CYCLIC CHANGES IN THE TENSION OF THE CONTRACTILE VACUOLE COMPLEX MEMBRANE CONTROL ITS EXOCYTOTIC CYCLE

The freshwater protozoan Paramecium multimicronucleatum maintains a co nstant cytosolic osmolarity through the exocytotic activity of its con tractile vacuole complex. The contractile vacuole (CV) expels the exce ss cytosolic water, acquired osmotically, to the exterior of the cell at fairly regular intervals. In a single exocytotic cycle, the CV swel ls as the cytosolic water enters the vacuole, rounds up, and then rapi dly shrinks as the fluid is expelled through the pore of the CV. The e xocytotic activity of the CV complex can sometimes be slowed and disto rted 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 cap acitance and membrane potential of the organelle. We have clearly demo nstrated 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 ent ered the next fluid-filling phase without expelling its fluid. The rad ial arms remained severed from the CV during such prolonged rounding p hases 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 tensio n of the CV membrane. This suggested that the periodic development of increased tension in the CV membrane might be the primary event leadin g 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 fra gmented into vesicles when the cell had been mechanically ruptured in a salt solution. Many of the resulting in vitro vesicles showed period ic rounding and slackening which occurred at different times so that t hey were out of phase one with the other. This indicates that the memb ranes 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 asso ciated cytoskeletal structures) possesses a mechanism by which its ten sion is periodically increased. Such a periodic change in membrane ten sion may govern the exocytotic cycle of the contractile vacuole comple x 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 caus es closure of the pore and rejoining of the radial arms to the CV. Tra nsformation of the CV membrane into 40 nm tubules is assumed to be res ponsible for the development of tension in the planar CV membrane. The causes of this periodic tubulation must now be sought.