THE ROLE OF CALCIUM IN SELECTIVE CATION UPTAKE

BY PLANT ROOTS

II. E旺ectsof Temperature， De田rptionTreatment and Sodium Salt on Rubidium Uptake*

Toshio KA WASAKI and Shiro HORI紳

In the previous paper (1)， the effect of calcium on the selective uptake of monovalent cations was studied in relation to monovalent and divalent cation concentrations in the ab田 rptionsolution. Data prel:抱ntedin that paper showed that the uptake of rubidium by plant roots was stimulated in the presence of calcium at a relatively high concentration of monovalent cations in the absorption solution. However， the uptake of sodium was inhibited by calcium at both the low and high concentra-tion of monovalent cations. Further， it was suggested that the site of stimulation of calcium on rubidium uptake might be located in the metabolic process of ion uptake.

The pre田 ntinvestigation was undertaken to extend the results of the previous paper， and to study in more detail the mechanism(s) of the calcium effect on the selective uptake of monovalent cations by plant roots. For this purpose， the calcium effect was examined at different experimental temperatures， with and without the desorption treatment of roots， and in the presence and absence of sodium in the a凶orptionsolution.

MATERIALS AND METHODS

Pre抑:ration01 Root Mαt俳句ls

Excised roots of barley (Hordeum vulgare L.， variety Akashinriki) were used. Root materials were prepared according to the pr∞吋ure

described previously (1).

EゆerimentalPr侃 edure

To inv田 tigatethe effect of calcium or magn回 iumon the uptake of

rubidium and sodium， equimolar mixtures of rubidium chloride and

sodium chloride were used as the absorption solution. In experiments

on the effect of sodium on rubidium uptake， absorption solutions con-sisted of a constant concentration of rubidium chloride and varying

* Data presented in this paper were published in the Journal of the Science of Soi1 and Manure， Japan， Vo1. 41， p. 149-154 (1970)， in Japanese.

柿Departmentof Applied Biology， Radiation Center of Osaka Prefecture.

20 T. Kawasaki & S. Hori

∞ncentrations of sodium chloride. Absorption solutions were labeled with radioactive rubidium (86Rb) or radioactive sodium (22Na). The initial pH of the absorption solutions was adjusted to 5.4"，-，5.7.

The procedur田 forthe absorption experiment and the washing operation for roots were virtually indentical with those described in the previous paper (1). 1n a11 experiments. 0.5 g of excised r，∞ts were used in 500 ml of absorption solution. The absorption period was 60 minutes， during which the solutions were continuously aerated and kept at 25土0.10C or 2土O.50C.

At the end of an absorption period， the r，∞ts were田 paratedfrom the solution. Then the roots were blotted with filter paper and shaken for 30詑 condsin 30 ml of demineralized water. This washing operation was repeaぬdonce more.

R∞ts of certain plots underwent desorption treatment， iιafter washing the roots were shaken repeatedly for 30 minutes in 50 ml of d回orptionsolution. then were blotted with filter paper. D白 orptionsolutions used in the experiment on the effect of divalent cations on the uptake of monovalent cations had the回 mecomposition of monovalent cations as the absorption solution. 1n the experiment on the eff配 tof sodium on the uptake of rubidium， a rubidium chloride solution (1.0 mM) was used as the desorption solution. All desorption solutions we四non-radioactive， and no divalent cations were added to them.

Rσdioactive Assay

The pr∞edure for the radioactive as回 ywas identical with that d問 crib吋 inthe previous paper (1). Root samples were dried and ignited. One ml of dilute nitric acid solution was added to the ash and the samples were dried. Then， the samples were counted with a Geiger-Muller counter.

RESULTS

Influences 01 Experimental Temperature and Desorption Treatment on the Calcium Effect

Effects of divalent cations on the uptake of rubidium and sodium by excised barley r∞ts from equimolar mixtures of rubidium chloride and sodium chloride were investiga也dunder various conditions， with and without the desorption treatment of roots after an absorption period at 250C and 20C.

