Cholecystokinin (CCK) is one of the most abundant neuropeptides in brain (Rehfeld JF., 1978). The sulfated CCK-8 (CCK-8s) is the bioactive form of CCK that modulates various neuronal process, such as anxiety (Daugé and Léna., 1998), neuronal excitability (Wang et al., 2011) and inhibitory (Ma et al., 2006). If mice lost the CCK gene expression, it exhibits poor performance in a passive avoidance task and displays impaired spatial memory (Lo CM et al., 2008). CCK binds to CCK receptors to produce the subsequent neural plasticity. CCK receptors are a group of G-protein coupled receptors (Noble Florence et al., 1999) within two different subtypes named CCKA receptors and CCKB receptors that distribute in various regions of brain (M Dufresne et al., 2006). Previous study demonstrated that blocking CCK receptors suppresses conditioned fear (Tsutsumi T et al., 1999) and knocking out CCK-B receptor gene reduces anxiety-like behavior in rodents (Horinouchi Y et al., 2004). The publications in my laboratory have proved that CCK released from entorhinal cortex enables neural plasticity in the auditory cortex (Li X et al., 2014) which release triggered by N-methyl-D-aspartate (NMDA) receptors produces long-term potentiation (LTP) and sound-sound associated memory (Chen et al., 2019). CCK-8 expressed in GABAergic neurons shows the immunoreactivity in rat hippocampus and dentate gyrus (Kosaka Toshio et al., 1985). It confirmed the excitatory role of CCK in auditory cortex and its inhibitory role in hippocampus, however, there is very few discussions about the role of CCK in excitatory hippocampal neural plasticity. Hence, in this thesis I hope to study the excitatory roles of CCK in hippocampus and decode how CCK produces neural plasticity and the within underlying mechanism.

In this project, I performed in-vitro electrophysiological fEPSPs recording in auditory cortical slices and hippocampal slices by the commercial 4-slice 16/64-multiple channel recording system (MED64, Alpha-Med Sciences) to explore the connection between CCK and NMDA. And I investigated the in-vitro hippocampal dynamic changes of the presynaptic Ca²⁺ fluorescence and the real-time release of CCK via Two-wide field microscope to explore the role of presynaptic NMDA receptors on the secretion of CCK.

CCK is necessary for High-Frequency stimulation (HFS)-induced LTP at hippocampal synapse and auditory cortical synapse projected from the Entorhinal cortex. Hippocampal axonal GluN2A subunits of NMDA receptors mediate the LTP induction by triggering axonal Ca²⁺ elevation. The hippocampal HFS-induced LTP is CCKA receptors dependent, but CCKB receptors also contribute to hippocampal LTP induction. Genetic depletion of the GluN1 subunits of axonal NMDA receptors in entorhinal cortico-hippocampal CCK-Containing synapses could largely abolish the axonal Ca²⁺ elevation and disturb the secretion of CCK from axon terminals. It is indicated that functional presynaptic NMDA receptor is necessary for LTP induction via the axonal Ca²⁺ elevation at typical CCK-positive neurons, to induce CCK secretion by exocytosis, which supporting the conclusion that Activity-dependent LTP in the hippocampal synapse is CCK-dependent, and CCK secretion from the axon terminal is controlled by presynaptic NMDA receptors.

Conclusion: In this study, it demonstrated that auditory cortical and hippocampal long-term potentiation are involved in CCK signaling pathway and NMDA signaling pathway. And NMDA signaling pathway is up-stream while CCK signaling pathway is down-stream. CCK release triggered by NMDA receptors produces auditory cortical LTP, and hippocampal LTP which is GluN2A receptor dependent. Both CCKA receptors and CCKB receptors contribute to hippocampal LTP induction. Presynaptic NMDA receptors are essential for the production of hippocampal Activity-dependent LTP by controlling the release of CCK that induces hippocampal LTP.