Tree Physiology 21, 1215–1222
© 2001 Heron Publishing—Victoria, Canada
Responses of foliar δ13C, gas exchange and leaf morphology to reduced
hydraulic conductivity in Pinus monticola branches
LUCAS A. CERNUSAK1,2 and JOHN D. MARSHALL1
1
Department of Forest Resources, University of Idaho, Moscow, ID 83844-1133, USA
2
Present address: Environmental Biology Group and CRC for Greenhouse Accounting, Research School of Biological Sciences, Australian National
University, GPO Box 475, Canberra, ACT 2601, Australia
Received December 5, 2000
Summary We tested the hypothesis that branch hydraulic
conductivity partly controls foliar stable carbon isotope ratio
(δ13C) by its influence on stomatal conductance in Pinus monticola Dougl. Notching and phloem-girdling treatments were
applied to reduce branch conductivity over the course of a
growing season. Notching and phloem girdling reduced leafspecific conductivity (LSC) by about 30 and 90%, respectively. The 90% reduction in LSC increased foliar δ13C by
about 1‰ (P < 0.0001, n = 65), whereas the 30% reduction in
LSC had no effect on foliar δ13C (P = 0.90, n = 65). Variation in
the δ13C of dark respiration was similar to that of whole-tissues
when compared among treatments. These isotopic measurements, in addition to instantaneous gas exchange measurements, suggested only minor adjustments in the ratio of intercellular to atmospheric CO2 partial pressures (ci /ca) in response
to experimentally reduced hydraulic conductivity. A strong
correlation was observed between stomatal conductance (gs)
and photosynthetic demand over a tenfold range in gs. Although ci /ca and δ13C appeared to be relatively homeostatic,
current-year leaf area varied linearly as a function of branch
hydraulic conductivity (r 2 = 0.69, P < 0.0001, n = 18). These
results suggest that, for Pinus monticola, adjustment of leaf
area is a more important response to reduced branch conductivity than adjustment of ci /ca.
Keywords: carbon isotope ratio, intercellular CO2 concentration, leaf area.
Introduction
Stable carbon isotope ratios (δ13C) of plant tissue provide useful information about photosynthetic gas exchange (Ehleringer 1991, Marshall and Zhang 1994, Brugnoli and Farquhar
2000). For plants with the C3 photosynthetic pathway, discrimination against 13C during photosynthesis is controlled by
the CO2 partial pressure within the leaf intercellular spaces relative to that in the surrounding atmosphere (ci /ca) (Farquhar et
al. 1982). The ci /ca is a function of supply and demand for CO2
within leaf chloroplasts and can be related to photosynthetic
water-use efficiency (water loss per unit carbon gain) (Far-
quhar and Richards 1984, Farquhar et al. 1989). Understanding the sources of variation in ci /ca and δ13C is important for
predicting plant responses to environmental change (Ehleringer 1993a, Ehleringer and Cerling 1995, Marshall and Monserud 1996). In addition, variation in δ13C has been used to assess hydraulic limitations to maximum tree height (Yoder et
al. 1994).
The supply of CO2 to chloroplasts is controlled by stomatal
conductance (gs). Stomatal conductance, in turn, is partly controlled by a plant’s ability to conduct water from the soil to
sites of evaporation in leaves (Sperry and Pockman 1993,
Sperry et al. 1993, 1998, Bond and Kavanagh 1999). Excessive water tension in the soil-to-leaf hydraulic pathway can
lead to hydraulic failure associated with xylem cavitation and
embolism. Thus, when xylem tension is near the cavitation
threshold, plants may respond by decreasing gs.
Woody plants often exhibit large variations in δ13C, both
within individuals and among taxa (e.g., Craig 1954, Francey
et al. 1985, Leavitt and Long 1986, Gebauer and Schulze
1991, Yoder et al. 1994, Guehl et al. 1998, Martinelli et al.
1998). Meinzer et al. (1992, 1993) and Waring and Silvester
(1994) suggested that some of this variation reflects variation
in hydraulic conductivity. Subsequently, Panek (1996) presented a theory linking carbon isotope discrimination to conductance through the soil to leaf hydraulic pathway (expressed
per unit leaf area). She noted that branches make a proportionally large contribution to the resistance along this pathway and
suggested that δ13C should vary as a function of branch conductivity.
