Journal of Experimental Botany, Vol. 52, No. 359, pp. 1303-1313,
June 1, 2001
© 2001 Oxford University Press
Original Papers |
Stomatal oscillations at small apertures: indications for a fundamental insufficiency of stomatal feedback-control inherent in the stomatal turgor mechanism
Botanisches Institut der Christian-Albrechts-Universität zu Kiel, Olshausenstraße 40, D-24098 Kiel, Germany
Received 5 October 2000; Accepted 6 February 2001
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Abstract |
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Continuous measurements of stomatal aperture simultaneously with gas exchange during periods of stomatal oscillations are reported for the first time. Measurements were performed in the field on attached leaves of undisturbed Sambucus nigra L. plants which were subjected to step-wise increases of PPFD. Oscillations only occurred when stomatal apertures were small under high water vapour mole fraction difference between leaf and atmosphere (
W). They consisted of periodically repeated opening movements transiently leading to very small apertures. Measurements of the area of the stomatal complex in parallel to the determination of aperture were used to record volume changes of guard cells even if stomata were closed. Stomatal opening upon a light stimulus required an antecedent guard cell swelling before a slit occurred. After opening of the slit the guard cells again began to shrink which, with some delay, led to complete closure. Opening and closing were rhythmically repeated. The time-lag until initial opening was different for each individual stoma. This led to counteracting movements of closely adjacent stomata. The tendency to oscillate at small apertures is interpreted as being a failure of smoothly damped feedback regulation at the point of stomatal opening: Volume changes are ineffective for transpiration if stomata are still closed; however, at the point of initial opening transpiration rate rises steeply. This discontinuity together with the rather long time constants inherent in the stomatal turgor mechanism makes oscillatory overshooting responses likely if at high W the ‘nominal value’ of gas exchange demands a small aperture. Key words: Stomatal oscillations, stomatal aperture, feedback control, turgor mechanism.
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Introduction |
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The stomatal pore width of an illuminated leaf is regulated to satisfy the conflicting needs of maintaining a sufficient intercellular CO2 concentration (Ci) for photosynthesis on the one hand and of preventing excessive water loss by transpiration (E) on the other hand. Apart from direct responses, for example, to PPFD, negative feedback loops serve to maintain stomatal conductance for water vapour and CO2 in an appropriate range (Raschke, 1965
): A reduction of Ci due to photosynthetic CO2 assimilation stimulates stomatal opening (Mott, 1988), which increases CO2 diffusion into the leaf intercellular spaces. Stomata respond to changes in air humidity in a manner consistent with a feedback-regulation where an increase of transpiration acts as a negative stimulus (Franks et al., 1997; Monteith, 1995; Mott and Parkhurst, 1991). Despite the fundamental importance of these processes the physiological sensing and transduction involved in both feedback systems is barely understood (Cousson, 2000; Kearns and Assmann, 1993).
The control by negative feedback is most apparent when oscillations of stomatal aperture occur (Farquhar and Cowan, 1974; Raschke, 1979). These oscillations have fascinated many researchers, giving rise to numerous reports of oscillations observed by different methods (Barrs, 1971) with different plant species. Most often gas exchange techniques were used, but measurements of chlorophyll a fluorescence (Eckstein et al., 1996; Siebke and Weis, 1995), leaf temperature, leaf water potential or other methods concerning water relations (Herppich and von Willert, 1995; Lang et al., 1969; McBurney and Costigan, 1984; Naidoo and von Willert, 1994) were also used to record stomatal oscillations.
These methods integrate the responses of large numbers of stomata and therefore can convey only limited information on the actual responses at the single guard cell level. The synchronism or variability of the responses of individual stomata remains obscure. It is not possible to draw firm conclusions on the amplitude of the underlying stomatal movements unless the relationship between aperture and gS is known. This relationship has not been determined by parallel measurements of gas exchange and apertures in the majority of gas exchange studies. Even the general physical relation between aperture and stomatal conductance is still in doubt. Although it is commonly believed and supported by theoretical models (Lushnikov et al., 1994; Parlange and Waggoner, 1970), that gS is largely linearly related to aperture, recent measurements demonstrated a non-linear relationship, with the steepest increase of gS with aperture at small degrees of opening (Kaiser, 1999; Kaiser and Kappen, 2000).
The idea of this work was therefore to observe stomatal oscillations at the aperture level by a combination of microscopic observation and CO2/H2O gas exchange measurements. Measurements were performed on intact leaves of an undisturbed Sambucus nigra L. shrub on a field site. The conditions for the occurrence of oscillatory behaviour were explored and the responses described in detail. The results support an hypothesis which explains oscillations as the consequence of insufficient feedback-control inherent in the stomatal turgor mechanism and stomatal mechanics.
