Abstract:The a 2 -adrenoceptors (ARs) are important modulators of a wide array of physiological responses. As only a few selective compounds for the three a 2 -AR subtypes (a 2A , a 2B and a 2C ) have been available, the pharmacological profile of a new a 2C -selective AR antagonist ORM-10921 is reported. Standard in vitro receptor assays and antagonism of a 2 , and a 1 -AR agonist -evoked responses in vivo were used to demonstrate the a 2C -AR selectivity for ORM-10921 which was tested in established behavioural models related to schizophrenia and cognitive dysfunction with an emphasis on pharmacologically induced hypoglutamatergic state by phencyclidine or MK-801. The Kb values of in vitro a 2C -AR antagonism for ORM-10921 varied between 0.078-1.2 nM depending on the applied method. The selectivity ratios compared to a 2A -AR subtype and other relevant receptors were 10-100 times in vitro. The in vivo experiments supported its potent a 2C -antagonism combined with only a weak a 2A -antagonism. In the pharmacodynamic microdialysis study, ORM-10921 was found to increase extracellular dopamine levels in prefrontal cortex in the baseline conditions. In the behavioural tests, ORM-10921 displayed potent antidepressant and antipsychotic-like effects in the forced swimming test and prepulse-inhibition models analogously with the previously reported results with structurally different a 2C -selective AR antagonist JP-1302. Our new results also indicate that ORM-10921 alleviated the NMDA-antagonist-induced impairments in social behaviour and watermaze navigation. This study extends and further validates the concept that a 2C -AR is a potential therapeutic target in CNS disorders such as schizophrenia or Alzheimer's disease and suggests the potential of a 2C -antagonism to treat such disorders. a 2 -Adrenoceptors (a 2 -ARs) consist of three distinct a 2 -adrenergic receptor subtypes in mammals: a 2A , a 2B and a 2C . They are distributed widely, yet not homogenously, regulating a diverse range of physiological processes including sedation/ vigilance, anxiety, pain/analgesia and cardiovascular function [1][2][3]. In the CNS, the a 2C -ARs are expressed most extensively in the striatum and hippocampus [4][5][6][7]. Valuable information on the discrete functions of each a 2 -AR subtype has been gained by using gene-targeted mice [8,9], but the limited availability of suitable subtype selective ligands has hindered their pharmacological exploration [10].The discovery and pre-clinical profile of a chemically novel and exceptionally selective antagonist at the a 2C -AR, JP-1302, has been described [11]. Compared to other available a 2 -ARs, this agent displays striking (>100 times) selectivity. In the behavioural profiling, JP-1302 showed potent antidepressantand antipsychotic-like properties in the forced swimming test and prepulse inhibition (PPI) paradigms [9,12]. Although part of these results are consistent with those obtained with a 2 -AR knockout mice [9], the antipsychotic-like effect, that is, the reversal of impairme...

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The influence of protocol standardization between laboratories on their replicability of preclinical results has not been addressed in a systematic way. While standardization is considered good research practice as a means to control for undesired external noise (i.e., highly variable results), some reports suggest that standardized protocols may lead to idiosyncratic results, thus undermining replicability. Through the EQIPD consortium, a multi-lab collaboration between academic and industry partners, we aimed to elucidate parameters that impact the replicability of preclinical animal studies. To this end, 3 experimental protocols were implemented across 7 laboratories. The replicability of results was determined using the distance travelled in an open field after administration of pharmacological compounds known to modulate locomotor activity (MK-801, diazepam, and clozapine) in C57BL/6 mice as a worked example. The goal was to determine whether harmonization of study protocols across laboratories improves the replicability of the results and whether replicability can be further improved by systematic variation (heterogenization) of 2 environmental factors (time of testing and light intensity during testing) within laboratories. Protocols were tested in 3 consecutive stages and differed in the extent of harmonization across laboratories and standardization within laboratories: stage 1, minimally aligned across sites (local protocol); stage 2, fully aligned across sites (harmonized protocol) with and without systematic variation (standardized and heterogenized cohort); and stage 3, fully aligned across sites (standardized protocol) with a different compound. All protocols resulted in consistent treatment effects across laboratories, which were also replicated within laboratories across the different stages. Harmonization of protocols across laboratories reduced between-lab variability substantially compared to each lab using their local protocol. In contrast, the environmental factors chosen to introduce systematic variation within laboratories did not affect the behavioral outcome. Therefore, heterogenization did not reduce between-lab variability further compared to the harmonization of the standardized protocol. Altogether, these findings demonstrate that subtle variations between lab-specific study protocols may introduce variation across independent replicate studies even after protocol harmonization and that systematic heterogenization of environmental factors may not be sufficient to account for such between-lab variation. Differences in replicability of results within and between laboratories highlight the ubiquity of study-specific variation due to between-lab variability, the importance of transparent and fine-grained reporting of methodologies and research protocols, and the importance of independent study replication.