• mouse
• entorhinal cortex
• episodic-like-memory

The Entorhinal Cortex (EC) represents a crucial site for memory formation as it integrates spatial information processed from its medial subdivision (MEC) with non-spatial information processed from its lateral part (LEC). Specific behavioural tests, such as the novel object-place-context recognition test (OPCRT) have been designed to investigate episodic-like memory in mice. Using this task, it has been demonstrated that LEC superficial layer neurons are required for recognition of objects that have been experienced in a specific context and that the specific lesion of the LEC causes an impairment in the ability to discriminate either novel object/place or novel object/place/context associations without affecting the recognition of a novel object. However, it is still not clear whether LEC superficial layer neurons are merely involved in object information processing during the formation or are re-activated during retrieval of associative memory. To address this issue, we planned to assess the specificity of LEC activation induced by the OPCRT. We first investigated OPCRT-induced synaptic changes in layer II LEC intrinsic circuitry compared to the adjacent MEC and the main target circuitry (performant pathway to dentate gyrus). Then we compared the neuronal activation in superficial and deep layers of the LEC compared to the MEC and other brain areas, using the c-fos as a marker of neuronal activity. Experiments were conducted on male C57bl/6J mice (3 to 4 months of age) that were divided in the following groups: mice subjected to OPCRT, mice exposed only to the contexts (CNTX) and mice remained in their home cages (CTRL). The test consists in the presentation of the objects in association with a specific position and context in the sample trials, followed by a test trial in which mice discriminate between familiar and novel OPC associations. Mice memory performance was quantified after the analysis of the time spent in the exploration of the two objects and expressed as discrimination index (DI = time at novel - time at familiar object / time at novel + time at familiar object). Electrophysiological analyses revealed the capability of OPCRT to induce significant changes in the synaptic efficacy in the LEC intrinsic circuitry, but not in the MEC nor in the hippocampus. The analysis of c-fos immunostaining revealed a significant and selective increase of EC activation after the execution of the task. In order to dissect the specific role of LEC neurons we made use of inducible- DREADDs (Designer Receptors Exclusively Activated by Designer Drugs) that were stereotaxically injected in the LEC to specifically manipulate transfected neurons during the OPCRT. In particular, inhibitory G protein-coupled receptor (GPCR) in LEC neurons could be expressed in a tamoxifen-dependent manner, and then activated by administration of CNO during the recall phase of the associative memory task. Selective inhibition of LEC neurons previously engaged during memory formation, resulted in a significant loss of associative memory. Our findings suggest that activation of a selective population of EC neurons is necessary not only for the formation but also for the retrieval of associative memory.