Biomedical subjects

R G Boutilier

Publications and source records attributed to R G Boutilier.

At least 37 records · Page 2Linked to original sources

PubMed1998

The protective effects of metabolic rate depression in hypoxic cold submerged frogs.

Aerobic metabolic rates (MO2) and respiratory quotients (RQ = MCO2/MO2) were measured in hypoxic frogs, hibernating underwater for up to 90 days at 3 degrees C. After 3 months of submergence at a PO2 of 50 mmHg, MO2 levels were 25% of those seen prior to hibernation. In progressive hypoxia, a gradual shift in RQ from 0.85 to 1.0 signaled an increasing reliance upon carbohydrate metabolism. Moreover, the glycogen concentrations of skeletal muscle, liver and heart of hypoxic frogs were more rapidly depleted than in their normoxic counterparts. A plasma lactacidosis revealed that the hypoxic animals recruited anaerobiosis to fuel a large 'Pasteur effect'. Throughout all stages of hypoxia, cellular ATP levels were maintained homeostatic. The ability to depress metabolic rate such that ATP demands can be met by oxidative phosphorylation in an oxygen limited environment is the key to the frogs' overwintering survival.

Acid-Base Equilibrium↗

PubMed1998

Respiratory, metabolic, and acid-base correlates of aerobic metabolic rate reduction in overwintering frogs.

Aerobic metabolic rates (MO2) and respiratory quotients (RQ = CO2 production/MO2) were measured contemporaneously in hibernating frogs Rana temporaria (L.), submerged for 90 days at 3 degrees C. After 3 mo of submergence in fully aerated water, MO2 levels were 61% of those seen at the same temperature before hibernation. Over the first 40 days of hibernation, RQ values (< or = 0.82) favored a lipid-based metabolism that progressively shifted to an exclusively carbohydrate metabolism (RQ = 1.01) by 90 days of hibernation. Liver glycogen concentrations fell by 68% during the first 8 wk of submergence, thereafter exhibiting a less rapid rate of utilization. Conversely, muscle glycogen concentrations remained stable over the first 2 mo of the experiment before falling by 33% over the course of the remaining 2 mo, indicating that the frog was recruiting muscle glycogen reserves to fuel metabolism. Submerged frogs exhibited an extracellular acidosis during the first week of submergence, but over the course of the next 15 wk "extracellular pH" values were not significantly different from the values obtained from the control air-breathing animals. The initial extracellular acidosis was not mirrored in the intracellular compartment, and the acid-base state was not significantly different from the control values for the first 8 wk. However, over the subsequent 8- to 16-wk period, the acid-base status shifted to a lower intracellular pH-HCO3 concentration set point, indicative of a metabolic acidosis. Even so, there was no indication that the acidosis could be attributed to anaerobic metabolism, as both plasma and muscle lactate levels remained low and stable. Muscle adenylate energy charge and lactate-to-pyruvate and creatine-to-phosphocreatine ratios also remained unchanged throughout hibernation. The capacity for profound metabolic rate suppression together with the ability to match substrate use to shifts in aerobic metabolic demands and the ability to fix new acid-base homeostatic set points are highly adaptive, both in terms of survival and reproductive success, to an animal that is often forced to overwinter under the cover of ice.

Adipose Tissue↗

PubMed1997

Hypometabolic homeostasis in overwintering aquatic amphibians.

