Reversible Changes in the Hydraulic Permeability of Plant Cell Membranes

169

In view of the possibility that senescence may be a consequence of the deterioration of membrane compartments in the cells of leaves, calcium was studied as a possible agent which might defer senescence. The senescence of corn leaf discs was deferred by added calcium, and the effect was additive to the cytokinin deferral of senescence. Likewise, the senescence of Rumex leaf discs was deferred by added calcium, and the effect was additive to the gibberellin deferral of senescence.Detailed experiments with corn leaf discs established that the increase in apparent free space associated with senescence was completely prevented by calcium. An increase in hydraulic permeability during senescence was likewise demonstrated, and this increase was deferred by calcium; calcium plus benzyladenine was even more effective. Each of the measured functions of leaf senescence (chlorophyll content, protein decrease, apparent free space increase, and hydraulic permeability increase) was suppressed by calcium, and the interpretation is offered that the effects are a consequence of the calcium function in maintaining cellular membranes.Since the discovery by Richmond and Lang (17) growth chamber (2000 ft-c, 16 hr daily) at a temperature close to 22 C. At 8 to 10 days, leaf discs measuring 1 cm in size were taken from the primary leaf with the aid of a cork borer. The discs were floated in 10 ml of test solution, 10 discs per Petri dish in darkness at 25 C for 3 to 5 days.For the Rumex assay (Rumex obtusifolius L.), following the procedure of Whyte and Luckwill (27), old leaves which were uniformly green were selected from matured plants in the greenhouse, and they were allowed to stand overnight at 25 C in the dark with petiole in water. Leaf discs (1 cm) were cut with a cork borer and randomized, and 10 discs were floated in each Petri dish containing 10 ml of solution to be tested in the darkroom at 25 C for 4 or 5 days. Chlorophyll was extracted in ethanol. Ten leaf discs were transferred to 10 ml of ethanol in a closed vial and allowed to stand overnight, and absorbance was read at 665 nm. Experiments were repeated 4 to 6 times.For protein measurement 20 leaf discs were ground in a mortar and pestle with phosphate buffer at pH 6.5 and centrifuged at 800g to discard cellular debris. Protein was precipitated from the supernatant with 10% trichloroacetic acid and centrifuged at 10,000g for 10 min. The precipitate was dissolved in 5 ml of 0.1 N NaOH, and a 0.1-ml aliquot was used for protein analysis by the method of Lowry et al. (12).Apparent free space was determined by the method of Epstein (5). Twelve leaf discs were blotted on a paper towel to remove solution adhering to the surface. To initiate the absorption, the leaf discs were transferred to a salt solution (1 mm KCl) containing a radioactively labeled ion . At the end of the absorption period (90 min), the solution containing the radioactively labeled ion was decanted, and the leaf discs were blotted on a paper towel. Some leaf discs were used to determine the ini...

Section: Discussionmentioning

confidence: 95%

Deferral of Leaf Senescence with Calcium

Poovaiah¹,

Leopold²

1973

169

“…2) show only a slightly steeper slope for entry into living than into heat-killed tissue, which is consistent with the concept that the passage of water through tissues is only slightly hindered by membranes (cf. 3,15). Temperatures which cause chilling injury (16 C and below) do not bring about dramatic changes in the Arrhenius plot of water entry.…”

