Phosphate excretion during 24 h of hypoxia in conscious rats

The objective of this study was to investigate renal phosphate excretion during 24 h of hypoxia in conscious rats fed by total parenteral nutrition. Wistar rats weighing 190 g were exposed to hypoxia (inspired oxygen fraction = 0.10) or normoxia (inspired oxygen fraction = 0.21) for 24 h in a normobaric chamber. Renal clearance and hormonal studies were performed. The results showed a greater fractional excretion of phosphate (5.37 pL 0.07%, P < 0.05) and hypophosphataemia (7.40 pL 0.12 mg dL^-1, P < 0.01) in hypoxic rats (n = 10) than in normoxic rats (n = 13; 3.50 pL 0.37% and 8.02 pL 0.16 mg dL^-1, respectively). In addition, during hypoxia there was a significant decrease in the excretion of urinary adenosine 3′,5′-cyclic monophosphate per glomerular filtrate (2.97 pL 1.27 nmol dL^-1, P < 0.005), a parameter of the renal action of parathyroid hormone, and a stable level of serum parathyroid hormone (10.2 pL 2.6 ng mL^-1) (cf. normoxia: 8.57 pL 0.70 nmol dL^-1 and 8.0 pL 1.7 ng mL^-1, respectively). However, creatinine clearance and the renal adenosine triphosphate level, both of which affect adenosine 3′,5′-cyclic monophosphate excretion, were not different between the two groups. These data suggest that exposure of conscious rats to 24 h of hypoxia causes renal hyporesponsiveness to physiological levels of parathyroid hormone, which is manifested as a decrease in adenosine 3′,5′-cyclic monophosphate excretion. Phosphaturia is not a direct net effect of hypoxia and secondary hypocapnia on renal phosphate transport, which is known to be regulated by parathyroid hormone through adenosine 3′,5′-cyclic monophosphate.     

Rapid increase in guanosine 3',5'-cyclic-monophosphate interferon treated mouse leukemia L1210 cells: relationship to the development of the antiviral state and inhibition of cell multiplication

Treatment of mouse leukemia L1210 cells with electrophoretically pure mouse interferon caused a rapid (1 to 5 min) and transitory (duration 5 to 10 min) increase in the intracellular concentration of guanosine 3′,5′-cyclic monophosphate (cyclic GMP). No significant increase in the intracellular concentration of adenosine 3′,5′-cyclic monophosphate (cyclic AMP) was observed until 2 to 3 hr after the addition of interferon. Electrophoretically pure mouse interferon caused a dose-dependent increase in the intracellular concentration of cyclic GMP at concentrations ranging from 1.0 to 10⁴ unite/ml. The interferon-induced increase in cyclic GMP was abrogated by specific antibody to mouse interferon. Furthermore, interferon had no effect on the intracellular concentration of cyclic GMP in a line of L1210 cells resistant to the action of interferon. The interferon-induced increase in cyclic GMP was calcium dependent and was inhibited by indomethacin and aspirin at concentrations of 10^?7 to 10^?6 M. Depletion of intracellular calcium prior to the addition of interferon abrogated the interferon-induced increase in cyclic GMP without preventing either the development of the antiviral state or the inhibition of cell multiplication in interferon-treated L1210 cells. These results suggest that cyclic GMP may not mediate these two biologic effects of interferon but may rather reflect rapid changes in interferon-treated cells.     

Urinary cyclic AMP and vasopressin excretion in rat strains selected for their alcohol intake

Urinary excretion of adenosine 3′,5′-cyclic monophosphate (cAMP) and immunoreactive arginine vasopressin (AVP) were investigated after water loading and following ethanol loading in two rat strains selected for their voluntary ethanol intake. After ethanol loading ethanol preferring (AA) rats excreted more cAMP but less AVP than water preferring (ANA) rats. The results suggest that the strain difference in cAMP excretion is of renal origin and is not due to vasopressin or parathormone. Differences in the sympathetic nervous activity may be responsible for the difference in cAMP excretion.     

