NANOCORDIS

Chronic activation of ß-adrenergic receptor (ß-AR)/cAMP cascade is centrally involved in heart failure (HF) and sudden death, as demonstrated by the beneficial effect of ß-blocker therapy. ß-AR stimulation acts via activation of the cAMP-dependent protein kinase (PKA) which phosphorylates a number of key proteins, and most importantly the L-type Ca2+ channel (LTCC). LTTCs are responsible for Ca2+ entry into the myocardial cell, which triggers intracellular Ca2+ release from the sarcoplasmic reticulum (SR) via ryanodine receptor. Under acute condition, ß-AR activation of LTCCs leads to an increase in the force of contraction. However, chronic ß-AR activation as occurs in HF also leads to excessive Ca2+ entry and release: this causes intracellular Ca2+ overload, cell depolarisation, spontaneous action potentials, which eventually lead to ventricular arrhythmias and sudden death. Therefore, it is imperative to better understand how ß-ARs regulate LTCCs in ventricular cells, and in particular how they are compartmentalized within the cardiac cell membrane. One potential source of compartmentation in the highly structured ventricular cardiomyocyte is the physical arrangement of the sarcolemma. The most evident features are the transverse tubules (T-tubules), a network of deep invaginations of the surface membrane which form an interconnected network of tubes penetrating deep into the interior of the cardiomyocyte. T-tubules mainly run along the z-disks of sarcomeres, allowing the development of dyadic junctions with the SR that are crucial in Ca2+ transients regulating cardiomyocyte contraction. LTCCs and ß-ARs, as well as other membranes proteins, are located in both compartments but at different densities. The general goal of this project is to explore how location of LTCCs and ß-ARs in either surface or T-tubule membrane impacts on their function. We will use an innovative nanotechnology approach by combining the expertise of an INSERM laboratory specialized in cardiac cellular signaling (Partner 1) and a CNRS laboratory specialized in pharmaceutical sciences and drug delivery (Partner 2). Both belong to the University Paris-Sud and are located on the campus of the Faculty of Pharmacy in Châtenay-Malabry. The strategy used is based on the preparation of functionalized nanoparticles decorated with LTCC or ß-AR agonists or antagonists covalently bound to their surface. The nanoparticles will have a diameter of 100-150 nm to prevent them accessing T-tubule membranes due to steric hindrance. While free agonists will be used to activate both external and T-tubule LTCCs and ß-ARs, respectively, agonists immobilized on the surface of nanoparticles will be used to only activate those located in the external membrane. A free agonist applied in the presence of a saturating concentration of nanoparticle-immobilized antagonist will be used to only activate LTCCs or ß-ARs located in the T-tubule membrane. This strategy will be applied in adult rat ventricular myocytes isolated from either normal or HF animals to explore the specific alterations of the two subpopulations of LTCCs and ß-ARs during pathological remodeling. Since LTCCs and ß-ARs are relevant targets for therapeutic drugs, a better understanding of the role of the respective sub-populations of these major membrane proteins in physiological and pathological settings may lead to new drug delivery systems that would allow a specific targeting of the relevant sub-population.