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Lily pollen tubes possess a steep, tip-focused intracellular Ca2+ gradient and a tip-directed extracellular Ca2+ influx. Ratiometric ion imaging revealed that the gradient extends from above 3.0 microM at the apex to approximately 0.2 microM within 20 microns from the tip, while application of the Ca(2+)-specific vibrating electrode indicated that the extracellular influx measured between 1.4 and 14 pmol cm-2 sec-1. We examined the relationship between these phenomena and their role in tube growth by using different 1,2-bis(o-aminophenoxy)ethane N,N,N',N'-tetraacetic acid (BAPTA)-type buffers and hypertonic media. Injection of active BAPTA-type buffers or application of elevated levels of sucrose reversibly inhibited growth, destroyed tip zonation of organelles, and modified normal patterns of cytoplasmic streaming. Simultaneously, these treatments dissipated both the intracellular tip-focused gradient and the extracellular Ca2+ flux. Of the BAPTA-type buffers, 5,5'-dibromo-BAPTA (dissociation constant [Kd] is 1.5 microM) and 4,4'-difluoro-BAPTA (Kd of 1.7 microM) exhibited greater activity than those buffers with either a higher affinity (5,5'-dimethyl-BAPTA, Kd of 0.15 microM; BAPTA, Kd of 0.21 microM; 5,5'-difluoro-BAPTA, Kd of 0.25 microM) or lower affinity (5-methyl, 5'-nitro-BAPTA, Kd of 22 microM) for Ca2+. Our findings provide evidence that growing pollen tubes have open Ca2+ channels in their tip and that these channels become inactivated in nongrowing tubes. The studies with elevated sucrose support the view that stretching of the apical plasma membrane contributes to the maintenance of the Ca2+ signal.

The role of extracellular Ca(2+) in root-hair tip growth has been investigated in Arabidopsis thaliana (L.) Heynh. Root-hair length was found to be dependent on the concentration of Ca(2+) in the growth medium, with maximum length achieved at [Ca(2+)] of 0.3-3.0 mM. Using a non-intrusive calcium-specific vibrating microelectrode, an extracellular Ca(2+) gradient was detected at the tips of individual growing root-hair cells. The direction of the gradient indicated a net influx of Ca(2+) into root-hair cells. No gradient was detected near the sides of the root hairs or at the tips of non-growing root hairs. When root hairs were exposed to the Ca(2+)-channel blocker nifedipine, tip growth stopped and the extracellular Ca(2+) gradient was abolished. These results indicate that Ca(2+) influx through plasma-membrane Ca(2+) channels is required for normal root-hair tip growth.

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Detection of motion and position by the vestibular labyrinth depends on the accumulation of potassium within a central compartment of the inner ear as a source of energy to drive the transduction process. Much circumstantial evidence points to the vestibular dark cell (VDC) epithelium as being responsible for concentrating K+ within the lumen. We have used the vibrating probe technique to directly observe voltage and ion gradients produced by this tissue to put this assumption on a solid experimental footing. Relative current density (Isc,probe) over the apical membrane of VDC epithelium was measured with the vibrating voltage-sensitive probe, and this technique was validated by performing maneuvers known to either stimulate or inhibit the transepithelial equivalent short circuit current. Basolateral bumetanide (5 x 10(-5) M) and ouabain (1 x 10(-3) M) caused a decrease in Isc,probe by 55 +/- 6% and 39 +/- 3%, respectively while raising the basolateral K+ concentration from 4 to 25 mM caused an increase by 35 +/- 8%. A K+ gradient directed toward the apical membrane was detected with the vibrating K(+)-selective electrode, demonstrating that, indeed, the VDC epithelium secretes K+ under control conditions. This secretion was inhibited by bumetanide (by 94 +/- 7%) and ouabain (by 52 +/- 8%). The results substantiate the supposition that dark cells produce a K+ flux and qualitatively support the correlation between this flux and the transepithelial current.

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A vibrating probe technique for measuring the current density distribution over a corroding steel surface has been used to investigate corrosion inhibition by cerium salts. The probe vibrations enabled the current magnitude and direction in a plane perpendicular to the steel surface to be measured. Contrary to published data, it has been concluded that, at least for steels, cerium acts as an anodic inhibitor.