Retrograde blood flow in the inactive limb is enhanced during constant-load leg cycling in hypoxia

Purpose

This study aimed to elucidate the effects of hypoxia on the pattern of oscillatory blood flow in the inactive limb during constant-load dynamic exercise. We hypothesised that retrograde blood flow in the brachial artery of the inactive limb would increase during constant-load leg cycling under hypoxic conditions.

Methods

Three maximal exercise tests were conducted in eight healthy males on a semi-recumbent cycle ergometer while the subjects breathed a normoxic [inspired oxygen fraction (FIO₂) = 0.209] or two hypoxic gas mixtures (FIO₂ = 0.155 and 0.120). Subjects then performed submaximal exercise at the same relative exercise intensity of 60 % peak oxygen uptake under normoxic or the two hypoxic conditions for 30 min. Brachial artery blood velocity and diameter were recorded simultaneously during submaximal exercise using Doppler ultrasonography.

Results

Antegrade blood flow gradually increased during exercise, with no significant differences among the three trials. Retrograde blood flow showed a biphasic response, with an initial increase followed by a gradual decrease during normoxic exercise. In contrast, retrograde blood flow significantly increased during moderate and severe hypoxic exercise, and remained elevated above normoxic conditions during exercise. At 30 min of exercise, the magnitude of the change in retrograde blood flow during exercise was greater as the level of hypoxia increased (normoxia: ?18.7 ± 23.5 ml min^?1; moderate hypoxia: ?39.3 ± 21.4 ml min^?1; severe hypoxia: ?64.0 ± 36.3 ml min^?1).

Conclusion

These results indicate that moderate and severe hypoxia augment retrograde blood flow in the inactive limb during constant-load dynamic leg exercise.     

Myocardial adenosine stimulates release of cyclic adenosine monophosphate from capillary endothelial cells in guinea pig heart

Activation of coronary endothelial cell adenylate cyclase was studied in the isolated guinea pig heart by prelabelling endothelial adenine nucleotides using intracoronary infusion of [³H]-adenosine, and measuring the coronary efflux of [³H]-cyclic adenosine monophosphate (cAMP). Hypoxia (30 % O₂) caused a 4-fold increase in coronary release of [³H]-cAMP, which was decreased by 63 % by infusion of the adenosine receptor antagonist, theophylline (50 M). During normoxic control conditions, degrading adenosine to non-vasoactive inosine by intracoronary infusion of adenosine deaminase (1.7 U/ml) caused a 20 % decrease in the release of [³H]-cAMP. The effect of adenosine deaminase was reversed by a specific enzyme inhibitor erythro-9-(2-hydroxy-3-nonyl) adenine hydrochloride. Coronary efflux of [³H]-cAMP during intracoronary infusion of 1 M adenosine triphosphate (ATP), adenosine diphosphate or adenosine monophosphate (AMP) (plus adenosine deaminase 8 U/ml) was only 13 % of that due to 1 M adenosine. Adenosine receptor blockers theophylline and CGS 15943A caused equivalent inhibition of the coronary vasodilator actions of adenosine and ATP. Intracoronary infusion of prostaglandin E₁ and the ₂-adrenergic agonist procaterol caused parallel, dose-dependent increases in coronary conductance and the venous release of [³H] cAMP. It is concluded that (1) under both normoxic and hypoxic conditions, adenosine formed by the heart may activate endothelial cell adenylate cyclase via membrane adenosine receptors, (2) coronary receptors for adenosine and ATP share common ligand affinities but ATP receptors are not coupled to adenylate cyclase, and (3) other vasodilators known to activate endothelial adenylate cyclase in vitro cause parallel increases in coronary conductance and adenylate cyclase activity in the beating heart.     

Presence of a calcium2+-dependent activator of cyclic-nucleotide phosphodiesterase in rat carotid body: effects of hypoxia.

The carotid body of rats contains cyclic 3′,5′-adenosine monophosphate and cyclic 3′,5′-guanosine monophosphate phosphodiesterases. The phosphodiesterase acting on cyclic adenosine monophosphate in carotid body can be regulated by a Ca²⁺-dependent activator protein. It was found that the content of this activator was increased two-fold after exposure of rats to hypoxia. Kinetic analysis of endogenous cyclic adenosine monophosphate phosphodiesterase indicated that the enzyme exists in both a high- and a low-K_m form. Exposure to hypoxia resulted in an increase in V_max and an apparent conversion to a single low-K_m form. This shift in the molecular forms of phosphodiesterase was Ca²⁺-dependent. Disappearance of the high-K_m form of the enzyme was also obtained in vitro after incubation of carotid body homogenates under protein phosphorylating conditions. Transection of the carotid sinus nerve prevented the shift from the high-K_m to a low-K_m form elicited by hypoxia.     