Many amphibians encounter conditions each winter when their body temperature is so low that normal activities are suspended and the animals enter into a state of torpor. In ice-covered ponds or lakes, oxygen levels may also become limiting, thereby forcing animals to endure prolonged periods of severe hypoxia or anoxia. Certain frogs (e.g. Rana temporaria) can dramatically suppress their metabolism in anoxia but are not as tolerant as other facultative vertebrate anaerobes (e.g. turtle, goldfish) of prolonged periods of complete O2 lack. Many overwintering amphibians do, however, tolerate prolonged bouts of severe hypoxia, relying exclusively on cutaneous gas exchange. Rana temporaria overwintering for 2 months in hypoxic water (PO2 approximately 25 mmHg) at 3 degrees C progressively reduce their blood PCO2 to levels characteristic of water-breathing fish. The result is that blood pH rises and presumably facilitates transcutaneous O2 transfer by increasing Hb O2-affinity. Even after months of severe hypoxia, there is no substantial build-up of lactate as the animals continue to rely on cutaneous gas exchange to satisfy the requirements of a suppressed aerobic metabolism. Our recent experiments have shown that the skeletal muscle of frogs oxyconforms in vitro to the amount of O2 available. The cellular basis for the oxyconformation of skeletal muscle is unknown, but the hypothesis driving our continuing experiments theories that metabolic suppression at a cellular level is synonymous with suppressed ion leak across cellular membranes.

Adaptation, Physiological↗

PubMed1997

Balancing hypoxia and hypothermia in cold-submerged frogs.

Many animals respond to hypoxic stress by selecting cooler environments, the so-called 'behavioural hypothermia' response. Amphibians overwintering in ice-covered ponds and lakes offer an ecologically relevant test of this response since they must choose between the confounding metabolic effects of profound hypothermia or hypoxia; thermal and chemical conditions can vary from 0 degrees C and normoxic at the ice-water interface to 4 degrees C and markedly hypoxic at depths of 2-4 m. To mimic such environmental conditions, we constructed an experimental chamber that enabled continuous electronic surveillance of an animal's movement along a thermal gradient. When Rana temporaria pre-acclimated to 3.5 degrees C were placed in a normoxic thermal gradient ranging from 0.8 to 8 degrees C, they invariably favoured the warmer end of the chamber. Upon exposure to hypoxia, however, their preferred temperature shifted from a median of 6.8 degrees C (P02 = 158 mmHg; 1 mmHg = 0.133 kPa) to 1.9 degrees C (P02 = 25 mmHg). Metabolic rate measurements from animals exposed simultaneously to acute changes in water temperature and PO2 suggest that movement to colder conditions in hypoxia effects the greatest metabolic savings and prolongs the onset of a plasma lactacidosis.

Acid-Base Equilibrium↗

PubMed1995

Effects of carbonic anhydrase inhibition on the acid base status in lamprey and trout.

Inhibition of red cell carbonic anhydrase (CA) activity resulted in the rapid development of a respiratory acidosis (0.25 pH depression within 15 min post-injection) in the blood of trout. In the lamprey, however, the onset of the respiratory acidosis was delayed and its magnitude was less (0.18 pH depression at 6 h post-injection). Erythrocyte pH of both species decreased by about 0.12 units by 1 h after CA inhibition. These data, combined with the lack of rapid anion (Cl-/HCO3-) exchange in the red cells of agnathans but not in other lower vertebrates, support the hypotheses that (1) the majority of total CO2 in lamprey is transported within the erythrocyte, and (2) the limiting step in the evolution of a functioning Jacobs-Stewart cycle, and thus the evolution of the common mechanism of systemic CO2 transport in vertebrate blood, was the incorporation of the band-3 anion exchange protein into the membrane of the red cell.

Acetazolamide↗

PubMed1995

Genetic population structure and gene flow in the Atlantic cod Gadus morhua: a comparison of allozyme and nuclear RFLP loci.

High levels of gene flow have been implicated in producing uniform patterns of allozyme variation among populations of many marine fish species. We have examined whether gene flow is responsible for the limited population structure in the Atlantic cod, Gadus morhua L., by comparing the previously published patterns of variation at 10 allozyme loci to 17 nuclear restriction fragment length polymorphism (RFLP) loci scored by 11 anonymous cDNA clones. Unlike the allozyme loci, highly significant differences were observed among all populations at the DNA markers in a pattern consistent with an isolation-by-distance model of population structure. The magnitude of allele frequency variation at the nuclear RFLP loci significantly exceeded that observed at the protein loci (chi 2 = 24.6, d.f. = 5, P < 0.001). Estimates of gene flow from the private alleles method were similar for the allozymes and nuclear RFLPs. From the infinite island model, however, estimates of gene flow from the DNA markers were fivefold lower than indicated by the proteins. The discrepancy between gene flow estimates, combined with the observation of a large excess of rare RFLP alleles, suggests that the Atlantic cod has undergone a recent expansion in population size and that populations are significantly displaced from equilibrium. Because gene flow is a process that affects all loci equally, the heterogeneity observed among populations at the DNA level eliminates gene flow as the explanation for the homogeneous allozyme patterns. Our results suggest that a recent origin of cod populations has acted to constrain the extent of population differentiation observed at weakly polymorphic loci and implicate a role for selection in affecting the distribution of protein variation among natural populations in this species.

