Biased agonism for dopamine at alpha1A-adrenoceptors towards receptor internalization

[Speaker] Andre S. Pupo:1
[Co-author] Rodrigo R. Wandekin:1, Vanessa Lima:1
1:Pharmacology, Instituto de Biociiencias, UNESP, Botucatu, SP, Brazil

Introduction: Dopamine activates alpha1-adrenoceptors (alpha1-ARs) and is used to rise blood pressure in shock. However, there is loss of vasoconstriction upon prolonged exposure to dopamine (tachyphylaxis), which has been attributed to alpha1-AR desensitization. The aim of this study was to investigate the desensitization/internalization of alpha1A-ARs induced by dopamine.
Methods: The effects of dopamine and norepinephrine were investigated in HEK293 cells stably expressing N-terminally FLAG tagged human alpha1A-ARs (intracellular calcium increases and cell Elisa-type assay using anti-FLAG antibodies) and in the isolated rat vas deferens, a tissue in which contractions to adrenoceptor agonists result from the activation of alpha1A-ARs (classic tissue bath experiments).
Results: In HEK293 cells expressing alpha1A-ARs, dopamine was approximately 300-fold less potent than norepinephrine in increasing intracellular calcium and both agonists produced similar Emax. However, in an Elisa-type assay in intact cells, dopamine was as potent and efficacious as norepinephrine in inducing alpha1A-AR internalization. In the rat vas deferens, dopamine behaved as a full agonist in relation to noradrenaline; however, the potency for dopamine was approximately 60-fold lower than that of noradrenaline. Up to seven consecutive concentration-response curves (CRCs) to noradrenaline with intervals between each CRC ranging from 5 to 30 min showed no tachyphylaxis and presented similar pEC50 and Emax values. However, there was tachylaphylaxis in the contractions induced by dopamine when the intervals between each CRC was 5 and 10 min, as indicated by a reduction in both the potency (3 to 10-fold loss of potency) and efficacy (up to 30% loss in Emax). In addition, the treatment of the tissues with dopamine 100 µM/5 min resulted in a 3-fold loss of potency for noradrenaline, whereas the treatment with noradrenaline 10 µM/5 min was unable to affect the potency or efficacy of noradrenaline on a subsequent exposure. Therefore, there is tachyphylaxis in the contractions of the rat vas deferens in response to dopamine, but not in response to noradrenaline, indicating that dopamine desensitizes the alpha1A-ARs.
Conclusions: The ability of dopamine to desensitize alpha1A-ARs may result from a biased agonism for dopamine favoring receptor internalization.
Financial support: CAPES and FAPESP (08/50423-7).

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