Electrical field effects induced in membranes of developing chloroplasts

Etioplasts, etiochloroplasts, and chloroplasts of Avena sativa L. purified on a Percoll gradient were subjected to increasing electric field strengths in the orifice of a hydrodynamically focussing Coulter Counter. The change in resistance of the orifice when an organelle is present correlates well with the size of the plastid for field strengths up to about 3.5 kV cm^-1. Beyond this field strength, depending on the size of the organelle, the size is underestimated. The underestimation of the size is caused by the dielectric breakdown of the envelope membranes once a critical membrane potential has been exceeded. Beyond breakdown the signal of the particle is predominately determined both by the internal conductivity and the increased membrane conductivity. Measurements of the breakdown voltage of different developmental stages of the plastids reveal that the breakdown voltage decreases from 1.2 V in etioplasts to about 0.9 V in chloroplasts after 48 h illumination. The decrease in breakdown voltage can be explained in terms of increasing incorporation of proteins into the inner envelope membrane during development.This view is consistent with conclusions drawn by other authors from transport and biochemical studies. The underestimation of the size beyond breakdown is about 20% and increases to a constant value of about 40% during the first 3 h of illumination. The underestimation decreases again to about 10% when the chloroplast stage is reached. This result is consistent with the current view of chloroplast development. Mobilisation of glucans, the transformation of the prolamellar body of etioplasts into thylacoid membranes as well as an intensive synthesis of pigments and enhanced rates of ions transport in the first hour of illumination gives rise to an increased pool of ionic compounds within the plastid stroma.It should be noted that purification of the plastids on Percoll gradient leads to size distributions which are almost normally distributed over the whole field range, suggesting that the preparations are also electrically homogeneous (U. Zimmermann, F. Riemann and G. Pilwat: Biochim. Biophys. Acta 436, 460–474 (1976)). In contrast with results of Lürssen, K., Z. Naturforsch. 25b, 1113–1119 (1970) only a slight increase of the modal volume from the etioplast stage to the chloroplast stage is observed.