2 saturation and content, and lactate. As PIO2 decreased, maximal O2 uptake decreased from 3.98 +/- 0.20 l/min at sea level to 1.17 +/- 0.08 l/min at PIO2 43 Torr. This was associated with profound hypoxemia and hypocapnia; at 60 W of exercise at PIO2 43 Torr, arterial PO2 = 28 +/- 1 Torr and PCO2 = 11 +/- 1 Torr, with a marked reduction in mixed venous PO2 [14.8 +/- 1 (SE) Torr]. Considering the major factors responsible for transfer of O2 from the atmosphere to the tissues, the most important adaptations occurred in ventilation where a fourfold increase in alveolar ventilation was observed. Diffusion from alveolus to end-capillary blood was unchanged with altitude. The mass circulatory transport of O2 to the tissue capillaries was also unaffected by altitude except at PIO2 43 Torr where cardiac output was increased for a given O2 uptake. Diffusion from the capillary to the tissue mitochondria, reflected by mixed venous PO2, was also increased with altitude. With increasing altitude, blood lactate was progressively reduced at maximal exercise, whereas at any absolute and relative submaximal work load, blood lactate was higher. These findings suggest that although glycogenolysis may be accentuated at low work loads, it may not be maximally activated at exhaustion.     

Lactate during exercise at extreme altitude

Maximal exercise at extreme altitude results in profound arterial hypoxemia and, presumably, extreme tissue hypoxia. The best evidence available indicates that the resting arterial PO2 on the summit of Mount Everest is about 28 torr and that it falls even further during exercise. Nevertheless, some 10 climbers have now reached the summit without supplementary oxygen. Paradoxically, blood lactate for a given work rate at high altitude in acclimatized subjects is essentially the same as at sea level. Because work capacity decreases markedly with increasing altitude, maximal blood lactate also falls. Extrapolation of available data up to 6300 m indicates that a climber who reaches the Everest summit will have no increase in blood lactate. The cause of the low blood lactate during exercise at extreme altitude is not fully understood. One possibility is depletion of plasma bicarbonate in acclimatized subjects, which reduces buffering and results in large increases in H+ concentration for a given release of lactate. The consequent local fall in pH may inhibit enzymes, e.g., phosphofructokinase (EC 2.7.1.56), in the glycolytic pathway.     

Low P50 in deer mice native to high altitude

Whereas it is widely believed that animals native to high altitude show lower O2 partial pressures at 50% hemoglobin saturation (P50) than do related animals native to low altitude, that "fact" has not been well documented. Consequently, P50 at pH 7.4, PCO2(7.4), the CO2 Bohr effect, and the buffer slope (delta log PCO2/delta pH) were determined via the mixing technique in Peromyscus maniculatus native to a range of altitudes but acclimated to 340 or 3,800 m. PCO2(7.4) and buffer slope were substantially lower at high altitude. The change in P50(7.4) between acclimation altitudes was minimal (0.8% increase at 3,800 m), because of counterbalancing changes in PCO2, 2,3-diphospho-D-glycerate concentration, and perhaps other factors. At both acclimation altitudes there was a highly significant negative correlation between P50(7.4) and native altitude. Since pH in vivo probably increases slightly at high altitude, the data on P50 corrected to pH 7.4 are probably underestimates of the difference in in vivo P50 at low vs. high altitude. Hence these results corroborate theoretical predictions that low P50 is advantageous under severe hypoxic stress.     

Control of ventilation in extreme-altitude climbers

Ten climbers who participated in the Nepal-Japan Kangchenjunga Expedition (altitude, 8,478-8,586 m) in 1984 were examined for their hypercapnic and isocapnic hypoxic ventilatory responses (HCVR and HVR) at sea level before and after the expedition. Five climbers who reached an altitude higher than 8,000 m, [designated high-performance climbers (HPC)] exhibited significantly higher HVR than five climbers who did not [low-performance climbers (LPC)]. On the other hand, no significant difference in HCVR was seen between HPC and LPC. Our results were in agreement with the findings reported by Schoene et al. (J. Appl. Physiol. 56: 1478-1483, 1984) obtained in the American Medical Research Expedition to Everest in 1981. Significant depression in HVR in five climbers was found even 35-40 days after the expedition, which was accompanied by decreased arterial partial pressure of CO2 and increased pH at rest. Hence, the effect of altitude acclimatization in the climbers exposed to extreme altitude may have still persisted at the time of the postexpedition study. Our results confirmed that HRV evaluated at sea level may be used as an indicator of a climber's capability at great high altitude.     

