PHRC Article in Science Advances

Manoj Kumar — Research Assistant Professor in the Department of Otolaryngology and faculty at the Pittsburgh Hearing Research Center — is first author of a study published in the March 2023 issue of Science Advances. Entitled “A genetically encoded far-red fluorescent indicator for imaging synaptically released Zn2+,” the study reveals a new technology with important implications.

According to Kumar, the existence of zinc ions in the brain has been known for decades. Zinc ions help neurons to communicate with one another at neuron junctions called synapses and regulate sensory information processing in the brain. Changes to the amount of zinc ions at the synapses has been linked to numerous neurological diseases, such as stroke, epilepsy, depression, Alzheimer’s disease, and hearing disorders.

“Although the importance of zinc in the brain has been widely known, because of the lack of sophisticated technology, it was not possible to visualize how the zinc ions are being released at the synapses and mediate its actions in the brain,” Kumar said. “By utilizing genetic, molecular, and imaging techniques, we have now engineered and validated a technology that will allow the biomedical field to visualize and record zinc ion activity in the brain in real-time in acute brain-slice preparations.”

This was accomplished by engineering a zinc-sensitive fluorescent protein called Far-Red Fluorescent Indicator for Synaptic Zinc (FRISZ). Next, the team genetically modified FRISZ and packed it with viral vectors to express FRISZ over the outer cell membranes of brain neurons. As Kumar explained, “upon activation by zinc release at the neuron synapses during neurotransmission, FRISZ fluoresces sufficient brightness to be detected by sophisticated microscopes, providing a direct and robust measure of zinc signal intensity and location.”

Next, the team validated FRISZ to visualize zinc release in the auditory cortex, the brain region where sound information gets processed. In acute brain slices, electric stimuli elicit a robust increase in FRISZ fluorescence that was reduced with the application of a chemical compound that temporarily mops up zinc ions from the synapses.

 “Our study establishes a novel genetically encoded technology for studying the roles of zinc ions in the nervous system,” Kumar said.