Microscopic elements of electrical excitation in Chara: transient activity of Cl- channels in the plasma membrane

The plasma membrane of Chara corallina was made accessible for patch pipettes by cutting a small window through the cell wall of plasmolyzed internodal cells. With pipettes containing Cl- as Ca2+ or Ba2+ (50 or 100 mM), but not as Mg2+ or K+ salt, it was possible to record in the cell-attached mode for long periods with little channel activity, randomly interspersed with intervals of transient activation of two Cl- channel types (cord conductance at +50 mV: 52 and 16 pS, respectively). During these periods of transient channel activity, variable numbers (up to some 10) of the two Cl- channel types activated and again inactivated over several 100 msec in a coordinated fashion. Transient Cl- channel activity was favored by voltages positive of the free running membrane voltage (> -45 mV); but positive voltage alone was neither a sufficient nor a necessary condition for activation of these channels. Neither type of Cl- channel was markedly voltage dependent. A third, nonselective 4 pS channel is a candidate for Ca2+ translocation. The activity of this channel does not correlate in time with the transient activity of the Cl- channels. The entire set of results is consistent with the following microscopic mechanism of action potentials in Chara, concerning the role of Ca2+ and Cl- for triggering and time course: Ca2+ uptake does not activate Cl- channels directly but first supplies a membrane-associated population of Ca2+ storage sites. Depolarization enhances discharge of Ca2+ from these elements (none or few under the patch pipette) resulting in a local and transient increase of free Ca2+ concentration ([Ca2+]cyt) at the inner side of the membrane before being scavenged by the cytoplasmic Ca2+ buffer system. In turn, the transient rise in [Ca2+]cyt causes the transient activity of those Cl- channels, which are more likely to open at an elevated Ca2+ concentration.