No abstract

It is widely accepted that adenosine is released within tissues under hypoxic conditions. Accordingly, several studies in which adenosine receptor antagonists or adenosine deaminase have been used indicate that locally released adenosine makes important contributions to the respiratory and cardiovascular responses that are induced by systemic hypoxia. For example, adenosine has been shown to contribute to the central respiratory depression of systemic hypoxia, by actions within the brain, and to play a part in the bradycardia and myocardial depression by local actions on the sino-atrial node and ventricular muscle (see Neylon & Marshall, 1991; and references therein). In these ways the actions of adenosine tend to counteract the hyperventilation and tachycardia that are the reflex, neurally mediated responses to systemic hypoxia (Marshall, 1994). In addition, experiments on the rat have indicated that locally released adenosine is responsible for at least 50% of the vasodilatation that occurs in skeletal muscle during systemic hypoxia and which overcomes the reflex vasoconstriction evoked by sympathetic nerve activation 1. In anaesthetized rats we tested responses evoked by systemic hypoxia (breathing 8% Oµ for 5 min) and adenosine (i.a. infusion for 5 min) before and after administration of a selective adenosine AÔ receptor antagonist DPCPX (8-cyclopentyl-1,3-dipropylxanthine), or a selective adenosine A2A receptor antagonist ZM 241385. Arterial blood pressure, (ABP), heart rate (HR), femoral blood flow (FBF) and femoral vascular conductance (FVC: FBFÏABP) were recorded together with the K¤ concentration in arterial blood ([K¤]a) and in venous blood of hindlimb muscle ([K¤]v) before and at the 5th minute of hypoxia or agonist infusion. 2. In 12 rats, DPCPX reversed the fall in ABP and HR and the increase in FVC evoked by the selective AÔ agonist CCPA (2-chloro-N É-cyclopentyladenosine; i.a. infusion for 5 min). DPCPX also reduced both the increase in FVC induced by hypoxia and that induced by adenosine; the control responses to these stimuli were comparable in magnitude and both were reduced by •50%. 3. In 11 rats, ZM 241385 reversed the fall in ABP and increase in FVC evoked by the selective A2A agonist CGS 21680 (2-p-(2-carboxyethyl)-phenethylamino-5'-N-ethylcarboxamidoadenosine hydrochloride; i.a. infusion for 5 min). ZM 241385 also reduced the increase in FVC induced by adenosine by •50%, but had no effect on the increase in FVC induced by hypoxia. 4. In these same studies, before administration of DPCPX, or ZM 241385, hypoxia had no effect on the venous-arterial difference for K¤ ([K¤]v-a), whereas after administration of either antagonist, hypoxia significantly reduced [K¤]v-a suggesting an increase in hypoxiainduced K¤ uptake, or a reduction in K¤ efflux. 5. These results indicate that both AÔ and A2A receptors are present in hindlimb muscle and can mediate vasodilatation and that AÔ and A2A receptors contribute equally to dilatation induced by infused adenosine. However, they suggest that endogenous ade...

1. Direct observations have been made on the responses of individual vessels of the microcirculation of rat spinotrapezius muscle to stimulation of the sympathetic paravascular nerve fibres and to topically applied catecholamines. 2. All arteries and arterioles were constricted by sympathetic stimulation, the maximum response occurring at a stimulus frequency of 8‐10 Hz. Primary and secondary arterioles (13‐50 μm internal diameter) showed the greatest percentage change in diameter and remained constricted throughout the 1 min stimulation period whilst terminal arterioles (7‐13 μm internal diameter) constricted initially but then returned towards their control diameter before the stimulus ceased. 3. By contrast the venules and veins showed no response to sympathetic stimulation. 4. In accord with these observations, fluorescence histochemical studies revealed a network of noradrenergic nerve fibres on all arterial vessels but showed no innervation of any venous vessels. 5. Topically applied noradrenaline or adrenaline (10−10‐10−8 g/ml.) dilated the majority of arteries and arterioles while higher concentrations of either agent produced dose‐dependent constrictor responses. In addition, many venules dilated in response to adrenaline (10−9 g/ml.) while others constricted, but concentrations of either noradrenaline or adrenaline greater than 10−8 g/ml. produced dose‐dependent constriction of all venules and veins. 6. The behaviour of the more proximal and more distal arterioles during sympathetic stimulation is in accord respectively with the changes in muscle vascular resistance and in capillary surface area recorded in previous studies during sympathetic stimulation. 7. The observation that venous vessels are not influenced by sympathetic nerve fibres contrasts with the established view that venous vessels of skeletal muscle are strongly constricted during sympathetic stimulation. However, reappraisal of the evidence used to support this view indicates that such results may have been misinterpreted, while other available evidence supports the proposal that the present findings may be representative of skeletal muscle vasculature in general.

