Long-term potentiation (LTP) is a well-established experimental model used to investigate the synaptic basis of learning and memory. The present study concerns the induction of mossy fiber (MF) LTP in hippocampal brain slices. This form of synaptic plasticity is particularly interesting because it is NMDA receptor-independent and requires activation of pre-synaptic kainate receptors. Previous electrophysiological data ^[1] from our lab has demonstrated a role for intracellular Ca²⁺-induced Ca²⁺ release (CICR) in both the induction of MF LTP and the pronounced synaptic facilitation that occurs during high-frequency stimulation. These experiments also provided evidence that, under physiological conditions, Ca²⁺ permeation through kainate receptors acts as the trigger for intracellular CICR. Our present study extends this research by using a technique ^[2] that combines 2-photon microscopy, patch-clamp electrophysiology and calcium imaging to directly investigate the role of pre-synaptic calcium in MF LTP. Granule cells of the dentate gyrus are held in the whole cell patch-clamp configuration and experiments are conducted in current-clamp mode. The patch pipette contains a red morphological marker (to identify pre-synaptic giant MF boutons) and a calcium indicator dye. Cells are held for >2hrs prior to calcium imaging to allow for dye diffusion and equilibration. Pre-synaptic calcium signals are generated by evoking orthodromic action potentials (via current injection into the patch pipette: train of 5 pulses; 2ms; 25Hz) and are recorded by line-scanning individual MF boutons in the presence and absence of kainate receptor agonists and antagonists.