A Limited Vs a Transient AT1R Internalization Indicates that Distinct Mechanisms of AT1R Activation are Possible

Abstract

We have shown that a 15,000 kDa polymer of Ang II (Ang II‐POL) at maximal concentration upon a sustained action on the AT1R of rat coronary endothelial luminal membrane (CELM) produced sustained vasoconstriction and inotropic effects, and a lack of CELM AT1R internalization. In contrast, an equivalent concentration of the monomer Ang II (1 kDa) caused transient effects: desensitization, associated with the expected progressive internalization of CELM AT1R. It is accepted that AT1R Internalization into the cell, results from the CELM agonist‐receptor complex sequestration into vesicles which depends on the internalization proteins: β‐arrestin, clathrin and dynamin. Objective The purpose of this study was to study the effect of angII and AngII‐POL on the recruitment of internalizing proteins: β arrestin and clathrin, in rat CELM. Hypothesis The different rates of AT1R internalization induced by the monomer Ang II and the polymer Ang II‐POL indicates that each agonist recruits in a different way the internalization proteins. Hypothesis test In 2 groups hearts, time course inotropic Ang II or Ang II‐POL effects were measured. In another groups, CELM proteins were isolated by the silica pellicle technique at various times from; control hearts, during stimulation with either Ang II or Ang II‐POL, AT1R; β arrestina1/2 and clathrin quantified by Western blot. Results Ang II inotropic effects show desensitization, paralleled by CELM AT1R internalization, β‐arrestin1/2 and Clathrin recruitment into CELM. In contrast Ang II‐POL did not induce CELM AT1R internalization, it recruits considerable β‐arrestin1/2 into CELM, without clathrin recruitment. Conclusions The activation mechanisms of AT1R by the monomer Ang II and the polymer Ang II‐POL are distinct, because, in contrast to Ang II, the Ang II‐POL effect, lacks of AT1R internalization, triggers an enhanced CELM β‐arrestin1/2 recruitment with a much‐reduced Clathrin recruitment. The likely resultant poor formation of clathrin‐vesicles could explain the lack of AT1R internalization and suggest that clathrin density controls and limits the amount of β‐arrestin1/2 incorporated. Clearly, Ang II‐POL constitutes a powerful tool to understanding the mechanism of AT1R activation and internalization.