Polar localization of a dihydropyridine receptor on living Fucus zygotes.

We have used a fluorescently-labeled dihydropyridine (FL-DHP) to vitally stain living Fucus zygotes during the establishment of cell polarity. Localization of FL-DHP is primarily at the plasma membrane and FL-DHP binding is competitively blocked by an unlabeled dihydropyridine. Distribution of FL-DHP is initially symmetrical before fixation of the polar axis, but becomes asymmetrical in response to a unilateral light gradient. The distribution of FL-DHP receptors can be relocalized when the direction of the photopolarizing stimulus is changed. Treatment of cells with cytochalasin B prior to axis fixation reversibly prevents localization of FL-DHP receptors. Observation of FL-DHP labeling by time-lapse fluorescence microscopy indicates that the existing receptors are redistributed during polar axis formation. The asymmetric distribution of FL-DHP receptors coincides temporally and spatially with increased local intracellular calcium ion concentrations, as measured by calcium green dextran. Based on the site, timing, photo-reversibility, and actin dependence of the asymmetric localization of FL-DHP receptors, we conclude that FL-DHP is a vital probe for the later stage of polar axis formation in Fucus zygotes. Furthermore, we propose that FL-DHP receptors correspond to ion channels that are transported to the future site of polar growth to create the changes in local calcium concentration required for polarity establishment.