Morphine reprograms brain-derived extracellular vesicles from cargo linked to synaptic remodelling in the prefrontal cortex

Extracellular vesicles (EVs) released by neurons and glial cells mediate intercellular communication in the brain and regulate synaptic function, neuronal survival, and neuropathological processes. Although chronic opioid exposure induces widespread neuroadaptations, the contribution of brain-derived EVs (BDEVs) to these processes remains largely unknown. Here, we isolated BDEVs from the prefrontal cortex of rats chronically exposed to morphine and performed integrative transcriptomic and proteomic analyses of their molecular cargo. Total RNA sequencing combined with unbiased proteomics revealed that morphine profoundly reprograms the BDEV transcriptome and proteome, enriching pathways related to synaptic plasticity, endoplasmic reticulum (ER) stress, mitochondrial dysfunction, and neurodegeneration. Among the most prominent alterations, the synaptic regulator ARC was consistently modulated at the mRNA level, while the ER stress marker HSPA5 was altered at both mRNA and protein levels. Functional assays further demonstrated that BDEVs derived from morphine-treated rats were sufficient to reconfigure transcriptional programs in naïve cortical neurons, affecting genes associated with synaptic remodeling and excitability. Collectively, these findings provide the first evidence that chronic opioid exposure reprograms BDEV cargo in a brain region critical for addiction and that these vesicles can propagate transcriptional reorganization to recipient neurons. BDEVs thus emerge as active mediators of morphine-induced neuroadaptations and as potential targets for biomarker discovery and therapeutic intervention in opioid use disorder.