Functional Ecology 2007 21, 705–712
Forced depression of leaf hydraulic conductance in situ: effects on the leaf gas exchange of forest trees Blackwell Publishing Ltd
T. J. BRODRIBB*†‡ and N. M. HOLBROOK‡ *Department of Plant Science, University of Tasmania, PO Box 252-55, Australia 7001, and ‡Department of Organismic and Evolutionary Biology, Harvard University, 16 Divinity Avenue, Cambridge, MA, USA
Summary 1. Recent work on the hydraulic conductance of leaves suggests that maximum photosynthetic performance of a leaf is defined largely by its plumbing. Pursuing this idea, we tested how the diurnal course of gas exchange of trees in a dry tropical forest was affected by artificially depressing the hydraulic conductance of leaves (Kleaf). 2. Individual leaves from four tropical tree species were exposed to a brief episode of forced evaporation by blowing warm air over leaves in situ. Despite humid soil and atmospheric conditions, this caused leaf water potential (Ψleaf) to fall sufficiently to induce a 50–74% drop in Kleaf. 3. Two of the species sampled proved highly sensitive to artificially depressed Kleaf, leading to a marked and sustained decline in the instantaneous rate of CO2 uptake, stomatal conductance and transpiration. Leaves of these species showed a depression of hydraulic and photosynthetic capacity in response to the ‘blow-dry’ treatment similar to that observed when major veins in the leaf were severed. 4. By contrast, the other two species sampled were relatively insensitive to Kleaf manipulation; photosynthetic rates were indistinguishable from control (untreated) leaves 4 h after treatment. These insensitive species demonstrate a linear decline of Kleaf with Ψleaf, while Kleaf in the two sensitive species falls precipitously at a critical water deficit. 5. We propose that a sigmoidal Kleaf vulnerability enables a high diurnal yield of CO2 at the cost of exposing leaves to the possibility of xylem cavitation. Linear Kleaf vulnerability leads to a relatively lower CO2 yield, while providing better protection against cavitation. Key-words: dry tropics, leaf hydraulics, photosynthetic depression, cavitation, diurnal, assimilation Functional Ecology (2007) 21, 705–712 doi: 10.1111/j.1365-2435.2007.01271.x
Introduction
© 2007 The Authors. Journal compilation © 2007 British Ecological Society
A growing body of evidence points to the plumbing of plants as the functional nexus between water relations and photosynthetic performance. Xylem conduits carry the same water that exits the plant through the stomata, so in principle the propensity for leaves to lose water should match the capacity of the xylem to deliver the same water (Meinzer & Grantz 1990; Sperry 2000). It is widely accepted that the rates of transpiration (E ) and CO2 assimilation (A) in leaves are linked because optimization of the ratio A/E requires that the marginal cost of transpiration (δA/δE) remains constant (Cowan & Farquhar 1977). This feature of stomatal control effectively couples traits involved †Author to whom correspondence should be addressed. E-mail:
[email protected]
with water flow to those associated with primary production, and explains the observed correlation between hydraulic efficiency and photosynthetic maxima (Brodribb & Feild 2000; Hubbard et al. 2001). Co-ordination between these highly distinct physiological processes suggests that C3 land plants have converged on a common solution for optimizing water supply with respect to assimilation-linked water loss. The concept that changes in the hydraulic conductivity of the vascular system of individual plants might effect a corresponding depression in plant gas exchange has been confirmed by studies of drought (Saliendra et al. 1995; Vilagrosa et al. 2003), wounding (Sperry & Pockman 1993; Hubbard et al. 2001) and seasonal change (Brodribb & Holbrook 2003a). However, these factors influence plant conductivity over the long term, and all appear to impose an irreversible decline on the efficiency of water conduction through 705
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© 2007 The Authors. Journal compilation © 2007 British Ecological Society, Functional Ecology, 21, 705–712
