Neurons dynamically regulate neurotransmitter release through many processes known collectively as synaptic plasticity. Post-tetanic potentiation (PTP) is a widespread form of synaptic plasticity that lasts for tens of seconds, and which may have important computational roles and contribute to short-term memory. According to a leading mechanism, the high-frequency stimulation that is necessary to induce PTP increases presynaptic calcium, which activates calcium sensors to regulate the amplitude and time-course of increased release. At the “Synaptic Vesicle Fusion and Retrieval” symposium, I will discuss our investigations into the role of the calcium-sensitive protein kinase C (PKC) as an essential regulator of PTP at 3 mammalian excitatory synapses: the parallel fiber - Purkinje cell synapse in the cerebellar cortex, the calyx of Held in the auditory brainstem, and the CA3 - CA1 synapse in the hippocampus. I will introduce a conditional knock-in approach to selectively mutate all PKC phosphorylation sites in the prominent PKC phosphorylation substrate Munc18-1, an SM family protein that is essential for release. Finally, I will describe initial efforts using this new mouse model to delineate the molecular events underlying enhanced neurotransmission during PTP.