Impaired β-adrenergic inotropic reserve in organotypic cardiac slices of a murine cardiometabolic HFpEF model

Background/Introduction/Purpose:
Heart failure with preserved ejection fraction (HFpEF) is closely associated with inadequate adaptation of cardiac function to increased workload. The role of intrinsic myocardial contractile function in HFpEF is not well understood, and its investigation requires controlled in vitro conditions.  Isolated cardiomyocytes lack  active and passive cell-to-cell-communication and mechanical preload. Engineered heart tissues exhibit immature properties and were not exposed to the in vivo conditions of HFpEF-related systemic triggers. We therefore investigated the β-adrenergic and frequency-dependent functional reserve in adult organotypic mouse heart slices of a cardiometabolic HFpEF model.

Methods:
12 week old male C57Bl/6N mice were fed standard chow (control) or high fat diet and L-NAME (HFpEF) for 13 weeks. To confirm HFpEF phenotype animals were subjected to baseline and isoproterenol-induced (iso) stress echocardiography. Hearts were isolated, ventricles were embedded in 4% agarose and 300 µm short axis slices were prepared. Three slices of each heart were mounted on force transducers and subsequently used for contraction measurements. During an acclimatization phase, preload was gradually applied, calcium was reintroduced (1.8 mM final) and slices were gently rewarmed to 37° C at a stimulation of 3 Hz. Subsequently, incremental frequency pacing (1, 3, 5, 7 Hz) was performed followed by two isoprotenerol cycles at 3 Hz. Slices that showed no response to isoprotenerol stimulation were excluded.

Results:
In vivo echo revealed significantly impaired diastolic function in HFpEF animals (E/é: control: 20.6 ± 4.2; HFpEF: 35.9 ± 7.6; p <0.0001) with preserved ejection fraction (control 59.2 ± 7.8; HFpEF: 60.0 ± 5.8, p=0.0696). An increase in developed force following iso administration was observed in slices of all animals (Control: 11/11 mice, 29/33 slices; HFpEF: 11/11 mice, 28/33 slices, Fig.). While groups showed  comparable force development under baseline conditions  (3 Hz, same preload: control: 0.1585 ± 0.02352; HFpEF: 0.1520 ± 0.02079), developed force augmentation following β-adrenergic stimulation was significantly higher in control (Fig. ; Iso (1) p=0.0063; Iso (2) p=0.0003; Fig.). Moreover, relaxation was significantly impaired in HFpEF (time constant tau relaxation: Tau (1) control: 0.01692 ± 0.002; HFpEF: 0.01993 ± 0.001; p= 0.0033; Tau (2) control: 0.01548 ± 0.002; HFpEF: 0.01828 ± 0.003; p=0.0063).

   
Figure: force development after first and second administration of isoprotenerol in control and HFpEF mice (as percentage of force amplitude before iso)

Conclusions:
We show that functional reserve is limited and relaxation is slowed under β-adrenergic stress in organotypic slices in this mouse model of metabolic HFpEF. The methodology of functional cardiac slices provides a promising basis for the translation of previously obtained research results to a more complex system mimicking the physiological and pathophysiological environment of cardiomyocytes.