Plant Physiol. (1985) 77, 157-1610032-0889/85/77/0157/05/$0 1.00/0

Permeability of the Suberized Mestome Sheath in Winter Rye'Received for publication May 16, 1984 and in revised form September 19, 1984

CAROL A. PETERSON, MARILYN GRIFFITH2, AND NORMAN P. A. HUNER*Department ofBiology, University of Waterloo, Waterloo, Ontario, Canada N2L 3GJ (C.A.P.);Department ofPlant Sciences, University of Western Ontario, London, Ontario, Canada N6A 5B7 (M.G.,N.P.A.H.)

ABSTRACr

Mestome sheath cells of winter rye (Secal cereale L. cv Puma) depositsuberized lamellae in their secondary cell walls. Histochemical testsincluding acid digestion and staining with Sudan IV and Chelidoniummajus root extract were used to detect the presence of suberin in theprimary cell wall. There was no evidence of a Casparian band betweenadjacent mestome sheath cells. Fluorescent dye techniques were used totrace solute movement through the rye leaf apoplast. Calcofluor whiteM2R, a fluorescent dye which binds to cell walls as it moves apoplasti-cally, proved to be too limited in its mobility in leaves to test mestomesheath permeability. Trisodium 3-hydroxy-5,8,10 pyrene trisulfonate, afluorescent dye which is mobile in the apoplast, moved easily up thevascular bundles in the transpiration stream, and diffused outward fromthe veins to the epidermal cell walls within minutes of reaching aparticular level in the leaf. We conclude that the suberized mestomesheath of rye leaves is freely permeable to solutes moving apoplasticallythrough radial primary cell walls.

Vascular bundles in winter rye leaves are encircled by twobundle sheaths. The outerbundle sheath, termed the parenchymasheath, consists of large, thin-walled, photosynthetic (C3) cells.The inner bundle sheath, or mestome sheath, is characterized bysmall, nonphotosynthetic cells. The cell walls of the mestomesheath are asymmetrically thickened along their inner tangentialand radial walls (6), and contain suberin lamellae which com-pletely encircle each mestome sheath cell in the secondary cellwall (7).The structure of the mestome sheath is reminiscent of the

structure of a state III root endodermis in which the secondarycell wall is thickened, suberized, and often lignified, and portionsof the primary wall are suberized to form Casparian bands (16).These Casparian bands are hydrophobic and prohibit the apo-plastic movement of water and solutes into the vascular systemof roots. Although Strugger (23) reported that the dyes berberinesulfate, eosin, sulforhodamine G, and PTS3 moved from theveins of rye and wheat leaves to the epidermis, there are indica-tions from later work (3, 4, 17, 24) that the mestome sheath mayimpede apoplastic movement. With the use of histochemicaltechniques to detect Casparian bands in root hypodermal and

'Supported by funds from the Natural Science and Engineering Re-search Council of Canada.

2Present address: Agricultural Experiment Station, University ofAlaska, Fairbanks, AK 99701.

3Abbreviations: PTS, trisodium, 3-hydroxy-5,8, 10 pyrene trisulfonate;RNH, unhardened rye; RH, cold-hardened rye; CFW, calcofluor whiteM2R.

endodermal cell walls (18, 20) and fluorescent dyes, mobile (PTS)and immobile (Calcufluor white M2R) within the apoplast (8,19, 21, 23, 25), the present study was undertaken to clarifywhether the mestome sheath cell walls could restrict apoplasticmovement from the vascular system.

MATERIALS AND METHODS

Plant Materials. Winter rye (Secale cereale L. cv Puma) wasgrown in vermiculite watered with modified Hoagland solutionin growth chambers programmed for a temperature regime of20°C/16°C (day/night) with a light intensity of 200 ,uE m-2 s-'during the 16-h photoperiod (11). Plants grown under theseconditions for 4 weeks are termed nonhardened rye (RNH).Plants transferred after 1 week to a growth chamber set at 5C(day/night) for an additional 7 weeks have become cold hardenedrye (RH). Comparative growth kinetics indicated that the RNHand RH plants were of similar physiological age (13).Leaf Anatomy. In order to study leaf anatomy, RNH and RH

leaves were cleared using a modification ofthe method describedby Arnott (1). Pigments were extracted by boiling leaves in 95%ethanol, and proteins were extracted by treatment with 2% (w/v) NaOH at 60°C for 6 h followed by treatment with saturatedaqueous chloral hydrate at 6O°C for 18 h. The cleared leaveswere rinsed in water for 10 min, stained in 2% (w/v) safranin for15 to 30 min, and destained in brief rinses ofwater, 30% ethanol,50% ethanol, and 30% ethanol. To enhance the contrast of thered-stained lignified cells for photography, the specimens wereilluminated with transmitted light which had passed through awide band green filter (520-550 nm).