Alternatively, plants may respond to variation in hydraulic
conductivity by modifying the relationship between conducting area and leaf area, such that the balance between CO2 supply and demand is unaffected by changes in hydraulic conductivity. Thus ci /ca and δ13C may be homeostatically maintained,
whereas other parameters such as leaf area (Becker et al. 2000)
and leaf morphology (Niinemets and Kull 1995) vary in response to alterations in hydraulic conductivity.
To determine whether ci /ca and δ13C operate as homeostatic
set points with respect to hydraulic conductivity, we investigated responses of foliar δ13C, gas exchange, leaf area, and
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CERNUSAK AND MARSHALL
leaf morphology to reduced hydraulic conductivity in
branches of Pinus monticola Dougl. (western white pine). To
do this, we experimentally altered branch conductivity on
trees growing at a common site, thereby avoiding complications associated with sampling trees exposed to large variations in soil water content and atmospheric vapor pressure
deficit.
Materials and methods
Site description
The study site is located at the University of Idaho Experimental Forest (46°50′ N, 116°43′ W) at an altitude of 915 m a.s.l.
The climate at the site is characterized by cool, mild winters
and warm, dry summers. Mean annual temperature and precipitation are approximately 7 °C and 720 mm, respectively.
Most precipitation occurs between November and April, with
the majority falling as snow. Soils on the site are deep, welldrained silt loams formed in volcanic ash. Sampled trees grew
in a Pinus monticola plantation established after clear felling
in 1982. At the time of the study in 1999, the trees averaged
4 to 5 m in height. Selected study trees were apparently free of
insects and disease and experienced minimal shading by
neighboring trees.
Experimental design
Three treatments were applied to sample branches: notching,
phloem girdling, and control. In the notching treatment, a razor blade was forced through the bottom half of each branch
near the branch base. This effectively reduced the branch
cross-sectional area by a factor of 0.5 in the plane encompassing the blade insertion. The razor blades were sterilized by
rinsing with ethanol before insertion. Once inserted, the blades
remained in place throughout the experiment, creating a permanent barrier to xylem water flux to the foliage distal to the
point of insertion.
In the phloem-girdling treatment, bark (periderm, cortex,
and phloem) was removed from a 5-cm branch section near
each branch base. This was achieved by inserting a razor blade
through the bark until it met the hard, outer xylem. Two such
incisions were made around the circumference of each branch
and were joined by a straight line. The isolated section of bark
was peeled away. The girdled branch sections were immediately wrapped in several layers of Parafilm to prevent desiccation. Parafilm was reapplied periodically during the experiment. The girdling treatment was intended to block phloem
transport out of the branch, which was expected to result in assimilate accumulation in the foliage and feedback-inhibition
of photosynthesis (Stitt 1991, Myers et al. 1999). Control
branches were left untreated.
A total of 25 trees were included in the experiment. On each
tree, three branches within one whorl were selected from either the third, fourth, or fifth whorl from the treetop. The three
selected branches had the most southerly exposures in the
whorl. Each treatment was randomly applied to one of the
three sample branches on each tree, for a total of 75 experi-
mental branches. Treatments were applied between May 6 and
11, 1999, about 3 weeks before bud break occurred.
Gas exchange
Photosynthesis, gs, and ci /ca were measured on current-year
foliage (foliage produced after the application of the treatments). Gas exchange measurements were performed with an
LI-6200 portable photosynthesis system (Li-Cor, Inc., Lincoln, NE) equipped with a 0.25-l cuvette. Two or three needle
fascicles (10 to 15 needles) were detached from each branch
and placed in the cuvette. Gas exchange was measured within
5 min of needle detachment (Meng and Arp 1993). Gas exchange measurements were conducted between 0800 and
1100 h on August 3 and 4, 1999. Each of the three experimental branches was measured sequentially on each tree; thus
there was no bias toward any of the treatments resulting from
the time of measurement. Irradiance exceeded 1000 µmol m –2
s –1 of photosynthetically active radiation (PAR) during all
measurements. We assumed that this irradiance was sufficient
to saturate photosynthesis in Pinus monticola, as was the case
for Pinus strobus L. (eastern white pine) (Maier and Teskey
1992).
Projected leaf area of foliage measured for gas exchange
was estimated from a regression equation relating needle
length to projected leaf area (J.D. Marshall, unpublished data).
Samples were frozen following gas exchange measurements
and stored for determinations of needle length. Following
needle length measurements, samples were air-dried at 70 °C
and weighed to the nearest 0.1 mg. Specific leaf area (SLA) of
each sample was determined by dividing projected leaf area by
leaf dry weight.
