Model Of Agonism Research Articles

Effect of disease states on α1-adrenoceptor binding and signal transduction parameters in isolated perfused heart: quantification by pharmacokinetic-pharmacodynamic modelling

To employ a pharmacokinetic-pharmacodynamic modelling approach for analysing the effect of experimental endotoxemia and mild hypoxia on α1 -adrenoceptor (α1 AR) binding and signal transduction. In Langendorff-perfused rat hearts, phenylephrine was continuously infused, and [(3) H]-prazosin was injected as single dose (infused over 1 min). Simultaneous analysis of the time courses of prazosin outflow concentration and inotropic response (left ventricular developed pressure) using an agonist-antagonist interaction model and nonlinear regression allowed to estimate receptor affinity, as well as the parameters of the operational model of agonism. Both endotoxemia and hypoxia, significantly reduced the maximum response achievable in the system to 67% and 49% of the control group mean, respectively. In addition, endotoxemia decreased the efficiency of stimulus-response coupling and increased the steepness of the stimulus-response curve. In both disease models, no change in receptor affinity and density were found. The results revealed the causes of reduced α1 AR-mediated inotropic responsiveness in endotoxemia and hypoxia. In contrast with traditional dose-response studies, it was possible to quantify separately the underlying changes in α1 AR binding and signal transduction.

Cyclic AMP- and (Rp)-cAMPS-induced Conformational Changes in a Complex of the Catalytic and Regulatory (RIα) Subunits of Cyclic AMP-dependent Protein Kinase

We took a discovery approach to explore the actions of cAMP and two of its analogs, one a cAMP mimic ((S(p))-adenosine cyclic 3':5'-monophosphorothioate ((S(p))-cAMPS)) and the other a diastereoisomeric antagonist ((R(p))-cAMPS), on a model system of the type Iα cyclic AMP-dependent protein kinase holoenzyme, RIα(91-244)·C-subunit, by using fluorescence spectroscopy and amide H/(2)H exchange mass spectrometry. Specifically, for the fluorescence experiments, fluorescein maleimide was conjugated to three cysteine single residue substitution mutants, R92C, T104C, and R239C, of RIα(91-244), and the effects of cAMP, (S(p))-cAMPS, and (R(p))-cAMPS on the kinetics of R-C binding and the time-resolved anisotropy of the reporter group at each conjugation site were measured. For the amide exchange experiments, ESI-TOF mass spectrometry with pepsin proteolytic fragmentation was used to assess the effects of (R(p))-cAMPS on amide exchange of the RIα(91-244)·C-subunit complex. We found that cAMP and its mimic perturbed at least parts of the C-subunit interaction Sites 2 and 3 but probably not Site 1 via reduced interactions of the linker region and αC of RIα(91-244). Surprisingly, (R(p))-cAMPS not only increased the affinity of RIα(91-244) toward the C-subunit by 5-fold but also produced long range effects that propagated through both the C- and R-subunits to produce limited unfolding and/or enhanced conformational flexibility. This combination of effects is consistent with (R(p))-cAMPS acting by enhancing the internal entropy of the R·C complex. Finally, the (R(p))-cAMPS-induced increase in affinity of RIα(91-244) toward the C-subunit indicates that (R(p))-cAMPS is better described as an inverse agonist because it decreases the fractional dissociation of the cyclic AMP-dependent protein kinase holoenzyme and in turn its basal activity.

An operational model of pharmacological agonism: the effect of E/[A] curve shape on agonist dissociation constant estimation

An operational model of pharmacological agonism: the effect of E/[A] curve shape on agonist dissociation constant estimation

Towards a Thermodynamic Definition of Efficacy in Partial Agonism: I Concentration-Response Theory for Explicit Agonist and Antagonist Complexes

The pharmacological characteristics of a partial agonist, namely its binding and efficacy, are transposed, here, to give more insight into the mode of binding of the requisite agonist and antagonist bound complexes. Simple proportionality relationships of the efficacy to the overall binding define the agonist and antagonist binding constants. The advantage of such description lies in determining the thermodynamic profiles of the respective complexes by simple temperature variation. The enthalpic difference between agonist and antagonist complexes becomes a function of the efficacy, e alone, the thermodynamic scale ratio, e / (1-e), yielding this value directly. The concentration-response theory for two bound ligand conformer complexes, one of which yields an agonist response is examined using a simple adaptation of the Black and Leff model of agonism. A selected tissue response, the chronotropic in guinea-pig right atria, dominated by a single receptor, the β1 -adrenergic under normal signal amplification conditions, was selected. The chronotropic response accurately obeys a hyperbolic dose-response form, a condition for the model’s applicability and component profiles are exemplified at 30.0°C. The precision was found sufficient to evaluate the thermodynamics of the respective complexes over a 15°C range of temperature. in a second paper.

