Effect of nitrovasodilators and inhibitors of nitric oxide synthase on ischaemic and reperfusion function of rat isolated hearts

Abstract

1 The functional role of the nitric oxide (NO)/guanosine 3':5'-cyclic monophosphate (cyclic GMP) pathway in experimental myocardial ischaemia and reperfusion was studied in rat isolated hearts. 2 Rat isolated hearts were perfused at constant pressure with Krebs-Henseleit bu er for 25 min (baseline), then made ischaemic by reducing coronary¯ow to 0.2 ml min 71 for 25 or 40 min, and reperfused at constant pressure for 25 min. Drugs inhibiting or stimulating the NO/cyclic GMP pathway were infused during the ischaemic phase only. Ischaemic contracture, myocardial cyclic GMP and cyclic AMP levels during ischaemia, and recovery of reperfusion mechanical function were monitored. 3 At baseline, heart rate was 287+12 beats min 71 , coronary¯ow was 12.8+0.6 ml min 71 , left ventricular developed pressure (LVDevP) was 105+4 mmHg and left ventricular end-diastolic pressure 4.6+0.2 mmHg in vehicle-treated hearts (control; n=12). Baseline values were similar in all treatment groups (P40.05). 4 In normoxic perfused hearts, 1 mM N G -nitro-L-arginine (L-NOARG) signi®cantly reduced coronarȳ ow from 13.5+0.2 to 12.1+0.1 ml min 71 (10%) and LVDevP from 97+1 to 92+1 mmHg (5%; P50.05, n=5). 5 Ischaemic contracture was 46+2 mmHg, i.e. 44% of LVDevP in control hearts (n=12), una ected by low concentrations of nitroprusside (1 and 10 mM) but reduced to *30 mmHg (*25%) at higher concentrations (100 or 1000 mM; P50.05 vs control, n=6). Conversely, the NO synthase inhibitor L-NOARG reduced contracture at 1 mM to 26+3 mmHg (23%), but increased it to 63+4 mmHg (59%) at 1000 mM (n=6). Dobutamine (10 mM) exacerbated ischaemic contracture (81+3 mmHg; n=7) and the cyclic GMP analogue Sp-8-(4-p-chlorophenylthio)-3',5'-monophosphorothioate (Sp-8-pCPT-cGMPS; 10 mM) blocked this e ect (63+1 mmHg; P50.05 vs dobutamine alone, n=5). 6 At the end of reperfusion, LVDevP was 58+5 mmHg, i.e. 55% of pre-ischaemic value in control hearts, signi®cantly increased to *80% by high concentrations of nitroprusside (100 or 1000 mM) or L-NOARG at 1 mM, while a high concentration of L-NOARG (1000 mM) reduced LVDevP to *35% (P50.05 vs control; n=6). 7 Ischaemia increased tissue cyclic GMP levels 1.8 fold in control hearts (P50.05; n=12); nitroprusside at 1 mM had no sustained e ect, but increased cyclic GMP *6 fold at 1000 mM; L-NOARG (1 or 1000 mM) was without e ect (n=6). Nitroprusside (1 or 1000 mM) marginally increased cyclic AMP levels whereas NO synthase inhibitors had no e ect (n=6). 8 In conclusion, the cardioprotective e ect of NO donors, but not of low concentrations of NO synthase inhibitors may be due to their ability to elevate cyclic GMP levels. Because myocardial cyclic GMP levels were not a ected by low concentrations of NO synthase inhibitors, their bene®cial e ect on ischaemic and reperfusion function is probably not accompanied by reduced formation of NO and peroxynitrite in this model.