the plant. Of potentially greater impact is the proposition we address here, that reversible, short-term changes in plant hydraulic conductivity might influence diurnal gas-exchange dynamics by changing the amount of water that can be transpired while maintaining leaf water potential homeostasis. This type of xylem–stomatal interaction is expected, assuming the stomatal aperture is under hydro-mechanical control, driven (in the light) by the impact of leaf water potential on guard cell turgor (Buckley et al. 2003). Several studies have linked declining hydraulic conductivity with reductions in gas exchange (Lo Gullo et al. 2003; Stiller et al. 2003), but it remains unknown how the dynamics of xylem dysfunction and repair might affect diurnal patterns of gas exchange in the natural environment. Brodribb & Holbrook (2004) made parallel measurements of leaf hydraulic conductance and gas exchange in a Costa Rican forest tree and showed a correlated depression of both parameters at midday. These observations led the authors to conclude that midday depression of gas exchange was linked to a reversible depression of leaf hydraulic conductance (Kleaf). The possibility that dynamic changes in the hydraulic conductance of the plant body might be involved in the depression of photosynthetic function on a daily basis is exciting, as it casts the water transport system in a vital role, patterning the process of leaf gas exchange. In seeking to test the proposition of a diurnal communication between fluctuating hydraulic conductance and gas exchange, we focus on leaves as the most likely location for such a dynamic interaction. Due to its location at the dry end of the hydraulic network, the leaf vascular tissue is most openly exposed to diurnal fluctuations in hydraulic supply and demand, leading to the likelihood of rapid changes in leaf water potential. These conditions are potentially inhibitory to the conductive function of the leaf vascular system, given that leaves seem to be equally or more susceptible than stems to depression of hydraulic conductivity under water deficit (Nardini et al. 2001; Stiller et al. 2003). Indeed, it is not unusual for stomatal closure to be initiated only after the incipient stages of Kleaf depression (Salleo et al. 2001; Brodribb & Holbrook 2004). Furthermore, a recent study suggested that Kleaf in many species is a linear function of leaf water potential (Ψleaf) (Brodribb & Holbrook 2006). In these species, Kleaf was maximal only when the leaf was fully hydrated, and declined proportionally with Ψleaf. The conditions under which Kleaf depression is most likely to lead to reduced gas exchange are those where evaporative demand is high and somewhat variable, yet soil water potential is sufficient to avoid nonreversible cavitation of the stem or root xylem. These prerequisites are best satisfied by the environment of the seasonally dry tropics. Our field site on the Pacific coast of northern Costa Rica is an ideal location for the study of dynamism in the physiology of water relations and gas exchange. The climate of this region is
characterized by 6 months of high rainfall and 6 months of zero rainfall (Janzen 1983). As a result, the tree flora is dominated by deciduous species that spend most of the dry season leafless. Most conveniently of all, the wet season is interrupted for a period of 6– 8 weeks, during which time dry wind and falling soil water potential lead to strong oscillations in stomatal conductance in some species, and uniform midday stomatal closure in others. We chose this period to test the effects of a forced depression in Kleaf on the diurnal course of gas exchange in four species with contrasting leaf characteristics. The aim of this study was first, to determine if the hydraulic conductivity of leaves could be manipulated in situ on trees in the field, and second, to ascertain whether a reduction in the hydraulic conductivity of the leaves alone was sufficient to induce a response in A and gs (stomatal conductance). The means of manipulating Kleaf was to force a rapid drop in Ψleaf by driving transpiration in individual leaves to supernatural levels. A recent study showed that, by forcing excised leaves to transpire in this fashion, Ψleaf could be driven down to a level where Kleaf was markedly reduced (Brodribb & Holbrook 2006). Employing the same technique on leaves attached to trees in the field, it was supposed that Ψleaf could be forced to even lower levels, thus inflicting a significant depression of Kleaf. We used four angiosperm tree species from this previous study, chosen for their contrasting physiological and ecological character. Two species exhibit a typical sigmoid relationship between Kleaf and Ψleaf, while Kleaf in the other two species was a linear function of Ψleaf. By monitoring gas exchange in the daylight hours following a short burst of induced high transpiration, we were able to measure the impact of reduced Kleaf on the diurnal course of gas exchange in these four species under natural forest conditions.