P50 values in hypoxic albino rats of first and second generation.

We determined the "in vivo" (arterial pH and PCO2) and standard (pH = 7.4, PCO2 = 40 mm Hg) PO2 at 50% O2 saturation of hemoglobin (P50, vv and P50, st) in Wistar albino rats when living in a normobaric hypoxic environment. Two generations of hypoxic rats were observed for changes in their P50, vv, P50, st, (n50) 2,3-diphosphoglycerate (2,3-DPG), hemoglobin (Hb) and DPG-Hb ratio: the first generation (H1) and the second generation (H2). A few hours after birth, the H1 rats were placed and raised in a normobaric hypoxic environment (10% O2 in N2). The H2 rats were born from hypoxic parents of first generation and were raised in the same hypoxic environment. The control group had a normoxic environment. The P50, st was significantly higher in H1 rats than both H2 and controls. P50, st was similar in H2 and control rats. The P50, vv was significantly higher in H1 rats than both H2 and controls but it was significantly lower in H2 when compared with both controls and H1. Hb and 2,3-DPG had values significantly greater for both H1 and H2 when compared with their controls. However, the values of H2 were significantly lower than H1. The effectiveness of an increase in Hb-O2 affinity as an adaptive mechanism in H2 rats is discussed.     

Pulmonary vasoconstrictor response to acute decrease in blood P50

The O2 sensor that triggers hypoxic pulmonary vasoconstriction may be sensitive not only to alveolar hypoxia but also to hypoxia in mixed venous blood. A specific test of the blood contribution would be to lower mixed venous PO2 (PvO2), which can be accomplished by increasing hemoglobin-O2 affinity. When we exchanged transfused rats with cyanate-treated erythrocytes [PO2 at 50% hemoglobin saturation (P50) = 21 Torr] or with Créteil erythrocytes (P50 = 13.1 Torr), we lowered PvO2 from 39 +/- 5 to 25 +/- 4 and to 14 +/- 4 Torr, respectively, without altering arterial blood gases or hemoglobin concentration. Right ventricular systolic pressure increased from 32 +/- 2 to 36 +/- 3 Torr with cyanate erythrocytes and to 44 +/- 5 Torr with Créteil erythrocytes. Cardiac output was unchanged. Control exchange transfusions with normal rat or 2,3-diphosphoglycerate-enriched human erythrocytes had no effect on PvO2 or right ventricular pressure. Alveolar hypoxia plus high O2 affinity blood caused a greater increase in right ventricular systolic pressure than either stimulus alone. We concluded that PvO2 is an important determinant of pulmonary vascular tone in the rat.     

Hematological alterations and response to acute hypobaric stress.

Exposure of rats to simulated altitude (15,000 ft) for 1 day and 3 and 9 wk produced progressive polycythemia, elevated 2,3-diphosphoglycerate levels and raised P50 values; the latter two parameters decreased toward control values after 9 wk. Carbon monoxide (38-43% HbCO) exposure produced polycythemia after 3- and 9-wk exposure, no change in 2,3-DPG and a fall in P50 value. Ten days' treatment with sodium cyanate produced a large decrease in 2,3-DPG and P50. Survival during 90 min of acute hypobaria (0.3 atm) under Nembutal anesthesia was highest with NaOCN (75%), intermediate with 3- and 9-wk exposure to altitude and CO (56-58%) lower in 1-day altitude exposure (44%) and lowest in controls (5%). Heart and ventilation rate was monitored during this hypobaric test and response patterns established for each exposure/treatment. In states of extreme oxygen deprivation the results suggest, in order of importance, the survival value of 1) increased oxygen-hemoglobin affinity, and 2) polycythemia.     

