Calcium sensitisation impairs diastolic relaxation in post-ischaemic myocardium: implications for the use of Ca(2+) sensitising inotropes after cardiac surgery

Abstract

Background: Calcium sensitising inotropes are increasingly being used in cardiac surgical patients. Theoretically, increasing contractile protein sensitivity to Ca(2+) prevents the Ca(2+) elevation associated arrhythmogenicity and potentiates the inotropic effect of catecholamines. On the other hand, we hypothesised that Ca(2+) sensitisation exacerbates post-ischaemic myocardial stunning by impairing diastolic relaxation, which might have deleterious effects in postoperative cardiac surgical patients.

Methods: In an isolated rabbit heart model, 45 min normothermic ischaemia with potassium-induced cardioplegic arrest was followed by 120 min reperfusion. Isovolumetric left ventricular (LV) function and myocardial oxygen consumption (MvO(2)) were measured, and cytosolic Ca(2+) was monitored by rhod-2 surface spectrofluorometry. During reperfusion, ORG 30029 (250 microM) and levosimendan (0.5 microM) were used as Ca(2+) sensitisers (ORG, n=6, Levo, n=6), Ca(2+) de-sensitisation was induced with butanedione-monoxime (5mM, BDM, n=6), and dopamine (20 nM) served as a representative catecholamine (n=6). To counteract the PDE III inhibiting properties of ORG and Levo, IGF-1 (0.1 microM) and parathyroid hormone (0.05 microM) were used.

Results: As expected, ischaemia/reperfusion induced moderate cytosolic calcium overload. Dopamine increased LV contractility and MvO(2) by augmenting the amplitude of the Ca(2+) transient, but relaxation was unchanged due to faster diastolic Ca(2+) removal. Dopamine-induced Ca(2+) handling was unchanged after uncoupling the Mg-ATPase with BDM, and MvO2 decreased in proportion with the reduced LV mechanical work load. ORG improved contractility without apparent effects on Ca(2+) handling, and MvO(2) remained constant despite increased contractile work. Conversely, ORG induced a rightward shift of the diastolic pressure-volume relationship in post-ischaemic hearts (diastolic pressure at 0.8 ml balloon volume 14.3+/-5 mmHg, p=0.01 vs control), but not in non-ischaemic control hearts. With levosimendan, the Ca(2+) sensitising effects were less pronounced (7.6+/-3 mmHg, p=0.4 vs control). By counteracting the PDE inhibiting effects of ORG and Levo using parathyroid hormone and IGF-1, the negative lusotropic effects of Ca(2+) sensitisation were unmasked.

Conclusions: Calcium sensitisation improves systolic function and energetic efficiency. However, Ca(2+) sensitisers should be used with caution during post-ischaemic reperfusion, as they may exacerbate myocardial stunning and thus impair cardiac output.

Fig. 1

Schematic depiction of the experimental protocol. LVP, left ventricular pressure; PTH, parathormone; IGF-1, insulin-like growth factor-1.

Fig. 2

(A) Isovolumetric systolic pressure-volume relationship (developed pressure, devP) in non-ischaemic isolated rabbit hearts treated with the calcium de-sensitiser butanedione monoxime (BDM), dopamine, and the calcium sensitisers ORG 30029 (ORG) and levosimendan (Levo), n = 6 per group. The LV balloon was inflated in increments of 0.1 ml. (B) Diastolic pressure-volume relationship in the same non-ischaemic hearts. The p-values are based on the comparison of the slopes of the pressure-volume relationships by ANOVA with Dunnet’s post hoc test for multiple comparisons (unpaired), with ‘control’ as the reference group.

Fig. 3

(A) Isovolumetric systolic pressure-volume relationship (developed pressure, devP) in post-ischaemic isolated rabbit hearts treated with the calcium desensitiser butanedione monoxime (BDM), dopamine, and the calcium sensitisers ORG 30029 (ORG) and levosimendan (Levo), n = 6 per group. The LV balloon was inflated in increments of 0.1 ml. (B) Diastolic pressure-volume relationship in the same post-ischaemic hearts. The p-values are based on the comparison of the slopes of the pressure-volume relationships by ANOVA with Dunnet’s post hoc test for multiple comparisons (unpaired), with ‘control’ as the reference group.

Fig. 4

Minimum diastolic pressure at a filling volume of 0.8 ml in post-ischaemic hearts treated with the calcium desensitiser butanedione monoxime (BDM), dopamine, and the calcium sensitisers ORG 30029 (ORG) and levosi-mendan (Levo), n = 6 per group. The boxplots indicate minimum, maximum (whiskers), interquartile range (box) and median (bar). The p-values represent the results of ANOVA with Dunnet’s post hoc test for multiple comparisons (unpaired), with ‘control’ as the reference group.

Fig. 5

Mean cytosolic Ca²⁺ concentration in post-ischaemic rabbit hearts treated with butanedione monoxime (BDM), dopamine, and the calcium sensitisers ORG 30029 (ORG) and levosimendan (Levo), n = 6 per group. Control, untreated post-ischaemic hearts. The p-values represent the results of ANOVA with Dunnet’s post hoc test for multiple comparisons (unpaired), with ‘control’ as the reference group.

Fig. 6

(A) Calcium-force relationship in post-ischaemic isolated rabbit hearts treated with a standard catecholamine (dopamine) or the calcium sensitiser ORG 30029, n = 6 per group. Depicted is systolic developed pressure as a function of free cytosolic Ca²⁺ measured by rhod-2 spectrofluorometry. The Ca²⁺ sensitising effect of ORG is indicated by the rightward shift of the curve, i.e. more force is produced for a given amount of Ca²⁺. (B) Metabolic efficiency of post-ischaemic isolated rabbit hearts treated with a standard catecholamine (dopamine) or the calcium sensitiser ORG 30029. Depicted is myocardial work (rate-pressure product) as function of myocardial oxygen consumption (MvO₂). The rightward shift of the curve in response to ORG indicates that contractile efficiency is increased, whereas there is energy wasting under the influence of dopamine. The p-values are based on the comparison of the slopes of the pressure-volume relationships by ANOVA with Dunnet’s post hoc test for multiple comparisons (unpaired), with ‘dopamine’ as the reference group.

Fig. 7

Minimum diastolic pressure at a balloon filling volume of 0.8 ml in post-ischaemic hearts treated with the calcium sensitisers ORG 30029 (ORG, A) or levosimendan (Levo, B). n = 5 per group to counteract their PDE III-inhibiting effect, parathyroid hormone (PTH) or insulin-like growth factor-1 (IGF-1) were given at the end of the reperfusion period. Thus, the impact of Ca²⁺ sensitisation on diastolic relaxation was unmasked. The p-values represent the results of ANOVA with Dunnet’s post hoc test for multiple comparisons (unpaired), with ‘Levo’ or ‘ORG’ as the respective reference group.