In dogs and rats in vivo brief coronary occlusions at the beginning of reperfusion after a myocardial ischaemia protect against ischemia/reperfusion (I/R) damage. The procedure is called ischaemic postconditioning (Post-C). This study was planned a) to compare the protection exerted by Post-C with that due to ischemic preconditioning (IP); b) to understand the mechanism of protection. Wistar rats were anesthetized with urethane and decapitated. The hearts were excised, attached to a perfusion apparatus and perfused at constant flow with oxygenated Krebs-Henseleit buffer. Left ventricular pressure (LVP)was measured. The hearts were paced at 280 bpm and kept at 37°C. Coronary flow was monitored with an electromagnetic flow-probe placed along the perfusion line. Developed LVP was taken as an index of contractility. Infarct size was determined with nitro-blue tetrazolium technique. Five groups of 10 hearts were studied. In all groups I/R was carried out with 30 min of global ischaemia and 120 min of reperfusion. Group 1 was used as a control of I/R damage. In Group 2, the hearts were postconditioned starting 15 s after the end of ischaemia. Post-C consisted of four progressively shorter (20, 15, 10 and 5 s) flow interruptions separated by increasing periods of reperfusions (10, 15 and 20 s). In Group 3, IP was made before I/R. It was obtained with 3 cycles of 3 min of occlusion separated by 5 min of reperfusion. No Post-C was carried out in this group. In Group 4, during Post-C and reperfusion the NO synthase (NOS) inhibitor L-NAME (100 μM) was added to the buffer. In Group 5, guanylyl-cyclase inhibitor ODQ (10 μM) replaced L-NAME. A comparison of Group 2 vs Group 1 revealed that Post-C improved developed LVP during reperfusion by about 61% and reduced the infarct size from 68±3% to 22±1% (p<0.01). In Group 3 IP reduced infarct size to 39±5 (P<0.01) only, i.e. to a value which was significantly higher (p<0.05) than that observed after Post-C. In Group 4, NO-inhibition only blunted the effect of Post-C, so that infarct size was reduced (p<0.01) to 37±4% only, i.e to an extent similar to that induced by IP. In Group 5 guanylyl-cyclase inhibition suppressed the effect of Post-C. L-NAME and ODQ totally abolished the Post-C mediated improvement of developed LVP after reperfusion, which did not occur after IP. It can be concluded that Post-C is more effective than IP in reducing I/R damage in isolated rat hearts. Moreover Post-C is totally achieved through cGMP, while NOS activity plays only a partial role. It may be suggested that in Post-C the activation of cGMP may occur also through a pathway independent of NOS mechanism.
GUANYLYL-CYCLASE IN ISCHAEMIC POSTCONDITIONING IN THE ISOLATED RAT HEART
PAGLIARO, Pasquale;PENNA, Claudia;CAPPELLO, SANDRA;MACERAUDI, Donatella;LOSANO, Giovanni
2004-01-01
Abstract
In dogs and rats in vivo brief coronary occlusions at the beginning of reperfusion after a myocardial ischaemia protect against ischemia/reperfusion (I/R) damage. The procedure is called ischaemic postconditioning (Post-C). This study was planned a) to compare the protection exerted by Post-C with that due to ischemic preconditioning (IP); b) to understand the mechanism of protection. Wistar rats were anesthetized with urethane and decapitated. The hearts were excised, attached to a perfusion apparatus and perfused at constant flow with oxygenated Krebs-Henseleit buffer. Left ventricular pressure (LVP)was measured. The hearts were paced at 280 bpm and kept at 37°C. Coronary flow was monitored with an electromagnetic flow-probe placed along the perfusion line. Developed LVP was taken as an index of contractility. Infarct size was determined with nitro-blue tetrazolium technique. Five groups of 10 hearts were studied. In all groups I/R was carried out with 30 min of global ischaemia and 120 min of reperfusion. Group 1 was used as a control of I/R damage. In Group 2, the hearts were postconditioned starting 15 s after the end of ischaemia. Post-C consisted of four progressively shorter (20, 15, 10 and 5 s) flow interruptions separated by increasing periods of reperfusions (10, 15 and 20 s). In Group 3, IP was made before I/R. It was obtained with 3 cycles of 3 min of occlusion separated by 5 min of reperfusion. No Post-C was carried out in this group. In Group 4, during Post-C and reperfusion the NO synthase (NOS) inhibitor L-NAME (100 μM) was added to the buffer. In Group 5, guanylyl-cyclase inhibitor ODQ (10 μM) replaced L-NAME. A comparison of Group 2 vs Group 1 revealed that Post-C improved developed LVP during reperfusion by about 61% and reduced the infarct size from 68±3% to 22±1% (p<0.01). In Group 3 IP reduced infarct size to 39±5 (P<0.01) only, i.e. to a value which was significantly higher (p<0.05) than that observed after Post-C. In Group 4, NO-inhibition only blunted the effect of Post-C, so that infarct size was reduced (p<0.01) to 37±4% only, i.e to an extent similar to that induced by IP. In Group 5 guanylyl-cyclase inhibition suppressed the effect of Post-C. L-NAME and ODQ totally abolished the Post-C mediated improvement of developed LVP after reperfusion, which did not occur after IP. It can be concluded that Post-C is more effective than IP in reducing I/R damage in isolated rat hearts. Moreover Post-C is totally achieved through cGMP, while NOS activity plays only a partial role. It may be suggested that in Post-C the activation of cGMP may occur also through a pathway independent of NOS mechanism.
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