Norberto C. Gonzalez

Contractility in Isolated Mammalian Heart Muscle after Acid-Base Changes

In-vitro experiments performed in cat papillary muscles and strips of rat right ventricle suggest that the changes in myocardial contractility that follow acid-base disturbances are not a function of extracellular pH. Simultaneous changes in Pco2 and NaHCO3 concentration, with extracellular pH constant, decreased developed tension and maximal rate of rise of the tension (dT/dt) without significant changes in the time to peak tension when the muscle was exposed to the solution with higher Pco2 and NaHCO3 concentration. At an extracellular pH of 7.40, developed tension decreased 0.51 ± 0.13 g/mm2 (P < 0.02) and dT/dt decreased 1.29 ± 0.50 g/sec (P < 0.05) with no significant change in time to peak tension (0.038 ± 0.022 sec). Changes in pH produced by increasing Pco2 at constant NaHCO3 concentration were followed by a significant decrease in contractility. A change of Pco2 from 20 to 90 mm Hg that produced a change in extracellular pH from 7.60 to 7.00 was accompanied by a decrease in developed tension of 0.67 ± 0.14 g/mm2 (P < 0.01), in dT/dt of 2.63 ± 0.54 g/sec (P < 0.01) with no changes in time to peak tension (0.0017 ± 0.10 seconds). We were unable to show significant variations in contractility when extracellular pH was changed at a constant Pco2 of approximately 21 mm Hg (NaHCO3 7.5, 15, and 30 mM) or at a Pco2 of approximately 95 mm Hg (NaHCO3 15, 30, 60, 80 and 120 mM). Only when extracellular pH reached a value as high as 8.0 (Pco2 21 mm Hg, NaHCO3 80 mM) a small but significant increase in contractility was evidenced. Either Pco2 or intracellular pH could be the major determinants of the changes in myocardial contractility that follow acid-base alterations.

Leukocyte-endothelial interactions in environmental hypoxia

Hypoxia induced by reducing inspired PO2 (PIO2) to 70 Torr, promotes a rapid microvascular response characterized by increased leukocyte rolling and adherence to the venular endothelium, leukocyte emigration to the perivascular space and increased vascular permeability. This appears to be a generalized response since it is observed in venules of the mesentery, cremaster muscle and pial microcirculations. After three weeks of acclimatization to hypoxia (barometric pressure 380 Torr, PIO2 70 Torr), the initial microvascular response resolves and exposure to even lower PIO2 (50 Torr) fails to elicit a microvascular response. The initial response is accompanied by a reversible increase in the generation of reactive oxygen species (ROS) and is blocked by antioxidants and by interventions that increase the tissue levels of nitric oxide (NO). In contrast to ischemia/reperfusion, ROS levels increase during hypoxia and return towards pre-hypoxic values after return to normoxia. Acclimatization involves upregulation of inducible NO synthase (iNOS): inhibition of iNOS using two different antagonists results in increased leukocyte-endothelial interactions and increased ROS generation. The results suggest that hypoxia initially leads to an alteration of the ROS/NO balance which is eventually restored during the acclimatization process. This phenomenon may have relevance to the microcirculatory alterations associated with hypoxic exposure, including acute mountain sickness and high altitude pulmonary and cerebral edema.

Peripheral oxygen transport and utilization in rats following continued selective breeding for endurance running capacity

Untrained rats selectively bred for either high (HCR) or low (LCR) treadmill running capacity previously demonstrated divergent physiological traits as early as the seventh generation (G7). We asked whether continued selective breeding to generation 15 (G15) would further increase the divergence in skeletal muscle capillarity, morphometry, and oxidative capacity seen previously at G7. At G15, mean body weight was significantly lower (P < 0.001) in the HCR rats (n = 11; 194 +/- 3 g) than in LCR (n = 12; 259 +/- 9 g) while relative medial gastrocnemius muscle mass was not different (0.23 +/- 0.01 vs. 0.22 +/- 0.01% total body weight). Normoxic (Fi(O(2)) = 0.21) Vo(2max) was 50% greater (P < 0.001) in HCR despite the lower absolute muscle mass, and skeletal muscle O(2) conductance (measured in hypoxia; Fi(O(2)) = 0.10) was 49% higher in HCR (P < 0.001). Muscle oxidative enzyme activities were significantly higher in HCR (citrate synthase: 16.4 +/- 0.4 vs. 14.0 +/- 0.6; beta-hydroxyacyl-CoA dehydrogenase: 5.2 +/- 0.2 vs. 4.2 +/- 0.2 mmol.kg(-1).min(-1)). HCR rats had approximately 36% more total muscle fibers and also 36% more capillaries in the medial gastrocnemius. Because average muscle fiber area was 35% smaller, capillary density was 36% higher in HCR, but capillary-to-fiber ratio was the same. Compared with G7, G15 HCR animals showed 38% greater total fiber number with an additional 25% decrease in mean fiber area. These data suggest that many of the skeletal muscle structural and functional adaptations enabling greater O(2) utilization in HCR at G7 continue to progress following additional selective breeding for endurance capacity. However, the largest changes at G15 relate to O(2) delivery to skeletal muscle and not to the capacity of skeletal muscle to use O(2).

