Cyclic changes in the tension of the contractile vacuole complex membrane control its exocytotic cycle

The freshwater protozoan Paramecium multimicronucleatum maintains a constant cytosolic osmolarity through the exocytotic activity of its contractile vacuole complex. The contractile vacuole (CV) expels the excess cytosolic water, acquired osmotically, to the exterior of the cell at fairly regular intervals. In a single exocytotic cycle, the CV swells as the cytosolic water enters the vacuole, rounds up, and then rapidly shrinks as the fluid is expelled through the pore of the CV. The exocytotic activity of the CV complex can sometimes be slowed and distorted by microelectrode impalement. Using a CV with exocytotic activity slowed in this way makes it possible to determine more precisely the time course of changes in the exocytotic-activity-related membrane capacitance and membrane potential of the organelle. We have clearly demonstrated that the radial arms of the CV were always severed after the CV had exhibited rounding. Microelectrode impalement sometimes caused a failure of the CV pore to open after rounding up, so that the CV entered the next fluid-filling phase without expelling its fluid. The radial arms remained severed from the CV during such prolonged rounding phases and then rejoined the CV at the start of the next fluid-filling phase. The rounding of the CV corresponds to an increase in the tension of the CV membrane. This suggested that the periodic development of increased tension in the CV membrane might be the primary event leading to periodic severing of the radial arms and the opening of the pore. We then observed that the CV and its radial arms sometimes became fragmented into vesicles when the cell had been mechanically ruptured in a salt solution. Many of the resulting in vitro vesicles showed periodic rounding and slackening which occurred at different times so that they were out of phase one with the other. This indicates that the membranes of the CV and the radial arms maintain their ability to develop a periodic increase in tension even after the cell has been ruptured. We propose the hypothesis that the CV membrane (together with its associated cytoskeletal structures) possesses a mechanism by which its tension is periodically increased. Such a periodic change in membrane tension may govern the exocytotic cycle of the contractile vacuole complex by increasing the tension to a point that the radial arms sever from the CV and the pore opens. Conversely, a decrease in the tension causes closure of the pore and rejoining of the radial arms to the CV. Transformation of the CV membrane into 40 nm tubules is assumed to be responsible for the development of tension in the planar CV membrane. The causes of this periodic tubulation must now be sought.