State

Studies with a2adrenergic, and many other GPCRs, indicate that they are capable of activating different types of G proteins. According to a two-state receptor model (i.e. where the receptor can only reside in an inactive or an active conformation), the role of agonists is to increase the number of activated receptors. Each agonist will have its own intrinsic efficacy (e) for a given type of receptor (Figure 181). Hence, the amount of activated receptors (which may be regarded to represent the 'stimulus, S') is a unique property of the agonist-receptor combination and this also implies that a given agonist increases the likelihood of the receptor encountering and activating distinct types of G proteins in the same way. Hence, according to a strict two-state model for receptor activation, and in the absence of a 'receptor reserve', the same pharmacological profile (i.e. ligand potency and intrinsic efficacy orders) will be obtained irrespective of the G protein-mediated response pathway.

Figure 181 One receptor-two G proteins interaction model (Kenakin, 1995a). It must be emphasized that in a strict two-state model, R* and AR* will bind with the same affinities to a given G protein (i.e. R and AR*/R* represent uniquely defined conformational states). Reprinted from Trends in Pharmacological Science, 16, Kenakin, T., Agonist-receptor efficacy. I: Mechanisms of efficacy and receptor promiscuity, 188-192. Copyright (1995), with permission from Elsevier.

Figure 181 One receptor-two G proteins interaction model (Kenakin, 1995a). It must be emphasized that in a strict two-state model, R* and AR* will bind with the same affinities to a given G protein (i.e. R and AR*/R* represent uniquely defined conformational states). Reprinted from Trends in Pharmacological Science, 16, Kenakin, T., Agonist-receptor efficacy. I: Mechanisms of efficacy and receptor promiscuity, 188-192. Copyright (1995), with permission from Elsevier.

In models dealing with the interaction between an activated receptor and distinct G proteins (i.e. G: and G2) (Figure 181), the former is often regarded as acting as a 'ligand' and the latter as distinct 'receptors' (Kenakin, 1995; Kukkonen et al., 2001). According to such models, the amount of R*G: and R*G2 complexes will depend on:

• The ratio between the concentrations of the receptor and each type of G protein (and, hence, also on the ratio between the concentrations of each type of G protein).

• The affinity of the activated receptor for each type of G protein.

Simulations according to such models suggest that increased receptor expression may lead to receptor-G protein promiscuity (Figure 182). When two G proteins have different affinities for the activated receptor, the receptor-G protein selectivity is strictly preserved when levels of receptor are low. However, when the receptor expression level exceeds that of the G protein with the highest affinity, all of them may be solicited to form AR*G complexes. In the same vein, provided that the receptor concentration is sufficiently elevated, high efficacy agonists will produce sufficiently activated receptors to couple to both G proteins while partial agonists (i.e. with low efficacy) will just produce enough activated receptors to couple to the G protein with the highest affinity.

Although the accumulation of AR*G complexes provides a fair view of what is going on between receptors and distinct G proteins, such a process is only allowed to take place in the absence of GTP, such as in membrane preparations. In intact cells, these complexes will almost immediately fall apart because of the ongoing GDP/GTP exchange. Moreover, when more distant effects/responses are measured, agonist dose-response curves will also depend on how the different G proteins interact with their effector, as well as on the solicited response pathways. In this respect, two situations can be distinguished, i.e.:

Figure 182 Computer-simulated AR*G1 and AR*G2 concentrations for increasing agonist concentrations. The receptor expression level is 60% (A) or 200% (B) of the G protein expression level. The activated receptor has higher affinity for G1. Reprinted from Trends in Pharmacological Science, 16, Kenakin, T., Agonist-receptor efficacy. I: Mechanisms of efficacy and receptor promiscuity, 188-192. Copyright (1995), with permission from Elsevier.

Figure 182 Computer-simulated AR*G1 and AR*G2 concentrations for increasing agonist concentrations. The receptor expression level is 60% (A) or 200% (B) of the G protein expression level. The activated receptor has higher affinity for G1. Reprinted from Trends in Pharmacological Science, 16, Kenakin, T., Agonist-receptor efficacy. I: Mechanisms of efficacy and receptor promiscuity, 188-192. Copyright (1995), with permission from Elsevier.

EC50-Gs>EC50_Gi o v

EC50-Gs>EC50_Gi v

Ratio

Gs/Gi

Agonist concentration In (Log(M)

Figure 183 Computer-simulated adenyl cyclase activities for increasing agonist concentrations. Left panel, the stimulated receptor displays the same affinities for Gj and Gs. Right panel, the stimulated receptor displays 10-fold higher affinity for Gi. Reprinted from Biochemistry and Pharmacology, 62, Nasman, J., Kukkonen, J. P., Ammoun, S. and Akerman, K. E., Role of G protein availability in differential signalling by alpha 2-adrenoceptors, 913-922. Copyright (2001), with permission from Elsevier.

Ratio

Gs/Gi

Agonist concentration In (Log(M)

Figure 183 Computer-simulated adenyl cyclase activities for increasing agonist concentrations. Left panel, the stimulated receptor displays the same affinities for Gj and Gs. Right panel, the stimulated receptor displays 10-fold higher affinity for Gi. Reprinted from Biochemistry and Pharmacology, 62, Nasman, J., Kukkonen, J. P., Ammoun, S. and Akerman, K. E., Role of G protein availability in differential signalling by alpha 2-adrenoceptors, 913-922. Copyright (2001), with permission from Elsevier.

• The pathways produce two independently measurable responses.

• The pathways can recombine to modulate one measurable response (Figure 183). An obvious example is the opposite regulation of the adenyl cyclase activity by both Gas and Ga^ In practice, this appears to happen for a2B receptors: they produce pertussis toxin-sensitive adenyl cyclase inhibition (a Gj-mediated effect) as well as pertussis toxin-insensitive adenyl cyclase stimulation (a Gas-mediated effect). Biphasic/bell-shaped agonist dose-response curves can even be seen in certain a2B receptor-containing cell types. Computer-assisted simulations reveal that such a situation may take place when oppositely acting G proteins are involved. In general, the shape of such curves will depend on the G protein concentration ratio and on their relative affinities for the activated receptor.

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