Synaptically released zinc has neuromodulatory capabilities that could result in either inhibition or enhancement of neuronal excitability. To determine the net effects of vesicular zinc release in the brain in vivo, we examined seizure susceptibility and seizure-related neuronal damage in mice with targeted disruption of the gene encoding the zinc transporter, ZnT3 (ZnT3-/- mice). ZnT3-/- mice, which lack histochemically reactive zinc in synaptic vesicles, had slightly higher thresholds to seizures elicited by the GABA(A) antagonist, bicuculline, and no differences in seizure threshold were seen in response to pentylenetetrazol or flurothyl. However, ZnT3-/- mice were much more susceptible than wild-type mice to limbic seizures elicited by kainic acid, suggesting that the net effect of hippocampal zinc on acute seizures in vivo is inhibitory. The hippocampi of ZnT3-/- mice showed typical seizure-related neuronal damage in response to kainic acid, demonstrating that damage to the targets of zinc-containing neurons can occur independently of synaptically released zinc. Mice lacking the neuronal zinc-binding protein metallothionein III (MT-III) are also more susceptible to kainic acid-induced seizures. Double knockout (ZnT3 and MT3) mice show the same response to kainic acid as ZnT3-/- mice, suggesting that ZnT3 and MT-III function in the same pathway.