Abstract
Disruption of extracellular pH and proton-sensing can profoundly impact cellular and protein functions, leading to developmental defects. To visualize changes in extracellular pH in the developing embryo, we generated a zebrafish transgenic line that ubiquitously expresses the ratiometric pH-sensitive fluorescent protein pHluorin2, tethered to the extracellular face of the plasma membrane using a glycosylphosphatidylinositol (GPI) anchor. Monitoring of pHluorin2 with ratiometric fluorescence revealed dynamic and discrete domains of extracellular acidification over the first 72 h of embryonic development. These included acidification of the notochord intercalations, transient acidification of the otic placode, and persistent acidification of the extracellular space of the myotome at distinctly different pH from that within the T-tubules. Knockdown of centronuclear myopathy genes Bin1b (OMIM: 255200) and MTM1 (OMIM: 310400), which disrupt T-tubule formation, also disrupted myotome acidification. In this study we visualize extracellular acidic microdomains in the tissues of whole live animals. This real-time reporter line for directly measuring changes in extracellular pH can be used to illuminate the role of extracellular pH in normal physiological development and disease states.