Inverse Agonism Detected at the Level of the Receptor

FRET-based detection of receptor conformational changes which are directly related to activation proved to be a useful tool to determine the action of inverse agonists on GPCRs. Both inverse agonists for a2A- adrenergic receptors, as well as those acting on P1-adrenergic receptors have been studied using this approach [32, 78]. All inverse agonists tested induced a FRET change in the opposite direction compared to agonists. In the case of a2A- adrenergic receptor, the introduction of a point mutation which substantially increases the constitutive activity of the receptor allowed a detailed analysis of receptor activation [ 78]. As expected, inverse agonists induce FRET changes in the opposite direction compared to agonists. However, two features were found when using FRET that were not expected. The first surprise was the extent of inverse agonist-mduced changes were much larger than expected, based on the degree of inhibition of basal receptor activity [78]. This finding suggests that inverse agonists not only change the equilibrium toward inactive receptor conformations, but also induce larger conformational changes of the receptor. Unlike partial agonists, for which the degree of partial agonism correlated roughly with the detected FRET amplitude, the amplitude of FRET changes induced by inverse agonists did not match differences in receptor activity. For example, with P1- adrenergic receptor, the clinically relevant inverse agonist carvedilol exerted a much more dramatic increase in FRET compared to the equally potent inverse agonist bisoprolol [32] . Notably, a frequent occurring polymorphism of P1- adrenergic receptor (Arg389) exhibited a substantially larger FRET increase specifically for carvedilol but not bisoprolol [32]. Some indications exist that the population which carries this polymorphism also respond particularly well therapeutically to carvedilol. The FRET study may be a hint at how even clinical pharmacology can benefit from FRET-based studies of receptor activation. The second surprise was that the kinetics of inverse agonist action on receptors were much slower than the kinetics of agonist-induced receptor activation [73, 78]. So far, we do not understand the underlying molecular basis for this observation. This might change in the future, as more structural information on the receptors is becoming available, and computational approaches to model dynamics of conformational movements are forthcoming.

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