Introduction of oxygen function into isoprenoid systems by means of Botrytis cinerea (Persoon)

The introduction of oxygen functions into (±)- 1 -( 2′,2′,3′ -trimethyl- 3′ -cyclopenten-1′ -yl)-propan-2-one [(±)- 1a ], (±)- 1- ( 2′,2′,3′ ,-trimethyl-3′-cyclopenten-1′ -yl)-butan-2-one [(±)- 1b ], and (±)- 1 - (2′, 3′, 3′-trimethyl-3′ -cyclopenten-1′ -yl)-pentan-2-one [(±)-1c)] by Botrytis cinerea was carried out by hydroxylation in α position with respect to the double bond, or by epoxidation of the double bond itself. The main products had the hydroxy group at the methyl on the double bond. Simultaneously, a hydroxylation at the C′-5 position in 1a and 1b was observed. In 1e , an additional hydroxylation occurred at the C-4 position of the side chain. All the reactions were enantiospecific.     

Human erythropoietin response to hypocapnic hypoxia,normocapnic hypoxia,and hypocapnic normoxia

This study investigated the human erythropoietin (EPO) response to short-term hypocapnic hypoxia, its relationship to a normoxic or hypoxic increase of the haemoglobin oxygen affinity, and its suppression by the addition of CO₂ to the hypoxic gas. On separate days, eight healthy male subjects were exposed to 2 h each of hypocapnic hypoxia, normocapnic hypoxia, hypocapnic normoxia, and normal breathing of room air (control experiment). During the control experiment, serum-EPO showed significant variations (ANOVAP = 0.047) with a 15% increase in mean values. The serum-EPO measured in the other experiments were corrected for these spontaneous variations in each individual. At 2 h after ending hypocapnic hypoxia (10% O₂ in nitrogen), mean serum-EPO increased by 28% [baseline 8.00 (SEM 0.84) U · 1⁻¹, post-hypoxia 10.24 (SEM 0.95) U · 1⁻¹, P = 0.005]. Normocapnic hypoxia was produced by the addition of CO₂ (10% Co₂ with 10% O₂) to the hypoxic gas mixture. This elicited an increased ventilation, unaltered arterial pH and haemoglobin oxygen affinity, a lower degree of hypoxia than during hypocapnic hypoxia, and no significant changes in serum-EPO (ANOVAP > 0.05). Hypocapnic normoxia, produced by hyperventilation of room air, elicited a normoxic increase in the haemoglobin oxygen affinity without changing serum-EPO. Among the measured blood gas and acid-base parameters, only the partial pressures of oxygen in arterial blood during hypocapnic hypoxia were related to the peak values of serum-EPO (r = −0.81,P = 0.01). The present human EPO responses to hypoxia were lower than those which have previously been reported in rodents and humans. In contrast with the earlier rodent studies, it was found that human EPO production could not be triggered by short-term increases in pH and haemoglobin oxygen affinity per se, and the human EPO response to hypoxia could be suppressed by concomitant normocapnia without acidosis.     

Acute effect of hypobaria and hypoxia on renal nerve activity in anaesthetized rabbits

To determine the acute effect of reduced barometric pressure and hypoxia on renal nerve activity, urethane-anaesthetized and mechanically ventilated rabbits were randomly exposed to the following four separate conditions in a decompression chamber: hypoxic hypoharia (n= 71 , hypoxic normobaria (n= 5), normoxic hypoharia (n= 8) and slow normoxic hypobaria (n= 7). A combination of rapid decompression and simultaneous adjustment of inspired PO₂ was used to simulate an altitude of 6600 m, and renal nerve activity and haemodynamics, such as systemic blood pressure and heart rate, were measured. During both hypoxic hypobaria and hypoxic normobaria, there were significant and similar increases in renal nerve activity at 6600 m (54 ± 7% and 61 ± 13% from each baseline, respectively). However, there were no changes in renal nerve activity during normoxic hypoharia or slow normoxic hypoharia with decompression rates of 1000 or 400 m min^-l, respectively. From these results, we conclude that a reduction in barometric pressure without hypoxia does not affect renal nerve activity in anaesthetized rabbits.     

