Gabapentin lactam induces neurodevelopmental deficits via calcium-mitochondria-ROS cascade

Abstract

Gabapentin lactam (GBP-L), a transformation product of the anticonvulsant gabapentin, persists in surface waters at concentrations reaching μg/L levels due to its recalcitrance to conventional treatment processes. Despite growing concerns about pharmaceutical transformation products, their mechanistic toxicity pathways remain poorly characterized. Here, we used zebrafish embryo-larval models to investigate the neurodevelopmental effects of environmentally relevant GBP-L concentrations (8, 80, and 800 μg/L). Exposure to GBP-L had no significant effect on the survival rate of zebrafish embryos, but significantly impaired the hatching process, increased the malformation rate, and inhibited body length development—with a 3.86 % reduction in body length compared to the control group at 72 hpf and a 3.70 % reduction at 144 hpf. Motor assessment revealed a concentration-dependent reduction in swimming activity. Meanwhile, transgenic imaging indicated a decrease in neuronal fluorescence intensity by 9.46 %, alongside a reduction in axonal length of 7.84 %. Mechanistic investigations demonstrated that GBP-L activates calcium signaling pathways, triggering intracellular Ca²⁺ overload that leads to mitochondrial dysfunction, increased reactive oxygen species (ROS) production, and disrupted neurotransmitter homeostasis. Co-exposure with the calcium channel blocker verapamil significantly mitigated these effects, confirming calcium signaling as the molecular initiating event in GBP-L neurotoxicity. Transcriptomic analyses further validated the calcium-mitochondria-ROS cascade as the critical toxicity pathway. This study provides the first evidence that GBP-L, a persistent pharmaceutical transformation product, can disrupt neurodevelopment by perturbing calcium signalling at environmentally relevant concentrations, highlighting the need to establish a comprehensive risk assessment framework that includes transformation products.