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Experimental site and plant material
Experiments were made on leaves of an approximately 5 m high shrub of Sambucus nigra growing on the border of a small stand of shrubs and young trees in the New Botanical Gardens in Kiel (FRG). The site was irrigated during dry periods whenever a tensiometer monitored soil water potentials less than -0.5 MPa. The experiments presented here were performed at soil water potentials between -0.04 and -0.46 MPa. S. nigra continuously developed new leaves throughout the summer; thus by using only young but fully expanded leaves, interference from senescence effects could be avoided. Leaves were enclosed in the gas-exchange cuvette for about 1 week, during which several experiments on the same set of stomata were performed. Except during the experimental periods between 09.00 h and 15.00 h the cuvette received ambient light and tracked the external humidity and temperature conditions.
Gas exchange and aperture measurements
A combination of microscopic in situ observations and CO2/H2O gas exchange measurements on the same leaf was used. The technical details have been described previously (Kaiser and Kappen, 1997, 2000). In short, it consists of an inverted video microscope (long distance objective 40x, Zeiss, FRG) inserted in the bottom of a gas-exchange cuvette. A leaf is attached above the microscopic objective in a leaf holder, which is driven by a motorized remote controlled microscopic stage. By computer control it was possible repeatedly to relocate selected stomata and to capture time series of digitized video-images for image analysis. Observation of stomata at low light or in darkness were enabled by the transmitted light of a GaAlAs emitter infrared diode (model OD880F, Optek, US). The images of the stomata were taken with the microscope focused to the narrowest part lower down in the pore. Measurements of the pore area on digitized images were made manually by delineating the pore edge with the cursor. Measurement errors as determined by randomly repeated measurements of a set of images ranged between 0.5 and 3 µm2 (±standard deviation), depending on the image quality. The relative error is larger at small apertures. Apertures of very slightly opened stomata cannot be exactly determined. A ‘zero’ reading therefore cannot completely rule out the presence of a small stomatal opening, which nevertheless can significantly contribute to gas exchange (Kerstiens, 1996). Although uncommon in the literature, stomatal opening has been expressed in terms of pore area instead of pore width (aperture). This does not affect the interpretation of the results, because area and width of the elliptic pore are linearly related if the pore length stays constant during movements. This was confirmed for S. nigra. Apertures of stomata of different longitudinal extension were compared by a relative measure, the ‘degree of opening’, which expresses pore width as a percentage of pore length.
In some experiments the stomatal complex area (SCA) between the dorsal cell walls was also measured.
Either 15 or 50 stomata were observed in each experiment, usually randomly sampled from an area of about 2 cm2 in the centre of the leaf. The entire leaf surface could not be inspected as the petiole was not flexible enough and was then bent by the movements of the leaf holder.
The gas exchange equipment (Walz, FRG) is an open flow system which measures transpiration every minute by means of the bypass principle and CO2 exchange every 2 min with an infrared gas analyser. Humidity and temperature in the cuvette were regulated to a constant value during the experiments.
Leaf temperature was measured by a 0.2 mm thermocouple attached to the lower leaf surface. The internal fan of the cuvette was set to a moderate speed, producing a boundary layer conductance of about 600 mmol m-2 s-1 which was measured with a water-saturated filter paper sealed on one side. Gas exchange calculations were performed according to Ball (Ball, 1987). The relatively large cuvette volume (c. 4500 cm3) necessary to enclose the mechanical components for microscopic inspection caused some cuvette lag. The calculations described previously (Küppers et al., 1993) were used to estimate the effective cuvette volume and to calculate a time-corrected signal of CO2 gas exchange resulting in a temporal resolution of 2 min.
Microclimatic measurements
To measure the PPFD incident on the observed leaf, a small GaAsP Photodiode (model G2711–01, Hamamatsu, Japan, calibrated against a Li-Cor quantum sensor), was mounted inside the cuvette about 12 mm distant from the observed leaf region. Soil water potential in the root zone of S. nigra was measured at a depth of 40 cm by a pressure transducer tensiometer (Kappen et al., 2000). Microclimate and gas exchange data were recorded by a datalogger (model 21 XL, Campbell, Shepshed, UK) which was supplemented by a multiplexer (model AM 416, Campbell, UK).
Experiments
Stomatal and gas exchange responses to step-wise increases in PPFD at different levels of W were observed. Irradiance was provided by a fibre optic illuminator (Kaltlicht-Fiberleuchte FL-400, with Spezial Fiberoptik 400-F, Walz, Effeltrich, FRG) or five halogen cold-light lamps (50 W) emitting a PPFD of up to 750 µmol m-2 s-1 through a diffuser. During the experiments ambient light was nearly completely kept off by covering the cuvette with a black cloth.
Either ‘full’ steps from darkness to about 750 µmol m-2 s-1 were applied, or PPFD was increased step by step (e.g. from 0 to 60 to 130, 270, 510, 750 µmol m-2 s-1). PPFD was always held constant until a leaf conductance was at steady state or, in case of oscillations, at least for 1 h. In the ‘full step’ experiments a set of 15 stomata was observed every 4–10 min. In the step by step experiments 50 stomata were measured only once at the end of each PPFD step. During subsequent days the leaves were subjected to the identical light treatment under different levels of W and always the same set of stomata was observed. Experiments were performed always at the same time of the day, to avoid interference with diurnal rhythms.