Diurnal changes of the 2,3-diphosphoglycerate concentration in human red cells and the influence of posture.

The 2,3-diphosphoglycerate (2,3-DPG) concentration, oxygen half saturation pressure at pH 7.4 (P50), pH in plasma and red cells, and mean corpuscular hemoglobin concentration (MCHC) of venous blood were determined during unrestricted daily activity (series I) throughout 24 hrs as well as during prolonged bed rest until noon (series II). In series I almost synchronous dirunal behavior of P50 2,3-DPG, and plasma pH as well as red cell pH became significantly apparent with highest values in the afternoon. The [2,3-DPG] yielded most pronounced alterations, which made up to 13.5% of the average day value. During prolonged recumbency the [2,3-DPG] showed a nonsignificant tendency to decline; the P50 remained unchanged throughout that period. The possible reason for the missing [2,3-DPG] increase is a reduced change of red cell pH in series II. An influence of a posture dependent aldosterone secretion either directly on the 2,3-DPG metabloism of indirectly via mediating the red cell pH and thus ruling the formation of this organic PHOSPHORIS COMPOUND IS DISCUSSED.     

Altitude-induced erythrocytic 2,3-DPG and hemoglobin changes in rats of various ages.

Rats of various ages (2, 12, 24, and 40 mo of age) were exposed for 4 wk to either a simulated high altitude of 23,000 ft or to a Peoria, Ill., altitude of 650 ft above sea level. Hematocrit ratios, hemoglobin, and erythrocytic 2,3-diphospho-glycerate (2,3-DPG) concentrations were measured. Hematocrit and hemoglobin determinations revealed a decrease in erythrocytic content with increasing age, and the augmented erythropoietic response was seen in all age groups of animals as a result of altitude exposure. The maximal erythrocytic content of hemoglobin in the 40-mo-old animals was significantly lower than that of all other age groups. Erythrocytic 2,3-DPG levels were significantly changed by aging alone. In the 40-mo-old group there was a 35% increase over the next highest sea-level value. However, while erythrocytic 2,3-DPG content increased significantly in all other age groups following altitude exposure, it decreased 46% in the 40-mo-old group.     

Operation Everest II: elevated high-altitude pulmonary resistance unresponsive to oxygen

High altitude increases pulmonary arterial pressure (PAP), but no measurements have been made in humans above 4,500 m. Eight male athletic volunteers were decompressed in a hypobaric chamber for 40 days to a barometric pressure (PB) of 240 Torr, equivalent to the summit of Mt. Everest. Serial hemodynamic measurements were made at PB 760 (sea level), 347 (6,100 m), and 282/240 Torr (7,620/8,840 m). Resting PAP and pulmonary vascular resistance (PVR) increased from sea level to maximal values at PB 282 Torr from 15 +/- 0.9 to 34 +/- 3.0 mmHg and from 1.2 +/- 0.1 to 4.3 +/- 0.3 mmHg.l-1 X min, respectively. During near maximal exercise PAP increased from 33 +/- 1 mmHg at sea level to 54 +/- 2 mmHg at PB 282 Torr. Right atrial and wedge pressures were not increased with altitude. Acute 100% O2 breathing lowered cardiac output and PAP but not PVR. Systemic arterial pressure and resistance did not rise with altitude but did increase with O2 breathing, indicating systemic control differed from the lung circulation. We concluded that severe chronic hypoxia caused elevated pulmonary resistance not accompanied by right heart failure nor immediately reversed by O2 breathing.     

Studies on avian erythrocyte metabolism. Effect of organic phosphates on oxygen affinity of embryonic and adult-type hemoglobins of the chick embryo.

