ABSTRACT
N-methyl-D-aspartate (NMDA) receptor hypofunctionality is a well-studied hypothesis for schizophrenia pathophysiology, and daily dosing of the NMDA receptor co-agonist, D-serine, in clinical trials has shown positive effects in patients. Therefore, inhibition of D-amino acid oxidase (DAAO) has the potential to be a new therapeutic approach for the treatment of schizophrenia. TAK-831 (luvadaxistat), a novel, highly potent inhibitor of DAAO, significantly increases D-serine levels in the rodent brain, plasma, and cerebrospinal fluid. This study shows luvadaxistat to be efficacious in animal tests of cognition and in a translational animal model for cognitive impairment in schizophrenia. This is demonstrated when luvadaxistat is dosed alone and in conjunction with a typical antipsychotic. When dosed chronically, there is a suggestion of change in synaptic plasticity as seen by a leftward shift in the maximum efficacious dose in several studies. This is suggestive of enhanced activation of NMDA receptors in the brain and confirmed by modulation of long-term potentiation after chronic dosing. DAAO is highly expressed in the cerebellum, an area of increasing interest for schizophrenia, and luvadaxistat was shown to be efficacious in a cerebellar-dependent associative learning task. While luvadaxistat ameliorated the deficit seen in sociability in two different negative symptom tests of social interaction, it failed to show an effect in endpoints of negative symptoms in clinical trials. These results suggest that luvadaxistat potentially could be used to improve cognitive impairment in patients with schizophrenia, which is not well addressed with current antipsychotic medications.
Subject(s)
Antipsychotic Agents , Schizophrenia , Animals , Oxidoreductases , Rodentia , Schizophrenia/drug therapy , Antipsychotic Agents/pharmacology , Antipsychotic Agents/therapeutic use , Enzyme Inhibitors/pharmacology , Cognition , Serine/pharmacology , Amino Acids , Receptors, N-Methyl-D-AspartateABSTRACT
We report a significant decrease in transcription of the G protein-coupled receptor GPR39 in striatal neurons of Parkinson's disease patients compared to healthy controls, suggesting that a positive modulator of GPR39 may beneficially impact neuroprotection. To test this notion, we developed various structurally diverse tool molecules. While we elaborated on previously reported starting points, we also performed an in silico screen which led to completely novel pharmacophores. In vitro studies indicated that GPR39 agonism does not have a profound effect on neuroprotection.
Subject(s)
Pyrimidines/pharmacology , Receptors, G-Protein-Coupled/agonists , Allosteric Regulation/drug effects , Dose-Response Relationship, Drug , Humans , Molecular Structure , Pyrimidines/chemical synthesis , Pyrimidines/chemistry , Receptors, G-Protein-Coupled/metabolism , Structure-Activity RelationshipABSTRACT
Neuroinflammation, specifically the NLRP3 inflammasome cascade, is a common underlying pathological feature of many neurodegenerative diseases. Evidence suggests that NLRP3 activation involves changes in intracellular K+. Nuclear Enriched Transcript Sort Sequencing (NETSseq), which allows for deep sequencing of purified cell types from human post-mortem brain tissue, demonstrated a highly specific expression of the tandem pore domain halothane-inhibited K+ channel 1 (THIK-1) in microglia compared to other glial and neuronal cell types in the human brain. NETSseq also showed a significant increase of THIK-1 in microglia isolated from cortical regions of brains with Alzheimer's disease (AD) relative to control donors. Herein, we report the discovery and pharmacological characterisation of C101248, the first selective small-molecule inhibitor of THIK-1. C101248 showed a concentration-dependent inhibition of both mouse and human THIK-1 (IC50: â¼50 nM) and was inactive against K2P family members TREK-1 and TWIK-2, and Kv2.1. Whole-cell patch-clamp recordings of microglia from mouse hippocampal slices showed that C101248 potently blocked both tonic and ATP-evoked THIK-1 K+ currents. Notably, C101248 had no effect on other constitutively active resting conductance in slices from THIK-1-depleted mice. In isolated microglia, C101248 prevented NLRP3-dependent release of IL-1ß, an effect not seen in THIK-1-depleted microglia. In conclusion, we demonstrated that inhibiting THIK-1 (a microglia specific gene that is upregulated in brains from donors with AD) using a novel selective modulator attenuates the NLRP3-dependent release of IL-1ß from microglia, which suggests that this channel may be a potential therapeutic target for the modulation of neuroinflammation in AD.
Subject(s)
Alzheimer Disease , Inflammasomes , Potassium Channels, Tandem Pore Domain , Animals , Humans , Mice , Alzheimer Disease/metabolism , Brain/metabolism , Inflammasomes/metabolism , Microglia , Neuroinflammatory Diseases , NLR Family, Pyrin Domain-Containing 3 Protein/metabolism , Potassium Channels, Tandem Pore Domain/antagonists & inhibitorsABSTRACT
Multielectrode arrays (MEAs) are grids of substrate-integrated microelectrodes that allow for electrophysiological interrogation of dissociated cell cultures or tissue slices. Here we discuss the use of nonimplantable electrodes for studies. The methods described attempt to provide a starting point for researchers new to the field who wish to begin to utilize this powerful, but daunting technology and quickly apply the basic principles to their own research interests.
Subject(s)
Action Potentials , Nerve Net/physiology , Neurons/physiology , Tissue Array Analysis/instrumentation , Animals , Cell Culture Techniques/instrumentation , Cell Culture Techniques/methods , Electrophysiology/instrumentation , Electrophysiology/methods , Equipment Design , Hippocampus/cytology , Hippocampus/physiology , Mice , Microelectrodes , Nerve Net/cytology , Neurons/cytology , Organ Culture Techniques/instrumentation , Organ Culture Techniques/methods , Rats , Tissue Array Analysis/methodsABSTRACT
Motor neurons (MNs) and astrocytes (ACs) are implicated in the pathogenesis of amyotrophic lateral sclerosis (ALS), but their interaction and the sequence of molecular events leading to MN death remain unresolved. Here, we optimized directed differentiation of induced pluripotent stem cells (iPSCs) into highly enriched (> 85%) functional populations of spinal cord MNs and ACs. We identify significantly increased cytoplasmic TDP-43 and ER stress as primary pathogenic events in patient-specific valosin-containing protein (VCP)-mutant MNs, with secondary mitochondrial dysfunction and oxidative stress. Cumulatively, these cellular stresses result in synaptic pathology and cell death in VCP-mutant MNs. We additionally identify a cell-autonomous VCP-mutant AC survival phenotype, which is not attributable to the same molecular pathology occurring in VCP-mutant MNs. Finally, through iterative co-culture experiments, we uncover non-cell-autonomous effects of VCP-mutant ACs on both control and mutant MNs. This work elucidates molecular events and cellular interplay that could guide future therapeutic strategies in ALS.