Carbon isotope ratios
The δ13C was determined for the foliage samples measured for
gas exchange in early August as well as for current-year foliage sampled from treated branches in late August. All foliage
samples were air-dried at 70 °C and ground to a fine powder in
a rotating ball mill. Subsamples of about 1 mg were combusted in an elemental analyzer (NC2500, CE Instruments,
Milan, Italy); combustion products were swept by a helium
carrier gas and continuous-flow interface into an isotope ratio
mass spectrometer (Delta Plus, Finnigan MAT, Bremen, Germany) operated by the Idaho Stable Isotope Laboratory, University of Idaho. All carbon isotope ratios are expressed in
delta notation relative to the Pee Dee Belemnite standard. The
analytical precision of the analyses, based on repeated measurements of a working standard (Idaho flour), was ± 0.08‰
(standard deviation). Foliar carbon and nitrogen concentrations were determined from peak areas obtained from mass
spectrometric measurements.
Although we applied our treatments well before bud break
occurred, it is possible that current-year foliage was partially
constructed from stored photosynthate. If this were the case,
whole-tissue δ13C would not provide an accurate measure of
the response of isotopic discrimination to the experimental
treatments, whereas δ13C of dark-respired CO2 should provide
TREE PHYSIOLOGY VOLUME 21, 2001
δ13C RESPONSE TO REDUCED CONDUCTIVITY
a measure of the isotopic composition of more recently fixed
photosynthate.
We collected dark-respired CO2 from whole current-year
shoots in late August 1999. The procedure used for CO2 collection and isotopic analysis has been described (Cernusak et
al. 2001). Briefly, current-year shoots were harvested and
transported to the laboratory; on the same day, shoots were
placed in a 1-l cuvette attached to the Li-Cor LI-6200. The
closed system was then scrubbed free of CO2 and allowed to
refill with respired CO2. When the CO2 concentration within
the system reached 360 µmol mol –1, a 1-ml air sample was extracted with a gas-tight, locking syringe (VICI Precision Sampling, Baton Rouge, LA). Samples of about 400 µl of air were
injected into an automated trace-gas condensing device
(Precon, Finnigan MAT, Bremen, Germany). The condensate
was swept by a helium carrier gas through a 50-m POROPLOT Q column, which separated N2O from CO2, and into the
isotope ratio mass spectrometer. The analytical precision,
based on repeated injections of a standard gas (ISU-720C,
Oztech Trading Corp., Dallas, TX; δ13C = –10.98‰), was
± 0.2‰.
Hydraulic conductivity
Hydraulic conductivity (K h) was determined for a randomly
selected subsample of the treated branches (six trees, 18
branches) in early September 1999. Branch sections of approximately 50 cm in length were excised near the branch
bases. For notched and girdled branches, the section included
the region of treatment. Once in the laboratory, the branch sections were recut under water and the bark removed to prevent
resin from bleeding into and occluding the severed tracheids.
The K h was measured as described by Sperry et al. (1988). The
flow rate of purified water (filtered to 0.2 µm and acidified to
pH 2 with HCl) through branch sections was determined
gravimetrically. Flow was generated by pressure from a hydraulic head and K h was calculated as the water flux divided
by the pressure gradient multiplied by the branch length.
Branch sections were kept underwater during flow measurements to prevent evaporation from the wood surface. The flow
rate under zero hydraulic head was measured and subtracted
from the pressurized flow rate as described by Stiller and
Sperry (1999).
To estimate projected leaf area distal to the branch sections,
we determined SLA of five needle fascicles on each branch as
described for the gas exchange measurements. Specific leaf
area was then used to convert whole-branch leaf dry mass to
projected leaf area. Leaf-specific hydraulic conductivity
(LSC) was calculated as the quotient of K h and whole-branch
projected leaf area.
We analyzed treatment-related variation in measured parameters by analysis of variance. Individual branches were
considered to be independent experimental units; thus we assumed branches to be autonomous with respect to fluxes of
carbon and water. When treatment was a significant term in
the analysis of variance, individual treatments were compared
by Tukey’s method for pair-wise comparisons. All statistical
1217
analyses were performed with SYSTAT 9.0 (SPSS Inc., Chicago, IL).
