Tetrahydropalmatine is associated with attenuation of fentanyl exposure related myocardial and cardiomyocyte changes

Objective Fentanyl exposure is accompanied by myocardial and cellular alterations, but the exact mechanisms underlying cardiomyocyte injury and effective intervention strategies remain to be fully elucidated. This study aims to investigate the association between fentanyl exposure, changes in autophagy related markers, mitochondrial structural alterations, and PI3K/Akt/mTOR pathway marker changes in mouse myocardial tissue and human cardiomyocyte (AC16), while evaluating subcellular marker changes following Tetrahydropalmatine (THP) co-exposure. Methods In vivo mouse models of fentanyl exposure and in vitro AC16 injury models were established. Myocardial histological changes were assessed using hematoxylin and eosin (H&E) staining and immunohistochemistry. Myocardial cell ultrastructure and mitochondrial morphology were evaluated using transmission electron microscopy (TEM). Mitochondrial membrane potential fluorescence, MitoTracker Green (MTG) signals, and intracellular Ca 2+ fluorescence were quantitatively assessed using JC-1, MTG, and Fluo-4-AM staining. The phosphorylation ratios of PI3K/Akt/mTOR markers and autophagy related markers (total ULK1, Beclin-1, LC3-II/I protein ratio,and p62 protein; Map1lc3b for transcript measurement) was analyzed by Western blot, immunohistochemistry, immunofluorescence, and RT-qPCR. Results Fentanyl exposure was accompanied by myocardial histological shifts in mice, alongside elevated Beclin-1, LC3-II/I ratio, and total ULK1 expression and reduced p62 levels in myocardial tissue and AC16 cells (P < 0.05). THP co-exposure was accompanied by attenuation of these myocardial histological changes and autophagy related marker alterations (P < 0.05). TEM and fluorescent staining showed that fentanyl exposure was accompanied by mitochondrial structural changes, reduced MTG fluorescence intensity, increased intracellular Ca 2+ fluorescence signals, and altered mitochondrial membrane potential fluorescence (P < 0.05). Conversely, THP co-exposure was accompanied by lower Ca 2+ fluorescence signals, a higher JC-1 red/green fluorescence intensity ratio, and reduced mitochondrial structural alterations (P < 0.05). Similarly, while fentanyl exposure was accompanied by lower phosphorylation levels and mRNA expression of Pi3k, Akt, and Mtor, whereas THP co-exposure was accompanied by higher phosphorylation ratios and mRNA expression of these pathway markers (P < 0.05). Conclusion Fentanyl exposure is accompanied by alterations in autophagy related markers, mitochondrial structural changes, changes in intracellular Ca 2+ fluorescence, mitochondrial membrane potential fluorescence changes, and abnormal phosphorylation ratios of PI3K/Akt/mTOR pathway markers. THP co-exposure was associated with shifts in biological markers relative to fentanyl exposure alone. These findings provide preliminary baseline data for evaluating Corydalis yanhusuo alkaloids in opioid associated cardiomyocyte models.