Comparison of automated video tracking systems in the open field test : ANY-Maze versus EthoVision XT

This project included funding from the Innovative Medicines Initiative 2/EFPIA, European Quality in Preclinical Data (EQIPD) consortium under grant agreement number 777364. We would also like to acknowledge the staff of the Medical Research Facility for their support with animal care, handling and behavioural experiments.Peer reviewedPostprin

GABA(B) receptor positive allosteric modulators with different efficacies affect neuroadaptation to and self-administration of alcohol and cocaine

Drugs of abuse induce widespread synaptic adaptations in the mesolimbic dopamine (DA) neurons. Such drug-induced neuroadaptations may constitute an initial cellular mechanism eventually leading to compulsive drug-seeking behavior. To evaluate the impact of GABA(B) receptors on addiction-related persistent neuroplasticity, we tested the ability of orthosteric agonist baclofen and two positive allosteric modulators (PAMs) of GABA(B) receptors to suppress neuroadaptations in the ventral tegmental area (VTA) and reward-related behaviors induced by ethanol and cocaine. A novel compound (S)-1-(5-fluoro-2,3-dihydro-1H-inden-2-yl)-4-methyl-6,7,8,9-tetrahydro-[1,2,4]triazolo[4,3-a]quinazolin-5(4H)-one (ORM-27669) was found to be a GABA(B) PAM of low efficacy as agonist, whereas the reference compound (R,S)-5,7-di-tert-butyl-3-hydroxy-3-trifluoromethyl-3H-benzofuran-2-one (rac-BHFF) had a different allosteric profile being a more potent PAM in the calcium-based assay and an agonist, coupled with potent PAM activity, in the [S-35] GTP gamma S binding assay in rat and human recombinant receptors. Using autoradiography, the high-efficacy rac-BHFF and the low-efficacy ORM-27669 potentiated the effects of baclofen on [S-35] GTP gamma S binding with identical brain regional distribution. Treatment of mice with baclofen, rac-BHFF, or ORM-27669 failed to induce glutamate receptor neuroplasticity in the VTA DA neurons. Pretreatment with rac-BHFF at non-sedative doses effectively reversed both ethanol- and cocaine-induced plasticity and attenuated cocaine i.v. self-administration and ethanol drinking. Pretreatment with ORM-27669 only reversed ethanol-induced neuroplasticity and attenuated ethanol drinking but had no effects on cocaine-induced neuroplasticity or self-administration. These findings encourage further investigation of GABA(B) receptor PAMs with different efficacies in addiction models to develop novel treatment strategies for drug addiction.Peer reviewe

Systematic assessment of the replicability and generalizability of preclinical findings: Impact of protocol harmonization across laboratory sites.

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

Inverse agonism at α2A adrenoceptors augments the hypophagic effect of sibutramine in rats

Because the use of monoamine reuptake inhibitors as weight-reducing agents is limited by adverse effects, novel antiobesity drugs are needed. We studied acute effects of the noradrenaline (NA) and serotonin (5-HT) reuptake inhibitor sibutramine (SIB), alone and after pretreatment with α1- and α2-adrenoceptor (AR), and 5-HT1/2/7, 5-HT1B and 5-HT2C receptor antagonists in order to determine which ARs and 5-HT receptors act downstream of SIB on feeding and locomotion. Acute effects on caloric and water intake, meal microstructure and locomotion were assessed, using an automated weighing system and telemetry in male rats with restricted 18-h access to Western style diet. SIB 3 mg/kg reduced meal size and frequency, which suggests enhanced within- and postmeal satiety. Imiloxan (α2B-AR), WB4101 (α1-AR), SB-224289 (5-HT1B), and modestly BRL 44408 (α2A/D-AR) attenuated SIB's effect on meal size, suggesting that α2B- and α1-ARs and 5-HT1B receptors mediate within-meal satiety, with a modest role for α2A/D-ARs. Only prazosin (α1/2B/2C-AR) counteracted SIB's effect on meal frequency. At 3 mg/kg, SIB modestly increased locomotion. This effect was blocked by metergoline (5-HT1/2/7), WB4101 (α1-AR), and RX821002 (α2-AR). Interestingly, the α2-AR antagonists atipamezole and RX821002 enhanced SIB's effect on caloric intake, probably due to inverse agonistic actions at α2A-autoreceptors that further enhanced release of NA that regulates caloric intake. Thus, an inverse agonist of presynaptic α2A-ARs might beneficially enhance SIB's weight-reducing effect and offer novel treatment for obesity. All in all, the present data supports the ARs and 5-HT receptors involved in the effects of SIB on different aspects of caloric intake and locomotio