Figs. 1 and 2. respectively， show the rat田 ofrubidium uptake from absorption solutions containing 0.001 and 1.0 mM of monovalent cations. Num加rson bars indicate the ratios of the results when calcium chlorde or magnesium chloride was added to that when there was no addition

m -2 L

4回

....'" -_， 吾て 0.15 ω 』。

国 Eミえ

Divllent cations in abaorption solution

Role of Calcium in Selective Cation Uptake (11) 21

Temperature :25 'c Temperature :2t

0.30ト 100 100 0.10 ~ 100 100

77 73 74 71

0.05

a Mg

: 0 0.5 0.5 0 0.5 0.5 mM 、--旬申ーーJ 、ーーー一一~N開・ Desorption desorption

。、-ーーーー__，Non-de岡市tion

、一一一一_JDeaorption

Fig. 1. Effects of divalent cations on the rubidium uptake with or without the desorption treatment. (Rubidium chloride and sodium chlorlde in absorption solution: O.∞1 mM)

E

。E国的

.0¥z-。Eミ

-ωA

・】-az』国

Divalent c・tIonsin absorption solution

Temperature: 25.C

5.0 138 Temperature:2'C

2.0ト10095

165 119

100

クイ先.t.(.

c. Mg Ca Mg

o 0.5 0.5 .0 0.5 0.5 mM 、一ーーーー一一一_/、ーー一ーーー一ー'-./N輔・ De・orptiond・回rption

86 3.0

1.0

1.0

H5JI

Mo--刷

a5一

巾

co-r附

-O則

。一N・曲

Ca Mg o 0.5 0.5 mM 、ーー一一一一一一一」Desorption

Fig. 2. Effects of divalent cations on the rubidium uptake with or without the desorption treatment. (Rubidium chloride and sodium chlorlde in absorption釦 lution:1.0 mM)

of divalent cations. The same system is used in the figures d白 cribedbelow， except for Figs. 5 and 6. At the lower concentration (0.∞1mM) of monovalent cations in the absorption solution， the rubidium uptake decrl聞記d in the prl田 enぽ ofcalcium or magn偲 iumchloride， with or without the d回 orptiontrl回 tmentat 250 and 20C. At the relatively high concentration (1.0 mM) of monovalent cations， the rubidium uptake increased in the presen句 ofcalcium and magn田 iumchlorides both with and without the desorption treatment at 250C. At 20C， however， the rubidium uptake decreased in the pr回 enceof calcium and magnesium chlorides when there was no desorption treatment， whereas it incr，回記dwith desorption treatment. Further， the stimulating effect of calcium chloride on rubidium uptake was larger than that of magn田 iumchloride.

22 T. Kawasaki & S. Hori

The rates of sodium uptake from absorption solutions containing 0.001 and 1.0 mM of monovalent cations are shown in Figs. 3 and 4，

aa.'冨

ay

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υ

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Temperature :2"C 100

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m

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h

・・i唱

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と土日さと土日二5皿MN帽・ Do.orption dOlOrption

Fig. 3. Effects of divalent cations on the sod.ium uptake with or without the desorption treatment. (Rubidium chloride and sod.ium chloride in a凶orptionsolution: 0.001 mM)

.. 2 h

d岡

京.:1 ZE

"l

Div・lentutions」μ1ME Ca ME in _ ab.orption :ρ0.5・0.5 ‘o0.5 0.5.mM S刷 utlOn '-一一一一一ー }一一一一一ーNon- Dosorption

do.orption

Temperature: 25 "c FU-

--n4-

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1

100 100

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1.5

Ca Mg Ca Mg o 0.5 0.5 0 0.5 0.5皿M¥一一一一一一ー__/~ーーーーー一.C..JNon- Do.orption desorption

100

0.4

Fig. 4. Effects of divalent cations on the sαiium uptake with or without the de田 rptiontreatment. (Rubidium chloride and sod.ium chloride in a回orptionsolution: 1.0 mM)

respectively. Sodium uptake decr回 sedin the pre田 nceof calcium and magnesium chlorid田 underall the conditions of the experiments; at the low and high concentrations of monovalent cations in the absorption solutions， with or without the d田 orptiontreatment of roots after the absorption period， and at bothお。 and20C.

lnfluence 01 Sodium Salt on Rubid.um Uptake in Relation to the Presence and Absence 01 Calcium

The effects of increasing concentrations of sodium chloride on rubi-dium uptake from a constant concentration of rubidium chloride solution were examined in the absence of calcium.