The Systemic Inflammation of Alveolar Hypoxia Is Initiated by Alveolar Macrophage–Borne Mediator(s)

Alveolar hypoxia produces widespread systemic inflammation in rats. The inflammation appears to be triggered by activation of mast cells by a mediator released from alveolar macrophages, not by the reduced systemic partial pressure of oxygen (PO2). If this is correct, the following should apply: (1) neither mast cells nor tissue macrophages should be directly activated by hypoxia; and (2) mast cells should be activated when in contact with hypoxic alveolar macrophages, but not with hypoxic tissue macrophages. We sought here to determine whether hypoxia activates isolated alveolar macrophages, peritoneal macrophages, and peritoneal mast cells, and to study the response of the microcirculation to supernatants of these cultures. Rat mesenteric microcirculation intravital microscopy was combined with primary cultures of alveolar macrophages, peritoneal macrophages, and peritoneal mast cells. Supernatant of hypoxic alveolar macrophages, but not of hypoxic peritoneal macrophages, produced inflammation in mesentery. Hypoxia induced a respiratory burst in alveolar, but not peritoneal macrophages. Cultured peritoneal mast cells did not degranulate with hypoxia. Immersion of mast cells in supernatant of hypoxic alveolar macrophages, but not in supernatant of hypoxic peritoneal macrophages, induced mast cell degranulation. Hypoxia induced release of monocyte chemoattractant protein-1, a mast cell secretagogue, from alveolar, but not peritoneal macrophages or mast cells. We conclude that a mediator released by hypoxic alveolar macrophages activates mast cells and triggers systemic inflammation. Reduced systemic PO2 and activation of tissue macrophages do not play a role in this phenomenon. The inflammation could contribute to systemic effects of diseases featuring alveolar hypoxia.

Effect of chronic hypoxia on hemodynamics, organ blood flow and O2 supply in rats

Aortic blood flow, heart rate, blood pressure and blood flow distribution were measured in 10 chronically hypoxic rats (3 weeks, PB 370-380 Torr) breathing 10% O2 (chronic hypoxia) and after 30 min of breathing air (acute normoxia). Controls were 10 normoxic littermates breathing air (normoxia) and 10% O2 for 30 min (acute hypoxia). Acute hypoxia resulted in increased aortic blood flow and heart rate, and decreased total peripheral resistance. Blood flow and oxygen supply to vital organs increased, indicating that blood flow redistribution plays an important role in oxygen supply. In chronic hypoxia, aortic blood flow and heart rate remained elevated, and total peripheral resistance remained decreased. Blood flow distribution returned towards normoxia levels. Oxygen supply was maintained via increased arterial oxygen concentration. Acute normoxia resulted in decreased aortic blood flow and heart rate, and increased blood pressure and total peripheral resistance. Blood flow distribution was similar to that of chronic hypoxia except skeletal muscles, in which blood flow decreased markedly. Oxygen supply remained unchanged or increased.

Monocyte Chemoattractant Protein–1 Released from Alveolar Macrophages Mediates the Systemic Inflammation of Acute Alveolar Hypoxia

Alveolar hypoxia produces rapid systemic inflammation in rats. Several lines of evidence suggest that the inflammation is not initiated by low systemic tissue partial pressure of oxygen (Po(2)) but by a mediator released into the circulation by hypoxic alveolar macrophages. The mediator activates tissue mast cells to initiate inflammation. Monocyte chemoattractant protein-1/Chemokine (C-C motif) ligand 2 (MCP-1/CCL2) is rapidly released by hypoxic alveolar macrophages. This study investigated whether MCP-1 is the mediator of the systemic inflammation of alveolar hypoxia. Experiments in rats and in alveolar macrophages and peritoneal mast cells led to several results. (1) Alveolar hypoxia (10% O(2) breathing, 60 minutes) produced a rapid (5-minute) increase in plasma MCP-1 concentrations in conscious intact rats but not in alveolar macrophage-depleted rats. (2) Degranulation occurred when mast cells were immersed in the plasma of hypoxic intact rats but not in the plasma of alveolar macrophage-depleted rats. (3) MCP-1 added to normoxic rat plasma and the supernatant of normoxic alveolar macrophages produced a concentration-dependent degranulation of immersed mast cells. (4) MCP-1 applied to the mesentery of normoxic intact rats replicated the inflammation of alveolar hypoxia. (5) The CCR2b receptor antagonist RS-102895 prevented the mesenteric inflammation of alveolar hypoxia in intact rats. Additional data suggest that a cofactor constitutively generated in alveolar macrophages and present in normoxic body fluids is necessary for MCP-1 to activate mast cells at biologically relevant concentrations. We conclude that alveolar macrophage-borne MCP-1 is a key agent in the initiation of the systemic inflammation of alveolar hypoxia.

