AbstractFebrile seizures (FS) are the most common seizure syndrome and are potentially a prelude to more severe epilepsy. Although zinc (Zn²⁺) metabolism has previously been implicated in FS, whether or not variation in proteins essential for Zn²⁺ homeostasis contributes to susceptibility is unknown. Synaptic Zn²⁺ is co-released with glutamate and modulates neuronal excitability. SLC30A3 encodes the zinc transporter 3 (ZNT3), which is primarily responsible for moving Zn²⁺ into synaptic vesicles. Here we sequenced SLC30A3 and discovered a rare variant (c.892C > T; p.R298C) enriched in FS populations but absent in population-matched controls. Functional analysis revealed a significant loss-of-function of the mutated protein resulting from a trafficking deficit. Furthermore, mice null for ZnT3 were more sensitive than wild-type to hyperthermia-induced seizures that model FS. Together our data suggest that reduced synaptic Zn²⁺ increases the risk of FS and more broadly support the idea that impaired synaptic Zn²⁺ homeostasis can contribute to neuronal hyperexcitability.