Materials and methods plant material and study site This investigation was conducted in the Santa Rosa National Park, located on the northern Pacific coast of Costa Rica (10°52′ N, 85°34′ W, 285 m a.s.l.). Mean annual rainfall in the park is 1528 mm; however, more than 90% of this falls between May and December. Rain falls on most days of the wet season, except for a window of dry weather in July and August, during which time low relative humidity and high irradiance result in a high evaporative demand. Diurnal and seasonal temperature ranges are relatively small, with a mean annual temperature of 28 °C. Vegetation in the park comprises a heterogeneous mosaic consisting of various stages of regeneration from former pastures, as well as some small areas of primary forest. Evergreen and deciduous species can be found at all successional stages, and on the particular site chosen for this study (a small open copse of trees 50 × 50 m), the vegetation
707 Forced depression of leaf hydraulics
cover was 50% evergreen Quercus oleoides and 50% early successional deciduous and brevi-deciduous tree species. We chose four tree species; Byrsonima crassifolia (Malpighiaceae), Curatella americana (Dilleniaceae), Genipa americana (Rubiaeae) and Rehdera trinervis (Verbenaceae) for their contrasting ecological and physiological characteristics. Byrsonima crassifolia and C. americana retain leaves for most of the dry months, while G. americana and R. trinervis remain leafless throughout the dry season. All species are small trees generally 40 °C on sunny days. A test of the response of A to leaf temperature was undertaken using three leaves of each species over the temperature range 30– 45 °C, and these data indicated that leaves of all four species could be heated to 38 °C without more than 10% decrease in assimilation below maximum (T.J.B., unpublished data). Hence, leaf temperature during blow-drying was limited to a maximum of 38 °C. Mature, healthy leaves were selected that would experience uniform sunlight throughout the day. During mid-morning, a pair of fine wire thermocouples were attached to the adaxial surface of the leaf and a hairdryer was used to direct a vigorous flow of warm air onto the abaxial (stomatal) surface of the leaf, while maintaining the leaf exposed to the sun and leaf temperature within the range 36–38 °C. Leaves were exposed to blow-drying for 3 min (Brodribb & Holbrook 2006). A non-desiccating heat treatment was used to control for the effect of leaf heating during blow-drying. Ten heated control leaves were established on two trees by wrapping five leaves on each tree with transparent plastic to prevent transpiration. These leaves were immediately exposed to the sun at a controlled angle such that leaves heated to 38 ± 0·5 °C for 3–4 min. The plastic was then removed and leaves allowed to re-equilibrate for 90 min before gas exchange in these leaves was measured at 12.00 h and compared with untreated controls. On the day prior to measurements of gas-exchange dynamics, five leaves of the target species were exposed to the forced evaporation protocol, then immediately excised and put in vapour-tight bags for measurement of Ψleaf in the pressure chamber. The mean Ψleaf from these measurements was assumed to be close to the Ψleaf in the target leaf, and the degree of Kleaf depression induced by the heating protocol could be estimated from the recently published vulnerability curves for each of these species (Brodribb & Holbrook 2006). The species most sensitive to forced evaporation, R. trinervis, was used to compare the effects of blowdrying with the hydraulic perturbation caused by cutting major leaf veins. Leaves from a single (10-m-tall) tree were manipulated over a period of 3 days. Four leaves per day from a single sunlit branch were subjected to the above-mentioned blow-drying treatment between 09.00 and 10.00 h (producing a total of 12 blow-dried leaves). The severity of desiccation was varied between leaves by setting the target temperature between 30 and 38 °C. At the same time, four leaves per day were injured by cutting two to three of the three to five major veins that supply water to the lamina, using a clean razor blade to cut only the vein,
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leaving the lamina undamaged. The treated leaves and four control leaves were left unmolested for a period of 2–3 h, at which time gas exchange was measured in all leaves using the above procedure. Immediately following gas-exchange measurements, leaves were excised and bagged for determination of Ψleaf in a pressure chamber. At the same time, four leaves that had been covered with plastic and foil in the predawn were also harvested for determination of stem water potential (Ψstem). Using the mean Ψstem for the measured branch, it was possible to calculate Kleaf using equation 1: Kleaf = νE/(Ψstem – Ψleaf)
(eqn 1)
where ν = kinematic viscosity of water at the measured leaf temperature. Relatively high ambient wind speed around sampled leaves meant that E, as measured in a ventilated cuvette, was a reliable measure of water loss in situ (Brodribb & Holbrook 2003a).