Results
Leaf structure
Leaves produced after the experimental manipulations varied
in structure among treatments. Bud break occurred at the study
site on about May 27, 1999. Needle length was measured on
August 3 and 4, 1999 and again between August 26 and 28,
1999. Current-year foliage increased in length between the
two measurement periods for all three treatments (P < 0.0001
for control, P < 0.001 for notched, and P = 0.03 for girdled);
however, the increase was less for girdled branches than for
control or notched branches (Figure 1A). On both measurement dates, needle length of girdled branches was less than
needle length of control branches (P < 0.001 for early August,
P < 0.0001 for late August), whereas needle lengths of control
and notched branches did not differ (P = 0.83 for early August,
P = 0.27 for late August).
Patterns of change in SLA between early and late August
were opposite to those in needle length (Figure 1B). Specific
leaf area was unchanged between early and late August for
control (P = 0.18) and notched (P = 0.21) branches, but de-
Figure 1. Needle length (A) and specific leaf area (B) of Pinus
monticola foliage collected in either early or late August 1999. Treatments were applied in late May 1999. Error bars represent one standard error.
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CERNUSAK AND MARSHALL
creased for girdled branches (P < 0.0001). On both sampling
dates, SLA of girdled branches was less than that of controls
(P < 0.0001 for early and late August), whereas SLA of
notched branches did not differ from controls (P = 0.96 for
early August, P = 0.87 for late August).
Current-year foliage of girdled branches also differed in
chemical composition from controls (Table 1). Foliar nitrogen
and carbon concentrations, expressed per unit mass, were less
for girdled branches than for control branches. Foliage of
notched branches, on the other hand, did not differ in nitrogen
or carbon concentration from that of control branches.
Gas exchange
Maximum photosynthetic rates of current-year foliage were
strongly related to gs for all treatments combined (Figure 2).
The relationship between gs and photosynthesis was approximately linear for gs values ranging from 20 to near 150 mmol
H2O m –2 s –1; more than 90% of the data fell within this range.
The intercept of the linear portion of this relationship was approximately zero. Maximum photosynthetic rates ranged from
less than 2 to more than 12 µmol CO2 m –2 s –1. Photosynthetic
rates of girdled branches were less than half those of controls;
whereas those of notched branches did not differ from controls
(Figure 3A). Mean gs of notched branches was similar to that
of controls (Figure 3B), whereas mean gs of girdled branches
was substantially less than the control value. We observed no
difference among treatments in ci /ca as measured by instantaneous gas exchange (Figure 3C).
Figure 2. Light-saturated photosynthetic rates plotted against stomatal conductance for Pinus monticola foliage measured in early August 1999 between 0800 and 1100 h local time at irradiances exceeding 1000 µmol m –2 s –1. Measurement of each treatment spanned the
full range of measurement times.
branches did not differ from that of controls (Figure 4B). In
general, the δ13C of dark respiration was enriched over that of
whole tissue by about 2‰. This difference did not vary among
treatments (Figure 4C).
Stable isotope ratios
Whole-tissue δ13C of current-year foliage collected in late August was higher (less negative) by 0.88‰ for girdled branches
than for controls. In contrast, notched branches did not differ
from controls (Figure 4A). The δ13C of foliage collected in
early August did not differ from that collected in late August
for control or notched branches (P = 0.74 for control branches,
P = 0.56 for notched branches). In contrast, foliar δ13C of girdled branches increased by 0.15‰ between early and late August (P = 0.002). Values shown in Figure 4 are those measured
in late August.
Variation among treatments in δ13C of respired CO2 was
similar to that of whole tissues. The δ13C of dark respiration
from girdled branches was 0.90‰ heavier than that from control branches, whereas δ13C of dark respiration from notched
Table 1. Nitrogen and carbon concentrations ([N] and [C], respectively) of current-year Pinus monticola foliage. Treatments were applied to branches in late May 1999, and foliage samples were collected in late August 1999. Means within a row followed by different
letters are significantly different at P < 0.05. Values in parentheses are
one standard error.
[N] (mg g –1)
[C] (mg g –1)
Control
Notched
Girdled
14.6 (0.19) a
484 (1.75) a
14.5 (0.26) a
485 (1.81) a
8.9 (0.25) b
477 (2.23) b
Figure 3. Net photosynthesis (A), stomatal conductance (B), and ci /ca
ratio (C) for current-year foliage of treated Pinus monticola branches.
Measurements were conducted in early August 1999. Bars within a
panel followed by different letters are different at P < 0.05. Error bars
represent one standard error.
