We have generated genetically-engineered mice to suppress specific subset of neurons activated during learning. We found that the suppression of neuronal ensembles that are naturally activated during learning results in a failure of the memory retrieval. We further found that the suppression selectively inhibits relearning without disrupting the ability to acquire and retrieve a memory for distinct context. These results indicate that there is a mechanism ensuring that the same neuronal ensemble is engaged for the same learning to strengthen the memory, and it is not substitutable after the ensemble is allocated for the initial learning. Our results provide substantial insights into the machinery underlying how the brain allocates individual memories to discrete neuronal ensembles and how it ensures that repetitive learning strengthens memory by reactivating the same neuronal ensembles. Our findings could be of some help in understanding the mechanism underlying memory impairments associated with aging and psychiatric disorders.