Postischemic Recovery and Oxidative Stress Are Independent of Nitric-Oxide Synthases Modulation in Isolated Rat Heart

“…D-NAME 100 M n ϭ 14 122.9 Ϯ 2.0 138.9 Ϯ 6.0 29.5 Ϯ 6.8 20 Ϯ 4 L-NAME 100 M n ϭ 15 127.5 Ϯ 4.9 107.9 Ϯ 9.2* 32.6 Ϯ 7.2 38 Ϯ 9 D-arginine 3 mM n ϭ 6 124.2 Ϯ 5.6 114.3 Ϯ 13.0 23.5 Ϯ 5.7 22 Ϯ 7 L-arginine 3 mM n ϭ 6 132.8 Ϯ 5.9 123.7 Ϯ 8.3 11.2 Ϯ 3.8 9 Ϯ 3 Heart rate (beats/min) D-NAME 100 M n ϭ 14 329.3 Ϯ 5.69 283.8 Ϯ 9.1 221.5 Ϯ 8.8 L-NAME 100 M n ϭ 15 326.7 Ϯ 7.4 258.7 Ϯ 10.9 196.9 Ϯ 13.9 D-arginine 3 mM n ϭ 6 323. du Toit et al, 1998), whereas concentrations above 100 M would be detrimental. Moreover, when using vasoactive concentrations of NOS inhibitors, the result is considered as beneficial in terms of postischemic recovery (Zweier et al, 1995;Wang and Zweier, 1996), or deleterious when expressed in net values (Brunner et al, 1997;du Toit et al, 1998), as was experienced in our study.…”

“…du Toit et al, 1998), whereas concentrations above 100 M would be detrimental. Moreover, when using vasoactive concentrations of NOS inhibitors, the result is considered as beneficial in terms of postischemic recovery (Zweier et al, 1995;Wang and Zweier, 1996), or deleterious when expressed in net values (Brunner et al, 1997;du Toit et al, 1998), as was experienced in our study. Therefore, because L-NAME treatment does not modify the duration of postischemic rhythm disturbances, we propose that the administration of 100 M L-NAME before 30 min of global ischemia and throughout reperfusion does not afford protection to the heart against postischemic injury.…”

“…The evolution of contractility was not different between L-or D-NAMEtreated hearts during reperfusion, but the recovery of preischemic RPP was slightly higher for the L-NAME (40 Ϯ 11%) than for the D-NAME groups (29 Ϯ 11%). Various competitive inhibitors of NOS have been shown to reduce reperfusion injury in various settings (Depré et al, 1995;Naseem et al, 1995;Zweier et al, 1995;Wang and Zweier, 1996;du Toit et al, 1998;Zhang et al, 2001). In contrast to the above-mentioned studies, some experiments reported that NOS antagonists can enhance postischemic myocardial injury (Brunner et al, 1997;du Toit et al, 1998).…”

“…Various competitive inhibitors of NOS have been shown to reduce reperfusion injury in various settings (Depré et al, 1995;Naseem et al, 1995;Zweier et al, 1995;Wang and Zweier, 1996;du Toit et al, 1998;Zhang et al, 2001). In contrast to the above-mentioned studies, some experiments reported that NOS antagonists can enhance postischemic myocardial injury (Brunner et al, 1997;du Toit et al, 1998). The differences may be due to the experimental models, but also to the concentration of NOS antagonists, because nonvasoactive concentrations seem to protect the heart against postischemic injury (Depré TABLE 2 Evolution of myocardial functional parameters of NAME (100 M)-or arginine (3 mM)-treated hearts for the groups receiving PBN (3 mM) as a spin trap Data are presented as means Ϯ S.E.M.…”

“…All three isoforms of NOS may be expressed in the heart (for review, see Shah and MacCarthy, 2000), albeit in a cell-specific manner, and the numerous physiological effects of ⅐ NO on cardiac function have been reviewed (Kelly et al, 1996). During myocardial reperfusion, the role of ⅐ NO in the development of myocardial injury has been extensively studied in different experimental models, using either NOS antagonists (Depré et al, 1995;Naseem et al, 1995;Zweier et al, 1995;Wang and Zweier, 1996;Brunner et al, 1997;du Toit et al, 1998;Zhang et al, 2001), L-arginine (Takeuchi et al, 1995;Engelman et al, 1996;Brunner et al, 1997;Wang et al, 1997;Mizuno et al, 1998), ⅐ NO donors (Brunner et al, 1997;du Toit et al, 1998), or NOS-knockout mice models (Flögel et al, 1999;Kanno et al, 2000). However, there are contradictory results concerning the possible protective or deleterious role of ⅐ NO during ischemia and reperfusion.…”