Pulmonary gas exchange during hypoxic exercise in the rat

Pulmonary gas exchange and O2 transport were studied at rest and during maximal treadmill exercise in rats in acute hypoxia (PIO2 approximately 71 Torr), and in littermates acclimatized to PB = 380 Torr (PIO2 approximately 71 Torr) for 3 weeks (chronic hypoxia). To obtain valid estimates of blood gas partial pressures, particularly during exercise, the temperature coefficients of blood pH, PO2 and PCO2 were determined (Appendix). In both acute and chronic hypoxia, the following changes were observed: alveolar and arterial PO2 increased considerably, but the difference, A-aPO2, did not change significantly; arterial O2 concentration (CaO2) decreased, and apparent pulmonary diffusing capacity for O2, Dapp, increased. The increase in Dapp, together with hyperventilation, may prevent further drop in CaO2 due to a large rightward shift in the blood-O2 equilibrium curve caused by lactic acidosis in conjunction with a large Bohr coefficient characteristic of this species. Comparison with corresponding results obtained in man reveals that during hypoxic exercise, the rat shows a larger increase in PAO2, an increase, instead of a decrease, in PaO2, and a larger increase in Dapp.

Alveolar macrophages initiate the systemic microvascular inflammatory response to alveolar hypoxia

Alveolar hypoxia occurs as a result of a decrease in the environmental [Formula: see text] , as in altitude, or in clinical conditions associated with a global or regional decrease in alveolar ventilation. Systemic effects, in most of which an inflammatory component has been identified, frequently accompany both acute and chronic forms of alveolar hypoxia. Experimentally, it has been shown that acute exposure to environmental hypoxia causes a widespread systemic inflammatory response in rats and mice. Recent research has demonstrated that alveolar macrophages, in addition to their well known intrapulmonary functions, have systemic, extrapulmonary effects when activated, and indirect evidence suggest these cells may play a role in the systemic consequences of alveolar hypoxia. This article reviews studies showing that the systemic inflammation of acute alveolar hypoxia observed in rats is not initiated by the low systemic tissue [Formula: see text] , but rather by a chemokine, Monocyte Chemoattractant Protein-1 (MCP-1, or CCL2) released by alveolar macrophages stimulated by hypoxia and transported by the circulation. Circulating MCP-1, in turn, activates perivascular mast cells to initiate the microvascular inflammatory cascade. The research reviewed here highlights the extrapulmonary effects of alveolar macrophages and provides a possible mechanism for some of the systemic effects of alveolar hypoxia.

Alveolar hypoxia, alveolar macrophages, and systemic inflammation

Diseases featuring abnormally low alveolar PO2 are frequently accompanied by systemic effects. The common presence of an underlying inflammatory component suggests that inflammation may contribute to the pathogenesis of the systemic effects of alveolar hypoxia. While the role of alveolar macrophages in the immune and defense functions of the lung has been long known, recent evidence indicates that activation of alveolar macrophages causes inflammatory disturbances in the systemic microcirculation. The purpose of this review is to describe observations in experimental animals showing that alveolar macrophages initiate a systemic inflammatory response to alveolar hypoxia. Evidence obtained in intact animals and in primary cell cultures indicate that alveolar macrophages activated by hypoxia release a mediator(s) into the circulation. This mediator activates perivascular mast cells and initiates a widespread systemic inflammation. The inflammatory cascade includes activation of the local renin-angiotensin system and results in increased leukocyte-endothelial interactions in post-capillary venules, increased microvascular levels of reactive O2 species; and extravasation of albumin. Given the known extrapulmonary responses elicited by activation of alveolar macrophages, this novel phenomenon could contribute to some of the systemic effects of conditions featuring low alveolar PO2.

Continued artificial selection for running endurance in rats is associated with improved lung function

Previous studies found that selection for endurance running in untrained rats produced distinct high (HCR) and low (LCR) capacity runners. Furthermore, despite weighing 14% less, 7th generation HCR rats achieved the same absolute maximal oxygen consumption (Vo(2max)) as LCR due to muscle adaptations that improved oxygen extraction and use. However, there were no differences in cardiopulmonary function after seven generations of selection. If selection for increased endurance capacity continued, we hypothesized that due to the serial nature of oxygen delivery enhanced cardiopulmonary function would be required. In the present study, generation 15 rats selected for high and low endurance running capacity showed differences in pulmonary function. HCR, now 25% lighter than LCR, reached a 12% higher absolute Vo(2max) than LCR, P < 0.05 (49% higher Vo(2max)/kg). Despite the 25% difference in body size, both lung volume (at 20 cmH(2)O airway pressure) and exercise diffusing capacity were similar in HCR and LCR. Lung volume of LCR lay on published mammalian allometrical relationships while that of HCR lay above that line. Alveolar ventilation at Vo(2max) was 30% higher, P < 0.05 (78% higher, per kg), arterial Pco(2) was 4.5 mmHg (17%) lower, P < 0.05, while total pulmonary vascular resistance was (insignificantly) 5% lower (30% lower, per kg) in HCR. The smaller mass of HCR animals was due mostly to a smaller body frame rather than to a lower fat mass. These findings show that by generation 15, lung size in smaller HCR rats is not reduced in concert with their smaller body size, but has remained similar to that of LCR, supporting the hypothesis that continued selection for increased endurance capacity requires relatively larger lungs, supporting greater ventilation, gas exchange, and pulmonary vascular conductance.