Mechano-regulation of β2AR signalling pathway of ventricular cardiomyocytes in healthy and failing hearts

Myocardial stretch occurs during haemodynamic overload and may be physiological (e.g., exercise) or pathological (e.g., heart failure). Understanding the effects of stretch on signalling pathways associated with mechanosensitive structures, such as caveolae, is therefore essential. Caveolar microdomains regulate the distribution of signalling proteins, including β-adrenoceptors (βARs), and may act as membrane reserves, protecting cells during the mechanical stretch. Phosphodiesterase 4 (PDE4), particularly subtypes PDE4B and PDE4D, have been shown to localise in caveolae-rich membranes, where they tightly regulate βAR-induced cAMP production. Regional differences in membrane structure and βAR signalling also exist across the myocardium. This study investigates the role of caveolae in βAR-signalling in response to membrane deformation, the specific contributions of PDE4B and PDE4D, and regional differences in this regulation. We further assess how this signalling is altered in the context of heart failure. To induce membrane stretch-like stimuli, we used osmotic swelling in left ventricular cardiomyocytes isolated from healthy and failing (16 weeks post-myocardial infarction) rat hearts, as well as from human donor and failing (dilated cardiomyopathy) hearts. The βAR response was measured using a Förster Resonance Energy Transfer (FRET) reporter for the second messenger cyclic adenosine monophosphate (cAMP) and using CytoCypher for the measurement of cell contractility. Our study reveals a stretch-regulation of the β2AR signalling pathway that is dependent on functional caveolae and PDE4 activity, particularly in the basal region of the myocardium; this regulation is lost in heart failure. These findings indicate that caveolae are mechanosensitive membrane domains that undergo structural and functional changes in response to membrane deformation, thereby mediating the mechanical regulation of caveolae-associated signalling pathways.