Studying the movement of cellular channels under the microscope

Master’s student Haifei You (pictured; Swayne Lab) has received a Canada Graduate Scholarship – Master’s (CGS-M) from the Natural Sciences and Engineering Research Council of Canada (NSERC) to study how pannexin 1 (PANX1) moves from the outer surface of neural cells to membranes inside those cells.

PANX1 is a channel protein that forms doorways in membranes that surround cells. These doorway proteins enable the transmission of signals telling the cell how to change in response to its environment, and they also act as scaffolds to bring key signalling proteins together. These functions allow PANX1 to conduct a variety of roles in the brain, both in development and disease.

The Swayne Lab previously reported that extracellular ATP triggers internalization of PANX1 in a model neural cell line, called Neuro2a, thereby increasing the levels of PANX1 inside those cells. This model was used in the foundational work because these cells are relatively uncomplicated compared to neurons, facilitating the tracking of PANX1. For her CGS-M project, Haifei is exploring the cellular mechanism underlying this internalization process. Based on prior work, some of which the lab published in a preprint article, she suspects that it may be via a process known as macropinocytosis, or “cell-drinking.” This is when a cell internalizes large amounts of fluid, molecules, and membrane in large vesicles aptly called macropinosomes.

Haifei is using imaging techniques, including super-resolution and confocal microscopy, to determine if the vesicles containing PANX1 are located at the same sites as the macropinosomes. “If we observe this correlation, it would suggest that the mechanism of internalization is macropinocytosis,” she says. She presented her preliminary findings at the 16^th Canadian Neuroscience Meeting in Montreal in May.

Haifei will also use the same techniques to determine if PANX1 is also internalized by macropinocytosis in cultured neurons. These experiments will be more technically challenging given that neurons have a relatively more complex shape, but Haifei will have garnered ample experience from her initial work with Neuro2a cells. “I’m excited to apply these imaging techniques to neurons and to confirm exactly how PANX1 is getting inside cells. The outcomes of this work might also provide clues as to what PANX1 might be doing on the inside and how its various functions in cells could be linked.”

CGS-M awards are administered jointly by Canada’s three granting agencies: NSERC, the Canadian Institutes of Health Research, and the Social Sciences and Humanities Research Council. These awards are highly competitive and are designed to support research excellence, innovation, and impact in the health field.