TREE PHYSIOLOGY VOLUME 21, 2001
δ13C RESPONSE TO REDUCED CONDUCTIVITY
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between current-year leaf area on the main axis of sample
branches and K h of the measured sections at the bases of
branches (Figure 6). Note that in this analysis, K h was expressed independently of the amount of subtending leaf area.
Discussion
Figure 4. Whole-tissue carbon isotope ratio for foliage collected in
late August (A), carbon isotope ratio of CO2 respired in late August
(B), and the difference between carbon isotope ratios of whole-tissue
and respired CO2 (C) for current year foliage of treated Pinus
monticola branches. Bars within a panel followed by different letters
are different at P < 0.05. Error bars represent one standard error.
Foliar δ13C decreased with increasing gs for all treatments
combined (data not shown). The relationship between the two
parameters was curvilinear and was best described by an exponential decay function (δ13C = –26.3 + 2.06exp(–0.009gs);
r 2 = 0.41, n = 59). In addition, foliar δ13C declined linearly
with SLA for all treatments combined. The two were related
by the equation δ13C = –23.2 – 0.05SLA (r 2 = 0.39, P <
0.0001, n = 65).
We tested the hypothesis that foliar δ13C varies in response to
reduced hydraulic conductivity in Pinus monticola branches.
As predicted by theory (Panek 1996), δ13C increased with decreasing LSC; however, the increase in δ13C was small. A
more than 90% reduction in LSC led to an average increase in
foliar δ13C of about 1‰, whereas a 30% reduction in LSC had
no effect on foliar δ13C. Both isotopic and instantaneous measurements suggested only minor adjustments in ci /ca over a
large range of hydraulic conductivities, despite large changes
in gs. Although ci /ca and δ13C appeared to be fairly homeostatic parameters, leaf area and leaf morphology were more
plastic. In particular, current-year leaf area varied linearly as a
function of branch conductivity. Thus, it appears that, for
Pinus monticola, adjustment of leaf area may be a more likely
response to alterations in branch conductivity than adjustment
of ci /ca.
Relationships between foliar δ13C and branch conductivity
have been investigated previously over a naturally occurring
climate gradient in Oregon, USA (Panek and Waring 1995,
Panek 1996). The shape of the relationship between δ13C and
LSC that we observed (Figure 5) is similar to that reported by
Panek (1996). However, the range of δ13C values was slightly
less in our study. We induced variation in LSC by experimentally treating branches at a common site, whereas in the Oregon study sample trees were exposed to varying degrees of soil
water stress and atmospheric vapor pressure deficit, in addition to variation in LSC. Because LSC decreased with increas-
Hydraulic conductivity
Leaf specific hydraulic conductivity (LSC) of control
branches was 1.12 × 10 – 4 ± 0.15 × 10 – 4 kg m –1 s –1 MPa –1
(mean ± SE). The LSC of notched branches was reduced by
about 30% relative to controls (LSC = 0.77 × 10 – 4 ± 0.09 ×
10 – 4 kg m –1 s –1 MPa –1), with the difference between the two
being moderately significant (P = 0.07, n = 18). For girdled
branches, LSC was reduced by more than 90% when compared with the control value (LSC = 0.06 × 10 – 4 ± 0.02 ×
10 – 4 kg m –1 s –1 MPa –1), and the difference was highly significant (P < 0.0001, n = 18).
The δ13C of current-year foliage was negatively related to
LSC when compared across all treatments combined (Figure 5). In addition, we observed a strong, linear relationship
Figure 5. Whole-tissue carbon isotope ratio of current-year Pinus
monticola foliage collected in late August plotted against the leaf-specific hydraulic conductivity (LSC) of the supporting branch. Data are
for all treatments combined. Conductivity was measured in early September. Conductivity values are expressed as LSC × 10 5. The equation relating δ13C to LSC is y = –24.8x0.016 with r 2 = 0.61, P < 0.0001,
and n = 18.
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CERNUSAK AND MARSHALL
Figure 6. Variation in current-year leaf area on the main terminus of
Pinus monticola branches expressed as a function of branch hydraulic
conductivity (K h). Both conductivity and leaf area were measured in
early September. Data are for all treatments combined. Conductivity
values are expressed as K h × 106. Leaf area is related to conductivity
by the equation y = 256 + 9.1x with r 2 = 0.69, P < 0.0001, and n = 18.
ing aridity across the Oregon transect, stomatal constraints
arising from factors other than LSC may have contributed to
the observed variation in δ13C.