Section: Resultsmentioning

confidence: 99%

Temperature Effects on Soybean Imbibition and Leakage

Leopold¹

1980

As a part of an analysis of the nature of chilling injury to seeds, measurenents were made of the nitial linear rates of water entry into and solute leakage out of cotyledons of soybean at various temperatures.Arrhenlus plots were approximatel linear for water entry Into both lving and dead cotyedons, with the slpe (and activation energy) for entry Into iving celis being insipificantly higher than for dead cells, suggesting lttle effect of membrane barriers on water entry. The plots for solute leakage showed 10-fold lower leakage rates from lving than from dead tissues; a reversal of slo in the Arrhenlus plot at temperatures below 15 C reflected increasing leakage rates, interpreted as a quantitative disruptio of membrane reorganization at the temperatures associated with chilng injury.peratures between 1 and 40 C were maintained by immersing the imbibing system in a water bath.The time course of solute leakage was measured as the rise in conductance of 25 ml of water containing 10 cotyledons. Conductance was measured with a Markson ElectroMark analyzer at 5-min intervals.Rates were determined from the slope of the 10-to 40-mm periods, using calculated regression lines. The logarithms of these slope values were plotted as Arrhenius plots, and activation energies were then calculated using the equation of Stein (13): 2.3 log k= ( I where Ad is the activation energy, k' and k" are log rates for 40 and 5 C (T' and T", respectively), and R is the gas constant. RESULTSEvidence has been accumulating that chilling injury to seeds during imbibition might be attributed to disruptive effects on the reorganization of membranes as water enters the dry seed (1, 9). A quantitative means of analyzing the barrier effectiveness of membranes could utilize the initial linear rates of water entry and of solute leakage from soybean cotyledons, and their analysis through Arrhenius plots.The time course of initial water entry into dry cotyledons shows a period of rapid, nonlinear entry lasting 5-10 min, followed by a period of linear water entry lasting 30 min or more (7,11). The initial time course of solute leakage from the imbibing cotyledon also shows a period of rapid, nonlinear leakage, followed by a linear phase beginning at about the same time that water entry becomes linear. It has been suggested that the initial rapid phases of each of these events represented a period during which the membranes were relatively disorganized, and the linear phases represented steady-state processes through the reorganized membranes (7,8,11). If this interpretation is correct, linear rates of water entry and of solute leakage should reflect the effectiveness of membranes in resisting the passage of water and of solutes, respectively.The experiments reported here measure the linear rates of water entry and solute leakage of cotyledons at various temperatures to permit construction of Arrhenius plots and estimation of the energy barriers provided by the reconstituted membranes. MATERIALS AND METHODSWater entry and solute leakage were mea...

“…Changes in leakage and in photosynthesis (Figs. 7,8) are apparent at similar degrees of dehydration. Since structural integrity of the chloroplast membranes is required for efficient photophosphorylation, the reduction in photosynthesis may be associated with changes in permeability.…”

Section: ) Amentioning

confidence: 59%

“…3). Furthermore, influx is measured from distilled water, thus meeting two of the criteria which were required to demonstrate the role of Lp in determining the flow of water (8) (Table III). (e) Changes in temperature are followed by rapid transitions of the rehydration rates (Fig.…”

Section: ) Amentioning

confidence: 99%

See 1 more Smart Citation

Dehydration, Water Fluxes, and Permeability of Tobacco Leaf Tissue

Graziani¹,

Livné²

1971

Removal of the lower epidermis from a tobacco leaf allows a faster and wider range of water fluxes, without damaging the mesophyll. It also permits a more direct examination of the photosynthetic potential of the tissue at various levels of hydration.The rehydration rate of leaf discs is essentially linear. It decreases with leaf age and is correlated with the rate of dehydration, but it is independent of the tissue's water potential, as estimated by the isopiestic method. The hydraulic permeability coefficient of water influx is directly related to water potential of the tissue, suggesting a mechanism for the regulaton of the hydration level of the leaf tissue. The "energy of activation" of rehydration amounts to about 9 kilocalories per mole at intermediate dehydration, but it greatly declines following water loss in excess of 600 milligrams per gram fresh weight. The excessive dehydration is also characterized by a major increase in permeability (monitored by efflux of ions and materials absorbing ultraviolet light) and by a parallel decrease in photosynthetic activity. The interrelationship of these effects of excessive dehydration is discussed.The effects of dehydration of plants (see reviews: 4, 7, 12, 15, 18, 31, 32) Dehydration and Rehydration. For dehydration the discs with the stripped side up were exposed to an atmosphere of 45 to 60% relative humidity under a light intensity of 2500 lux (Sylvania Gro-Lux lamps) at a room temperature of 24 to 27 C. The tissue was weighed on an analytical balance at frequent intervals (Fig. 1). The rate of dehydration was computed on the basis of the time required for the tissue to lose a known weight of water (commonly 350 mg per g fresh wt).When the desired degree of dehydration had been attained, rehydration was started by floating the discs, with the lower side down, on distilled water at 22 C in the light. Where indicated, other temperatures were employed. At 2-to 3-min intervals, the discs were taken out, blotted once with a paper tissue, and weighed. Additional blotting had a negligible effect on the weight of the discs. Comparative measurements showed that the interruptions for the weighing (up to 8 times) did not alter the rate or the extent of rehydration, which were dependent on the net time of exposure of the tissue to water.The rate of rehydration was derived from the initial rate of water uptake. For example, in Figure 2 the rate of rehydration was 37 mg per g fresh wt per min, based on the linear portion of the curve (up to 6 min).Water Potential. The water potential of the leaf discs was found by estimation of the water potential of a sucrose solution (29) in which the weight of the tissue did not change. The validity of this isopiestic technique has been questioned (1,9) because of the possibility of diffusion either of sucrose or of its hydrolytic products into the tissue. To minimize such uncertainties, the discs were floated on graded solutions (0-0.6 M sucrose) at 22 C, and the linear changes in weight of the tissue within 4 min were use...