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Results |
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Oscillations were visible in both gas-exchange responses and movements of individual stomata. They could be easily elicited by step-wise increases of PPFD at
W higher than about 10 mmol mol-1. The tendency to oscillate was quite variable. It ranged between damped oscillations with one single overshooting response and sustained oscillations over a time period of more than 2 h (Figs 1, 2, 4, 7).
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The conditions for the different oscillation patterns were explored in a series of experiments at different levels of
W. Oscillations were elicited either by increase of PPFD from zero to 850 µmol m-2 s-1 in one single step (Figs 1
, 4, 7) or by step-wise increases (Fig. 2). Oscillations were generally more likely to occur at higher W (Fig. 2). This figure also shows that the initial opening as well as the closing reaction is faster the drier the air is. Another determining factor for oscillations appeared in the experiments with step-wise increases of PPFD. The tendency to oscillate was maintained as long as the average degree of opening was quite low and was diminished when stomata opened further (Fig. 2). This suggested a restriction of oscillations to small apertures, which was further explored by summing up all PPFD-step experiments (Fig. 3). This demonstrates that pronounced oscillatory responses were restricted to both, W above 10 mmol mol-1 and small apertures below 3–4% average degree of opening. This also entails that a large proportion (about 50%) of stomata was closed at any time. One condition alone was not sufficient to produce oscillations: If stomata were opened wider than 3–4% (due to, for example, high PPFD), light changes did not elicit oscillations even if W was 15 mmol mol-1. Damped oscillations with only one single peak were observed in a larger range of apertures and W.
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The detailed observation of aperture changes allows an analysis of the processes during stomatal oscillations (Figs 4, 5, 7). Figure 4 shows the responses of leaf conductance and net photosynthetic rate together with the responses of 15 stomata. The period of gas-exchange oscillations amounted to about 40 min. After PPFD was increased the individual stomata needed between 10–40 min to open. The aperture was then oscillating between a slight opening (2–5%) and complete closure. In absolute terms the aperture changes were rather slight. This is illustrated in Fig. 5
which shows the image of the maximum observed aperture in the experiment of Fig. 4. It was remarkable that some stomata reacted phase shifted to the overall course of leaf conductance. Phase shift even occured between stomata which were only a few mm apart. Stomata furthermore displayed individually different frequencies of the oscillations. A more vivid demonstration of the spatio-temporal action of the responses displayed in Fig. 4 is given in a video-clip which is available at JXB online as supplementary material.
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To record stomatal responses (volume changes) in the closed state the area of the stomatal complex between the dorsal cell walls of the guard cells was measured. As this measurement, as far as is known, has not previously been used to record guard cell activity, the close relationship to stomatal aperture was first confirmed by simultaneously measuring pore area and guard cell complex area (SCA) of individual stomata over a period of several days under different conditions (Fig. 6
). The linear relationship to pore area confirms that SCA is a suitable indirect measurement for guard cell swelling. There is, however, a certain measurement error associated with both area measurements, which introduces some scatter. Interestingly SCA in the closed state was observed to fall to about 10% below the threshold SCA at the point of stomatal opening, giving an idea of the extent of the ‘Spannungsphase’.
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Using simultaneous measurements of SCA and pore area, the events during stomatal oscillations were observed in detail (Fig. 7
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After the increase of PPFD the guard cells began to swell. Swelling proceeded for some time without the appearance of a pore. This introduced a time lag individually different for each stoma before initial pore opening. As soon as the pore was slightly open the swelling stopped and the guard cells deflated again, which in turn led to complete closure. Deflation thereafter proceeded for some time until a new swelling of the guard cells began. These processes were repeated periodically by some stomata (e.g. stoma C in Fig. 7). This experiment also revealed a phase shift between closely adjacent stomata (2–4 mm). This is apparently brought forth by the individually different lag time (e.g. 1 h in stoma A, 0.5 h in stoma B) before initial opening.
As the conditions at small apertures were obviously decisive for the development of oscillations, the relationship between pore area and leaf conductance was explored by simultaneously measuring gas exchange and apertures of 50 randomly selected stomata during opening responses after transfer to saturating PPFD (Fig. 8). This brought forth a typical saturation curve with a steeply rising slope at small apertures which can be fitted quite well by a hyperbolic function. As data were collected over three consecutive days it can also be concluded that this relationship is repeatable with respect to time. The dependence of Amax under saturating PPFD on pore area displayed an even steeper increasing slope at small apertures, which demonstrates that in S. nigra even small apertures are sufficient to allow non-limiting CO2-supply of the mesophyll.
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The controlling effect of aperture changes on transpiration can be described best by the slope of the dependence of E on pore area (
E/A) which is demonstrated in Fig. 9 using the experimentally determined hyperbolic function (Fig. 8). It is obvious that during continual changes of guard cell volume the effect on transpiration is most pronounced at high W at the state of transition from closed to slightly open.
