Background:
With ageing populations heart failure (HF) remains a major burden for global health. Due to structural and electrophysiological remodelling patients are vulnerable to develop lethal arrhythmias. Here, the chronically overstimulated β-adrenergic pathway plays a key role by producing elevated levels of cAMP which lead to dysregulated Ca2+-homeostasis and can result in strong arrhythmogenic triggers. The Phosphodiesterase 2 (PDE2) has the unique property to be enhanced in its hydrolytic activity towards cAMP by cGMP and can therefore lower cAMP levels. Interestingly PDE2 is upregulated in HF indicating at a potential protective mechanism. 5,6-DM-cBIMP (cBIMP) is a cell permeant cGMP analogue capable of activating PDE2. This study aims to investigate whether the direct activation of PDE2 with cBIMP reduces arrhythmogenic triggers in isolated cardiomyocytes (CM) from mice with HF.
Methods:
With a high fat diet and the NO-synthase inhibitor L-NAME in the drinking water (0.5 g/L) over 5 weeks the HF was induced and verified by echocardiography. Experiments were performed in isolated CM from PDE2 wildtype (WT) mice as well as from cardio-specific PDE2 knockout (KO) mice. Protein expression was quantified with Western blots. cAMP levels were analysed using a competitive ELISA. Inwards L-type Calcium current (ICaL) was measured using the patch-clamp technique. Using calcium imaging, spontaneous calcium waves (SCW) out of the sarcoplasmic reticulum (SR) were quantified with Fura-2-AM and calcium sparks with Fluo-4. Sarcomere contraction properties were analysed with video-based contraction measurements.
Results:
Compered to healthy mice, there was a significant decrease in the expression of the β1-adrenoreceptor in mice with HF. Indicating at a desensitized signalling cascade we also found higher doses of isoprenaline (ISO) were needed to provoke calcium sparks in HF compared to healthy mice. Interestingly, we also found an increase in the expression of PDE2 in mice with HF compared to non-failing controls. The β-adrenergic stimulation with (ISO) significantly increased intracellular cAMP levels in isolated CM, which could significantly be lowered by simultaneous cBIMP incubation. The ICaL amplitude was clearly enhanced upon ISO stimulation, which was again prevented by cBIMP. This effect was reversed after specific PDE2 inhibition with BAY 60-7550 and moreover was not observed in CM from PDE2 KO mice. Importantly, cBIMP markedly reduced the number of ISO-induced arrhythmogenic SCW of the SR. However, cBIMP did not decrease the cellular basal and ISO stimulated contraction as characterised by contraction/relaxation velocity and contraction amplitude in WT or KO mice.
Conclusion:
By lowering elevated cAMP level via direct activation of PDE2, 5,6-DM-cBIMP significantly reduced arrhythmogenic calcium triggers without negatively affecting single cell contractility in isolated CM of mice with HF offering a promising strategy for antiarrhythmic therapy in HF.