Maternal hypoxia developmentally programs low podocyte endowment in male,but not female offspring

Fetal hypoxia is a common complication of pregnancy. We have previously reported that maternal hypoxia in late gestation in mice gives rise to male offspring with reduced nephron number, while females have normal nephron number. Male offspring later develop proteinuria and renal pathology, including glomerular pathology, whereas female offspring are unaffected. Given the central role of podocyte depletion in glomerular and renal pathology, we examined whether maternal hypoxia resulted in low podocyte endowment in offspring. Pregnant CD1 mice were allocated at embryonic day 14.5 to normoxic (21% oxygen) or hypoxic (12% oxygen) conditions. At postnatal day 21, kidneys from mice were immersion fixed, and one mid-hilar slice per kidney was immunostained with antibodies directed against p57 and synaptopodin for podocyte identification. Slices were cleared and imaged with a multiphoton microscope for podometric analysis. Male hypoxic offspring had significantly lower birth weight, nephron number, and podocyte endowment than normoxic male offspring (podocyte number; normoxic 62.86 ± 2.26 podocytes per glomerulus, hypoxic 53.38 ± 2.25; p < .01, mean ± SEM). In contrast, hypoxic female offspring had low birth weight but their nephron and podocyte endowment was the same as normoxic female offspring (podocyte number; normoxic 62.38 ± 1.86 podocytes per glomerulus, hypoxic 61.81 ± 1.80; p = .88). To the best of our knowledge, this is the first report of developmentally programmed low podocyte endowment. Given the well-known association between podocyte depletion in adulthood and glomerular pathology, we postulate that podocyte endowment may place offspring at risk of renal disease in adulthood, and explain the greater vulnerability of male offspring.     

Erythropoietin response to acute hypobaric or anaemic hypoxia in gentamicin-administered rats

In an attempt to assess the erythropoietin (Epo) production site(s) in rat kidney, Epo response to hypoxia and renal histopathological changes were studied in rats administered with graded doses of gentamicin. Male Sprague-Dawley rats of 9 to 11 weeks old were used. Following a 14-day subcutaneous administration (67.5 or 33.8 mg kg^-1 day^-1) of gentamicin, a nephrotoxic aminoglycoside, selective proximal tubular lesions were produced. These gentamicin-administered rats were compared with normal rats with respect to Epo response to hypoxia. Two different kinds of hypoxic load, either 0.35 atm hypobaric hypoxia (P_IO2= 46 torr) or acute anaemia (Ht: 29.3 ± 0.2% and [Hb]: 9.7 ± 0.3 g dl^-1) by withdrawing of blood corresponding to 1–2% of body weight was used. During the hypoxic period of up to 48 h, the peak renal venous plasma Epo titres of 3.1 ± 0.6 and 4.3 ± 0.6 U ml^-1 was observed at the 6th h in normal hypobaric hypoxic and anaemic rats, compared with the prehypoxic value of 0.7 ± 0.1 U ml^-1. The Epo titres then declined gradually. In the rats which were administered gentamicin, Epo response pattern was the same as that observed in the normal rats, but the peak value decreased significantly to 0.8 ± 0.3 and 1.1 ± 0.4 U ml^-1 in hypoxic and anaemic rats (P < 0.05). Histological examination revealed the selective damage to renal proximal convoluted tubules. The Epo response was reduced by the tissue damages, and restoration of the gentamicin-induced tissue injury was accompanied with restored Epo response to hypoxia. The results suggest that renal proximal convoluted tubules are involved in Epo production under hypoxia.     