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Discussion |
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These observations of individual stomata add important information not already contained in the numerous existing observations of oscillations based on methods which integrate responses of several stomata. The observation that in S. nigra stomatal oscillation developed only at very small pore widths and that the aperture changes in absolute terms were quite small, points to an important role of the low aperture range near stomatal closure. Alternative explanations for the lack of oscillations at higher apertures could be ruled out: in the experiments with the step-wise increases of PPFD the decreasing tendency to oscillate could also have been explained by an inhibiting action of high PPFD (Fig. 2
). However, this seems unlikely, as in other experiments stomata oscillated at high PPFD if other conditions (e.g. high W) led to a very small aperture (Figs 4, 7). A decreasing tendency to oscillate in the afternoon due to the well-known endogenous rhythmicity of stomata is also unlikely as stomata showed no damping of oscillations in experiments extending into the afternoon hours (Fig. 4).
The general validity for other species of the assertion that small apertures are an essential precondition for oscillatory responses is uncertain. The experimental treatments leading to oscillations as, for example, the increase of W, blocking of water supply, increase of PPFD at high W, changing CO2 concentration of the air, treatment with abscisic acid (Cardon et al., 1994; Eckstein et al., 1996; Naidoo and von Willert, 1994; Raschke, 1965; Siebke and Weis, 1995) all tend to induce stomatal closure. It thus appears that the formerly observed stomatal oscillations mostly occurred in the lower aperture range. There are, however, some reports demonstrating oscillations at higher apertures with larger amplitudes (Barrs and Klepper, 1968; Bunce, 1987). However, even if oscillations may occur at higher degrees of opening in other species, the observed tendency to oscillate at the threshold of stomatal opening in S. nigra suggests a phenomenon of general importance.
The reasons for oscillations are clarified by the observation that many of the observed stomata cycled between the slightly open and the closed state. The instability of the reaction is obviously caused by the fact that here small movements have a large effect on gas exchange, making it likely that feedback responses surpass the narrow appropriate aperture range leading either to complete closure or to a too large aperture.
This interpretation is supported by measurements of guard cell activity in the periods when the pores were closed. This was implemented by measurements of the area of the stomatal complex (SCA). Although similar measures such as ‘peristomatal groove distance’ (Eckstein, 1997; Lawson et al., 1998) and ‘width of stomatal complex’ (Stålfelt, 1929b, 1963) have already been used as a surrogate measure for stomatal aperture, SCA has hitherto not been used to measure guard cell movements continuously. Therefore, it had to be confirmed first that a close and reproducible relationship between SCA and pore area exists (Fig. 6), and that continuous changes of SCA could be measured in the closed state, as can be seen in Fig. 7. These measurements illustrate the two phases of the stomatal responses first described by Stålfelt as the ‘Spannungsphase’ with a guard cell turgor not sufficient to open the pore and as the ‘Motorphase’, the status when the opened pore changes its aperture according to the turgor changes (Stålfelt, 1929a).
The continuous observation of SCA together with pore area shows that the transition between swelling and deflation phases is closely linked to the moments of opening and closing of the pore: light-induced swelling continues until the pore opens, which after some delay is followed by deflation. This deflation continues until the pore is closed, after which it again swells. This behaviour can be explained by the action of negative feedback loops at the level of single guard cell complexes: the opening of the pore(s) increases local intercellular CO2 concentration (Ci) and transpiration which act as closing stimuli. Due to the lag time of stomatal responses, which is inherent in the relatively slow turgor mechanism, these opening and closing responses periodically shoot beyond the appropriate ‘nominal value’ of stomatal aperture. It cannot definitely be concluded from these experiments to what extent the two feedback loops are involved in the oscillations. It appears, however, that the feedback related to transpiration plays a major role, because in some experiments stomata oscillated at low PPFD although Ci was relatively high and nearly constant (H Kaiser, unpublished data).
In addition, there exists an hydraulic effect, which increases the tendency of stomata to oscillate: any increase of transpiration, for example by an increase of W, lowers epidermis turgor more than guard cell turgor (Shackel and Brinckmann, 1984) and thus supports stomatal opening (Raschke, 1970). The following increase of transpiration in turn facilitates further opening by a positive feedback loop. The reverse effect acts when stomata close. This physical effect acts quite fast and precedes the slower, reversely directed physiological feedback response (Kappen et al., 1987). The strength of this hydraulic effect is correlated to W and causes the different opening and closing rates at different air humidities, which have previously been observed in different species (Assmann and Grantz, 1990; Barradas et al., 1994; Kaiser and Kappen, 2000) and also appear in the current results with S. nigra (Fig. 1). This component of positive feedback should be strongest in the lowest aperture range, where aperture changes have the largest effect on transpiration. For stomata at the transition from ‘Spannungsphase’ to ‘Motorphase’ this means that the opening movement is accelerated and an overshooting response becomes likely. Conversely, shrinking of the guard cells is accelerated and may proceed over the point of complete closure, because the decline of transpiration boosts epidermis turgor more than guard cell turgor.
