Unexpected decrease in in vivo binding of [3H]QNB in the mouse cerebral cortex in the developing brain - A comparison with [11C]NMPB

Introduction: Significant discrepancies between in vitro and in vivo binding of themuscarinic receptor ligand – 3HlabeledQuinuclidinyl Benzilate (QNB) – have been well documented. Discernable in vivo cerebellar [3H]QNBbinding has been observed in mouse brain, despite the maximum number of binding sites (Bmax) being low. Inorder to understand this unique in vivo binding phenomenon, the binding of two muscarinic receptor ligands– [3H]QNB and N-[11C]methylpiperidyl Benzilate ([11C]NMPB) – were compared in vivo and in vitro in 3- and8-week-old mice.Method: In vitro binding parameters of [3H]QNB were determined using brain homogenates. The time course ofradioactivity concentration (TACs) in the cerebral cortex and cerebellum was measured following injection of[3H]QNB and [11C]NMPB with or without 3 mg/kg of carrier QNB in 3- and 8 week old mice using a dual traceradministration technique. A graphical method was employed for the quantitative analysis of in vivo binding ofthese radioligands.Results: In vitro, the available number of binding sites for cerebral cortical muscarinic receptors increased by 17%during the developmental period studied. Paradoxically, in vivo, we observed a decrease of [3H]QNB binding inthe cerebral cortex, while [11C]NMPB binding was markedly increased. In vivo saturation analysis of [3H]QNBin 3-week-old mice revealed an apparent positive cooperativity of binding in the cerebral cortex.Conclusions: Our results support the hypothesis that microenvironmental factors proximal to muscarinic receptorscause a local decrease in the cortical free-ligand concentration of [3H]QNB and that this ‘ligand barrier’ ismodulated during brain development.Advances in knowledge: The present study demonstrates that the combined use of radiolabeled QNB and NMPBhas the potential to reveal the important effects of receptor microenvironmental factors on receptor functionin the living brain