Effects of ATP on rat renal haemodynamics and excretion: role of sodium intake, nitric oxide and cytochrome P450

Aim: Adenosine‐5′‐triphosphate (ATP) affects intrarenal vascular tone and tubular transport via P2 receptors; however, the actual role of the system in regulation of renal perfusion and excretion remains unclear and is the subject of this whole‐kidney study. Methods: Effects of suprarenal aortic ATP infusion, 0.6–1.2 mg kg⁻¹ h⁻¹, were examined in anaesthetised rats maintained on low‐ (LS) or high‐sodium (HS) diet. Renal artery blood flow (RBF, transonic flow probe) and the perfusion (laser‐Doppler flux) of the superficial cortex (CBF) and outer and inner medulla (OM–BF, IM–BF) were measured, together with sodium and water excretion and urine osmolality. Results: Adenosine‐5′‐triphosphate did not change arterial pressure, RBF or CBF while the effects on medullary perfusion depended on sodium intake. In LS rats ATP increased IM–BF 19 ± 6%, the effect was prevented by inhibition of nitric oxide (NO) with N‐nitro‐l ‐arginine methyl ester. In HS rats ATP decreased OM–BF 16 ± 3% and IM–BF (7 ± 4%, not significant); previous inhibition of cytochrome P450 with 1‐aminobenzotriazol blunted the OM–BF decrease and reversed the previous decrease of IM–BF to a 13 ± 8% increase. Inhibition of P2 receptors with pyridoxal derivative (PPADS) abolished medullary vascular responses to ATP. In HS rats pre‐treated with PPADS, ATP increased tubular reabsorption, probably via adenosine formation and stimulation of P1 receptors. Conclusion: The data indicate a potential role of ATP in the selective control of renal medullary perfusion, different in sodium depleted and sodium replete rats. The action of ATP appears to be mediated by the NO system and the cytochrome P450 dependent vasoactive metabolites.     

Diurnal normobaric moderate hypoxia raises serum erythropoietin concentration but does not stimulate accelerated erythrocyte production

This study was performed to examine the effect of diurnal normobaric hypoxia on hematological parameters. Eleven healthy male volunteers were randomly selected to be in either the hypoxic group (n=6) or the control group (n=5). The hypoxic group was exposed to 8 h of normobaric hypoxia in hypoxic tent systems that elicited a target peripheral O₂ saturation of 81±2% on three consecutive days. The control group spent three consecutive 8-h days in modified tent systems that delivered normoxic air into the tent. Venous blood samples were collected before the exposure (days –5, 0), after each day of the exposure (days 1, 2, 3), and for 3 weeks after the exposure (days 7, 10, 13, 17, 24). Serum erythropoietin concentration significantly increased from 9.1±3.3 U·L⁻¹ to 30.7±8.6 U·L⁻¹ in the hypoxic group. Although there were significant increases in hematocrit (4%), hemoglobin concentration (5%), red blood cell count (4%) on day 7 in the hypoxic group, these observations were likely due to dehydration or biological variation over time. There was no significant change in early erythropoietic markers (reticulocyte counts or serum ferritin concentration), which provided inconclusive evidence of accelerated erythroid differentiation and proliferation. The results suggest that the degree of hypoxia was sufficient to stimulate increased erythropoietin production and release. However, the duration of hypoxic exposure was insufficient to propagate the erythropoietic cascade.     

The effect of hypoxia on performance during 30 s or 45 s of supramaximal exercise