The reasons for the promotion of oscillations by high W can now be expressed more precisely: On the one hand high W leads to a higher controlling impact of aperture on transpiration (Farquhar and Cowan, 1974), on the other hand the aperture which is required to arrive at the ‘nominal value of gas-exchange’ is shifted to very low pore areas, where volume changes have the largest impact on gas exchange (Fig. 9) and closing movements may even completely close the pore. This causes feedback responses to surpass the narrow appropriate aperture range. The situation becomes worse by the component of positive feedback, brought forth by the previously mentioned hydraulic effect, which is also strongest at small apertures.
These unfavourable characteristics of regulation are inseparably linked to the physiological and physical mechanisms of stomatal functioning and, therefore, represent a basically inavoidable insufficiency of the stomatal feedback regulation.
The counteracting movements even of closely adjacent stomata point to the action of a spatially limited feedback system within a distance of less than 2 mm. It could even be possible that individual stomata act as autonomous feedback systems, independent of the gas exchange through the adjacent stomata. But to confirm this, further experiments are required. The action of small-scale responses to humidity changes has been described previously (Lange et al., 1971). This does not exclude possible feedback mechanisms acting on larger scales. Co-ordinated oscillations of stomata have been observed by chlorophyll a fluorescence imaging on the level of ‘stomatal patches’ (Cardon et al., 1994; Siebke and Weis, 1995). Even oscillations at the whole plant level were observed (McBurney and Costigan, 1984; Naidoo and von Willert, 1994). These were apparently co-ordinated by hydraulic signals from the xylem which are believed to act mainly in woody plants and as a signal in the feedback regulation of whole plant water status (Fuchs and Livingston, 1996; Saliendra et al., 1995; Whitehead et al., 1996). A very sensitive stomatal response to high W reduces the risk of xylem embolism in stenohydric woody species (Vogt, 1998; Vogt and Lösch, 1999) in limiting whole plant transpiratory water loss. Obviously S. nigra is of the type of plant that makes use of this feedback mechanism.
These observations, however, were made on single attached leaves in the cuvette on a well-watered plant and oscillations occurred even in cool and cloudy weather at probably high xylem potential. They thus show that, in addition to a possible large-scale feedback regulation of water status, a feedback control of gas exchange is located on a very small spatial scale. Integration of stomatal responses into the regulation of whole plant water status obviously occurs within a wide range of spatial scales.
An interesting phenomenon is that an apparently coherent gas exchange response was produced by non-synchronous stomatal movements (Fig. 4), which indicates a more or less synchronous action of the bulk of stomata. This could be caused by some co-ordination between the stomata. A more realistic, stochastic explanation is, however, that the statistical distribution of stomatal properties led to a simultaneous action without functional synchronization.
As a result, the actual stomatal reponses as presented in Fig. 4 can by no means be deduced solely from the gS response. This raises some doubt as to the data base on which previous attempts to analyse the stomatal control system (Cowan, 1972; Jarvis et al., 1999) were founded. These analytical approaches were based on time series of gS which were assumed to be a reasonable measure for the average stomatal response. This requires that the responses must be largely uniform and that the relation of gS to stomatal aperture is known. Results in this study show that both prerequisites cannot be taken for granted. Uniformity can only be proven by visual observations of apertures, otherwise it is impossible to exclude counteracting movements concealed by the gas exchange. Uniformity cannot simply be deduced from the absence of conditions favouring stomatal patchiness (Jarvis et al., 1999), because variability on the micro-scale has to be taken into account, as has been demonstrated here. The assumption of a linearity between gS and apertures which was demonstrated analytically is contradicted by the experimental data presented in Fig. 8 which show a strongly non-linear relationship (see also Kaiser and Kappen, 2000; Nonami et al., 1990). This non-linearity makes it difficult to conclude directly from gS on aperture because the shape of the aperture distribution (Laisk et al., 1980) has to be considered, which is not the case when linearity is assumed. In particular, changes of gS do not directly reflect stomatal activity when a large fraction of stomata is closed, because then only the responses of the still open stomata contribute to changes of gS, while the action of closed stomata, being in the ‘Spannungsphase’, is completely ignored. These results demonstrate, however, that these responses are of high importance for the understanding of the stomatal control system.
It is concluded that any analysis based on integrating methods such as measurements of gS should be more cautiously interpreted with respect to responses of the guard cells. While important for the understanding of responses on leaf level and their ecophysiological effect, they may probably convey only limited information on the feedback processes acting in single guard cells.
