During insulin secretion, pancreatic a-cells are exposed to Zn2+ released from insulin-containing secretory granules. Although maintenance of Zn2+ homeostasis is critical for cell survival and glucagon secretion, very little is known about Zn2+-transporting pathways and the regulation of Zn2+ in a-cells. To examine the effect of Zn2+ on glucagon secretion and possible mechanisms controlling the intracellular Zn2+ level ([Zn2+]i) we employed a glucagon-producing cell line (a-TC6) and mouse islet where non-beta cells were identified using islets expressing green fluorescent protein (GFP) exclusively in b-cells. In this study, we first confirmed that Zn2+ treatment resulted in the inhibition of glucagon secretion in a-TC6 cells and mouse islets in vitro. The inhibition of secretion was not likely via activation of KATP channels by Zn2+. We then determined that Zn2+ was transported into a-cells and was able to accumulate under both low and high glucose conditions, as well as upon depolarization of cells with KCl. The nonselective Ca2+ channel blocker Gd3+ partially inhibited Zn2+ influx in a-TC cells, while the L-type voltage-gated Ca2+ channel inhibitor nitrendipine failed to block Zn2+ accumulation. To investigate Zn2+ transport further we profiled a-cells for Zn2+ transporter transcripts from the two families, SLC39 (ZIP/ZRT1-related cytoplasmic Zn2+ uptake transporters) and SLC30 (ZnTs, Zn2+ export transporters). We observed that Zip1, Zip10, and Zip14 were the most abundantly expressed ZIPs and ZnT4, ZnT5, and ZnT8 the dominant ZnTs. Because the redox state of cells is also a major regulator of [Zn2+]i we examined the effects of oxidizing agents on Zn2+ mobilization within a-cells. DTDP (SH-group oxidant), menadione (superoxide generator) and SIN-1 (peroxinitrite generator), all increased [Zn2+]i in a-cells. Together these results demonstrate that Zn2+ inhibits glucagon secretion and it is transported into a-cells in part through Ca2+ channels. Zn2+ transporters and the redox state also modulate [Zn2+]i .