Cross sections of rye leaves were prepared for the observationof transverse veins. Small pieces of RH leaf tissue were cut into0.07 M K-phosphate (pH 6.8) and fixed overnight in 4% (v/v)glutaraldehyde. The tissue was vacuum infiltrated with fixativefor 1 h, dehydrated in a graded acetone series, infiltrated withSpurr's resin, and polymerized at 60°C for 2 d (22). Thicklongitudinal sections (1-2 mm) were cut with glass knives andstained with 0.05% (w/v) Toluidine blue 0 in 1% (w/v) sodiumtetraborate (15).

Histochemistry. Freehand cross-sections taken midway alongthe laminas ofwinter rye plants were used to test for the presenceof a Casparian band. Sections were either stained for lipid withSudan IV or subjected to acid digestion using concentratedH2SO4 (12). Other sections were cleared using NaOH and stainedwith Chelidonium majus root extract, a technique which allowedvisualization ofCasparian bands in hypodermal cells ofcorn andonion roots (20).Dye Tracer Experiments. Leaves were cut from warm- and

cold-grown rye plants and stood in a 0.01% (w/v) aqueoussolution of Calcofluor white M2R (CFW) for 1 h, by which timethe dye had ascended to the leaf tip. Leaves were rinsed byplacing the cut bases in distilled H20 for 2 h. Freehand cross-sections were cut midway along the lamina, mounted in glycerin,

157

Dow

nloaded from https://academ

ic.oup.com/plphys/article/77/1/157/6082874 by guest on 07 D

ecember 2021

Plant Physiol. Vol. 77, 1985

and observed immediately with violet light (exciting wavelengths,361-435 nm) provided by a Nikon apophot epifluorescencemicroscope. Leaves of Phaseolus vulgaris, excised with a shortpiece of petiole attached to the blade, were treated, sectioned,and viewed in the same manner. In other experiments, 0.01%(w/v) CFW was applied to ab- and adaxial leaf surfaces whichhad been scarified with 150 grit-powder carborundum, orthrough a wound produced by stripping a subepidermal groupof fibers from the abaxial leaf surface, or by cutting the leafparallel to the venation. In all cases where CFW was appliedexternally, the leaves were rinsed 1 h in H20 prior to sectioningthe area where the dye had been applied.Another fluorescent apoplastic dye, PTS was applied as a 0.1%

(w/v) aqueous solution for 1 min through the base of an excisedlamina, followed by a 20-h rinse in H20. Segments about 50 mmlong were removed from the middle of the lamina and mountedwhole on a slide using tape on the ends to prevent leaf curling.Both ab- and adaxial leaf surfaces were viewed under violet light.Similar experiments were performed in which PTS was appliedto excised or wounded leaves as a 0.01 % aqueous solution for20 h. In other experiments, leaves still attached to the plant were

taped to a horizontal support, so that the abaxial surface wasuppermost. An area 10 mm in length was abraded with 150 grit-powder carborundum, encircled with vaseline to form a well,and treated with 0.01 % (w/v) PTS for 21 h. The excess treatmentsolution was then removed, and the leaf was excised and viewedas described above.Time sequence experiments were also conducted using PTS.

In these experiments, an excised leaf was mounted on the epi-fluorescence microscope stage so that the leaf base stood in awell and the leaf blade was parallel to a microscope slide. Thewell was initially filled with water which was replaced with 0.01%(w/v) PTS to start the experiment. The leaf blade was viewedunder violet light to follow movement of the fluorescent dye.To test for PTS toxicity in rye, healthy RNH and RH leaves

were excised, weighed, and placed in small vials containing 2 mlPTS at one of the following concentrations: 0, 0.1, 0.2, 0.5, 1.0,2.0, 5.0, and 10.0% (w/v). Each treatment was replicated fourtimes. The vials were covered to reduce evaporation from theliquid surface and weighed. The leaves were then allowed totranspire for 24 h at room temperature before reweighing thevials. The volume of liquid transpired was determined by weightloss. PTS toxicity was manifested by leaf wilting, curling, anddrying of the leaf tips.

