ZIP-ZnT1 complexes mediate a local Zn(2+)-cycle regulating neuronal Zn²⁺ transport.

ZnT1 is the main transporter mediating Zn²⁺ efflux from the cytoplasm to the extracellular space. Paradoxically, due to relatively low Zn(2+) affinity of ZnT1, physiological levels of cytoplasmic free Zn(2+) are seemingly insufficient to support ZnT1 function. However, physiological modulation of NMDA receptor responses by synaptically released Zn(2+) requires a Zn(2+) rise in the postsynaptic neurons, accompanied by ZnT1 activity. We hypothesized that Zn(2+) import from the extracellular space could generate intracellular Zn(2+) microdomains in the vicinity of ZnT1 to enable its activity, thus forming a localized "Zn(2+)-cycle." To identify the functional machinery that may drive this process, we investigated the expression and function of ZIP proteins that import Zn(2+) into the neuronal cytoplasm. We focused on the dorsal cochlear nucleus (DCN) and hippocampus, regions where synaptic Zn(2+) modulates postsynaptic NMDA responses and synaptic plasticity. We demonstrate that ZIP3 is expressed on DCN cartwheel cells and required for postsynaptic Zn(2+) influx. Importantly, ZIP3 physically interacts with ZnT1, suggesting that it is an integral component of the synaptic Zn(2+)-cycle machinery. In the hippocampus, ZIP1, but not ZIP3, mediates Zn(2+) import into postsynaptic CA3 cells and we find that ZIP1 interacts with ZnT1 in this brain region. Importantly, Zn(2+) efflux rates are enhanced in SH-SY5Y cells co-expressing ZnT1 and either ZIP3 or ZIP1, compared to rates in cells expressing ZnT1 alone. Our findings indicate that ZnT1-dependent Zn(2+) efflux is facilitated by ZIP-ZnT1 complexes, which potentially induce local intracellular Zn(2+) microdomains supporting a Zn(2+)-cycle capable of modulating synaptic signaling.