Adenosine, prostaglandins (PG) and nitric oxide (NO) have all been implicated in hypoxia-evoked vasodilatation. We investigated whether their actions are interdependent. In anaesthetised rats, the PG synthesis inhibitors diclofenac or indomethacin reduced muscle vasodilatation evoked by systemic hypoxia or adenosine, but not that evoked by iloprost, a stable analogue of prostacyclin (PGI 2 ), or by an NO donor. After diclofenac, the A 1 receptor agonist CCPA evoked no vasodilatation: we previously showed that A 1 , but not A 2A , receptors mediate the hypoxia-induced muscle vasodilatation. Further, in freshly excised rat aorta, adenosine evoked a release of NO, detected with an NO-sensitive electrode, that was abolished by NO synthesis inhibition, or endothelium removal, and reduced by ~50 % by the A 1 antagonist DPCPX, the remainder being attenuated by the A 2A antagonist ZM241385. Diclofenac reduced adenosine-evoked NO release bỹ 50 % under control conditions, abolished that evoked in the presence of ZM241385, but did not affect that evoked in the presence of DPCPX. Adenosine-evoked NO release was also abolished by the adenyl cyclase inhibitor 2‚,5‚-dideoxyadenosine, while dose-dependent NO release was evoked by iloprost. Finally, stimulation of A 1 , but not A 2A , receptors caused a release of PGI 2 from rat aorta, assessed by radioimmunoassay of its stable metabolite, 6-keto PGF 1a , that was abolished by diclofenac. These results suggest that during systemic hypoxia, adenosine acts on endothelial A 1 receptors to increase PG synthesis, thereby generating cAMP, which increases the synthesis and release of NO and causes muscle vasodilatation. This pathway may be important in other situations involving these autocoids.

SUMMARY1. In rats anaesthetized with Saffan, we have further analysed the respiratory, cardiac and regional vascular responses induced by 3 min periods of graded hypoxia (breathing 15, 12, 8 or 6% 02 in N2).2. Frequently, hypoxia evoked an episode, lasting 1 15 min, of tachycardia, renal and mesenteric vasoconstriction and skeletal muscle vasodilatation. The tachycardia and muscle vasodilatation persisted after vagotomy indicating they were not initiated by pulmonary stretch receptors secondary to hyperventilation. We propose that such episodes represented the cardiovascular components of the alertingdefence response initiated by activation of the brain stem defence areas by peripheral chemoreceptors.3. Each of these episodes was superimposed upon gradual hyperventilation, tachycardia. fall in arterial pressure and vasodilatation in renal, mesenteric and muscle circulation the magnitudes of which at 2 min were generally graded with the level of hypoxia. In the 3rd minute, respiration and heart rate tended to wane below control lev,els.4. Vagotomy had little effect on the heart rate changes and only slightly reduced the peripheral vasodilatation allowing the conclusion that the gra(lual tachycardia and peripheral vasodilatation was not a reflex initiated by pulmonary stretch receptors.5. Guanethidine given after vagotomy abolished the tachycardia indicating it was sympathetically mediated; possible initiating factors are discussed. But the secondary bradycardia persisted indicating it reflected the direct effect of hypoxia on cardiac pacemaker tissue.

1. In anaesthetized rats, systemic hypoxia evoked hyperventilation, tachycardia, a fall in arterial pressure, vasodilatation in skeletal muscle and increases in K+ concentration measured in arterial plasma ([K+]a), venous efflux from muscle ([K+]v) and in right atrial plasma ([K+]at). The ATP-sensitive potassium (K+ATP) channel inhibitor glibenclamide (10 or 20 mg kg-1 i.v.) reduced the muscle vasodilatation and increase in [K+]v, but had no significant effect on the other changes. 2. The adenosine receptor antagonist, 8-phenyltheophylline (8-PT, 10 mg kg-1 i.v.) had similar effects to glibenclamide. 3. Glibenclamide reduced the muscle vasodilatation evoked by the adenosine analogue, 2-chloroadenosine given i.v. (30 micrograms kg-1). 4. Infusion of adenosine (0.3 mg kg-1 min-1 for 5 min) into the hindlimb evoked muscle vasodilatation and an increase in [K+]v, both of which were abolished by 8-PT. 5. We propose that during systemic hypoxia, part of the muscle vasodilatation that can be attributed to adenosine is due to the action of K+, which is released from skeletal muscle fibres through glibenclamide-sensitive K+ channels (possibly K+ATP channels) that are activated by adenosine. This may be a general mechanism for the vasodilator influence of adenosine.