RESULTS

Leaf Anatomy. Clearings of mature rye leaves revealed thepresence of both longitudinal veins of varying diameters andnarrow transverse veins (Fig. 1). Cross-sectional views of trans-verse veins were observed in longitudinal sections of plastic-imbedded leaf tissue. The transverse veins consisted of a singletracheary element and a few phloem cells, and did not containan encircling mestome sheath.

Histochemical Tests. Histochemical tests were performed tosee if suberized regions existed in the primary cell walls of themestome sheath. Since suberized cell walls are resistant to aciddigestion, rye leaf cross-sections were treated with concentratedH2SO4. Mestome sheath cells from both RNH and RH leavesseparated easily in the acid, thus suggesting that no suberizedarea existed in the primary cell wall. Mestome sheath cell wallsdid not stain red with Sudan IV, a lipid stain which usually reactswith suberin-associated lipids. Staining of the mestome sheathcell walls with Chelidonium root extract revealed no obviousCasparian bands; however, intense staining of the secondarylignified cell walls of the mestome sheath may have obscuredany staining in the primary cell walls.

Apoplastic Movement of Calcofluor White. When CFW was

7WU -I i

FIG. 1. Cleared rye leaf showing parallel veins, transverse veins (ar-rowheads) and numerous hairs. x30.

FIG. 2. Abaxial surface of a nonhardened rye leaf following PTStreatment, epifluorescence optics. x30.

introduced at the cut leaf base, the dye was taken up in thetranspiration stream. CFW fluorescence was visible in xylem cellwalls in a rye leaf cross-section. The dye did not appear topenetrate either phloem or mestome sheath cell walls in eitherRNH or RH rye plants. As a control, the movement ofCFWwas also followed in P. vulgaris leaves which lack a suberizedbundle sheath. In these leaves too, CFW fluorescence was seenonly in xylem cell walls, and not in the surrounding mesophylltissue.CFW was also introduced to rye leaves using various proce-

dures for wounding the epidermal layer. In most cases, the dyebound to cell walls within a few cell layers of the wound. Whensufficient dye was introduced via an epidermal wound, the dyemoved into the mestome sheath cell walls. However, the mobilityof CFW through the apoplast of rye leaves appeared to be toolimited to properly test the permeability of the mestome sheath.

Apoplastic Movement of PTS. After cut leaves had been placedin 0.2% PTS for 1 min followed by water for 20 h, dye was

visible in the leaf blade as a series of parallel lines of irregular

158 PETERSON ET AL.

Dow

nloaded from https://academ

ic.oup.com/plphys/article/77/1/157/6082874 by guest on 07 D

ecember 2021

PERMEABILITY OF THE SUBERIZED MESTOME SHEATH

Sr i2As' Sw~~~~

FI.3.HgermgIfctoofalfasi Fiur 2..:.7

FIG. 4. Adaxial surface of a hardened rye leaf 5 min after PTSapplication. The dye produces a bright fluorescence (J) in the vein (whichis faintly autofluorescent [a]). Brightly autofluorescent hairs are visibleon either side of the vein. Epifluorescence optics. x70.

width interconnected by transverse lines at infrequent intervals(Fig. 2). The same dye pattern was visible on both the adaxialand abaxial sides of the leaf. Closer inspection of the adaxial sideshowed that each longitudinal strip of dye was located in one ofthe conspicuous ridges of the leaf and was centered around thelongitudinal vein which is positioned in the center of the ridge.The transverse lines of dye corresponded in position and fre-quency to the transverse veins. By focussing on the sides of twoadjacent ridges, at higher magnification, dye could be seen in thewalls of individual cells, some of which were distant from theunderlying vascular bundle (Fig. 3). Dye associated with a trans-verse vein is seen below the focal plane in the center of Figure 3.The same dye pattern was seen when cut leaves were treated for20 h with 0.01% PTS or intact leaves were treated with 0.1%PTS on an abraded area.PTS movement during short periods oftime could be followed

by treating the base ofthe lamina on the stage ofan epifluorescentmicroscope. When viewed from the adaxial leaf surface, the first

