Effects of La3+ on Surface Charges, Dielectrophoresis, and Electrofusion of Barley Protoplasts 1

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When dielectrophoresis and electrofusion of barley (Hordeum vulgare var Moor) leaf protoplasts were assayed in the presence of 0.1 to 1 millimolar lanthanum ion (La3+) in the basal medium (0.7 molar mannitol, 1 millimolar piperazine-N, N-bis[2-ethanesulfonic acid]-Na [pH 6.7], 0.1 millimolar CaCl2), dielectrophoresis and induction of electrofusion were strongly inhibited. The latter remained inhibited and the former recovered by about 60% after washing the La3+ -treated protoplasts without EDTA. These inhibitions were almost completely abolished by washing the La3+ -treated protoplasts with 1 millimolar EDTA. Inductively coupled plasma atomic emission spectroscopic analysis revealed that protoplasts retained a considerable amount of La3+ after washing without EDTA and released most of the bound La3+ by washing with 1 millimolar EDTA. This tightly bound La3+ seemed responsible for the inhibition of electrofusion and dielectrophoresis that was observed in the La3+ -treated protoplasts after washing. ζ-potentials of protoplasts were -39.0±3.2 millivolts, -16.7 ± 2.6 millivolts, and virtually zero in media containing 0, 0.1, and 0.3 millimolar La3+ (I = 7.2 millimolar), respectively, and had a positive value (+ 14.2 ± 2.2 millivolts) in the presence of 1 millimolar La3+. These effects of La3+ on ζ-potentials were easily abolished by washing without EDTA. This indicates that charged species located at the surface of plasma membrane of protoplasts cannot account for the sites at which La3+ exerts its inhibition of dielectrophoresis and electrofusion. In contrast, the promotion of spherical fusion and the reduction of broken fusion products observed in the presence of La3+ were almost completely abolished by washing without EDTA. Our results also indicate that the initial induction and development of electrofusion can be studied independently.

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