Brain transcriptomic analysis after the reinstatement of cocaine-seeking behavior
To further understand the role of PLCB1 in the molecular mechanisms involved in this cocaine relapse-related phenotype, we analyzed the transcriptomic profiles of mPFC and HPC immediately after the reinstatement of cocaine-seeking behavior in WT and Plcb1 +/- mice trained with cocaine. We identified 2,115 protein-coding genes differentially expressed (DEGs) in mPFC (1,231 downregulated and 943 upregulated) and only 12 in HPC, when comparing Plcb1+/- and WT animals (Supplementary Tables S2-3). In accordance, PCA analysis and heatmap plots revealed that individuals with the same genotype plotted together only in mPFC, but not in HPC (Supplementary Figure S3). This suggested a more prominent role of mPFC, so further studies were carried out only with DEGs of this brain area. Analysis of functional group over-representation identified several processes previously related to cocaine addiction, including dopaminergic synapse, learning, long-term potentiation (LTP), neurotransmitter secretion, and axon guidance, as well as other relevant signaling pathways such as MAPK, mTOR, and neurotrophin (Figure 3A-B and Supplementary Tables S3-5). We focused on the dopaminergic synapse pathway (enriched in the mPFCs DEGs;P Adj<0.05), in which phospholipase c (such as Plcb1) is directly participating in signal transmission (Figure 4). Interestingly, many genes coding for proteins in this pathway are differentially expressed in Plcb1+/- mice after cue-induced reinstatement (Figure 4 and Supplementary Table S6). Furthermore, we found that several TFBS were over‐represented in the DEGs of mPFC, including YY1, MYOD, NRF1, ERR1, FREAC2, NFY and E4F1 (complete list of TFBS in Supplementary Table S7). Then, we filtered the DEGs on mPFC based on fold-change (FC>|1.2|) and obtained a list of 238 genes that we used for gene network construction. This analysis showed a highly scored network (score=62, Figure 3C) that includes 31 DEGs involved in ”cellular development, cellular growth and proliferation, nervous system development and function”.