Cell age‐related differences in the interaction of a potential‐sensitive fluorescent dye with nuclear envelopes of Acetabularia mediterranea

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Abstract
To study whether an electrical potential difference exists across the nuclear envelope or inner nuclear membrane of plant cells, the authors have used an optical probe of membrane potential, the cationic fluorescent dye, DiOC6(3) (MW = 572.5). This dye was microinjected into the nucleoplasm of isolated Acetabularia nuclei (which are still surrounded by a thin layer of cytoplasm) and its subnuclear localization visualized by fluorescence microscopy. Striking differences, which seemed to be correlated with the developmental stage of the isolated nucleus, were observed. In nuclei isolated from cells at the stage of early cap stage formation, the dye was restricted to the nuclear envelope. In nuclei isolated from cells with intermediate or fully developed caps, there was increased nucleoplasmic staining, and the staining of the envelope was frequently diminished or abolished. In all nuclei, the dye remained within the nucleus after injection. Cytoplasmic staining was only observed when nuclei isolated from cells at the stage of early cap formation were incubated in a hyper‐ or hypo‐tonic medium. Various ionophores, injected before the dye into the nucleoplasm, had no effect on the subsequent nuclear localization of DiOC6(3), although they did rapidly induce nucleolar condensation in nuclei isolated from cells at the stage of early cap formation. The results suggested that the electrical properties of Acetabularia nuclear envelopes or inner nuclear membranes change during cell maturation. Furthermore, the retention of the dye in the nucleoplasm under isotonic conditions indicated that the nuclear pores were not open channels for molecules of this size.