The responses observed in these experiments may also suggest a comparison with the dynamics of patchy stomatal oscillations as observed by chlorophyll a fluorescence imaging (Cardon et al., 1994; Eckstein et al., 1996; Haefner et al., 1997; Siebke and Weis, 1995). The patterns of patchwise counteracting fluorescence changes were interpreted as a spatially co-ordinated movement of stomata. A direct comparison of the direct observations used here with this indirect as well as integrating method is difficult. The first question is whether the amplitude of the underlying stomatal movements is comparable with these results. A simultaneous recording of chlorophyll a fluorescence images and stomatal apertures indicated that patchy fluorescence only develops at very small apertures (Kaiser et al., 1999). This is supported by a calculation of gS from images of chlorophyll fluorescence, with gS being in the range from 0–40 mmol m-2 s-1 (Meyer and Genty, 1998), which certainly implies quite a low aperture level. This is not surprising if the type of relationship between aperture and photosynthetic capacity (Fig. 9) is considered, which points to the fact that Ci should be sufficiently reduced to increase non-photochemical quenching only at very small apertures. Therefore the oscillating fluorescence patterns are likely to be based on small aperture differences at a generally low degree of opening. This suggests that patchy stomatal closure for similar reasons as stomatal oscillations may be functionally linked to the particular properties of the lowest aperture range.
The results of this study indicate a functional explanation for oscillations which has already been proposed (Hopmans, 1971). The question arises, however, are the supposedly unavoidable oscillations advantageous or harmful? As most of the time transpiration and photosynthetic rate are far from the supposedly ‘optimal’ nominal value, oscillations should generally impair plant performance. However, calculations were presented which showed that oscillations may improve water use efficiency under certain conditions (Upadhyaya et al., 1988). This contradicts the assertion that ‘optimality’ is necessarily linked to steady-state responses and raises the question whether stomatal oscillations may even serve a ‘purpose’ (Cowan, 1972). However, even if stomatal oscillations in some cases are regarded as beneficial, the advantage is obviously restricted to moderate amplitudes. It seems necessary to avoid large amplitudes with wide fluctuations of gas exchange. While there appears to be no simple escape from the described insufficiency of feedback control at single stoma level, the responses of populations of stomata could be dampened by a high variability of stomatal responses. In these results it is obvious that individual stomata react differently as can be seen by phase shift and different frequences. The phase shift was caused by a variation in the lag-time necessary to overcome the ‘Spannungsphase’ (Laisk et al., 1980), leading to different starting times for the oscillations (Fig. 7). It is the variation in the properties ‘degree of initial guard cell swelling’ and ‘rate of volume increase’ which is transduced into a temporal variation and causes non-synchronous initial opening and subsequent phase-shifted oscillations. It is therefore proposed that stomatal variability reduces negative effects on gas exchange by avoiding synchronous closure and opening of all stomata. Variability should reduce the tendency to oscillate by lowering possible resonance effects caused, for example, by hydraulic coupling. Obviously, stomatal variability does not completely prevent oscillations observable at the leaf level. The observed oscillation in gL (Fig. 4) is presumably already levelled off and would have been much more pronounced if opening and closing movements of stomata had been synchronous.
The variability of stomatal responses has hitherto been mainly treated with respect to the effect on steady-state gas exchange (Laisk, 1983; Mott, 1995), as a phenomenon of biological variation (Weyers and Lawson, 1997) or a methodological problem annoying researchers with the need to use large samples (Kubinova, 1994; Weyers and Meidner, 1990). It now appears that this variation may also be advantageous to the plant by improving the regulation characteristics of the stomatal feedback system.
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Acknowledgments |
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The authors wish to thank the staff of the Botanical Garden of the University of Kiel for generous practical support of the experiments, Mr Thomas Walter for excellent technical assistance and Mrs Shamim Lenz for correcting the manuscript. The work was supported by a grant from the Deutsche Forschungsgemeinschaft (Ka 390/12-1).
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1 To whom correspondence should be addressed. Fax: +49 431 880 1522. E-mail: hkaiser{at}bot.uni\|[hyphen]\|kiel.de
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References |
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Assmann SM, Grantz DA. 1990. Stomatal response to humidity in sugarcane and soybean: effect of vapour pressure difference on the kinetics of the blue light response. Plant, Cell and Environment13, 163–169.
Ball JT. 1987. Calculations related to gas exchange. In: Zeiger E, Farquhar GD, Cowan JR, eds. Stomatal function. Stanford, California: Stanford University Press, 445–476.
Barradas VL, Jones HG, Clark JA. 1994. Stomatal responses to changing irradiance in Phaseolus vulgaris L. Journal of Experimental Botany45, 931–936.
Barrs HD. 1971. Cyclic variation in stomatal aperture, transpiration and leaf water potential under constant environmental conditions. Annual Review of Plant Physiology22, 223–236.
Barrs HD, Klepper B. 1968. Cyclic variations in plant properties under constant environmental conditions. Physiologia Plantarum21, 711–730.
Bunce JA. 1987. In-phase cycling of photosynthesis and conductance at saturating carbon dioxide pressure induced by increases in water vapour pressure deficit. Journal of Experimental Botany38, 1413–1420.
