The opioid receptor pharmacology of GSK1521498 compared to other ligands with differential effects on compulsive reward-related behaviours

Abstract

Rationale: The novel opioid receptor antagonist, GSK1421498, has been shown to attenuate reward-driven compulsive behaviours, such as stimulant drug seeking or binge eating, in animals and humans. Here, we report new data on the receptor pharmacology of GSK121498, in comparison to naltrexone, naloxone, 6-β-naltrexol and nalmefene.

Objectives: To determine whether the novel opioid antagonist, GSK1521498, is an orthosteric or allosteric antagonist at the μ opioid receptor (MOPr) and whether it has neutral antagonist or inverse agonist properties.

Methods: A combination of radioligand binding assays and [(35)S]GTPγS binding assays was employed.

Results: GSK1521498 completely displaced [(3)H]naloxone binding to MOPr and did not alter the rate of [(3)H]naloxone dissociation from MOPr observations compatible with it binding to the orthosteric site on MOPr. GSK1521498 exhibited inverse agonism when MOPr was overexpressed but not when the level of MOPr expression was low. In parallel studies under conditions of high receptor expression density, naloxone, naltrexone, 6-β-naltrexol and nalmefene exhibited partial agonism, not inverse agonism as has been reported previously for naloxone and naltrexone. In brain tissue from mice receiving a prolonged morphine pre-treatment, GSK1521498 exhibited slight inverse agonism.

Conclusions: Differences between GSK1521498 and naltrexone in their effects on compulsive reward seeking are arguably linked to the more selective and complete MOPr antagonism of GSK1521498 versus the partial MOPr agonism of naltrexone. GSK1521498 is also pharmacologically differentiated by its inverse agonist efficacy at high levels of MOPr expression, but this may be less likely to contribute to behavioural differentiation at patho-physiological levels of expression.

Fig. 1

Inhibition of Met-Enk-stimulated [³⁵S]GTPγS binding to membranes prepared from CHO cells overexpressing MOPr. Concentration-response curve for GSK1521498 (black square), naloxone (black circle), naltrexone (white circle), 6β-naltrexol (black up-pointing triangle) and nalmefene (white square) for inhibiting 10-nM Met-Enk-stimulated [³⁵S]GTPγS binding in CHO cells expressing MOPr. Mean data are expressed as percentage of the stimulation produced by Met-Enk and are from three experiments each performed in duplicate. For each curve, the maximum was constrained to 100 % and the slope to unity

Fig. 2

Displacement of [³H]naloxone binding to MOPr. a–c Competition displacement binding curves for the displacement of ³H-naloxone binding by GSK1521498, naltrexone, and 6-β-naltrexol to membranes prepared from HEK293 cells expressing the human MOPr. d Kinetics of [³H]naloxone dissociation from MOPr in the absence and presence of GSK1521498, naltrexone and 6-β-naltrexol (all at 1 μM). Mean t_1/2 values for [³H]naloxone dissociation in the presence of each drug are given with 95 % confidence limits

Fig. 3

Effect of antagonists on basal [³⁵S]GTPγS binding to membranes from CHO cells expressing MOPr. a Concentration-response curve for GSK1521498 (black square), naloxone (black circle), naltrexone (white circle), 6-β-naltrexol (black up-pointing triangle) and nalmefene (white square) in CHO cell membranes overexpressing MOPr. Mean data are expressed as percentage of the basal binding, determined in the absence of ligands. At concentrations greater than 0.4 nM, GSK1521498 produced a significant inhibition of basal [³⁵S]GTPγS binding (p < 0.001, ANOVA on log-transformed raw data, with treatment and plates as independent factors). b Drugs were tested at a final concentration of 1 μM on membranes prepared from CHO cells expressing a low level of MOPr. Values are expressed as a percentage of the basal value in the absence of drug, which was taken as 100 % in each experiment. Values shown as means ± SEM from five separate experiments in each case. None of the drugs had a significant effect on basal [³⁵S]GTPγS binding (one sample t test)

Fig. 4

Effect of antagonists on basal [³⁵S]GTPγS binding to membranes from whole mouse brain, following different morphine pre-treatments. Drugs were tested at a final concentration of 1 μM. Values are expressed as a percentage of the basal value in the absence of antagonist drug, which was taken as 100 % in each experiment. Values shown as means ± SEM from five to six separate experiments in each case, each performed in triplicate or quadruplicate.*p < 0.05 compared to 100 %, one-sample t test