11C-(+3)N-Methyl-3-piperidylbenzilate PET Study of Muscarinic Acetylcholine Receptor Occupancy by Antimuscarinic Agents

Abstract

Muscarinic acetylcholine receptor (mAChR) is implicated in various central and peripheral nervous system disorders. The in vivo imaging of brain receptors by positron emission tomography (PET) allows the estimation of the precise localization of muscarinic receptors and the pharmacological characterization of the antimuscarinics. 11C-(+3)N-methyl-3-piperidylbenzilate (11C-(+)3-NMPB) is a recently developed PET radioligand, which has preferential binding affinity to the M1 and M2 subtypes of mAChR. Additionally, it has more favorable kinetic properties (transient equilibrium). In the present study, we suggested a new semi-quantification method for simplified preclinical 11C-(+)3-NMPB PET studies, which we compared to a previous quantification method by investigating changes in mAChR occupancy by antimuscarinic agents. Ninety min dynamic 11C-(+)3-NMPB PET studies were performed in ICR mice (n = 22). The transient equilibrium was assumed on the difference of the time-activity curves between the region of interest and the reference tissue at the vehicle condition (n = 4). The binding potential (BP) of 11C-(+)3-NMPB was estimated using Logan graphical analysis with cerebellar reference tissue input function. The equilibrium area under the curve ratio (AUC ratio) between the target and reference tissue at the equilibrium state after the injection of 11C-(+)3-NMPB. The percent difference in BP (or occupancy) and AUC ratio in various cortical and subcortical regions between dose conditions (solifenacin [1, 3 and 10 mg/kg, i.v.] or oxybutynin [0.1, 0.3 and 1 mg/kg, i.v.]) were calculated compared to the vehicle. The relationship between the BP and AUC ratio was estimated by correlational analysis. The 50% effective dose (ED50) of the antimuscarinics was calculated from the degree of percentage in vivo specific 11C-(+)3-NMPB binding in brain regions. Finally, a set of statistical tests was performed for comparisons between BP and AUC ratio data. At 30 - 60 min after the injection of 11C-(+)3-NMPB, radioactivity was shown to transient equilibrium (mean of t* = 17.5 min) at the vehicle condition. 11C-(+)3-NMPB distribution in the cortical and subcortical regions were well visualized consistently with the known distribution of mAChR. The striatum showed the greatest changes in BP (36.9 - 80.5% after 1, 3, and 10 mg/kg solifenacin and 9.4 – 75.9% after 0.1, 0.3, and 1 mg/kg oxybutynin, respectively). The striatum showed the greatest changes in AUC30-60min ratio (40.9 – 66.1% after 1, 3, and 10 mg/kg solifenacin and 12.4 – 61.3% after 0.1, 0.3, and 1 mg/kg oxybutynin, respectively). BP and AUC30-60min ratio values decreased after the administration of solifenacin (F (9,36) = 29.2, P < 0.01 and F (9,36) = 29.4, P < 0.01) or oxybutynin (F (9,36) = 35.36, P < 0.01 and F (9,36) = 39.16, P < 0.01) in a dose-dependent manner. Excellent correlations between varying levels of BP and AUC30-60min ratio per solifenacin and oxybutynin doses were found across the regions. ED50 was estimated in the striatum (ED50 = 2.512 mg/kg and 3.162 mg/kg of solifenacin, and ED50 = 0.309 mg/kg and 0.312 mg/kg of oxybutynin, respectively) and the cortex (ED50 = 1.408 mg/kg and 1.092 mg/kg of solifenacin, and ED50 = 0.486 mg/kg and 0.471 mg/kg of oxybutynin, respectively). Data demonstrated that 11C-(+)3-NMPB PET may be applied to in vivo mAChR quantification studies. In this study, the results suggested that semi-quantification of 11C-(+)3-NMPB binding with the equilibrium AUC ratio method may extend the application of 11C-(+)3-NMPB PET techniques to assess the efficacy of various mAChR-targeting drugs.