Summary The purpose of this study was to evaluate the effect of hypoxia (10.8±0.6% oxygen) on performance of 30 s and 45 s of supramaximal dynamic exercise. Twelve males were randomly allocated to perform either a 30 s or 45 s Wingate test (WT) on two occasions (hypoxia and room air) with a minimum of 1 week between tests. After a 5-min warm-up at 120 W subjects breathed the appropriate gas mixture from a wet spirometer during a 5-min rest period. Resting blood oxygen saturation was monitored with an ear oximeter and averaged 97.8 ± 1.5% and 83.2 ± 1.9% for the air (normoxic) and hypoxic conditions, respectively, immediately prior to the WT. Following all WT trials, subjects breathed room air for a 10-min passive recovery period. Muscle biopsies from the vastus lateralis were taken prior to and immediately following WT. Arterialized blood samples, for lactate and blood gases, were taken before and after both the warm-up and the performance of WT, and throughout the recovery period. Opencircuit spirometry was used to calculate the total oxygen consumption (Vo₂), carbon dioxide production and expired ventilation during WT. Hypoxia did not impair the performance of the 30-s or 45-s WT.Vo₃ was reduced during the 45-s hypoxic WT (1.71±0.21 I) compared with the normoxic trial (2.16±0.261), but there was no change during the 30-s test (1.22±0.11 vs 1.04±0.171 for the normoxic and hypoxic conditions, respectively). Muscle lactate (LA) increased more during hypoxia following both the 30-s and 45-s WT (67.1±25.0 mmol· kg^–1 dry weight) compared with normoxia (30.8 ± 18.0 mmol · kg^–1 dry weight). Hypoxia did not influence the change in intramuscular adenosine triphosphate, creatine phosphate and glucose-6-phosphate. The performance of WT during hypoxia was associated with a greater decrease in muscle glycogen (P<0.06). Throughout the recovery period, blood LA was lower following the hypoxia (8.43±2.88 mmol · l^–1) comparedwith normoxia (9.15±3.06 mmol · 1^–1). Breathing the hypoxic gas mixture prior to the performance of WT increased blood pH to 7.44±0.03, compared with 7.39±0.03 for normoxia. Blood pH remained higher during the 10-min recovery period following the hypoxic WT trials (7.24±0.08) compared with the normoxic WT (7.22±0.06). BloodP _{CO 2} was reduced prior to and immediately following WT during hypoxia, but there were no differences between the normoxic and hypoxic trials during the 10 min recovery period. These data indicate that more energy was transduced from the catabolism of glycogen to lactate during the hypoxic WT trials, which offset the reduced O₂ availability and maintained performance comparable with normoxic conditions. It is suggested that the induced respiratory alkalosis associated with breathing the hypoxic gas could account for the increased rate of muscle LA accumulation.     

Carbon monoxide inhibits hypoxic pulmonary vasoconstriction in rats by a cGMP-independent mechanism

 Hypoxia activates erythropoietin-producing cells, chemoreceptor cells of the carotid body and pulmonary artery smooth muscle cells (PSMC) with a comparable arterial PO₂ threshold of some 70 mmHg. The inhibition by CO of the hypoxic responses in the two former cell types has led to the proposal that a haemoprotein is involved in the detection of the PO₂ levels. Here, we report the effect of CO on the hypoxic pulmonary vasoconstriction (HPV). Pulmonary arterial pressure (PAP) was measured in an in situ, blood-perfused lung preparation. PAP in normoxia (20% O₂, 5% CO₂) was 15.2±1.8 mmHg, and hypoxia (2% O₂, 5% CO₂) produced a ΔPAP of 6.3±0.4 mmHg. Addition of 8% or 15% CO to the hypoxic gas mixture reduced the ΔPAP by 88.3±2.7% and 78.2±6.1% respectively. The same levels of CO did not affect normoxic PAP nor reduced the ΔPAP produced by angiotensin II. The effect of CO was studied after inhibition of the NO-cyclic guanosine monophosphate (cGMP) cascade with N-methyl-l-arginine (5·10^–5 M) or methylene blue (1.4·10^–4 M). It was found that both inhibitors more than doubled the hypoxic ΔPAP without altering the effectiveness of CO to inhibit the HPV. In in vitro experiments we verified the inhibition of guanylate cyclase by measuring the levels of cGMP in segments of the pulmonary artery. Cyclic GMP levels were 1.4±0.2 (normoxia), 2.5±0.3 (hypoxia) and 3.3±0.5 pmole/mg tissue (hypoxia plus 8% CO); sodium nitroprusside increased normoxic cGMP levels about fourfold. Methylene blue reduced cGMP levels to less than 10% in all cases, and abolished the differences among normoxic, hypoxic and hypoxic plus CO groups. It is concluded that CO inhibits HPV by a NO-cGMP independent mechanism and it is proposed that a haemoprotein could be involved in O₂-sensing in PSMC. Received: 17 March 1997 / Received after revision: 10 June 1997 / Accepted: 11 July 1997     

Hypoxic inhibition of K+ currents in isolated rat type I carotid body cells: evidence against the involvement of cyclic nucleotides