Animals↗

PubMed1994

Beta-adrenoreceptors in the trout (Oncorhynchus mykiss) heart: characterization, quantification, and effects of repeated catecholamine exposure.

Specific binding of the hydrophilic radioligand [3H]CGP-12177 to cell surface (functional) beta-adrenoreceptors was quantified in ventricular micropunches (2 mm diameter, 350 microns thickness) from seawater-acclimated rainbow trout held at 7-9 degrees. Binding was stereospecific, saturable, of high affinity, and displaceable by appropriate agonists and antagonists. Phentolamine failed to displace [3H]CGP at concentrations up to 10(-4) M, indicating an absence of [3H]CGP binding to alpha-adrenergic receptors. Trout ventricular beta-adrenoreceptors are exclusively of the beta 2 type. This conclusion is based on (1) the IC50 value for the beta 2-antagonist ICI 118551 (2.9 x 10(-6) M); (2) the inability of the beta 1-antagonist atenolol to displace [3H]CGP from beta-adrenoreceptors; and (3) the order of agonist-binding affinity (isoproterenol > epinephrine >> norepinephrine). The Bmax and Kd values for [3H]CGP binding to myocardial tissue were approximately 0.04 fmol micrograms protein-1 and 0.25 nM, respectively. The Bmax value indicates that the density of cell surface (functional) beta-adreno-receptors in the ventricle was 12,000 sites per cell or 3.38 sites per microns 2 of sarcolemma. The Kd and Bmax values for [3H]CGP binding to ventricular beta-adrenoreceptors were unaffected by the in vivo administration of five bolus catecholamine injections (4.0 micrograms kg-1 epinephrine, 2.0 micrograms kg-1 norepinephrine). This suggests that stress-induced increases in plasma catecholamines are unlikely to cause the down-regulation of heart beta-adrenoreceptors in fish. The method described here represents a simple but powerful technique for the quantification and characterization of adrenergic receptors in the fish heart.

Adrenergic beta-Antagonists↗

PubMed1993

Relationship between blood buffering properties, erythrocyte pH and water content, in gray seals (Halichoerus grypus).

The separated plasma of gray seals (Halichoerus grypus) has an unusually high non-bicarbonate buffering capacity (beta NB = d[HCO3-]/dpH = -21 Slykes; Slyke = mmol l-1 pH unit-1). To examine whether this represents a trait of mammalian carnivora, or is related instead to deep diving, we examined blood from dog and killer whale, respectively. The beta NB of separated plasma of dog was typically low (-4 to -8 Slykes), as in other mammals, whereas that of the whale (-13.5 Slykes), while significantly higher than dog, was not nearly as high as in seal. As one would theoretically predict of a blood system with exceptionally high extracellular buffering and functional anion exchange across the erythrocyte membrane, there was little difference in the buffer slope between true plasma and whole blood of seal. This appears to be a novel pattern of CO2 carriage in mammals. Erythrocyte ion concentrations were similar to other mammalian carnivora, with intracellular sodium concentration being high and potassium low. The red cell water content was unaffected by changes in oxygen saturation and tended to decrease when extracellular pH decreased below 7.4. Apart from the comparatively elevated Bohr factor, -0.518 as a function of extracellular pH and -0.550 as a function of red blood cell pH, the blood oxygen binding properties were similar to terrestrial mammals of similar size.

