Along with the hallmark of α-synuclein deposition, neuroinflammation and iron accumulation have emerged as essential pathological features for dopaminergic neuron degeneration in PD patients and animal models. Preclinical studies have highlighted gentiopicroside's anti-inflammatory activities in treating arthritis, colitis and pancreatitis, and its neuroprotective effects on neurological diseases such as AD, chronic neuropathic pain and ischemia. However, the effects and mechanisms of gentiopicroside on PD-related conditions remain uncertain. Here, we evaluated the potential benefits of gentiopicroside using a unilateral 6-OHDA rat model and a MPP+-induced cell model. Our findings indicated that gentiopicroside improved motor deficits and restored nigral TH-positive neurons in vivo. Mechanistically, gentiopicroside ameliorated inflammatory responses of 6-OHDA-induced rats, decreased NF-κB and pro-inflammatory cytokines levels and reduced Iba-1-positive microglia in the substantia nigra. Furthermore, gentiopicroside regulated the levels of DMT1 and FPN1, thereby inhibiting iron accumulation in PD rats. In vitro, gentiopicroside preserved the viability of MPP+-treated SH-SY5Y cells and suppressed NF-κB activity and its downstream factors' levels. Meanwhile, gentiopicroside inhibited lipid peroxidation and ROS production, while it upregulated the expression of GPX4 in MPP+-treated cells. And these antiferroptosis effects were also linked to iron transporters regulation. Conclusively, gentiopicroside exhibits neuroprotective effects via alleviating neuroinflammation and iron-dependent ferroptosis, offering promise for PD treatment.© 2025 Nordic Association for the Publication of BCPT (former Nordic Pharmacological Society). Published by John Wiley & Sons Ltd.