1. In Saffan-anaesthetized rats, we have further investigated the mechanisms underlying the vasodilatation induced by adenosine in skeletal muscle by acute systemic hypoxia (breathing 8% O2 for 5 min). 2. In eleven rats the nitric oxide (NO) synthesis inhibitor nitro-L-arginine methyl ester (L-NAME, 10 mg kg-1, i.v.) reduced the increase in femoral vascular conductance (FVC) induced by hypoxia by approximately 50%. L-NAME had similar effects on the increase in FVC induced by intra-arterial (I.A.) infusion of adenosine (at 1.2 mg kg-1 min-1 for 5 min via the tail artery) and by ATP (I.A., 1 mg kg-1 min-1 for 5 min). Subsequent administration of the adenosine receptor antagonist 8-sulphophenyl theophylline (8-SPT, 20 mg kg-1, i.v.) virtually abolished the adenosine- and ATP-induced increase in FVC. 3. In a further nine rats, 8-SPT reduced the increase in FVC induced by hypoxia by approximately 50%. This remaining increase in FVC was substantially reduced by L-NAME. 4. In an additional nine rats, alpha,beta-methyleneADP (160 micrograms kg-1, i.v.) which inhibits the 5'-ectonucleotidase that degrades AMP to adenosine, reduced the peripheral vasodilatation (fall in arterial blood pressure, ABP) induced by ATP infusion, but had no effect on the increase in FVC or decrease in ABP evoked by systemic hypoxia. 5. These results provide the first evidence that the muscle vasodilatation induced by adenosine during systemic hypoxia is mainly dependent on NO synthesis. They also suggest that adenosine is released as such rather than being formed extracellularly from AMP. Given evidence that extraluminal adenosine acts in an NO-independent fashion we propose that hypoxia releases adenosine from the endothelium. Our results also indicate that hypoxia induces muscle vasodilatation that is adenosine independent but NO dependent: they allow the possibility that this is partly mediated by ATP released from the endothelium.

SUMMARY1. In rats anaesthetized with Saffan we have studied the effects of the adenosine receptor antagonists, theophylline and 8-phenyltheophylline, upon the respiratory and cardiovascular responses evoked by 5 min periods of systemic hypoxia.2. In the group of animals that were to receive theophylline (15 mg kg-' i.v.), arterial 02 pressure (Pal 02) fell from 83 + 2 mmHg during air breathing to 38 + 3 or 34 + 3 mmHg during the 5th minute of two different control periods of hypoxia, while in the group that were to receive 8-phenyltheophylline (10 mg kg-' i.v.), Pa 02 fell from 83 + 1 to 53 + 2 mmHg. Neither drug significantly altered the levels of Pa, 2 reached during hypoxia. 3. During the control periods of hypoxia respiration increased, but the increase evoked at the 5th minute was significantly less than that evoked at the 2nd minute of hypoxia. This secondary waning of the hyperventilation was abolished by both drugs.4. Similarly, both drugs attenuated the tendency for the hypoxia-induced tachyeardia to wane between the 2nd and 5th minute.5. Further, both drugs substantially reduced both the hypoxia-induced fall-in arterial pressure and the increases in vascular conductance in hindlimb muscle, carotid vasculature and kidney.6. Thus, we propose that in the rat the release of adenosine by hypoxic tissues makes a major contribution to the secondary decrease in respiration and heart rate that occurs during systemic hypoxia and to the accompanying vasodilatation in muscle and fall in arterial pressure. The effects of the adenosine antagonists on the carotid and renal vasculature are more equivocal and may be partly explained as a smaller autoregulatory dilatation to a smaller fall in systemic arterial pressure.7. These results and proposals are discussed in relation to the conditions that are known to cause release of adenosine and in relation to its known effects upon the respiratory and cardiovascular systems.