Analyzing PKA-dependent synaptic plasticity in the hippocampal mossy fiber synapse using novel optogenetic tools

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Synaptic plasticity plays an important role in brain activity and is thought to be one of the main cellular processes underlying learning and memory. While in many synapses long-term potentiation is mediated by post-synaptic mechanisms, several synapses, most notably the hippocampal mossy fiber (MF), the cerebellar parallel fiber and Drosophila neuromuscular-junction synapses, exhibit presynaptic plasticity, which manifests as changes in the probability of synaptic vesicle release. Interestingly, this form of pre-synaptic plasticity depends on presynaptic activation of PKA in all the aforementioned synapses. The molecular mechanism underlying this PKA-dependent synaptic plasticity is still largely unknown. Currently, the control over PKA-activation levels is limited to adenylyl cyclase agonists, such as forskolin (FSK), or metabotropic glutamate receptor agonists. These treatments affect both pre- and post-synaptic neurons, as well as non-neuronal cell populations, their effect is not restricted to the synapse and is usually irreversible, facts that make these pharmacological agents less-than-ideal candidates to study synapse-specific effects of PKA activation. In light of this, the ability to manipulate PKA in the synapses of a genetically-identifiable subset of cells is highly instrumental in studying presynaptic PKA-dependent mechanisms. Accordingly, the current project aims are; a) develop, optimize and characterize novel optogenetic tools to control the level of PKA-activation in a synapse-specific manner b) investigate the effects of these novel optogenetic tools on synaptic transmission in the MF synapse, c) Examine if local pre- or post-synaptic PKA activation is sufficient to potentiate synaptic transmission, d) elucidate changes in the molecular organization of the AZ machinery following PKA activation, using super resolution microscopy and e) investigate the involvement of PKA downstream targets on synaptic potentiation. We will join forces, capitalizing on the expertise in the lab of Prof. Nagel in developing novel light activated PKA tools and combining this with the genetic and physiological approaches developed in Prof. Ashery’s lab to target and activate PKA only in MF and examine its effect on synaptic plasticity and the molecular mechanisms.