Modulation of GABAA receptor subtypes by neurosteroids and anesthetics: Identification of molecular targets for neuroprotection

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Several naturally occurring neurosteroids possess neuroprotective, sedative, anxiolytic and anaesthetic properties. It is a fascinating concept that these compounds, synthetized in and released by neurons and astrocytes, act as powerful allosteric modulators of GABAA receptors. Similar to neurosteroids, benzodiazepines (BZDs) and general anaesthetics enhance GABAA receptor function, thereby causing sedation, anxiolysis, amnesia, muscle relaxation and unconsciousness. GABAA receptors are pentameric ion channels that can assemble from nineteen protein subunits, giving rise to a large number of receptor subtypes with different physiological functions. Receptors containing alpha1-subunits contribute to the sedative, alpha2-receptors to the anxiolytic and alpha5-receptors to the amnestic effects of BZDs. Endogenous neurosteroid synthesis in the brain is regulated by ligands of the mitochondrial translocator protein (TSPO). We recently showed that the TSPO agonist XBD173 enhances neurosteroidogenesis, increases GABAA receptor-mediated inhibition and causes anxiolysis in rats and humans in the absence of sedation and tolerance development. In another study we identified synergistic interactions between the neurosteroid allopregnanolone and propofol on GABAA receptor-mediated synaptic transmission. There is evidence in the literature that dissimilar GABAA receptor subtypes are not equally sensitive to neurosteroids, with delta-subunit containing receptors being among the very responsive ones. In addition, our observation that XBD173 produces anxiolysis in the absence of sedation prompts the hypothesis that alpha1-GABAA receptors are not a major target for endogenously synthesized neurosteroids. The primary goal of this application is to investigate how enhanced neurosteroidogenesis modulates the function of the most important GABAA receptor subtypes on the synaptic, cellular and network level. Specifically, XBD173 will be tested for its potential to alter inhibitory synaptic transmission and signal processing in the hippocampus. Furthermore, we aim to identify GABAA receptor subtypes contributing to potentially synergistic interactions between neurosteroids and anaesthetic drugs. By investigating the molecular and cellular mechanisms how neurosteroids reverse detrimental hypoxic effects on long-term potentiation, a cellular correlate for learning and memory, and apoptosis we intend to identify synaptic and extrasynaptic GABAA receptors involved in mediating neurosteroid-induced neuroprotection. To address these issues, we will employ optogenetics, optical and electrophysiological monitoring of neuronal in wild type and delta-subunit knockout mice and point-mutated mice expressing only one BZD-sensitive GABAA receptor subtype. Altogether, the present proposal intends to elaborate the molecular actions of neurosteroids and their interactions with anaesthetic agents in order to evaluate future applications in clinical anaesthesia.