Animals↗

PubMed1993

Metabolism and energetics in squid (Illex illecebrosus, Loligo pealei) during muscular fatigue and recovery.

The concentrations of intermediate and end products of anaerobic energy metabolism and of free amino acids were determined in mantle musculature and blood sampled from cannulated, unrestrained squid (Loligo pealei, Illex illecebrosus) under control conditions, after fatigue from increasing levels of exercise, and during postexercise recovery. Phosphagen depletion, accumulation of octopine (more so in Illex than in Loligo), and accumulation of succinate indicate that anaerobic metabolism contributes to energy production before fatigue. Proline was a substrate of metabolism in Loligo, as indicated by its depletion in the mantle. In both species, there was no evidence of catabolism of ATP beyond AMP. A comparison of the changes in the free and total levels of adenylates and the phosphagen indicates an earlier detrimental effect of fatigue on the energy status in Loligo. The acidosis provoked by octopine formation in Illex was demonstrated to promote the use of the phosphagen and to protect the free energy change of ATP such that the anaerobic scope of metabolism during swimming is extended and expressed more in Illex than in Loligo. In both species, there was no decrease in the sum of phospho-L-arginine, octopine, and L-arginine, and thus no release of octopine from the mantle, thereby supporting our earlier claim that octopine and associated protons are recycled in the mantle tissue. Overall, the metabolic strategy of Loligo is much less disturbing for the acid-base status. This strategy and the alternative strategy of Illex to keep acidifying protons in the tissue may be important for the protection of hemocyanin function in the two species.

Adenosine Diphosphate↗

PubMed1991

Acid-base regulation in exercising squid (Illex illecebrosus, Loligo pealei).

Squid (Illex illecebrosus, Loligo pealei) were cannulated in the vena cava and swum in a Beamish-type respirometer. Gas tensions and acid-base variables as well as octopine levels were estimated in samples of the mantle and of venous blood collected from quiescent, exercised, and recovered animals. When exhausted, both species exhibited a decrease in vena cava oxygen tensions and a slight alkalosis. With high swimming speeds prior to exhaustion in Illex a slight acidosis developed in the blood, which was linked to a severe intracellular acidosis. Generally, the drop in intracellular pH was linearly correlated with octopine accumulation in this species. Metabolic proton (and end-product) release from the mantle, however, was minimal, thus protecting arterial oxygen binding. High PCO2 values in the mantle of both species lead to the conclusion that the vena cava values analyzed in this and all literature studies on unrestrained cephalopods may not reflect the scope of respiratory acid-base changes in venous blood. Although metabolic changes in blood acid-base status are negligible, the respiratory acidification of venous mantle blood may allow for a classical function of Bohr and Haldane effects in these animals.

Acid-Base Equilibrium↗

PubMed1990

Determination of intracellular pH and PCO2 after metabolic inhibition by fluoride and nitrilotriacetic acid.

Mean intracellular pH (pHi) and PCO2 (PiCO2) have been analysed based on pH and total CO2 measurements in tissue homogenates. Tissues were sampled from undisturbed worms (Sipunculus nudus), squid (Illex illecebrosus), trout (Salmo gairdneri), toads (Bufo marinus), and rats. Homogenate metabolism was inhibited by the addition of potassium fluoride and nitrilotriacetic acid (NTA). Model calculations revealed that the influence of dilution, medium buffers, and contamination by extracellular fluids was negligible. In white muscle tissue the resulting pHi values were virtually the same as found in studies using DMO (dimethyloxazolidinedione). If large fractions of mitochondria were present (e.g. in heart muscle), DMO derived pHi values were considerably higher, probably representing overestimates. Homogenate derived pHi values are concluded to represent the effective mean pHi by taking into account pH gradients, and the volumes and buffering of cellular compartments. High time resolution and small variability make this method especially useful to assess rapid changes in pHi, e.g. in exercising animals.

Acid-Base Equilibrium↗

PubMed1990

Acid-base-electrolyte balance responses to catecholamine antagonists in Ambystoma tigrinum.

