Key result
WGA-pHRho successfully labeled the glycocalyx of mammalian cells without altering voltage gating, enabling spatiotemporal visualization of extracellular proton fluxes.
WGA-pHRho is a novel, photostable fluorescent sensor that enables real-time spatiotemporal visualization of extracellular proton fluxes in primary cells, including cardiomyocytes and neurons.
Provides a new tool for proton flux imaging in cardiomyocytes; leaves open translation to human cardiac electrophysiology.
Small-molecule fluorescent wheat germ agglutinin (WGA) conjugates are routinely used to demarcate mammalian plasma membranes, because they bind to the cell's glycocalyx. Here, we describe the derivatization of WGA with a pH-sensitive rhodamine fluorophore (pHRho; pKa = 7) to detect proton channel fluxes and extracellular proton accumulation and depletion from primary cells. We found that WGA-pHRho labeling was uniform and did not appreciably alter the voltage gating of glycosylated ion channels, and the extracellular changes in pH correlated with proton channel activity. Using single-plane illumination techniques, WGA-pHRho was used to detect spatiotemporal differences in proton accumulation and depletion over the extracellular surface of cardiomyocytes, astrocytes, and neurons. Because WGA can be derivatized with any small-molecule fluorescent ion sensor, WGA conjugates should prove useful to visualize most electrogenic and nonelectrogenic events on the extracellular side of the plasma membrane.
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Zhang et al. (2020) studied this question. WGA-pHRho vs. WGA-fluorescein was evaluated on Detection of extracellular proton accumulation and depletion. WGA-pHRho successfully labeled the glycocalyx of mammalian cells without altering voltage gating, enabling spatiotemporal visualization of extracellular proton fluxes.
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