Cardon ZG, Mott KA, Berry JA. 1994. Dynamics of patchy stomatal movements and their contribution to steady-state and oscillating stomatal conductance calculated using gas-exchange techniques. Plant, Cell and Environment17, 995–1007.
Cousson A. 2000. Analysis of the sensing and transducing processes implicated in the stomatal responses to carbon dioxide in Commelina communis L. Plant, Cell and Environment23, 487–495.
Cowan IR. 1972. Oscillations in stomatal conductance and plant functioning associated with stomatal conductance: observations and a model. Planta106, 185–219.
Eckstein J. 1997. Heterogene Kohlenstoffassimilation in Blättern höherer Pflanzen als Folge der Variabilität stomatärer Öffnungsweiten–Charakterisierung und Kausalanalyse des Phänomens ‘stomatal patchiness’. Dissertatio. Bayerische Ludwig-Maximilians-Universität Würzburg.
Eckstein J, Beyschlag W, Mott KA, Ryel RJ. 1996. Changes in photon flux can induce stomatal patchiness. Plant, Cell and Environment19, 1066–1074.
Farquhar GD, Cowan IR. 1974. Oscillations in stomatal conductance. The influence of environmental gain. Plant Physiology54, 769–772.
Franks P, Cowan I, Farquhar G. 1997. The apparent feedforward response of stomata to air vapour pressure deficit: information revealed by different experimental procedures with two rainforest trees. Plant, Cell and Environment20, 142–145.
Fuchs EE, Livingston NJ. 1996. Hydraulic control of stomatal conductance in Douglas fir (Pseudotsuga menziesii (Mirb.) Franco) and alder (Alnus rubra (Bong)) seedlings. Plant, Cell and Environment19, 1091–1098.
Haefner J, Buckley T, Mott K. 1997. A spatially explicit model of patchy stomatal responses to humidity. Plant, Cell and Environment20, 1087–1097.
Herppich WB, von Willert DJ. 1995. Dynamic changes in leaf bulk water relations during stomatal oscillations in mangrove species. Continuous analysis using a dewpoint hygrometer. Physiologia Plantarum94, 479–485.
Hopmans PAM. 1971. Rhythms in stomatal opening of bean leaves. Mededelingen van de Landbouwhoogeschool te Wageningen71, 1–86.
Jarvis A, Young P, Taylor C, Davies W. 1999. An analysis of the dynamic response of stomatal conductance to a reduction in humidity over leaves of Cedrella odorata. Plant, Cell and Environment22, 913–924.
Kaiser H. 1999. Die stomatäre Reaktion von Sambucus nigra und Aegopodium podagraria in Abhängigkeit von Licht und Luftfeuchte—in situ Beobachtungen und Gaswechselmessungen im Freiland. Dissertatio. Christian-Albrechts-Universität zu Kiel.
Kaiser H, Eckstein J, Beyschlag W, Kappen L. 1999. Variabilität stomatärer Aperturen bei Auftreten fleckenhafter Chlorophyllfluoreszenz. Bielefelder Ökologische Beiträge14, 246–252.
Kaiser H, Kappen L. 1997. In situ observations of stomatal movements in different light-dark regimes: the influence of endogenous rhythmicity and long-term adjustments. Journal of Experimental Botany48, 1583–1589.
Kaiser H, Kappen L. 2000. In situ observation of stomatal movements and gas exchange of Aegopodium podagraria L. in the understorey. Journal of Experimental Botany51, 1741–1749.
Kappen L, Andresen G, Lösch R. 1987. In situ observations of stomatal movements. Journal of Experimental Botany38, 126–141.
Kappen L, Schultz G, Gruler T, Widmoser P. 2000. Effects of N-fertilization on shoots and roots of rape (Brassica napus L.) and consequences for the soil matric potential. Journal of Plant Nutrition and Soil Science163, 481–489.
Kearns EV, Assmann SM. 1993. Update on guard-cells: the guard cell-environment connection. Plant Physiology102, 711–715.[Web of Science][Medline]
Kerstiens G. 1996. Diffusion of water vapour and gases across cuticles and through stomatal pores presumed closed. In: Kerstiens G, ed. Plant cuticles—an integrated functional approach. Oxford: Bios Scientific, 121–135.
Kubinova L. 1994. Recent stereological methods for measuring leaf anatomical characteristics: estimation of the number and sizes of stomata and mesophyll cells. Journal of Experimental Botany45, 119–127.
Küppers M, Schneider H, Swan AG. 1993. Leaf gas exchange of beech (Fagus sylvatica L.) seedlings in lightflecks: a system for measuring rapid changes in CO2 partial pressures. Photosynthesis Research35, 299–304.
Laisk A. 1983. Calculation of leaf photosynthetic parameters considering the statistical distribution of stomatal apertures. Journal of Experimental Botany34, 1627–1635.