Neotenic larval Ambystoma tigrinum were subjected to hypercapnia (3% CO2, 22 Torr) for 24 h under different conditions: alpha-adrenergic blockade using phentolamine, beta-adrenergic blockade using propranolol, and sham treatments. The sham animals were able to carry out a partial extracellular pH compensation that consisted of an increase in extracellular [HCO3-]. Animals treated with catecholamine antagonists did not compensate to the same extent. Analysis of plasma samples by high-performance liquid chromatography with electrochemical detection revealed a significant increase in circulating norepinephrine, but not epinephrine, during the high-CO2 exposure. Measurements of cutaneous ion transport showed that beta-antagonists block the increased Na+ influx associated with hypercapnia, whereas alpha-antagonists inhibited the decrease in cutaneous Cl- influx that is also associated with respiratory acidosis. Additionally, both alpha- and beta-blockers inhibited the increase in transcutaneous potential difference that accompanied the respiratory acidosis. The results are consistent with a role for circulating catecholamines in compensatory ion transport responses to respiratory acidosis in this species.

Acid-Base Equilibrium↗

PubMed1990

CO2 transport properties of the blood of a primitive vertebrate, Myxine glutinosa (L.).

The CO2 transport properties of the blood of the hagfish, Myxine glutinosa are markedly different from those previously demonstrated for the lamprey, Petromyzon marinus. As in most other vertebrates, the majority of the CO2 in Myxine blood is transported in the form of plasma bicarbonate. Erythrocyte bicarbonate does have some access to the plasma in the blood of Myxine although the documented bicarbonate movements were not sensitive to the chloride/bicarbonate exchange inhibitor, 4,4-diisothiocyanatostilbene-2,2-disulphonic acid (DIDS). The transmembrane pH gradient in hagfish erythrocytes is also quite different from the gradient previously demonstrated in other agnathans. As in other agnathans, significant differences do occur between the distribution ratios for protons, chloride and bicarbonate ions across the erythrocyte membrane, although the magnitude of these differences is smaller in Myxine. Finally, the absence of pH-dependent changes in erythrocyte water content appears to be a common feature of agnathan erythrocytes.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗

PubMed1989

Energy metabolism in trout red cells: consequences of adrenergic stimulation in vivo and in vitro.

beta-Adrenergic stimulation of salmonid red cells results in a rapid decrease (within 5 min) in the nucleotide triphosphate:haemoglobin ratio (NTP:Hb), which is thereafter maintained at a constant level, presumably through increased ATP turnover via matched aerobic metabolism and energy-consuming processes. Addition of the beta-adrenergic agonist isoproterenol to rainbow trout red cells in vitro leads to a rise in intracellular pH (pHi), a corresponding decrease in extracellular pH (pHe) and an increase in red cell oxygen consumption (MO2). Moreover, the extent to which red cell pHi is maintained constant in the face of an acute extracellular acidosis in vitro or in vivo is proportional to the adrenergically stimulated increase in red cell MO2. In the absence of oxygen, these red cells remain capable of pH regulation, but cannot maintain NTP:Hb constant. As a result, membrane and metabolic functions become uncoupled in the stimulated deoxygenated cells.

Aerobiosis↗

PubMed1989

Metabolic-membrane coupling in red blood cells of trout: the effects of anoxia and adrenergic stimulation.

Under oxygenated conditions, in vitro, the highly aerobic red cells of the rainbow trout (Salmo gairdneri) exhibit tight coupling between energy (i.e. nucleotide triphosphate, NTP)-consuming and NTP-producing metabolic activity, as shown by strict maintenance of red cell NTP:haemoglobin ratios. This coupling is maintained following adrenergic stimulation of oxygenated red cells when the increased NTP demands of ion transporting systems are met by enhanced energy production via aerobic metabolism. In unstimulated anoxic red cells, membrane-metabolic coupling is preserved via the arrest of NTP-consuming processes. Adrenergic stimulation of anoxic red cells, however, leads to a functional uncoupling of membrane metabolism with the result that NTP levels decline rapidly. At this time, cellular [NTP] is negatively correlated with [Na+]i and [Cl-]i and positively correlated with [K+]i. This, in addition to the fact that the pH of the intracellular compartment is also highly dependent on cellular NTP levels, provides evidence for the integration of energy and membrane metabolisms.

