Structural plasticity at mossy fiber-CA3 synapses

Neurons are connected via synapses, where the axon of one neuron forms a terminal contact with the dendrite of a second neuron. Information is transferred from the presynaptic neuron, through a synapse, to the postsynaptic neuron. The strength of a synapse can change over time, which we refer to as plasticity. The hippocampus is a brain region critical to the formation of new memories. Here mossy fibers from presynaptic granule cells contact postsynaptic CA3 neurons. This synapse can be transiently strengthened by a form of plasticity called post-tetanic potentiation (PTP), resulting from elevated activity in the presynaptic neuron. Once potentiated, CA3 neurons become full detonators, responding strongly to inputs from even a single presynaptic neuron. Such strong connections are a rare occurrence in the brain and are the focus of our study. During synaptic transmission, presynaptic electrical signals cause vesicles containing neurotransmitters to be released, carrying information across the synapse in chemical form. Using electrophysiological methods, we obtained preliminary data suggesting that the number of vesicles that are ready to be released (readily releasable pool, RRP) increases during PTP. To identify the mechanism of PTP, we aim to directly relate structure and function of the synapse at the level of synaptic vesicles. Electron microscopy (EM) is the technique of choice to visualize synaptic structure at high-resolution, but on its own provides a static picture. To capture the dynamic process of synaptic transmission, I thus combine EM with high-pressure freezing (HPF) and optogenetics, in a cutting-edge technique called Flash and Freeze: a pulse of light activates genetically modified granule cells, stimulating the presynaptic terminal and triggering synaptic transmission. After light stimulation the neurons are frozen within milliseconds, capturing specific time points of the synaptic transmission process. PTP at the MF-CA3 synapse was discovered decades ago, but its mechanism remains unclear. The DAG molecule binds to Munc13, a protein critical for vesicular synaptic transmission, and there is prior evidence that DAG interferes with refilling of the RRP after strong presynaptic activity. I will combine Flash and Freeze with traditional electrophysiology to test the hypothesis that PTP results from an increase in the RRP and depends on the DAG signaling pathway. The innovative approach of this proposal will elucidate presynaptic mechanisms of PTP that lead to plasticity-dependent detonation. This key synaptic function may well be essential for the storage or retrieval of memories.