In Alzheimer’s disease (AD) and other tauopathies, the cytosolic protein Tau misfolds and forms intracellular aggregates which accumulate within the brain leading to neurodegeneration. Clinical progression is tightly linked to the progressive spread of Tau pathology throughout the brain, and several lines of evidence suggest that Tau aggregates or “seeds” may propagate pathology by spreading from cell to cell in a “prion like” manner. Accordingly, blocking the spread of extracellular seeds with an antibody could be a viable therapeutic approach. However, as the structure of Tau seeds is unknown, it is only possible to rationally design therapeutic Tau antibodies by making a priori assumptions. To avoid this, we developed a robust and quantitative cell based assay and employed an unbiased screening approach to identify the antibody with the highest activity against human Tau seeds. The selected antibody (D), directed to the mid-region of Tau (amino acids 235–250), potently blocked the seeding of human AD Tau and was also fully efficacious against seeds from progressive supranuclear palsy. When we compared this antibody with previously described reference antibodies, we were surprised to find that none of these antibodies showed comparable efficacy against human pathological seeds. Our data highlight the difficulty of predicting antibody accessible epitopes on pathological Tau seeds and question the potential efficacy of some of the Tau antibodies that are currently in clinical development.Electronic supplementary materialThe online version of this article (10.1007/s00401-018-1911-2) contains supplementary material, which is available to authorized users.

Homeodomain containing transcription factors of the Hox family play critical roles in patterning the anteroposterior embryonic body axis, as well as in controlling several steps of organogenesis. Several Hox proteins have been shown to cooperate with members of the Pbx family for the recognition and activation of identified target enhancers. Hox proteins contact Pbx via a conserved hexapeptide motif. Previous biochemical studies provided evidence that critical amino acid substitutions in the hexapeptide sequence of Hoxa1 abolish its interaction with Pbx. As a result, these substitutions also abolish Hoxa1 activity on known target enhancers in cellular models, suggesting that Hoxa1 activity relies on its capacity to interact with Pbx. Here, we show that mice with mutations in the Hoxa1 hexapeptide display hindbrain, cranial nerve, and skeletal defects highly reminiscent of those reported for the Hoxa1 loss of function. Since similar hexapeptide mutations in the mouse Hoxb8 and the Drosophila AbdA proteins result in activity modulation and gain of function, our data demonstrate that the functional importance of the hexapeptide in vivo differs according to the Hox proteins.

Research in the epilepsy field is moving from a primary focus on controlling seizures to addressing disease pathophysiology. This requires the adoption of resource- and time-consuming animal models of chronic epilepsy which are no longer able to sustain the testing of even moderate numbers of compounds. Therefore, new in vitro functional assays of epilepsy are needed that are able to provide a medium throughput while still preserving sufficient biological context to allow for the identification of compounds with new modes of action. Here we describe a robust and simple fluorescence-based calcium assay to measure epileptiform network activity using rat primary cortical cultures in a 96-well format. The assay measures synchronized intracellular calcium oscillations occurring in the population of primary neurons and is amenable to medium throughput screening. We have adapted this assay format to the low magnesium and the 4-aminopyridine epilepsy models and confirmed the contribution of voltage-gated ion channels and AMPA, NMDA and GABA receptors to epileptiform activity in both models. We have also evaluated its translatability using a panel of antiepileptic drugs with a variety of modes of action. Given its throughput and translatability, the calcium oscillations assay bridges the gap between simplified target-based screenings and compound testing in animal models of epilepsy. This phenotypic assay also has the potential to be used directly as a functional screen to help identify novel antiepileptic compounds with new modes of action, as well as pathways with previously unknown contribution to disease pathophysiology.

Our results demonstrate that following in utero exposure to BA on GD 9, a disturbance of the expression of hox genes involved inthe specification of most anterior vertebrae is observed at GD 13.5. Based on their expression domain and on their implication in the definition of the cervicothoracic vertebral boundary, it is likely that the anteriorization of hoxc6 and hoxa6 reported here is correlated to the morphological phenotype observed in BA-exposed fetuses at GD 21.

Gamma amino butyric acid receptors (GABA) are major therapeutic targets for the development of drugs in neurological and psychiatric disorders. The new generation of GABA A modulators is targeting subtype selectivity and low/partial efficacy on the receptor to potentially overcome the adverse effects described for drugs with full agonist profile. We evaluated a screening approach to measure the relative efficacy of GABA A positive allosteric modulators (PAM) using automated patch clamp and fluorescence membrane potential assays. We determined that the use of an internal comparator (zolpidem), tested on each cell in parallel to the test compound, provides a reliable approach to measure and compare the relative efficacy of PAM ligands. Patch clamp recordings on recombinant GABA A receptors, using a multiple drug addition protocol, allows us to rank PAM ligands with different levels of efficacies. We observed that fluorescence membrane potential assays are not predictive of the relative efficacies of GABA A PAM ligands.