We characterized the transcriptional response accompanying maintenance of long-term sensitization (LTS) memory in the pleural ganglia of using microarray ( = 8) and qPCR ( = 11 additional samples). We found that 24 h after memory induction there is strong regulation of 1198 transcripts (748 up and 450 down) in a pattern that is almost completely distinct from what is observed during memory encoding (1 h after training). There is widespread up-regulation of transcripts related to all levels of protein production, from transcription (e.g., subunits of transcription initiation factors) to translation (e.g., subunits of eIF1, eIF2, eIF3, eIF4, eIF5, and eIF2B) to activation of components of the unfolded protein response (e.g., CREB3/Luman, BiP, AATF). In addition, there are widespread changes in transcripts related to cytoskeleton function, synaptic targeting, synaptic function, neurotransmitter regulation, and neuronal signaling. Many of the transcripts identified have previously been linked to memory and plasticity (e.g., Egr, menin, TOB1, IGF2 mRNA binding protein 1/ZBP-1), though the majority are novel and/or uncharacterized. Interestingly, there is regulation that could contribute to metaplasticity potentially opposing or even eroding LTS memory (down-regulation of adenylate cyclase and a putative serotonin receptor, up-regulation of FMRFa and a FMRFa receptor). This study reveals that maintenance of a "simple" nonassociative memory is accompanied by an astonishingly complex transcriptional response.

Most long-term memories are forgotten. What happens, then, to the changes in neuronal gene expression that were initially required to encode and maintain the memory? Here we show that the decay of recall for long-term sensitization memory in is accompanied both by a form of savings memory (easier relearning) and by persistent transcriptional regulation. A behavioral experiment ( = 14) shows that sensitization training produces a robust long-term sensitization memory, but that recall fades completely within 1 wk. This apparent forgetting, though, is belied by persistent savings memory, as we found that a weak reminder protocol reinstates a long-term sensitization memory only on the previously trained side of the body. Using microarray ( = 8 biological replicates), we found that transcriptional regulation largely decays along with recall. Of the transcripts known to be regulated 1 d after training, 98% (1172/1198) are no longer significantly regulated 7 d after training. Still, there is a small set of transcripts which remain strongly regulated even when recall is absent. Using qPCR ( = 11 additional biological replicates) we confirmed that these include the peptide transmitter FMRFamide, a transcript encoding a putative homolog of spectrin beta chain (Genbank: EB255259) , a transcript encoding a protein with a predicted EF-hand calcium-binding domain (Genbank: EB257711), and eight uncharacterized transcripts. To our knowledge, this is the first work to show that transcriptional changes evoked by learning can outlast recall. The small set of transcriptional changes that persist could mediate the rapid relearning of the memory (savings), or the decay of recall, or both, or neither.

Most long-term memories are forgotten, becoming progressively less likely to be recalled. Still, some memory fragments may persist, as savings memory (easier relearning) can be detected long after recall has become impossible. What happens to a memory trace during forgetting that makes it inaccessible for recall and yet still effective to spark easier re-learning? We are addressing this question by tracking the transcriptional changes that accompany learning and then forgetting of a long-term sensitization memory in the tail-elicited siphon withdrawal reflex of Aplysia californica. First, we tracked savings memory. We found that even though recall of sensitization fades completely within 1 week of training, savings memory is still detectable at 2 weeks post training. Next, we tracked the time-course of regulation of 11 transcripts we previously identified as potentially being regulated after recall has become impossible. Remarkably, 3 transcripts still show strong regulation 2 weeks after training and an additional 4 are regulated for at least 1 week. These long-lasting changes in gene expression always begin early in the memory process, within 1 day of training. We present a synthesis of our results tracking gene expression changes accompanying sensitization and provide a testable model of how sensitization memory is forgotten.