Branch length influences hydraulic conductance because it
is a component of the path length between soil and leaf. If
other parameters are held constant, an increase in branch
length should cause a decrease in leaf-specific conductance;
the decrease should be proportional to the amount of added resistance caused by the additional branch length relative to the
total resistance from soil to leaf. Observations of increasing
foliar δ13C with increasing branch length support the hypothesis that branch conductance influences foliar δ13C (Waring and
Silvester 1994, Panek and Waring 1995, Walcroft et al. 1996,
Warren and Adams 2000). However, a relationship between
branch length and foliar δ13C has not always been observed
(Heaton and Crossley 1995, Livingston et al. 1998, Warren
and Adams 2000). A lack of response of foliar δ13C to variation in branch length could result from reductions in leaf area
that compensate for the additional resistance conferred by increased path length. Although more data are necessary to determine the extent to which such compensation occurs, the
relationship between current-year leaf area and Kh observed in
this study (Figure 6) suggests the possibility. This suggestion
is further supported by observed reductions in the amount of
leaf area per unit sapwood area with increasing tree height
(Schafer et al. 2000).
Instantaneous measurements of ci /ca did not differ significantly among treatments, whereas δ13C, an integrated measure
of ci /ca, did. The instantaneous measurements were taken over
30 s. In contrast, the respired CO2 δ13C values should integrate
over time scales of hours to days, and whole-tissue δ13C values
should integrate over time scales of weeks to months (the period over which the leaf tissue was formed). A longer time period of integration should lead to a higher signal to noise ratio,
as any high-frequency variation in ci /ca will be averaged out to
a greater extent. The coefficients of variation for the three estimates confirm this suggestion. For the control treatment, coefficients of variation were 7.7, 2.4, and 1.6% for instantaneous
ci /ca, respired CO2 δ13C, and whole-tissue δ13C, respectively.
Thus, the power to detect differences among treatments was
greater for the isotopic data than for the instantaneous data.
Additionally, instantaneous ci /ca was measured in mid-morning when the atmospheric vapor pressure deficit was relatively
low. As the vapor pressure deficit increased during the day,
treatment related differences in ci /ca would have likely become more pronounced.
We observed a strong correlation between photosynthesis at
saturating irradiance and gs in treated branches (Figure 2). The
relationship was such that ci /ca was held nearly constant across
experimental treatments (Figure 3C). A similar relationship
between photosynthesis and stomatal conductance was observed in the C3 plant species Eucalyptus pauciflora Sieber ex
A. Spreng. and Gossypium hirsutum L. under wide-ranging
experimental conditions (Wong et al. 1978, 1979). More recently, analyses of photosynthesis in the leaves of several C3
species have indicated increasing inhibition of photosynthetic
metabolism with decreasing leaf water potentials (Kanechi et
al. 1996, Escalona et al. 1999, Tezara et al. 1999). These reductions in photosynthetic demand for CO2 in water-stressed
leaves appear to be caused by a slower regeneration of ribulose
bisphosphate leading to inhibition of ribulose bisphosphate
carboxylase/oxygenase activity (Tezara et al. 1999).
We suggest that correlated reductions in photosynthetic demand for CO2 and gs limit the response of foliar δ13C to perturbations in hydraulic conductivity. It was recently reported that
photosynthetic capacity was strongly related to LSC in seven
conifers and 16 angiosperm species (Brodribb and Feild
2000). Similarly, in a laboratory experiment, photosynthesis
and gs varied linearly with LSC in Pinus ponderosa Dougl. ex
Laws. seedlings (Hubbard et al. 2001). Correlated reductions
in gs and photosynthesis may be a general response to reduced
hydraulic conductivity among taxa. If this is the case, only
subtle shifts in δ13C should be expected in response to variation in LSC.
Phloem girdling caused a marked decrease in xylem hydraulic conductivity. The girdling treatment was initially intended to reduce phloem export, which was expected to reduce
photosynthetic demand while maintaining gs. Clearly the
treatment did not have the desired effect. It is possible that air
was trapped against the xylem when the girdled sections were
wrapped with Parafilm. If the xylem was “nicked” when the
bark was cut away, it would have provided an opening through
which air could enter the xylem conduits. Although the Parafilm was replaced regularly during the experiment, it may
have developed leaks allowing continued air entry. Alternatively, we considered the possibility that fungi invaded the girdled sections. However, attempts to culture and isolate fungi at
the conclusion of the experiment were unsuccessful. Because
we do not know the mechanism by which girdling reduced
conductivity, we are unsure of when the loss of conductivity
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δ13C RESPONSE TO REDUCED CONDUCTIVITY
occurred. However, differences in leaf area and structure suggest that it may have occurred early in the experiment.
