The hippocampus is broadly impacted by neuromodulations. However, how neuropeptides shape the function of the hippocampus and the related spatial learning and memory remains unclear. Here, we discover the crucial role of cholecystokinin (CCK) in heterosynaptic neuromodulation from the medial entorhinal cortex (MEC) to the hippocampus. Systematic knockout of the CCK gene impairs CA3-CA1 LTP and space-related performance. The MEC provides most of the CCK-positive neurons projecting to the hippocampal region, which potentiates CA3-CA1 long-term plasticity heterosynaptically in a frequency- and NMDA receptor (NMDAR)-dependent manner. Selective inhibition of MEC CCKergic neurons or downregulation of their CCK mRNA levels also impairs CA3-CA1 LTP formation and animals’ performance in the water maze. This excitatory extrahippocampal projection releases CCK upon high-frequency excitation and is active during animal exploration. Our results reveal the critical role of entorhinal CCKergic projections in bridging intra- and extrahippocampal circuitry at electrophysiological and behavioral levels.

The hippocampus is a spatial learning and memory hub underpinned by long-term potentiation (LTP). However, it remains unclear how neuropeptides shape the process. Dual-optogenetic stimulation enables us to differentiate the roles of different projections in LTP induction. Ex vivo recordings showed theta-burst stimulation (TBS) of neither Schaffer collateral (SC) nor medial entorhinal cholecystokinin (MECCCK) projection alone optogenetically could induce homosynaptic LTP in their own pathway. CA3-CA1 LTP could only be induced heterosynaptically when SC activation is paired with TBS of the MECCCK projections, which released CCK. MECCCK neurons showed higher activity and likely released CCK during exploration. Systematic knockout of CCK impaired LTP and spatial-related task performance. Selective inhibition of the MECCCK neurons or down-regulation of their Cck mRNA level impaired LTP formation and behavioral performance. Our results reveal an enabling role of MEC CCKergic modulation in TBS-induced CA3-CA1 LTP and spatial memory.

Cholecystokinin (CCK) is purported to be involved in neuroplasticity and memory encoding. Here we found that CCK from the medial entorhinal (MEC) to hippocampus projections facilitated CA3-CA1 long-term potentiation (LTP), which further enabled spatial memory transfer. CCK knockout mice lacked theta-burst stimulation-induced CA3-CA1 LTP and their spatial memory were compromised. Upon high-frequency optical activation, the MEC CCKergic terminals in the hippocampus released CCK and induced CA3-CA1 LTP, mainly through CCK A receptors. However, down-regulation of the MEC Cck expression impaired LTP. Stimulation of hippocampal CCK neurons, the Schaffer Collaterals, or the inhibitory MEC to hippocampus terminals, induced no CA3-CA1 LTP. Activation of a group of CA1 pyramidal neurons when the mouse visited a certain place of an arena, induced primed place cells (PPCs). The CCK-strengthened connectivity of two groups of PPCs enabled the transfer of fear memory from one place to another, implicating CCK's determining role in spatial memory encoding.

Human induced pluripotent stem cells (iPSC) can be used to understand the pathological mechanisms of human disease. These cells are a promising source for cell-replacement therapy. However, such studies require genetically defined conditions. Such genetic manipulations can be performed using the novel Transcription Activator-Like Effector Nucleases (TALENs), which generate site-specific double-strand DNA breaks (DSBs) with high efficiency and precision. Combining the TALEN and iPSC methods, we developed two iPS cell lines by generating the point mutation A5768G in the SCN1A gene, which encodes the voltage-gated sodium channel Nav1.1 α subunit. The engineered iPSC maintained pluripotency and successfully differentiated into neurons with normal functional characteristics. The two cell lines differ exclusively at the epilepsy-susceptibility variant. The ability to robustly introduce disease-causing point mutations in normal hiPS cell lines can be used to generate a human cell model for studying epileptic mechanisms and for drug screening.