Performance evaluation of a MIP for the MISPE-LC determination of p-[18F]MPPF and a potential metabolite in human plasma.

Within the family of serotonin (5-HT) receptors, the 5-HT1A subtype is particularly interesting as it may be involved in various physiological processes or psychological disorders. The p-[18F]MPPF, a highly selective 5-HT1A antagonist, is used for in vivo studies in human or animal by means of positron emission tomography (PET) [1]. In order to selectively extract p-[18F]MPPF and its main metabolites from plasma, molecularly imprinted polymer (MIP) was prepared against these compounds by using the p-MPPF as template. For the control of the selectivity, non-imprinted polymer (NIP) was also synthesized without template. The MIP sorbent, packed in disposable extraction cartridges (DECs), was then evaluated as molecularly imprinted solid-phase extraction (MISPE) prior to the LC determination. The conditions of extraction were evaluated in order to obtain the highest selective retention of the p-[18F]MPPF and its metabolites on this MIP. The MIP selectivity was exploited in the loading and washing steps by adjusting the pH of plasma samples at a suitable value and by selecting mixtures for the washing step to limit the contribution of non-specific interactions. Other important parameters involved in the conditioning and elution steps were also studied. Finally, a pre-validation was carried out with optimal extraction conditions to demonstrate the performance of this MISPE-LC method as a generic method in the context of evaluation of new MISPE for p-[18F]MPPF and its potential for metabolites extraction from human plasma.

Functional binding assays for estimation of the intrinsic efficacy of ligands at the 5-HT1A receptor: application for screening drug candidates.

INTRODUCTION: Determination of the intrinsic efficacy of ligands at the 5-HT1A receptor is important for selecting drug candidates, e.g. in the case of schizophrenia where partial agonism is a favorable property shared by different atypical antipsychotics. METHODS: Using seven ligands with different intrinsic efficacies and rat hippocampus synaptosomes, we compared critically three "functional" binding assays based on the ternary complex model that considers that the activated conformation of the receptor is the one coupled to G-protein. RESULTS: The Ki ratio method, based on the difference of affinity of the competing drug when using an antagonist vs. an agonist as radioligand, discriminated the ligands according to their intrinsic efficacies, with values from 77 for the full agonist 5-hydroxytryptamine to 0.09 for the inverse agonist WAY 100,635. The GTP-shift method, based on the decrease of affinity observed with agonists when GTP is added to the competition binding assay, was equally effective in classifying the drugs according to their intrinsic efficacy. The lower sensibility of the GTP-shift assay was investigated and explained by the different ionic conditions used in the two assays and the way competition curves were analyzed. Albeit more direct, the assay based on agonist-stimulated [(35)S]-GTPγS binding to G proteins was more expensive and of greater variability in our hands. DISCUSSION: We conclude that the GTP-shift procedure described herein for 5-HT1A receptors may expedite drug discovery efforts by predicting at the same time the affinity and intrinsic efficacy of ligands through a simple, rapid and economic ligand binding assay.

Influence of intrathecal injection of p-MPPF on the analgesic effects of isoflurane / 中国药理学通报

0. 05) . Compared with Iso analgesic group ( Iso group) ,the TFL or HPPT of co-administration groups ( Iso + M6 group,Iso + M3 group) shortened ( P 0. 05) . Conclusion These findings suggest that the surface analgesic effects of Iso are closely related to the excited 5-HT1A receptor in the spinal cord of mice.