In contrast to phloem girdling, notching had a relatively minor effect on hydraulic conductivity. This result is consistent
with a previous experiment in which foliar gas exchange and
LSC showed no response to notching in branches of young and
old Pinus ponderosa (Hubbard et al. 1999). Hubbard et al.
(1999) suggested that there was redundancy in the sapwood of
their pine branches. Because we treated each branch at only a
single point along its axis, it is also possible that the branches
formed a smaller portion of the total resistance along the pathway from soil to leaf than we anticipated.
Foliage from phloem-girdled branches differed from controls in both carbon and nitrogen concentration (Table 1). The
decrease in carbon concentration may have resulted from
starch accumulation in the foliage because phloem was removed from the branch bases, thus disrupting the pathway for
sugar transport out of the branches. An excess accumulation of
starch could have decreased the photosynthetic demand of the
foliage, thus decreasing foliar δ13C relative to controls. However, there was no correlation between foliar δ13C and carbon
concentration (P = 0.33, n = 65), or between photosynthesis
and carbon concentration (P = 0.12, n = 58). Therefore, we
suggest that photosynthetic demand was not limited by high
starch concentrations.
Different biochemical fractions in leaves frequently have
different isotopic signatures (Brugnoli and Farquhar 2000).
For this reason, it is often preferable to extract cellulose when
comparing δ13C among tissues that may have different biochemical compositions. The difference in carbon concentration between foliage of girdled and control branches suggests
possible differences in tissue composition. Although we chose
to compare whole-tissue δ13C rather than cellulose δ13C, we
also compared the δ13C of dark respiration. Assuming similar
substrates for respiration, the measurement of dark-respired
CO2 should be as effective as extracting cellulose in removing
variation associated with differences in tissue composition.
Measurements of whole-tissue δ13C and respired CO2 δ13C
yielded similar patterns among treatments.
Foliar nitrogen concentration is frequently related to photosynthetic capacity (Field and Mooney 1986). Ideally, such
comparisons should rely on measurements expressed on a
similar basis. In this study, the expression of nitrogen concentrations per unit leaf area led to a weak relationship with photosynthetic rates (r 2 = 0.13, P < 0.01, n = 58), and no relationship with foliar δ13C (P = 0.71, n = 65). A lack of correlation
between area-based photosynthesis and nitrogen concentration was also reported for several evergreen conifer species
grown in Wisconsin, USA (Reich et al. 1995). We suggest that
the response of foliar δ13C to reduced LSC in girdled branches
was not dampened by a decrease in photosynthetic capacity resulting from low foliar nitrogen concentrations. This suggestion is supported by the results of the notching treatment showing that the reduction in LSC occurred independently of any
change in foliar nitrogen concentration; a 30% reduction in
LSC had no effect on foliar δ13C.
In conclusion, the experimental data supported the hypothe-
1221
sis that foliar δ13C varies as a function of hydraulic conductivity in Pinus monticola. However, the response of foliar δ13C to
reductions in LSC appeared to be moderated by the strong correlation between stomatal conductance and photosynthetic demand for CO2. This result reinforces the notion that ci /ca is an
important set point for photosynthetic gas exchange (Wong et
al. 1979, Ehleringer 1993a, 1993b, Ehleringer and Cerling
1995). In contrast, leaf area may be more plastic in its response
to reduced hydraulic conductivity. Future studies may profit
from determining the extent to which variation in leaf area
compensates for reductions in hydraulic conductivity resulting from increased branch length or other long-term losses in
conducting capacity.
Acknowledgments
L.A.C. was supported by a fellowship from the Stillinger Foundation
at the University of Idaho. We are grateful to R. Brandon Pratt and
R. Alan Black for assistance with hydraulic measurements, and to
George Newcombe for assistance in attempting to culture fungi. We
thank Drs. Margaret Barbour, Graham Farquhar, Katy Kavanagh, Jeff
Miller and Jeanne Panek for critical comments on earlier versions of
the manuscript.
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