The influence of contraction, pH and enzyme inhibition on the release of adenosine from rat gracilis muscle

published_or_final_versionPhysiologyDoctoralDoctor of Philosoph

Intracellular adenosine formation and release by freshly-isolated vascular endothelial cells from rat skeletal muscle: effects of hypoxia and/or acidosis

Previous studies suggested indirectly that vascular endothelial cells (VECs) might be able to release intracellularly-formed adenosine. We isolated VECs from the rat soleus muscle using collagenase digestion and magnetic-activated cell sorting (MACS). The VEC preparation had >90% purity based on cell morphology, fluorescence immunostaining, and RT-PCR of endothelial markers. The kinetic properties of endothelial cytosolic 5’-nucleotidase suggested it was the AMP-preferring N-I isoform: its catalytic activity was 4 times higher than ecto-5’nucleotidase. Adenosine kinase had 50 times greater catalytic activity than adenosine deaminase, suggesting that adenosine removal in VECs is mainly through incorporation into adenine nucleotides. The maximal activities of cytosolic 5’-nucleotidase and adenosine kinase were similar. Adenosine and ATP accumulated in the medium surrounding VECs in primary culture. Hypoxia doubled the adenosine, but ATP was unchanged; AOPCP did not alter medium adenosine, suggesting that hypoxic VECs had released intracellularly-formed adenosine. Acidosis increased medium ATP, but extracellular conversion of ATP to AMP was inhibited, and adenosine remained unchanged. Acidosis in the buffer-perfused rat gracilis muscle elevated AMP and adenosine in the venous effluent, but AOPCP abolished the increase in adenosine, suggesting that adenosine is formed extracellularly by non-endothelial tissues during acidosis in vivo. Hypoxia plus acidosis increased medium ATP by a similar amount to acidosis alone and adenosine 6-fold; AOPCP returned the medium adenosine to the level seen with hypoxia alone. These data suggest that VECs release intracellularly formed adenosine in hypoxia, ATP during acidosis, and both under simulated ischaemic conditions, with further extracellular conversion of ATP to adenosine

Elevation of arterial potassium during acute systemic hypoxia is abolished by alkalosis

Background: Small elevations in plasma potassium evoke vasodilation in the peripheral circulation. Systemic hypoxia elevates arterial potassium and also modifies arterial pH. Aims: We examined the interaction between pH and potassium in blood during systemic hypoxia and the effect of pH on the uptake/release of potassium in the peripheral tissues. Methods: Anesthetized dogs were ventilated with air plus oxygen for normoxia or air plus nitrogen for hypoxia. Some animals received intravenous sodium bicarbonate to elevate pH by 0.1 units. Arterial plasma potassium concentration was measured in normoxia and hypoxia. A rat gracilis muscle was perfused with normoxic Krebs buffer and the potassium content of the venous outflow was compared during perfusion at pH 7.4, 6.8, or 7.8. Results: In dogs with an arterial pH of 7.40-7.45, systemic hypoxia elevated the arterial potassium by 1 mmol/L. An arterial pH of 7.55 did not alter the basal potassium concentration, but it abolished the hypoxia-induced increase. In rat muscle, reduction of the perfusate pH from 7.4 to 6.8 reduced arterial perfusion pressure from 8.73 to 7.32 kPa and venous potassium from 6.6 to 5.2 mM. Elevation of perfusate pH to 7.8 decreased the arterial perfusion pressure from 8.44 to 6.95 kPa but did not affect venous potassium. Conclusions: The hypoxia-induced elevation of arterial potassium is abolished by increasing the pH to 7.55. This is not due to enhanced potassium uptake into peripheral tissues at high pH. Red blood cells are suggested as the most likely source of the potassium released in hypoxia.link_to_subscribed_fulltex

Prevalence of and risk factors for chronic arthralgia and rheumatoid-like polyarthritis more than 2 years after infection with chikungunya virus

Abstract Objectives Chikungunya virus (CHIKV), transmitted to humans from infected mosquitoes, causes acute fever, arthralgia and rash. There is increasing evidence that it also causes longer-term rheumatic symptoms. In a circumscribed part of Mauritius where infectivity was high, a cohort of inhabitants was surveyed with the objectives of assessing the prevalence of and risk factors for chronic musculoskeletal symptoms and for a rheumatoid arthritis-like condition at 27.5 months after initial infection. Methods Participants were recruited May–November 2008 and invited to complete a questionnaire. CHIKV was diagnosed clinically. The primary outcomes for the analyses were (a) self-reported ongoing musculoskeletal symptoms and (b) fulfilment of modified diagnostic criteria for rheumatoid arthritis. Risk factors for these outcomes were explored in univariate analyses using logistic regression. Subsequently, multivariate logistic regression was used to identify factors that were independently associated with the outcomes. Results 173 individuals were identified with CHIKV, of whom 136 (78.6%) reported persisting musculoskeletal symptoms 27.5 months after infection. Persistent symptoms were associated with older age at time of infection, female gender and baseline symmetrical distribution of joint symptoms. We found that 5% of those infected with CHIKV fulfilled a modified version of the American College of Rheumatology criteria for rheumatoid arthritis 27.5 months after infection. Conclusions CHIKV is associated with a high prevalence of persistent rheumatic symptoms. Physicians need to be aware of CHIKV as a cause of acute and chronic rheumatic symptoms. </jats:sec