A Model for Agonism and Antagonism in an Ancient and Ubiquitous cAMP-binding Domain

The cAMP-binding domain (CBD) is an ancient and conserved regulatory motif that allosterically modulates the function of a group of diverse proteins, thereby translating the cAMP signal into a controlled biological response. The main receptor for cAMP in mammals is the ubiquitous regulatory (R) subunit of protein kinase A. Despite the recognized significant potential for pharmacological applications of CBDs, currently only one group of competitive inhibitor antagonists is known: the (R(p))-cAMPS family of phosphorothioate cAMP analogs, in which the equatorial exocyclic oxygen of cAMP is replaced by sulfur. It is also known that the diastereoisomer (S(p))-cAMPS with opposite phosphorous chirality is a cAMP agonist, but the molecular mechanism of action of these analogs is currently not fully understood. Previous crystallographic and unfolding investigations point to the enhanced CBD dynamics as a key determinant of antagonism. Here, we investigate the (R(p))- and (S(p))-cAMPS-bound states of R(CBD-A) using a comparative NMR approach that reveals a clear chemical shift and dynamic NMR signature, differentiating the (S(p))-cAMPS agonist from the (R(p))-cAMPS antagonist. Based on these data, we have proposed a model for the (R(p)/S(p))-cAMPS antagonism and agonism in terms of steric and electronic effects on two main allosteric relay sites, Ile(163) and Asp(170), respectively, affecting the stability of a ternary inhibitory complex formed by the effector ligand, the regulatory and the catalytic subunits of protein kinase A. The proposed model not only rationalizes the existing data on the phosphorothioate analogs, but it will also facilitate the design of novel cAMP antagonists and agonists.

Mechanism-based modeling of functional adaptation upon chronic treatment with midazolam.

A mechanism-based model is applied to analyse adaptive changes in the pharmacodynamics of benzodiazepines upon chronic treatment in rats. The pharmacodynamics of midazolam was studied in rats which received a constant rate infusion of the drug for 14 days, resulting in a steady-state concentration of 102 +/- 8 ng x ml(-1). Vehicle treated rats were used as controls. Concentration-EEG effect data were analysed on basis of the operational model of agonism. The results were compared to data obtained in vitro in a brain synaptoneurosomal preparation. The relationship between midazolam concentration and EEG effect was non-linear. In midazolam pre-treated rats the maximum EEG effect was reduced by 51 +/- 23 microV from the original value of 109 +/-15 microV in vehicle treated group. Analysis of this change on basis of the operational model of agonism showed that it can be explained by a change in the parameter tissue maximum (Em) rather than efficacy (tau). In the in vitro studies no changes in density, affinity or functionality of the benzodiazepine receptor were observed. It is concluded that the observed changes in the concentration-EEG effect relationship of midazolam upon chronic treatment are unrelated to changes in benzodiazepine receptor function.

Mechanism-based modeling of adaptive changes in the pharmacodynamics of midazolam in the kindling model of epilepsy.

A mechanism-based model is proposed for the analysis of adaptive changes in the pharmacodynamics of benzodiazepines in vivo. The pharmacodynamics of midazolam was studied in the kindling model of experimental epilepsy. Concentration-EEG effect data from kindled rats and their controls were fitted to the operational model of agonism. A stepwise procedure was used, allowing changes in the parameters efficacy (tau) and tissue maximum (Em) either separately or in combination. The results were compared to data obtained in vitro in a brain synaptoneurosomal preparation. The relationship between midazolam concentration and EEG effect was non-linear. In kindled rats the maximum EEG effect was reduced by 27+/-8.3 microV from the original value of 94+/-4.4 microV. Analysis on the basis of the operational model of agonism showed that this decrease could be explained by a difference in the parameter system maximum (Em) rather than efficacy (tau). In the in vitro receptor binding assay no changes in density, affinity or functionality of the benzodiazepine receptor were observed, consistent with the lack of a change in efficacy (tau). The operational model of agonism provides a mechanistic basis to characterise adaptive changes in the pharmacodynamics of midazolam.