Author ORCID Identifier

https://orcid.org/0000-0002-4498-932X

Semester

Fall

Date of Graduation

2024

Document Type

Dissertation

Degree Type

PhD

College

School of Medicine

Department

Not Listed

Committee Chair

Kevin Daly

Committee Member

Charles Anderson

Committee Member

Ekaterina Weil

Committee Member

Michelle Bridi

Committee Member

Martin Hruska

Abstract

Many regions of the brain are highly enriched with free zinc inside glutamatergic vesicles at presynaptic terminals. At these synapses, zinc transporter 3 (ZnT3) moves zinc into vesicles where it is co-released with glutamate. Alterations in ZnT3 are implicated in multiple neurodevelopmental disorders, and ZnT3 knock-out (KO) mice – which lack synaptic zinc – show behavioral deficits associated with autism spectrum disorder and schizophrenia. Shank3 is a synaptic scaffolding protein that assists in tethering and organizing structural proteins and glutamatergic receptors in the postsynaptic density of excitatory synapses. The localization of Shank3 at excitatory synapses and the formation of stable Shank3 complexes is regulated by the binding of zinc to the C-terminal sterile-alpha-motif (SAM) domain of Shank3. Mutations in the SAM domain of Shank3 result in altered synaptic function and morphology, and disruption of zinc in synapses that express Shank3 leads to a reduction of postsynaptic proteins important for synaptic structure and function. This suggests that zinc supports the localization of postsynaptic proteins via Shank3. Here we show that male and female ZnT3 KO mice have smaller dendritic spines and miniature excitatory postsynaptic current amplitudes than wildtype (WT) mice in the auditory cortex. Additionally, spine size deficits in ZnT3 KO mice are restricted to synapses that express Shank3. In WT mice, synapses that express both Shank3 and ZnT3 have larger spines compared to synapses that express Shank3 but not ZnT3. Together these findings suggest a mechanism whereby presynaptic ZnT3-dependent zinc supports postsynaptic structure and function via Shank3 in a synapse-specific manner. Additionally, cellular mechanisms that regulate the actions of zinc at synapses are of great importance for understanding the roles of zinc during synaptic signaling. Here, we identify the astrocytic zinc transporter protein ZIP12 as a candidate protein that contributes to zinc clearance from the synaptic cleft. We show that ZIP12 is expressed at cortical synapses, and that its expression is ZnT3-dependent. We identify novel small molecule compounds that specifically antagonize the function of ZIP12 in heterologous expression systems, and we use these compounds to increase the concentration of ZnT3-dependent zinc at synapses and inhibit the activity of AMPA and NMDA receptors in acute brain slices of mouse auditory cortex. These results identify a molecular pathway that can be targeted with pharmacological agents that modulate synaptic zinc signaling to advance our understanding of the roles of synaptic zinc in health and disease.

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