Platform for in vivo analysis of early GABA synapse function in neural circuitry formation

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In our funded NSERC Discovery Grant titled, "Crosstalk between inhibitory and excitatory neurotransmitter systems during brain development" we are investigating how interactions between different types of synaptic connections may help control the development and organization of neural circuitry. During development, neurons establish an intricate balance of excitatory and inhibitory synaptic connections and this balance is essential for properly encoding information in the brain. Too much inhibition is detrimental for brain plasticity, whereas over-excitation is toxic for neurons. Exactly how this equilibrium is achieved remains poorly understood. We have discovered that early signaling through the neurotransmitter GABA controls the number of excitatory synapses on developing hippocampal neurons. Using a combination of in vitro methods and high-resolution time-lapse confocal microscopy, we have made significant progress in determining how GABA transmission helps set the number of excitatory synapses during early development of neurons. We are now at a critical stage of the project where we need to understand how these mechanisms work in the living brain. In this grant, we propose to establish an advanced platform for in vivo experimental analysis in mice through combined in utero electroporation and patch clamp electrophysiology. In utero electroporation is needed to deliver novel molecular biology technologies in vivo and to precisely control the timing of gene expression of mutant GABA receptors, while patch clamp electrophysiology is needed to understand the functional impact of modifying early GABA transmission on synaptic organization. We have already secured in utero electroporation equipment, a patch-clamp amplifier, and manipulators, in part, through our Discovery Grant funds. However, to complete the platform, we would like to purchase a microscope that is compatible with electrophysiology. This microscope, when combined with the other components, will provide an unprecedented level of analysis for our experiments while, at the same time, offer a unique opportunity for students to use a cutting-edge approach to answer an important question in brain research.