 Whole-cell patch-clamp recordings were used to evaluate the effects of the cyclic nucleotides adenosine 3’,5’-cyclic monophosphate (cAMP) and guanosine 3’,5’-cyclic monophosphate (cGMP) on ionic currents in type I carotid body cells isolated from rat pups, and to investigate whether cyclic nucleotides are involved in K⁺ current inhibition by hypoxia. In the presence of 500 μM isobutylmethylxanthine, currents were not significantly modified by 8-bromo-cAMP (2 mM), dibutyryl-cAMP (5 mM) or 8-bromo-cGMP (2 mM). Currents were also unaffected by the phosphodiesterase (PDE)-resistant protein kinase A activators Sp-cyclic adenosine-3′,5′-monophosphorothioate (Sp-cAMPS) and Sp-8-bromoadenosine-3′,5′-monophosphorothioate (Sp-8-bromo-cAMPS) (50 μM), or by β-phenyl-1,N ²-ethenoguanosine-3′,5′-cyclic monophosphate (PET-cGMP) (100 μM) or the nitric oxide donor S-nitroso-N-acetylpenicillamine (SNAP; 500 μM). Ca²⁺ channel currents were also unaffected by Sp-8-Br-cAMPS, PET-cGMP and SNAP at the same concentrations. In the absence of cyclic nucleotide analogues, hypoxia (P_O2 17–23 mmHg) reversibly inhibited K⁺ currents. This degree of hypoxic inhibition was not significantly altered by the PDE-resistant protein kinase A inhibitors Rp-cyclic adenosine-3′,5′-monophosphorothioate (Rp-cAMPS) (50 μM) or Rp-8-bromoadenosine-3′,5′-monophosphorothioate (Rp-8-bromo-cAMPS) (200 μM). Similarly, PET-cGMP (100 μM) and SNAP (500 μM) did not alter the degree of inhibition caused by hypoxia. At the same concentrations used in type I cell experiments, Sp-8-bromo-cAMPS, PET-cGMP and SNAP completely relaxed isolated guinea-pig basilar arteries preconstricted with 20 mM K⁺-containing solutions. Our results indicate that cyclic nucleotides alone are not an important factor in the regulation by O₂ tension of K⁺ currents in rat type I carotid body cells. Received: 12 June 1996 / Accepted: 14 August 1996     

Combined effects of hypoxia and endurance training on lipid metabolism in rat skeletal muscle

Aim: To determine whether endurance training can counterbalance the negative effects of hypoxia on mitochondrial phosphorylation and expression of the long chain mitochondrial fatty acid transporter muscle carnitine palmitoyl transferase 1 (mCPT‐1). Methods: Male Wistar rats were exposed either to hypobaric hypoxia (at a simulated altitude of ≈4000 m, PIO₂ ≈ 90 mmHg) or to normoxia (sea level) for 5 weeks. In each environment, rats were randomly assigned to two groups. The trained group went through a 5‐week endurance training programme. The control group remained sedentary for the same time period. Muscle fatty acid oxidation capacity was evaluated after the 5‐week period on isolated mitochondria prepared from quadriceps muscles with the use of palmitoylcarnitine or pamitoylCoA + carnitine. Results: Chronic hypoxia decreased basal (V₀, ?31% with pamitoylCoA + carnitine and ?21% with palmitoylcarnitine, P < 0.05) and maximal (V_max, ?31% with pamitoylCoA + carnitine, P < 0.05) respiration rates, hydroxyacylCoA dehydrogenase activity (?48%, P < 0.05), mCPT‐1 activity index (?34%, P < 0.05) and mCPT‐1 protein content (?34%, P < 0.05). Five weeks of endurance training in hypoxia brought V₀, mCPT‐1 activity index and mCPT‐1 protein content values back to sedentary normoxic levels. Moreover, in the group trained in hypoxia, V_max reached a higher level than in the group that maintained a sedentary lifestyle in normoxia (24.2 nmol O₂· min^?1 · mg^?1 for hypoxic training vs. 19.9 nmol O₂ · min^?1 · mg^?1 for normoxic sedentarity, P < 0.05). Conclusion: Endurance training can attenuate chronic hypoxia‐induced impairments in mitochondrial fatty acid oxidation. This training effect seems mostly mediated by mCPT‐1 activity rather than by mCPT‐1 content.     

