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Aim In inflamed and damaged cardiovascular tissues, local extracellular adenosine concentrations increase coincidentally with activation of the transcription factor nuclear factor kappa B (NFκB). To investigate whether adenosine influences NFκB activation in vascular smooth muscle cells (VSMCs) and, if so, to examine the role of its receptors. Methods VSMCs were isolated from NFκB–luciferase reporter mice, cultured and then treated by lipopolysaccharide (LPS) to activate NFκB signalling. Adenosine, adenosine receptor agonists and antagonists, adenosine deaminase and uptake inhibitors were used together with LPS to evaluate the role of adenosine and its receptors on NFκB activation, which was assessed by luciferase activity and NFκB target gene expression. Results Adenosine potentiated LPS-induced NFκB activation. This was dependent on adenosine uptake and enhanced by an adenosine deaminase inhibitor, suggesting that intracellular adenosine plays an important role. Non-selective adenosine receptor agonists (2Cl-Ado and NECA) inhibited NFκB activation induced by LPS. Selective A1 or A2A antagonist given alone could not completely antagonize the NECA effect, indicating that the inhibitory effect was due to multiple adenosine receptors. The activation of the A3 receptor further increased LPS-induced NFκB activation. Conclusions Adenosine increases LPS-induced nuclear factor kappa B activation in smooth muscle cells via an intracellular mechanism and decreases it via actions on A1 and A2A receptors. These results provide novel insights into the role of adenosine as a regulator of inflammation-induced NFκB activation.

Aim: To investigate whether ischaemic post-conditioning (IPoC) combined with i.v. infusion of the nitric oxide (NO) substrate l-arginine at the onset of reperfusion exerts cardioprotective effect that is superior to either treatment given separately. Methods: Twenty-six anesthetized pigs were subjected to coronary artery (left anterior descending artery, LAD) ligation for 40 min followed by 4 h reperfusion. The pigs were randomized into five different groups receiving either i.v. vehicle, i.v. l-arginine, IPoC 4 × 60 s together with i.v. vehicle or IPoC together with i.v. l-arginine and a group with IPoC 8 × 30 s. All infusions were started 10 min before reperfusion. Results: The infarct size of the vehicle group was 82 ± 4% of the area at risk. l-Arginine alone (79 ± 8%), IPoC 4 × 60 s vehicle (86 ± 3%) or IPoC 8 × 30 s vehicle (94 ± 7%) did not affect infarct size. l-Arginine together with IPoC significantly reduced infarct size to 59 ± 4% (P

Aim: To examine the phenotype of mice that lack the adenosine A3 receptor (A3R). Methods: We examined the heart rate, body temperature and locomotion continuously by telemetry over several days. In addition, the effect of the adenosine analogue R-N6-phenylisopropyl-adenosine (R-PIA) was examined. We also examined heat production and food intake. Results: We found that the marked diurnal variation in activity, heart rate and body temperature, with markedly higher values at night than during day time, was reduced in the A3R knock-out mice. Surprisingly, the reduction in heart rate, activity and body temperature seen after injection of R-PIA in wild type mice was virtually eliminated in the A3R knock-out mice. The marked reduction in activity was associated with a decreased heat production, as expected. However, the A3R knock-out mice, surprisingly, had a higher food intake but no difference in body weight compared to wild type mice. Conclusions: The mice lacking adenosine A3 receptors exhibit a surprisingly clear phenotype with changes in diurnal rhythm and temperature regulation. Whether these effects are due to a physiological role of A3 receptors in these processes or whether they represent a role in development remains to be elucidated.

Aim Blood pressure is higher in A(1) receptor knock-out (A(1)R-/-) mice than in wild type litter mates (A(1)R+/+) and we have examined if this could be related to altered vascular functions. Methods Contraction of aortic rings and mesenteric arteries were examined. To examine if the adenosine A(1) receptor-mediated contraction of aortic muscle was functionally important we examined pulse pressure (PP) and augmentation index (AIX) using a sensor that allows measurements of rapid pressure transients. Results Contraction of aortic rings to phenylephrine and relaxation to acetylcholine were similar between genotypes. The non-selective adenosine receptor agonist N-ethyl carboxamido adenosine (NECA) enhanced the contractile response, and this was eliminated in aortas from A(1)R-/- mice. However, in mesenteric arteries no contractile response was seen and adenosine-mediated relaxation was identical between studied genotypes. A(2B) adenosine receptors, rather than A(2A) receptors, may be mainly responsible for the vasorelaxation induced by adenosine analogues in the examined mouse vessels. PP was higher in A(1)R-/- mice, but variability was unaltered. AIX was not different between genotypes, but the NECA-induced fall was larger in A(1)R-/- mice. Conclusions The role of adenosine A(1) receptors in regulating vessel tone differs between blood vessels. Furthermore, contractile effects on isolated vessels cannot explain the blood pressure in A(1) knock-out mice. The A(1) receptor modulation of blood pressure is therefore mainly related to extravascular factors.

Aims: Adenosine is involved in classic pre-conditioning (PC) in most species, acting through especially adenosine A 1 and A 3 receptors. We studied whether the adenosine A 1 receptor (A 1 R) was important for remote, delayed adaptation to ischaemia using a mouse with targeted deletion of the A 1 R gene. Methods: Remote, delayed adaptation was evoked by brain ischaemia (BIPC) through bilateral ligation of the internal carotid arteries. Through microdialysis probes placed in the brain and the abdominal aorta, we found that plasma adenosine increased following carotid artery ligation. Twenty-four hours after ligation, hearts were isolated, Langendorff perfused and subjected to 40 min global ischaemia and 60 min reperfusion. Hearts from sham operated and BIPC animals either with (A 1 R +/+ ) or without (A 1 R -/- ) the gene for the adenosine A 1 R were compared with each other. Results: In wild types, BIPC reduced infarct size and improved functional recovery during reperfusion, but BIPC did not protect hearts of A 1 R -/- mice. There were no significant differences between sham-operated A 1 R +/+ and A 1 R -/- in recovery of function or infarct size. The mitogen-activated protein kinases (MAPKs) extracellular signal-regulated protein kinasel/2 (ERK1/2), p38 and c-jun N-terminal kinase (JNK) were phosphorylated during reperfusion of sham treated hearts. The increase in ERK1/2 and p38 phosphorylation detected was attenuated in hearts of BIPC or A 1 R -/- animals. Conclusion: During BIPC adenosine acting on the A 1 R appears necessary for myocardial protection. MAPK signalling may possibly be involved in organ protection during the delayed phase of remote, delayed adaptation.