evidence of PTS arrival in a vein about 20 mm distal to the cutend usually occurred about 5 min after application of the dye. Itappeared as a thin, green streak at the same focal plane as thevein and with the same width as the faintly autofluorescent vein(Fig. 4). Subsequent movement of the dye is shown in a timeseries of photographs of one vein following PTS treatment (Fig.5). The width of the vein, as determined by its autofluorescenceprior to treatment, is indicated by a horizontal bar in Figure 5,a, b, d, and e. Within 2 min after its arrival in the vein, the dyebegan to spread outward from the vein so that the width of thegreen strip increased and the outlines of cells at a focal planehigher than that of the vein were visible (Fig. 5a). As the dyecontinued to move out of the vein (6 min after its first appear-ance), its intensity increased (Fig. Sb). At higher magnification,the dye could clearly be seen in the walls of the epidermis whichis composed of elongate, tabular epidermal cells altering withrectangular subsidiary cell-guard cell groups (Fig. Sc). Increasingmovement of the dye away from the vein was apparent 30 and60 min after its application to the leaf base (Fig 5, d and e,respectively). No further dye movement was apparent after 60min, by which time the pattern was the same as shown at lowermagnification following a 20 h incubation (Fig. 2). Regions ofthe leaf which had not been viewed at intervals with violet lightshowed the same results, indicating that the rapid dye movementwithin the leafwas not brought about by exposure to violet light.The rate and pattern of dye movement were similar in leaves ofRNH and RH plants.We also inferred apoplastic movement ofPTS inward through

the mestome sheath by applying PTS to an abraded area on thesurface of an intact leaf. In these experiments, PTS could beobserved moving upward in the vein above the point of appli-cation, and subsequently diffusing outward through the mestomesheath along the length of the veins.The results of PTS toxicity tests showed that RNH or RH

leaves invariably showed symptoms of injury when they hadtaken up 10 mg PTS/g leaf (fresh weight). Leaves were notinjured by doses of PTS less than 5 mg PTS/g leaf for RH and 8mg PTS/g leaf for RNH. At intermediate doses, some leavesexhibited a toxic response while others did not. In the PTSexperiments described above, the average RNH leaf treated with0.01% PTS for 20 h transpired 2.8 ml of treatment solution/gleaf, resulting in a total mean uptake of 0.28 mg PTS/g leaf.Similarly, the average RH leaf transpired 2.1 ml treatment solu-tion/g leaf, resulting in a mean uptake of 0.21 mg PTS/g leaf.RNH or RH leaves treated with 0.1% PTS for 1 min wouldtranspire about 2 1l of treatment solution, which representedonly 2 ,g PTS/g leaf. Thus, the amounts of PTS taken up by ryeleaves during the tracer experiments were far below the minimumtoxic dose of PTS.

DISCUSSIONSuberization of bundle sheath cell walls has been observed in

leaves of many grasses (9), yet the function of suberized bundlesheaths has yet to be elucidated. One possible function is toregulate solute transport by restricting apoplastic movement asobserved in the root endodermis. Several authors have attemptedto trace the apoplastic pathway of grass leaves using stains visiblein the electron microscope. Kuo and coworkers (14) observedthat ferric chloride was restricted to movement in the vasculartissue of both longitudinal and transverse veins in wheat leaves,and concluded that vessel walls restricted solute movement.Botha and coworkers (3) observed that the movement of ferrousions was restricted to vascular tissues and bundle sheath cells inThemeda triandra. They concluded that the suberin lamellae ofthe outer bundle sheath cell walls, but not the vessel walls,restricted movement ofsolutes into the mesophyll. In either case,the iron ions appear to have limited mobility in the leaf apoplast.

159

Dow

nloaded from https://academ

ic.oup.com/plphys/article/77/1/157/6082874 by guest on 07 D

ecember 2021

Plant Physiol. Vol. 77, 1985

FIG. 5. Series of photographs of the same area of the adaxial surface of a hardened rye leaf treated with PTS. Horizontal bars indicate the widthof the underlying vein. Epifluorescence optics. a, b, d, and e, x70; c, x 130. Time elapsed since start of PTS treatment: a, 7 min; b, I I min; c, 1 1.5min; d, 30 min; e, 60 min.