Role of adenosine in exercise‐induced human skeletal muscle vasodilatation

The role of adenosine in exercise‐induced human skeletal muscle vasodilatation remains unknown. We therefore evaluated the effect of theophylline‐induced adenosine receptor blockade in six subjects and the vasodilator potency of adenosine infused in the femoral artery of seven subjects. During one‐legged, knee‐extensor exercise at ～48% of peak power output, intravenous (i.v.) theophylline decreased (P < 0.003) femoral artery blood flow (F_aBF) by ～20%, i.e. from 3.6 ± 0.5 to 2.9 ± 0.5 L min^?1, and leg vascular conductance (VC) from 33.4 ± 9.1 to 27.7 ± 8.5 mL min^?1 mmHg^?1, whereas heart rate (HR), mean arterial pressure (MAP), leg oxygen uptake and lactate release remained unaltered (P = n.s.). Bolus injections of adenosine (2.5 mg) at rest rapidly increased (P < 0.05) F_aBF from 0.3 ± 0.03 L min^?1 to a 15‐fold peak elevation (P < 0.05) at 4.1 ± 0.5 L min^?1. Continuous infusion of adenosine at rest and during one‐legged exercise at ～62% of peak power output increased (P < 0.05) F_aBF dose‐dependently to level off (P = ns) at 8.3 ± 1.0 and 8.2 ± 1.4 L min^?1, respectively. One‐legged exercise alone increased (P < 0.05) F_aBF to 4.7 ± 1.7 L min^?1. Leg oxygen uptake was unaltered (P = n.s.) with adenosine infusion during both rest and exercise. The present findings demonstrate that endogenous adenosine controls at least ～20% of the hyperaemic response to submaximal exercise in skeletal muscle of humans. The results also clearly show that arterial infusion of exogenous adenosine has the potential to evoke a vasodilator response that mimics the increase in blood flow observed in response to exercise.     

Intermittent hypoxia improves atrial tolerance to subsequent anoxia and reduces stress protein expression

We tested the hypothesis that 21 days of intermittent hypoxia (IH) increases the tolerance of the spontaneously beating guinea‐pig double atria preparation to acute in‐vitro hypoxia, and reduces cardiac stress protein expression. A total of 28 guinea‐pigs were divided into four groups: (i) IH; (ii) IH + in‐vitro hypoxia (IH + IV); (iii) control (CON); (iv) control + in‐vitro hypoxia (CON + IV). The IH animals were exposed to 8% O₂/0.3% CO₂ for 12 h day^–1 for 21 days. Normoxic controls were exposed to room air for the same duration. Acute in‐vitro hypoxia (20, 10, 5 and 0% O₂ in 5% CO₂) was introduced into the atrial preparation. Heat shock protein (Hsp) 70 and Hsp90 content were determined by Western blotting. Intermittent hypoxia groups demonstrated typical responses to chronic hypoxic exposure, characterized by significantly (P < 0.05) lower body weights, reduced growth rates and increased heart weight/body weight ratios. In the CON + IV group, in‐vitro hypoxia reduced heart rate (20% O₂, –30 ± 8 beats min^–1; 10% O₂, –34 ± 8 beats min^–1; 5% O₂, –37 ± 9 beats min^–1 and 0% O₂, –51 ± 9* beats min^–1: *P < 0.05 vs. 20% O₂). At 0% O_2, the decrease in the rate response was significantly attenuated in the IH + IV (–30 ± 8 beats min^–1; n=10) compared with the CON + IV (–51 ± 9 beats min^–1; n=10). IH significantly reduced atrial Hsp70 and Hsp90 expression, however, levels of both proteins were unchanged in the ventricle. Furthermore, Hsp90 and to a lesser degree Hsp70 in the atria remained suppressed following in‐vitro hypoxia in the IH group. Our results show that the increased resistance of the isolated atria to anoxia following IH may contribute to the concomitant reductions in basal and hypoxia‐induced Hsp expression as the overall stress response is reduced.