Excised Albizzia leaflet pairs exposed to red (R) light close within 30–90 min after transfer to darkness. Interruption of darkness by far‐red (FR) light at any time after R inhibits closure within ca. 10 min. Similarly, irradiation with R at any time after prior FR promotes closure within ca. 10 min, and the increased rate of closure is independent of the time lapse between the FR and R irradiations. Closure in the dark is inhibited by NaN3 and DNP (5 X 10–4 m), by anaerobic conditions and by externally applied salts of monovalent cations, especially K; it is also temperature sensitive. Pulvinule cells are very high in K. Electron microprobe analysis of cryostated, lyophilized pulvinules reveals that during closure, K is lost from ventral cells and enters dorsal cells. FR before darkness inhibits the former but not the latter process. Thus, K flux appears to control the changes in volume of the pulvinule cells that control leaflet movement. While leaflet closure normally requires a dark period, salts of organic acids such as sodium acetate, propionate, and butyrate cause closure in the light.

Samanea leaflets usually open in white light and fold together when darkened, but also open and close with a circadian rhythm during prolonged darkness. Leaflet movement results from differential changes in the turgor and shape of motor cells on opposite sides of the pulvinus; extensor cells expand during opening and shrink during closure, while flexor cells shrink during opening and expand during closure but change shape more than size. Potassium in both open and closed pulvini is about 0.4 N. Flame photometric and electron microprobe analyses reveal that rhythmic and light-regulated postassium flux is the basis for pulvinar turgor movements. Rhythmic potassium flux during darkness in motor cells in the extensor region involves alternating predominance of inwardly directed ion pumps and leakage outward through diffusion channels, each lasting ca 12 h. White light affects the system by activating outwardly directed K+ pumps in motor cells in the flexor region.

ABSTRACILeaflet movements in Samanea saman are driven by the shrinking and swelling of cells in opposing (extensor and flexor) regions of the motor organ (pulvinus). Changes in cell volume, in turn, depend upon large changes in motor cell content of K+, Cl and other ions. We performed patch-clamp experiments on extensor and flexor protoplasts, to determine whether their plasma membranes contain channels capable of carrying the large K currents that flow during leaflet movement. Recordings in the "whole-cell" mode reveal depolarization-activated K+ currents in extensor and flexor cells that increase slowly (t½ = ca. 2 seconds) and remain active for minutes. Recordings from excised patches reveal a single channel conductance of ca. 20 picosiemens in both cell types. The magnitude of the K+ currents is adequate to account quantitatively for K+ loss, previously measured in vivo during cell shrinkage. The K+ channel blockers tetraethylammonium (5 millimolar) or quinine (1 millimolar) blocked channel opening and decreased light-and dark-promoted movements of excised leaflets. These results provide evidence for the role of potassium channels in leaflet movement.Leaflet movements in nyctinastic (night closure) plants often involve significant changes in the volume and up to severalfold variation in the ionic content of motor cells in the pulvinus (reviewed in Ref. 21). These variations may occur in response to light, darkness, and an endogenous biological clock. Cells in the extensor region of the pulvinus take up K+ and Cl-as they swell during leaflet opening, and lose both ions as they shrink during leaflet closure, while cells in the opposing (flexor) region behave in the reverse manner (12,22,23,25,30,32).We undertook this study to examine a possible role for K+ channels in leaflet movement-related K+ fluxes and changes in cell volume in the nyctinastic legume Samanea saman. K+ channels have already been described in giant algae (1, 5) and in protoplasts isolated from wheat mesophyll cells (13,14), Vicia faba guard cells (27,29) Protoplast Isolation. Terminal secondary pulvini from the fourth to ninth leaf (counting from the apex) were harvested 2 to 3 h after the beginning of the light period in the growth chamber, or 2 to 3 h after sunrise in the greenhouse. Protoplasts were prepared by enzymic digestion of slices of extensor or flexor tissue (pooled separately), as described in (7) but with the following modifications. (a) The osmotic pressure of the pre-digestion solution was raised in two steps to 680 mosmol to ensure plasmolysis. (b) Pectolyase Y-23 (Seishin Pharmaceutical, Tokyo, Japan) was added to the digestion solution (which contained cellulase, pectinase and Driselase) to a final concentration of 0.2% (w/v). (c) A second purification step was added, as follows: the cells collected from the Ficoll interface were layered again on a sucrose gradient (2), spun at 60 to IOOg for 5 min, and collected and kept on ice for up to 24 h for patch-clamp experiments.Forty to fifty protoplasts of each type, flexor and ...

Leaflets of Samanea saman open and close rhythmically, driven by an endogenous circadian clock. Light has a rapid, direct effect on the movements and also rephases the rhythm. We investigated whether light signals might be mediated by increased inositolphospholipid turnover, a mechanism for signal transduction that is widely utilized in animal systems. Samanea motor organs (pulvini) labeled with H3llinositol were irradiated briefly (5-30 sec) with white light, and membrane-localized phosphatidylinositol phosphates and their aqueous breakdown products, the inositol phosphates, were examined. After a 15-sec or longer light pulse, labeled phosphatidylinositol 4-phosphate and phosphatidylinositol 4,5-bisphosphate decreased and their labeled metabolic products inositol 1,4-bisphosphate and inositol 1,4,5-trisphosphate increased, changes characteristic of inositolphospholipid turnover. We conclude that inositolphospholipid turnover may act as a phototransduction mechanism in Samanea pulvini in a manner that is similar to that reported in animal systems.

A depolarization-activated K+ channel capable of carrying the large K+ currents that flow from shrinking cells during movements of Samanea saman leaflets has been described in the plasmalemma of Samanea motor cell protoplasts (N Moran et al [1988] Plant Physiol 88:643-648). We now characterize this channel in greater detail. It is selective for K+ over other monovalent ions, with the following order of relative permeability: K+ > Rb+ > Na+ = Cs+ Li+. It is blocked by Cs+ and by Ba2+ in a voltage dependent manner, exhibiting a 'long-pore' behavior, similarly to various types of K+ channels in animal systems. Cadmium, known for its blockage of Ca2+ channels in animal systems, and Gd3+, closely related to La3+, which also blocks Ca2+ channels in animal cells, both block K+ currents in Samanea in a voltage-independent manner, and without interfering with the kinetics of the currents. The suggested mechanism of block is either (a) by a direct interaction with the K+ channel, but external to its lumen, or, alternatively, (b) by blocking putative Ca2+ channels, and preventing the influx of Ca2 , on which the activation of the K+ channels may be dependent.that this channel plays a role in the passive efflux of K+ during cell shrinkage. The channel has the following characteristics: (a) it is voltage gated, opening upon membrane depolarization with a time constant of 1 to 2 s; (b) at moderate depolarizations, it does not inactivate for many minutes; (c) its conductance ranges from 15 to 40 pS at external K+ concentrations between 5 and 125 mm, respectively; (d) it can be blocked by TEA3 and quinine, compounds that block K'+ channels in membranes of other organisms; and (e) the channel is similar in both extensor and flexor cells.TEA and quinine also inhibited the movement of excised leaflets, thereby supporting the view that the flow of K+ current through these channels is necessary for leaflet movement. To characterize this channel in greater detail, we now extend our description of the channel's 'signature' to include its interaction with various inorganic ions: Li+, Na+, Rb+, Cs+, Ba2+, Cd2+, and Ga3+. Preliminary results appeared in abstract form ( 18).