The effects of 2,3 diphosphoglyceric acid (2,3-DPG), adenosine triphosphate (ATP), and inositol hexaphosphate (IHP) on the oxygen affinity of whole “stripped” hemoglobin (WSH), hemoglobin H (Hb-H), hemoglobin A (Hb-A) and hemoglobin D (Hb-D) isolated from 18-day chick embryo blood have been determined. The effect of the three organic phosphates upon the oxygen dissociation curves is similar and the following order of decreasing oxygen affinity of the organic phosphates was observed for each hemoglobin: 2,3-DPG < ATP < IHP. 2,3-DPG appears to have a slightly greater effect upon the P₅₀ of Hb-H than upon that of either of the two adult-type hemoglobins. However, this effect seems insufficient to suggest a preferential interaction of 2,3-DPG with Hb-H which would account for either the large amounts of 2,3-DPG in the erythrocytes of embryos or the higher oxygen affinity of the whole blood. The effects of the organic phosphates upon the Hill constant of the purified hemoglobins are variable. It is concluded that since the distribution of hemoglobins H, A, and D in the erythrocytes during the developmental period from 18-day embryos to 6-day chicks remains fairly constant, the previously described progressive decrease in oxygen affinity of the whole blood during this period results from changes in the total amount and distribution of the intraerythrocytic organic phosphates.²     

Relationship between the major phosphorylated metabolic intermediates and oxygen affinity of whole blood in the loggerhead (Caretta caretta) and the Green Sea Turtle (Chelonia mydas) during development.

(1) 2,3-Diphosphoglyceric acid (2,3-DPG) is present in the erythrocytes (RBC) of the 68-day loggerhead turtle embryo and 44-day green sea turtle embryo at levels of 7.4 and 5.5 μmoles/ml of RBC, representing the major organic phosphate during the latter period of embryonic development. (2) Inositol pentaphosphate (IPP) is absent in the red blood cells of the embryos of both the loggerhead and green sea turtle. (3) Near equimolar amounts of 2,3-DPG and IPP are present in the erythrocytes of the adult loggerhead and green sea turtle. The total concentration of these two organic phosphates is approximately 0.75 μmoles/ml of RBC in the adult of both species. (4) There is a switch from embryonic to adult hemoglobin during development of these two species of turtles; the two embryonic bands have identical electrophoretic mobilities, whereas the two adult bands migrate differently on cellulose acetate at pH 8.6. (5) The whole blood oxygen affinity of the adult loggerhead and green sea turtle is 60.3 and 32.6 Torr, respectively. (6) The stripped adult hemoglobins in these two species of turtles show no change in oxygen affinity upon addition of 2,3-DPG, ATP, or IPP. (7) It therefore appears unlikely that whole blood oxygen affinity is controlled by organic phosphate modulation of hemoglobin function in these species of turtles.     

Operation Everest II: metabolic and hormonal responses to incremental exercise to exhaustion.

The reasons for the reduced exercise capacities observed at high altitudes are not completely known. Substrate availability or accumulations of lactate and ammonium could have significant roles. As part of Operation Everest II, peak oxygen uptakes were determined in five normal male volunteers with use of progressively increasing cycling work loads at ambient barometric pressures of 760, 380, and 282 Torr. Decrements from sea level (SL) to 380 and 282 Torr occurred in peak power output (19 and 47%), time to exhaustion (19 and 48%), and oxygen uptake (41 and 61%), respectively. Arterial saturations after exhaustive exercise were decreased to 63% at 380 Torr and 39% at 282 Torr. At 380 and 282 Torr, postexercise plasma concentrations of glucose and free fatty acids were not increased, whereas plasma glycerol concentrations were decreased relative to SL (145 +/- 24 microM at 380 Torr and 77 +/- 10 microM at 282 Torr vs. 213 +/- 24 microM at SL). Preexercise plasma insulin concentrations were elevated at both 380 and 282 Torr (87 +/- 16 pM at 380 Torr and 85 +/- 18 pM at 282 Torr vs. 41 +/- 30 pM at SL). In general, postexercise concentrations of plasma catecholamines were decreased at altitude compared with SL. Preexercise lactate and ammonium concentrations were not different at any simulated altitude. From these data neither substrate availability nor metabolic product accumulation limited exercise capacity at extreme simulated altitude.