Role of Calcium in Selective Cation Uptake (11) 23

Fig. 5 shows the rates of rubidium uptake from an absorption solution which contained 0.001 mM of rubidium chloride and 0......1.0 mM of sodium chloride， at 250 and 20C. Fig. 6 shows the rat田 ofrubidium

Temperature: 25 'c 0.30

100 99 98 100 99 99

8

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京 0.15

-'"・=1 ミ

87 87

N.ce in .b.。叩tion: 0 O.ω1 0.01 0.1 1.0 0O.ω10.01 0.1 1.OmM .oluti酬 '--=-一一一一宇J ‘ ，

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N輔・

4・-.<<1蝿 Desorption

Fig.5. Eff，回tof increasing concentrations of sodium chloride on the rubidium uptake with or without the desorption treatment. (Rubidium chloride in aboorptionωlutlon: O.∞1 mM)

Temperatu~e: 25 'c

5.0

1ω 93

100 腕

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別冊・ Desorption 品-圃H踊

Fig. 6. Eff，田tof increasing concentrations of 5(出umchloride on the rubidium uptake with or without the de田，rptiontreatment. (Rubidium chlor1de in aboorption solution: 1.0 mM)

uptake from an absorption solution which contained 1.0 mM of rubidium chloride and 0-10.0 mM of sodium chloride. Numbers on bars indicate the ratios of the r，白u1tswhen varying concentrations of sodium chloride were added to the results of no addition. At both the low (0.001 mM) and relatively high (1.0 mM) concentrations of rubidium chloride， the rubidium uptake d配 rea田d with increasing concentrations of sodium chloride in the absorption回 lution，with or without the d缶 orptiontr，回tmentat 250 and 20C.

Subsequently， in order to determine the role of sodium in the calcium eff.配t，a compari回 n was made betw関 n the rubidium uptake from a single salt solution of rubidium chloride (1.0 mM) and from an equimolar

24 T. Kawasaki & S. Hori

mixture of rubidium and sodium chloride (1.0 mM each) in the presence and absence of calcium.

Figs. 7--9 show data from the experiments at various temperatur田.

These results showed that even when the temperature during the absorp-tion period was 250C. the calcium effect on the rubidium uptake from a single salt solution of rubidium chloride was ni1 or very small both with and without the d回 orptiontr回 tment. However， the calcium effect

円 . 126 100 100 B

z当5F2¥ i 5.0

::t

Ca.b。olsmuot凹rap。stnii。tin。n。f: Rb Rb+Na Rb Rb+Na

Noor desorption

Desorption

Fig. 7. Rubidium uptake from rubidium chloride and rubidium plus sodium chloride solutions in the presence and ab鑓 nceof calcium. (Rubidium chloride or sodium chloride ln 油田rptionsolution: 1.0 mM; Temperature: ab鈎 rption250C， washing 250C. desorption 250C)

c:=コ CaCl.in absorption solution: 0 mM ・・・・ CaClt in ab田 rption田 lution:0.5 m M

100 99 118

輔同

7.51 n・1・2・8 100. ・・I 144 f

三宅5.0

2.5 f.

::t

Composition 01 Rb absorption Rb+Na Rb Rb+Na

solution N輔・

CIe胸叩lIonDe剖rpti帽

Fig. 8. Rubidium uptake from rubidium chloride and rubidium plus sodium chlorideωlutions in the presence and absence of calcium. (Rubidium chloride or sodium chlorlde in absorption田 lution:1.0 mM; Temperature: absorption 250C. washing 20C. de田 rption20C)

仁二二コ CaClt in absorption solution: 0 mM ・・・ CaCl.in absorption solution: 0.5 mM

Role of Calcium in Selective Cation Uptake (11) 25

on the rubidium uptake from a mixture of rubidium and sodium chlかrides was clear. Table 1 summarizes the data shown in Figs. 7-9 based

Composition 01 absorption 8olution -J

N

一-"H

aT-

。

ー

M

・E

-D一品川

R一町-。-. -e

hu

一nUR一

・・4Mn-

+-m

b

-

U

R一

下-mro

同時

g

b

一Nh

RH

一

Fig. 9. Rubidium uptake from rubidium chloride and rubidium plus sodlum chloride solutions in the presence and abi割問ceof 伺 lcium. (Rubidium chloride or sodium chloride in 油田rption田 lution:1.0 mM; Temperature:油田rption20C. washing 20C. de!拘rption2・C)

E二コ CaClzin absorption solution: 0 mM ・・・・ CaCl. in absorption solution: 0.5 mM

TABLE 1

Repr田siveeffect of反xiiumon rubidium uptake in the pre田 U偲 andabsence of calcium

A国orption:250C Absorption: 250C Absorption: 20C Washing : 250C Washing : 20C Washing : 20C Desorption :話。C De釦rption:20C Desorption: 20C

Rb Rb+Na Rb Rb+Na Rb Rb+Na

100 66 1∞ 66 1∞ 75 Non-de田 rption

100 87 100 86 1∞ 88 +Ca

100 63 1∞ 66 1∞ 81 De田rption

+Ca 100 87 100 81 100 92

on the ratio of the rubidium uptake from a rubidium plus sodium chloride solution to that from a single salt solution of rubidium chloride.