forced transpiration in excised leaves Maximum transpiration rates induced by the hairdryer treatment were estimated in each species using a flow meter to measure transpiration rates in excised leaves. Between 08.00 and 10.00 h, leaves with stomata fully open were excised underwater from sample trees and rapidly connected to a flow meter that measured the transpiration stream as it was sucked into the leaf (Brodribb & Holbrook 2006). Leaves were exposed to forced evaporation conditions identical to those for leaves heated in situ on the tree, with hairdryer settings adjusted to produce a leaf temperature of 38 °C with leaves transpiring in full sun. After 3 min maximum transpiration, leaves were removed from the flow meter for determination of Ψleaf with a pressure chamber, and of leaf area with a digital camera. Ten leaves from each of the four species were sampled in this way to give a mean maximum vapour flux for each species during forced transpiration.
statistical analysis Diurnal courses of assimilation and stomatal conductance were estimated by fitting the lowest-order polynomial that yielded an r2 value of >0·85 to mean hourly gas-exchange measurements. Differences between means and treated values of gas exchange were tested for significance using a Student’s t-test.
Results © 2007 The Authors. Journal compilation © 2007 British Ecological Society, Functional Ecology, 21, 705–712
Excised leaves exposed to a current of warm air were induced to transpire at rates significantly higher than leaves measured under average midday field conditions (Fig. 1). When leaves attached to trees in the field were exposed to an identical blow-drying treatment, the high transpiration rate induced caused Ψleaf to
Fig. 1. Mean leaf transpiration and leaf water potential measured at midday (open bars) and at 10-00 h immediately following 3 min forced transpiration by blow-drying individual leaves in situ (shaded bars).
decline rapidly from values > –0·5 MPa, to minimum values that appeared to be conservative within species (Fig. 1). The post-treatment Ψleaf was significantly lower (by 0·4–0·6 MPa) than the mean minimum midday Ψleaf recorded for each species during the study period (Fig. 1). According to recently published data showing the response of Kleaf to Ψleaf in these trees (Brodribb & Holbrook 2006), the values of post-blowdrying Ψleaf generated here were sufficient to depress Kleaf by between 50 and 74% in all species (Fig. 2).
gas exchange Gas exchange in control leaves reached a maximum at 08.00 h, except in Genipa, which did not become light-saturated until 09.00 h. The proceeding course of gas exchange was slightly different in each species, and characterized by mild midday depression of gas exchange in leaves of B. crassifolia (Fig. 3), by stable midday gas exchange in R. trinervis (Fig. 3), or by a gradual decline of gas exchange in both G. americana and C. americana (Fig. 4). Although conditions of VPD were similar on each measurement day, minimum midday Ψleaf was lower in B. crassifolia and R.
709 Forced depression of leaf hydraulics
Fig. 2. The response of leaf hydraulic conductance (Kleaf) to leaf water potential (Ψleaf) in each species sampled (data from Brodribb & Holbrook 2006). Vertical lines show mean Ψleaf ± SD in five leaves harvested immediately after the blow-drying treatment. Distinct differences are evident between the sigmoid-shaped vulnerability of Rehdera trinervis and Byrsonima crassifolia compared with the linear response observed in the other two species sampled. These plots show the expected impact of blow-drying on Kleaf in each species.
© 2007 The single Authors. Fig. 3. For trees of Byrsonima crassifolia and Rehdera trinervis, three panels Journal compilation show data from simultaneous measurements of: (top panel) mean vapour-pressure © 2007(British deficit ) and leaf water potential (Ψleaf) () of control leaves; (lower panels) mean Ecological Society, instantaneous CO2 uptake and stomatal conductance of five control leaves (, polyFunctional Ecology,fitted to show trend) and individual treated leaves (open symbols, nomial regressions 21, 705–712 bold lines). Treated leaves were exposed to 3 min blow-drying at the time indicated by
trinervis (–1·3 MPa in both species) than C. americana and G. americana (–0·7 to –0·9 MPa, respectively). The response of leaves to blow-drying fell into two categories. First, leaves of B. crassifolia and R. trinervis were found to exhibit a strong and prolonged depression in A and gs after blow-drying. Leaf hydraulic conductivity in both these species responded in a sigmoidal fashion to Ψleaf (Fig. 2) and, as a result, Kleaf was highly sensitive to manipulation of Ψleaf below –1·3 MPa. In the case of B. crassifolia, post-blowdrying gas exchange fell over a period of 2–4 h, reaching a minimum around the middle of the day (12.00 h to 14.00 h), at which point A and gs were