Laisk A, Oja V, Kull K. 1980. Statistical distribution of stomatal apertures of Vicia faba and Hordeum vulgare and the Spannungsphase of stomatal opening. Journal of Experimental Botany31, 49–58.
Lang ARG, Klepper B, Cumming MJ. 1969. Leaf water balance during oscillation of stomatal aperture. Plant Physiology44, 826–830.
Lange OL, Lösch R, Schulze E-D, Kappen L. 1971. Responses of stomata to changes in humidity. Planta100, 76–86.[Web of Science]
Lawson T, James W, Weyers J. 1998. A surrogate measure of stomatal aperture. Journal of Experimental Botany49, 1397–1403.
Lushnikov AA, Vesala T, Kulmala M, Hari P. 1994. A semiphenomenological model for stomatal gas transport. Journal of Theoretical Biology171, 291–301.
McBurney T, Costigan PA. 1984. Rapid oscillations in plant water potential measured with a stem psychrometer. Annals of Botany54, 851–853.
Meyer S, Genty B. 1998. Mapping intercellular CO2 mole fraction (Ci) in Rosa rubiginosa leaves fed with abscisic acid by using chlorophyll fluorescence imaging—significance of Ci estimated from leaf gas exchange. Plant Physiology116, 947–957.
Monteith JL. 1995. A reinterpretation of stomatal responses to humidity. Plant, Cell and Environment18, 357–364.
Mott KA. 1988. Do stomata respond to CO2 concentrations other than intercellular? Plant Physiology86, 200–203.
Mott KA. 1995. Effects of patchy stomatal closure on gas exchange measurements following abscisic acid treatment. Plant, Cell and Environment18, 1291–1300.
Mott KA, Parkhurst DF. 1991. Stomatal response to humidity in air and helox. Plant, Cell and Environment14, 509–515.
Naidoo G, von Willert DJ. 1994. Stomatal oscillations in the mangrove Avicennia germinans. Functional Ecology8, 651–657.
Nonami H, Schulze E-D, Ziegler H. 1990. Mechanisms of stomatal movement in response to air humidity, irradiance and xylem water potential. Planta183, 57–64.[Web of Science]
Parlange J-Y, Waggoner PE. 1970. Stomatal dimensions and resistance to diffusion. Plant Physiology46, 337–342.
Raschke K. 1965. Die Stomata als Glieder eines schwingungsfähigen CO2-Regelsystems Experimenteller Nachweis an Zea mays L. Zeitschrift für Naturforschung 20b, 1261–1270.
Raschke K. 1970. Leaf hydraulic system: rapid epidermal and stomatal responses to changes in water supply. Science167, 189–191.
Raschke K. 1979. Movements of stomata. In: Haupt W, Feinlieb ME, eds. Encyclopedia of plant physiology, Vol. 7. Physiology of movements. Berlin: Springer Verlag, 383–441.
Saliendra NZ, Sperry JS, Comstock PJ. 1995. Influence of leaf water status on stomatal response to humidity, hydraulic conductance and soil drought in Betula occidentalis. Planta196, 357–366.
Shackel KA, Brinckmann E. 1984. In situ measurement of epidermal cell turgor, leaf water potential and gas exchange in Tradescantia virginiana L. Plant Physiology78, 66–70.
Siebke K, Weis E. 1995. Assimilation images of leaves of Glechoma hederacea: Analysis of non-synchronous stomata related oscillations. Planta196, 155–165.[Web of Science]
Stålfelt MG. 1929a. Die Abhängigkeit der Spaltöffnungsreaktionen von der Wasserbilanz. Planta8, 287–340.
Stålfelt MG. 1929b. Pulsierende Blattgewebe. Planta7, 720–734.
Stålfelt MG. 1963. Diurnal dark reactions in the stomatal movements. Physiologia Plantarum16, 756–766.
Upadhyaya SK, Rand RH, Cooke JR. 1988. Role of stomatal oscillations on transpiration, assimilation and water-use efficiency of plants. Ecological Modelling41, 27–40.
Vogt UK. 1998. Strukturell-funktionelle Koordination von Wasserleitung und Transpiration als Grundlage des hydroökologischen Konstitutionstyps bei Kräutern, Stauden und Sträuchern. Dissertatio. Heinrich-Heine-Universität Düsseldorf.
Vogt UK, Lösch R. 1999. Stem water potential and leaf conductance: a comparison of Sorbus aucuparia and Sambucus nigra. Physics and Chemistry of the Earth (B)24, 121–123.
Weyers JDB, Lawson T. 1997. Heterogeneity in stomatal characteristics. Advances in Botanical Research incorporating Advances in Plant Pathology26, 317–352.
Weyers JDB, Meidner H. 1990. Methods in stomatal research. Harlow: Longman.
Whitehead D, Livingston NJ, Kelliher FM, Hogan KP, Pepin S, et al. 1996. Response of transpiration and photosynthesis to a transient change in illuminated foliage area for a Pinus radiata D. Don tree. Plant, Cell and Environment19, 949–957.
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