It has been reported that reactive nitrogen oxide species can lead to ischaemic preconditioning (IP) (Pagliaro et al. 2001). However, it is unknown whether nitroxyl anion (NO^–), the one-electron reduction product of nitric oxide (NO), is involved in protection by IP. Since NO^– can be generated from chemical reactions that occur in vivo, which are influenced by tissue redox conditions, we tested whether NO^– may also trigger the protective effects of IP in isolated rat hearts.

Twenty-four male Wistar rats, 5-6 months old (450-550 g body weight), were decapitated after urethane (0.5 g I.P.) anaesthesia. Then the hearts were excised, suspended as Langendorff preparations, retrogradely perfused at constant flow (9 ± 1 ml min^-1 g^-1) with Tyrode oxygenated solution at 37 °C, and paced at 280 b.p.m. A polyvinyl chloride balloon was placed in the left ventricle via the mitral valve to record left ventricular pressure (LVP). Angeli’s salt (AS) was used to generate NO^– in solution.

The hearts were divided into three groups. All hearts underwent ischaemia/reperfusion (I/R) consisting of 30 min of global ischaemia and 30 min of reperfusion. In Group 1 (n = 9), I/R was preceded by 19 min of infusion of the vehicle used for AS; in Group 2 (n = 7) I/R was preceded by three cycles of 3 min of global ischaemia followed by 5 min reperfusion; in Group 3 (n = 8), I/R was preceded by 19 min of infusion of AS (1 mM). The size of the infarct produced by I/R was assessed on slices of left ventricle incubated in nitro-blue tetrazolium (0.1 % in phosphate buffer). Statistical analysis was performed by one-way ANOVA with Bonferroni correction. Data are means ± S.E.M.

As seen in vivo (Paolocci et al. 2001), AS per se exerted a positive inotropic effect revealed by a ¦10 % (P < 0.05) increase in developed LVP. However, post-ischaemic reduction in developed LVP was not different in the three groups. In contrast, during ischaemia end-diastolic LVP increased by 11 ± 5 mmHg in Group 1, by 8 ± 3 in Group 2 and only by 6 ± 3 in Group 3. The limited increase of end-diastolic LVP in preconditioned and in AS-pretreated hearts was considered an index of limited contracture due to increased resistance to ischaemia. In fact, while in Group 1 necrotic tissue was 30 ± 3 %, it was only 12 ± 3 % in Group 2 (P < 0.01 vs. Group 1), and 19 ± 2 % in Group 3 (P = n.s. vs. IP; P < 0.05 vs. Group 1). Our data are consistent with NO^– being a preconditioning agent and support the hypothesis that NO^– production is enhanced during IP and contributes to the triggering of protection.

Pagliaro, P., Gattullo, D., Rastaldo, R. & Losano, G. (2001). Life Sci. 69, 1-15.

Paolocci, N., Saavedra, F.W., Miranda, K.M., Martignani, C., Isoda, T., Hare, J.M., Espey, M.G., Fukuto, J.M., Feelisch, M., Wink, D.A. & Kass, D.A. (2001). Proc. Natl Acad. Sci. USA 98, 10463-10468.