Dynamics of membranous organelles in the mitotic apparatus of two cell types: A live-cell, laser-scanning confocal study

Abstract

Membranous organelles comprise a large proportion of the living cell's cytoplasmic volume. In interphase, the endoplasmic reticulum (ER), mitochondria, and Golgi apparatus are characterized by a typical distribution in the cytoplasm. When cells enter M-phase, however, the cytoplasmic architecture undergoes dramatic rearrangements, as the nuclear envelope (NE) breaks down and the mitotic spindle assembles. During this process, the typical distribution of the membrane systems must be disrupted, so as to accommodate the cytoplasmic rearrangements. The role of membranous organelles within and around the mitotic apparatus has been a source of active investigation, as well as controversy. It is not known if membranes are an ubiquitous structural element of the spindle. Here, we present a laser-scanning confocal study of membrane dynamics in living mitotic cells' demonstrated by the fluorescent vital membrane dye 3,3″-dihexyloacarbocyanine iodide (DiOC6 (3)). By exploiting the thin focal planes achieved by the confocal microscope, the precise distribution of membranes within the spindle was attained in the two epithelial cell lines; PtK2 and LLC-PK. This study demonstrates that the distribution and quantity of cytoplasmic organelles within the spindle of living cells is cell-type specific.