Synaptic Structure and Transcriptomic Profiling of Reward and Sensory Brain Areas in Male Mice of Fentanyl Addiction

Junli Feng,1,* Ningsi Xu,1,* Linhua Wang,2 Haixing Wang,3 Yi Zhou,3 Qing Shen4,5 1Collaborative Innovation Center of Seafood Deep Processing, Zhejiang Province Joint Key Laboratory of Aquatic Products Processing, Institute of Seafood, Zhejiang Gongshang University, Hangzhou, 310012, People’s Republic of China; 2Hangzhou Linping Hospital of Traditional Chinese Medicine, Linping, Zhejiang, 311106, People’s Republic of China; 3Key Laboratory of Drug Monitoring and Control of Zhejiang Province, National Narcotics Laboratory Zhejiang Regional Center, Hangzhou, 310053, People’s Republic of China; 4The Joint Innovation Center for Health & Medicine, The Quzhou Affiliated Hospital of Wenzhou Medical University, Quzhou People’s Hospital, Quzhou, 324000, People’s Republic of China; 5Laboratory of Food Nutrition and Clinical Research, Zhejiang Gongshang University, Hangzhou, 310012, People’s Republic of China*These authors contributed equally to this workCorrespondence: Yi Zhou; Qing Shen, Email [email protected]; [email protected]; [email protected]: Opioid-based medications are powerful analgesics commonly prescribed for pain management, but they are also highly addictive. The over-prescription of opioids analgesics has triggered current opioid crisis, which now has expanded to heroin and illicit synthetic opioids like fentanyl and its analogues. The side effects of fentanyl abuse have been well recognized, yet the underlying molecular adaptations across brain regions upon fentanyl exposure remain elusive.Methods: The transmission electron microscopy (TEM) and next-generation RNA-sequencing (RNA-seq) were used to investigate the ultrastructure synaptic alterations and transcriptional profiling changes of reward and sensory brain regions in mice after fentanyl exposure.Results: The naloxone-precipitated acute withdrawal symptoms were observed in mice exposed to fentanyl. Results of TEM showed an increase in the number of synapses, widening of synaptic gaps, and thickening of postsynaptic density in the NAc of the fentanyl addiction mice, accompanied by obvious mitochondrial swelling. RNA-seq identified differentially expressed genes (DEGs) in prefrontal cortex of mice brains after fentanyl exposure, and the expression of some addiction-related genes such as Calm4, Cdh1, Drd1/2/3/4, F2rl2, Gabra6, Ht2cr, Oprk1 and Rxfp3 showed the most striking changes among experimental groups. KEGG enrichment analysis indicated that these DEGs were related to the development of addiction behavior, dopaminergic/GABAergic/serotonergic synapse, synapse assembly/synaptic plasticity/synaptic vesicle cycle, cAMP/MAPK signaling pathway, neuroactive ligand-receptor interactions. These transcriptomic changes may be correlated with the structural and behavioral changes observed in fentanyl-exposed mice.Discussion: The findings of this study contribute to a better understanding of the molecular mechanism of addiction behavior, which is essential for the development of optimized therapy strategies for addicts.Keywords: fentanyl, addiction, synaptic plasticity, gene expression, transcriptional profiles