Modified compound 511 reverses nicotine addiction by normalizing the PKA-CREB-GluN2B-CaMKII signaling pathway in the ventral tegmental area.

Nicotine addiction is a chronic and relapsing disorder, but current therapies have limited effect with severe side effects. Modified Compound 511 (M511), optimized from the anti-addiction formula Compound 511, was innovatively employed in this study to evaluate its therapeutic intervention effects in nicotine addiction. This study aimed to explore the efficacy of M511 in nicotine addiction intervention and elucidate the underlying mechanisms. The therapeutic effect of M511 on nicotine addiction was evaluated using the conditional place preference (CPP) paradigm, with Varenicline as the positive control. Hematoxylin and eosin (HE) staining was performed for histopathological examination of visceral organs. c-Fos immunofluorescence was utilized to screen responsive brain regions. Network pharmacology and RNA sequencing (RNA-seq) of ventral tegmental area (VTA) tissue were integrated to predict key signaling pathways and targets, which were subsequently validated by Western blot and RT-qPCR. The PKA inhibitor H-89 was employed for target validation via CPP, Western blot, immunofluorescence, Nissl staining, and Golgi staining. LC-MS/MS was employed for chemical characterization of M511. M511 (6 g/kg, medium dose) most effectively blocked nicotine-induced CPP in mice and reversed nicotine-associated weight loss, attenuated spontaneous locomotor activity, and ameliorated multi-organ toxicity. c-Fos immunofluorescence identified the nucleus accumbens (NAc) and VTA as key responsive brain regions. Integrated analysis of network pharmacology and RNA-seq of the VTA collectively demonstrated that M511 likely acts by modulating chemical synaptic transmission and synaptic plasticity. Mechanistically, M511 inhibited PKA activation in the VTA, thereby downregulating CREB phosphorylation and subsequent GluN2B-CaMKII signaling, reducing synaptic marker proteins PSD-95 and Synapsin-Ⅰ expression, and ultimately ameliorating nicotine-induced synaptic structural and functional abnormalities. Using the PKA inhibitor H-89 mimicked the effects of M511, effectively intervening in addiction. LC-MS/MS identified 27 characteristic components, predominantly alkaloids and their derivatives, as well as flavonoids. M511 reverses nicotine addiction by normalizing the PKA-CREB-GluN2B-CaMKII signaling cascade in the VTA and ameliorating nicotine-induced synaptic plasticity abnormalities.