Evaluation of nitric oxide release in the coronary effluent by a novel EPR technique: a study on isolated rat hearts subjected to cold cardioplegia and reperfusion.
Aim of this study was to investigate the cardiac release of nitric oxide (NO) before and after cold cardioplegia by a novel electron paramagnetic resonance (EPR) technique. Isolated rat hearts were perfused for 20 min in a Langendorff apparatus and then subjected to 3 hours potassium-hypotermic cardioplegia, followed by 20 min reperfusion. The coronary effluent was collected in a flask containing ferrous-bis-diethyldithiocarbamate as a spin trap of NO. Since the trapping agent was not delivered to the heart with the perfusion medium, we avoided that an abnormal extraction of NO from the tissue could inhibit its biological activity. The EPR signal was well detectable after equilibration (25.6 +/- 3.0 nmol/L +/- S.E.M.) and significantly increased following perfusion with 10 micromol/L serotonin (41.1 +/- 3.2 nmol/L) or 10 micromol/L nitroprusside (43.5 +/- 2.9 nmol/L). The basal level of NO did not change after reperfusion, but serotonin administration was not able to stimulate its release. Serotonin failure to stimulate NO production was not due to a loss of endothelial NO synthase, since its protein expression was not modified after reperfusion. The perfusion pressure increased by 51% after reperfusion and was quite completely restored following serotonin or nitroprusside treatment, with respect to the non-stimulated equilibration condition. Therefore, we suggest that the coronary spasm following a cold cardioplegic arrest is not due to an impaired production of basal NO and that NO-donors can be effective in relaxing vascular smooth muscle cells.
Muscari
Claudio
C
Department of Biochemistry, University of Bologna, Via Irnerio 48, 40126 Bologna, Italy. claudio.muscari@unibo.it
Grossi
Loris
L
Giordano
Emanuele
E
Ferrari
Daniela
D
Bonafe
Francesca
F
Guarnieri
Carlo
C
Caldarera
Claudio Marcello
CM
D000818:Animals; D015153:Blotting, Western; D003080:Cold Temperature; D004578:Electron Spin Resonance Spectroscopy; D006324:Heart Arrest, Induced; D066298:In Vitro Techniques; D015425:Myocardial Reperfusion; D009569:Nitric Oxide / Q000378:metabolism*; D051381:Rats; D013055:Spectrophotometry, Infrared
D009569:Nitric Oxide