Biased M1-muscarinic-receptor-mutant mice inform the design of next-generation drugs.

Cholinesterase inhibitors, the current frontline symptomatic treatment for Alzheimer's disease (AD), are associated with low efficacy and adverse effects. M1 muscarinic acetylcholine receptors (M1 mAChRs) represent a potential alternate therapeutic target; however, drug discovery programs focused on this G protein-coupled receptor (GPCR) have failed, largely due to cholinergic adverse responses. Employing novel chemogenetic and phosphorylation-deficient, G protein-biased, mouse models, paired with a toolbox of probe molecules, we establish previously unappreciated pharmacologically targetable M1 mAChR neurological processes, including anxiety-like behaviors and hyper-locomotion.

Preclinical profile of a dopamine D1 potentiator suggests therapeutic utility in neurological and psychiatric disorders.

DETQ, an allosteric potentiator of the dopamine D1 receptor, was tested in therapeutic models that were known to respond to D1 agonists. Because of a species difference in affinity for DETQ, all rodent experiments used transgenic mice expressing the human D1 receptor (hD1 mice). When given alone, DETQ reversed the locomotor depression caused by a low dose of reserpine.

A validated LC-MS/MS method for neurotransmitter metabolite analysis in human cerebrospinal fluid using benzoyl chloride derivatization.

Background: Human cerebrospinal fluid (CSF) is often acquired in Phase I clinical trials to assess the CNS penetration of new pharmacological agents and to search for biomarkers associated with PD effects. Robust methods for neurotransmitter metabolites in CSF have proven elusive, in part due to inadequate reversed phase LC retention.

Results: Benzoyl chloride derivatization was used to promote retention for LC-MS/MS for a panel of neurotransmitter metabolites while delivering a concise method for sample preparation.

Glucose metabolic trapping in mouse arteries: nonradioactive assay of atherosclerotic plaque inflammation applicable to drug discovery.

Background: (18)F-Fluorodeoxyglucose (FDG)-positron emission tomography (PET) imaging of atherosclerosis in the clinic is based on preferential accumulation of radioactive glucose analog in atherosclerotic plaques. FDG-PET is challenging in mouse models due to limited resolution and high cost. We aimed to quantify accumulation of nonradioactive glucose metabolite, FDG-6-phosphate, in the mouse atherosclerotic plaques as a simple alternative to PET imaging.

Synthesis, ex vivo evaluation, and radiolabeling of potent 1,5-diphenylpyrrolidin-2-one cannabinoid subtype-1 receptor ligands as candidates for in vivo imaging.

We have reported that [methyl- (11)C] (3 R,5 R)-5-(3-methoxyphenyl)-3-[(R)-1-phenylethylamino]-1-(4-trifluoromethylphenyl)pyrrolidin-2-one ([(11)C] 8, [(11)C]MePPEP) binds with high selectivity to cannabinoid type-1 (CB 1) receptors in monkey brain in vivo. We now describe the synthesis of 8 and four analogues, namely, the 4-fluorophenyl (16, FMePPEP), 3-fluoromethoxy (20, FMPEP), 3-fluoromethoxy- d 2 (21, FMPEP- d 2), and 3-fluoroethoxy analogues (22, FEPEP), and report their activity in an ex vivo model designed to identify compounds suitable for use as positron emission tomography (PET) ligands. These ligands exhibited high, selective potency at CB 1 receptors in vitro (K b < 1 nM).

Positron emission tomography imaging using an inverse agonist radioligand to assess cannabinoid CB1 receptors in rodents.

[11C]MePPEP is an inverse agonist and a radioligand developed to image cannabinoid CB1 receptors with positron emission tomography (PET). It provides reversible, high specific signal in monkey brain. We assessed [11C]MePPEP in rodent brain with regard to receptor selectivity, susceptibility to transport by P-glycoprotein (P-gp), sensitivity to displacement by agonists, and accumulation of radiometabolites.

The PET radioligand [11C]MePPEP binds reversibly and with high specific signal to cannabinoid CB1 receptors in nonhuman primate brain.

The cannabinoid CB(1) receptor is one of the most abundant G protein-coupled receptors in the brain and is a promising target of therapeutic drug development. Success of drug development for neuropsychiatric indications is significantly enhanced with the ability to directly measure spatial and temporal binding of compounds to receptors in central compartments. We assessed the utility of a new positron emission tomography (PET) radioligand to image CB(1) receptors in monkey brain.

Comparison of rat dopamine D2 receptor occupancy for a series of antipsychotic drugs measured using radiolabeled or nonlabeled raclopride tracer.

Preclinical brain receptor occupancy measures have heretofore been conducted by quantifying the brain distribution of a radiolabeled tracer ligand using either scintillation spectroscopy or tomographic imaging. For smaller animals like rodents, the majority of studies employ tissue dissection and scintillation spectroscopy. These measurements can also be accomplished using liquid chromatography coupled to mass spectral detection to measure the brain distribution of tracer molecules, obviating the need for radioligands.

The relationship of in vivo central CB1 receptor occupancy to changes in cortical monoamine release and feeding elicited by CB1 receptor antagonists in rats.

Rationale: Cannabinoid type 1 (CB(1)) receptor antagonists are reportedly effective in reducing food intake both preclinically and clinically. This may be due in part to their effects on monoamine release in the brain. The level of central CB(1) receptor occupancy underlying these neurobiological effects is unclear.

Use of LC/MS to assess brain tracer distribution in preclinical, in vivo receptor occupancy studies: dopamine D2, serotonin 2A and NK-1 receptors as examples.

High performance liquid chromatography combined with either single quad or triple quad mass spectral detectors (LC/MS) was used to measure the brain distribution of receptor occupancy tracers targeting dopamine D2, serotonin 5-HT2A and neurokinin NK-1 receptors using the ligands raclopride, MDL-100907 and GR205171, respectively. All three non-radiolabeled tracer molecules were easily detectable in discrete rat brain areas after intravenous doses of 3, 3 and 30 microg/kg, respectively. These levels showed a differential brain distribution caused by differences in receptor density, as demonstrated by the observation that pretreatment with compounds that occupy these receptors reduced this differential distribution in a dose-dependent manner.

Fluoxetine, but not other selective serotonin uptake inhibitors, increases norepinephrine and dopamine extracellular levels in prefrontal cortex.

Rationale: The selective serotonin uptake inhibitor (SSRI) fluoxetine has been shown to not only increase the extracellular concentrations of serotonin, but also dopamine and norepinephrine extracellular concentrations in rat prefrontal cortex. The effect of other SSRIs on monoamine concentrations in prefrontal cortex has not been thoroughly studied.

Objective: The aim of this study was to compare the ability of five systemically administered selective serotonin uptake inhibitors to increase acutely the extracellular concentrations of serotonin, norepinephrine and dopamine in rat prefrontal cortex.