Aerobiosis↗

PubMed1989

The effects of forced and voluntary diving on plasma catecholamines and erythrocyte pH in the aquatic anuran, Xenopus laevis.

The lactacidosis and hypoxaemia associated with enforced diving in Xenopus leads to a marked increase in circulating catecholamines, a phenomenon not observed during periods of voluntary diving. The magnitude of the catecholamine surge (predominantly noradrenaline) during an enforced dive appears to be matched to the severity of the hypoxaemia and/or acid-base disturbance. Indeed, a strong linear correlation was found between the decrease in arterial pH and the rise in plasma catecholamines that resulted from an enforced 60 min dive. During that time, the levels of plasma adrenaline and noradrenaline showed a strong inverse exponential correlation with arterial oxygen tensions (PaO2). Simultaneous measurements of red cell pH showed that Xenopus does not regulate its erythrocyte pH in the face of a plasma acidosis, unlike the adrenergically mediated pH regulation of red cells of certain teleost fish.

Anaerobiosis↗

PubMed1988

Acute exposure to graded levels of hypoxia in rainbow trout (Salmo gairdneri): metabolic and respiratory adaptations.

We have studied the mechanisms of acute hypoxia tolerance in rainbow trout (Salmo gairdneri). Fish held at 9 degrees C were exposed to various levels of hypoxia for 24 h. At an environmental PO2 of 30 Torr, the fish showed an initial plasma acidosis probably of metabolic origin which was subsequently offset such that blood pH returned to normal within about 4 h. Over this time period, red cell pH was maintained constant. Comparing the effects of different levels of hypoxia following 24 h exposure, oxygen consumption of the animal remained unchanged over a broad range of inspired oxygen tensions but declined by over 30% of normoxic values at inspired water PO2 levels of 80 Torr. This appeared to be a true metabolic depression because signs of increased anaerobic metabolism did not occur until there was a further reduction in water oxygen levels. Rainbow trout appear to be able to maintain a relatively high energy status in their white muscle during 24 h exposure to severe hypoxia (water PO2 = 30 Torr). As the level of hypoxia was intensified, there was a reduction in the oxygen gradient across the gills, probably facilitated in part by the release of catecholamines into the blood. The erythrocytic ATP: Hb4 molar ratios declined with increasing hypoxic stress as did the pH gradient between the erythrocyte and plasma. The overall effect was no change in Hb O2-affinity after 24 h exposure to severe hypoxia.

Adenosine Triphosphate↗

PubMed1988

Metabolic energy production during adrenergic pH regulation in red cells of the Atlantic salmon, Salmo salar.

Whole blood from Atlantic salmon was incubated anaerobically at 10 degrees C so as to measure the metabolic activity of the nucleated erythrocytes. An acute extracellular acidosis was produced by adding either an acid solution (sham) or an acid solution with adrenaline (final concentration, 5 x 10(-4) M). The extracellular acidosis produced by the sham solution was transferred to the erythrocytes, whereas with adrenaline, intracellular pH actually increased in the face of a plasma acidosis. Indeed, the extracellular acidosis in the adrenaline-treated blood was significantly higher than that of the sham as a result of net H+ excretion from the erythrocyte. This pH response of the erythrocyte was accompanied by a proportional increase in the O2 consumption of the blood, with no change in lactate production. In comparison to sham-treated cells, the content of erythrocytic nucleotide triphosphates initially decreased upon addition of adrenaline but was thereafter maintained at a constant NTP/Hb ratio presumably due to an increased ATP turnover. In conclusion, it appears that the aerobic rather than anaerobic metabolism of erythrocytes is accelerated upon addition of adrenaline to blood, and that this increased metabolism is involved in fueling the membrane transport processes involved in adrenergic pH regulation of salmonid red cells.

Acid-Base Equilibrium↗