At the level of the light microscope, O'Brien and Carr (17)reported that an aqueous solution of basic fuchsin movedthrough the vascular bundle of wheat leaves, and then spreadlaterally to subepidermal fibers within minutes. They found thatdyes such as alcian blue, fast green FCF, and acid fuchsin causedwilting and made transport studies difficult.We also obtained conflicting results using fluorescent dyes. If

we used CFW, a fluorescent dye which moves apoplastically andbinds to cell walls, then we found that the dye was restricted tomovement through only a few cell layers. While we did observerapid uptake of the dye in the transpiration stream, we did notsee lateral movement of CFW into either bundle sheath ormesophyll cell walls. We concluded that mobility ofCFW in theapoplast was too limited to properly test movement of solutesout of the transpiration stream. PTS, on the other hand, is anonbinding fluorescent dye which is mobile within the apoplast.Although this mobility restricts observation to intact leaves ratherthan cross-sections, the movement of PTS through cells can beeasily detected due to its bright green fluorescence. PTS movedrapidly through the veins of rye leaves. Within minutes ofreaching a particular level in the leaf, PTS would diffuse laterally

from the vascular bundles to the epidermal cell walls. We ob-served movement of PTS from vascular bundles of transverseveins which have no suberized mestome sheath, from longitudi-nal vascular bundles of rye leaves grown at 20C which have asuberized mestome sheath, and from longitudinal vascular bun-dles of rye leaves grown at 5°C which have thicker mestomesheath cell walls and greater amounts of suberin (7). In all cases,rates of PTS movement through the veins and diffusion outwardto the epidermal cells were the same. Our results with rye do notagree with those of Tanton and Crowdy (24) who reported that,in wheat, the transverse veins allowed lead chelate to move intothe mesophyll whereas the longitudinal veins did not. However,our results are in complete agreement with those of Strugger (23)who followed the movement of several apoplastic dyes in ryeand wheat leaves. We could also confirm his observation thatwhen PTS first appeared in the epidermal anticlinal walls, thedye was brighter in some portions of the walls than others.Strugger (23) correlated the positions of the bright spots with thepositions of the anticlinal walls of the underlying palisade cellsthrough which the PTS apparently moved on its way to theepidermis.

160 PETERSON ET AL.

Dow

nloaded from https://academ

ic.oup.com/plphys/article/77/1/157/6082874 by guest on 07 D

ecember 2021

PERMEABILITY OF THE SUBERIZED MESTOME SHEATH

The apoplastic pathway for movement of solutes through themestome sheath must be through the radial primary cell walls.In plants such as corn and T. triandra where the suberin lamelladoes not completely encircle the cell but is present on the radialand outer tangential walls, tracer entering the secondary wallsfrom the vein may become trapped and eventually concentratednear the suberin lamellae. Tracer moving within the primarywall would not be trapped and would be much less conspicuous.This could explain why Botha et al. (3) located ferrous iron insidethe suberin lamellae in T. triandra whereas, in our preliminaryexperiments, PTS movement in corn leaves was the same as inrye.Using histochemical techniques, we found no evidence for the

presence of suberin in the primary cell walls as often observedwith Casparian bands in the root endodermis. Bocher and Olesen(2) documented "outbulgings in walls of bundle sheath cellsresembling Casparian strips" at one stage of development ofSporobolus rigens; however, we did not observe these structuresin the mestome sheath cell walls in rye leaves. O'Brien and Carr(17) reported that suberized areas did exist in the radial primarycell walls between mestome sheath cells; however, they treatedleaf sections with silver hexamine without pretreatment withperiodate oxidation. Eleftheriou and Tsekos (5) have reportedthat the silver hexamine stain is only specific for suberin whensections are pretreated with periodate. In winter rye leaf sectionsstained with silver proteinate, an ultrahistochemical stain specificfor suberin (10), suberin lamellae were observed in the secondarywall of each mestome sheath cell, yet no evidence of suberizationof the primary cell wall was observed (7). We conclude that themestome sheath ofwinter rye leaves does not contain a Casparianband and is freely permeable to solutes moving apoplasticallythrough the radial primary cell walls.

Acknowledgments-We thank Ann Eastman for excellent technical assistance;Dr. G. McLeod, Agriculture Canada, Swift Current, Saskatchewan, for providingPuma rye seed; and the Bayer Co. for their gift of PTS.

LITERATURE CITED

1. ARNOTT HJ 1959 Leaf clearings. Turtox News 37: 192-1942. BOCHER TW, P OLESEN 1978 Structural and ecophysiological pattern in the

xero-halophytic C4 grass, Sporobolus rigens (Tr.). Desv Biolog Skrift 22: 483. BOTHA CEJ, RF EVERT, RHM CROSS, DM MARSHALL 1982 The suberin

lamella, a possible barrier to water movement from the veins to the mesophyllof Themeda triandra Forsk. Protoplasma 112: 1-8

4. CROWDY SH, TW TANTON 1970 Water pathways in higher plants. I. Free

space in wheat leaves. J Exp Bot 2: 102-1115. ELEFTHERIOU LP, I TSEKOS 1979 Development of mestome sheath cells in

leaves ofAegilops comosa var. thessalica. Protoplasma 100: 139-1536. GRIFFITH M, GN BROWN 1982 Cell wall deposits in winter rye Secale cereale

L. 'Puma' during cold acclimation. Bot Gaz 143: 486-4907. GRIFFITH M, NPA HUNER, KE ESPELIE, PE KOLATTUKUDY 1985 Lipid poly-

mers accumulate in walls ofepidermal and mestome sheath cells during lowtemperature development of winter rye leaves. Protoplasma. In press

8. GUNNING BES, JE HUGHES 1976 Quantitative assessment of symplastic trans-port of prenectar into the trichomes of Abutilon nectaries. Aust J PlantPhysiol 3: 619-637

9. HATrERSLEY PW, AJ BROWNING 1981 Occurrence of the suberized lamella inleaves of grasses of different photosynthetic types. I. In parenchymatousbundle sheaths and PCR ("Krantz") sheaths. Protoplasma 109: 371-401

10. HOLLOWAY PJ, CA BROWN, J WATrENDORF 1981 Ultrahistochemical detectionof epoxides in plant cuticular membranes. J Exp Bot 32: 1051-1066

11. HUNER NPA, FDH MACDOWALL 1976 Chloroplastic proteins of wheat andrye grown at warm and cold-hardening temperatures. Can J Biochem 54:848-853

12. JOHANSEN D 1940 Plant Microtechnique. New York: McGraw-Hill13. KROL M, M GRIFFITH, NPA HUNER 1984 An appropriate physiological control

for environmental temperature studies. Comparative growth kinetics ofwinter rye. Can J Bot 62: 1026-1068

14. Kuo J, TP O'BRIEN, S-Y ZEE 1971 The transverse veins ofthe wheat leaf. AustJ Biol Sci 25: 721-737

15. MILLONIG G 1976 Staining solutions for light microscopy. In CP Vercelli, ed,Laboratory Manual of Biological Electron Microscopy. Mario Saviolo, Italy,pp 57-59

16. MYLIUs G 1913 Das Polyderm. Eine Vergleichende Untersuchung uber diephysiologischen Scheiden: Polyderm, Periderm und Endodermis. BibliothBot 18: 1-119

17. O'BRIEN TP, DJ CARR 1970 A suberized layer in the cell walls of the bundlesheath of grasses. Aust J Biol Sd 23: 275-287

18. PETERSON CA, RL PETERSON, AW ROBARDS 1978 A correlated histochemicaland ultrastructural study of the epidermis and hypodermis of onion roots.Protoplasma 96: 1-21

19. PETERSON CA, ME EMANUEL, GB HUMPHREYS 1981 Pathway ofmovement ofapoplastic fluorescent dye tracers through the endodermis at the site ofsecondary root formation in corn (Zea mays) and broad bean (Viciafaba).Can J Bot 59: 618-625

20. PETERSON CA, ME EMANUEL, C. WILSON 1982 Identification of a Casparianband in the hypodermis of onion and corn roots. Can J Bot 60: 1529-1535

21. PETERSON CA, CJ PERUMALLA 1984 Development of the hypodermal Caspar-ian band in corn and onion roots. J Exp Bot 35: 51-57

22. SPURR AR 1969 A low-viscosity epoxy resin embedding medium for electronmicroscopy. J Ultrastruct Res 26: 31-43

23. STRUGGER S 1939 Studien uber den Transpirationsstrom im Blatt von Secalecereale and Triticum vulgare. Z Bot 35: 97-113

24. TANTON TW, SH CROWDY 1972 Water pathways in higher plants III. Thetranspiration stream within leaves. J Exp Bot 23: 619-625

25. WEERDENBURG CA, CA PETERSON 1984 Effect of secondary growth on theconformation and permeability of the endodermis of broad bean (Viciafaba), sunflower (Helianthus annuus) and garden balsam (Impatiens balsa-mina). Can J Bot 62: 907-9 10

161

Dow

nloaded from https://academ

ic.oup.com/plphys/article/77/1/157/6082874 by guest on 07 D

ecember 2021