Under all the ex関 rimentalconditions， the rubidium uptake decreased on the addition of sodium to the absorption solution. However， the repressive effect of sodium on the rubidium uptake was less when cal-

cium was pre田 ntin the absorption solution than when it was absent.

26 T. Kawasaki & S. Hori

DISCUSSION

Before discussing the above data， we should consider the character-istics of the exchang朗 bleand non-exchangeable fractions of ions in plant roots. Some studies have been done on the passive and active proce話回

of ion uptake by plant r∞ts (2， 3). From the results， it has generally been accepted that ions gained by the active proc回 sof ion uptake are largely non-exchangeable with ambient ions of the same or other species (4). Thus the non-exchangeable fraction of ions taken up by plant roots must be related to the active (metabolic) pr∞ess of ion uptake，

and the exchang伺 blefraction to the passive (non-metabolic) proc田s. In the present paper， the ions retained in the roots after the desorption treatment correspond to the non-exchangeable fraction， and the ions taken up in the r∞ts without the desorption treatment after washing corres卯 ndto the sum of the non慌 changeableand exchangeable fractions.

It has 国enreported that calcium incrl回 sedthe affinity between pota罰 ium(or rubidillm) ion and its carrier， based on the ion-carrier complex thωry (5-7). In the results of this investigation also， the stimulation of rubidium uptake by divalent cations (es開ciallythe calcium ion) was larger with the desorption treatment than without the desorp-tion treatment (Fig. 2). Since the non-exchangeable fraction of ions taken up by plant r，∞ts is closely related to the active (metabolic) pr∞ess of ion uptake， the stimulating e旺'ectof calcium on the rubidium uptake is thought to be dependent on the act1ve (metabolic) pr∞ess of ion uptake. Furthermore， since the metabohc activity of living cells decreases at low temperatures， the fact that the calcium effect on rubidium uptake was reduced or abolished by low temperature treatment (20C) supports the above hypoth田 is(Fig. 2).

On the other hand， it was found that the calcium effect on the rubidium uptake from a rubidium plus sodium chloride solution was greater than that from a single salt solution of rubidium chloride (Figs. 7"，9). These results suggest that sodium plays an important role in the mechanism(s) of the calcium e百ecton rubidium uptake. Although the rubidium uptake decreased on the addition of sodium， whether in the pr'田 enceor absence of calcium (Figs. 7--9)， this repression of the rubidium uptake by sodium was smaller in the presen句 ofcalcium than in its absence (Table 1). Jaco凶onet al. (8， 9) and Waisel (10) suggested that the stimulating effect of calcium on potassium or rubidium uptake was due田sentiallyto blockage of interfering ions.

Though the uptake of potassium or rubidium was reduced by the addition of sodium (11)， both carriers (or carrier sites) for potassium or rubidium were posited， which were diffieultly and easily combined by sodium (12-14). In this paper， the rubidium uptake decreased with

Ro1e of Ca1cium in Se1ective Cation Uptake (Il) 27

incI官~asing concentrations of sodium chloride， but the inhibition by sodium was not equivalent to rubidium. In addition， some reports showed that the sodium uptake decreased markedly in the pre田 nceof calcium (1， 9， 10， 14-16). Accordingly， it is reasonable to consider that， as a cau田of the stimulation of rubidium uptake by calcium， calcium reversed the reprl白 sionof rubidium uptake by sodium.

SUMMARY

Effects of calcium on the田lectiveabsorption of monovalent cations were studied under various conditions. The role of sodium in the stimu-lating effect of calcium on rubidium uptake was also inv田 tigated.

1) At a low concentration (0.001 mM) of monovalent cations in an equimolar mixture of rubidium chloride and sodium chloride， the rubid-ium and田 diumuptake decreased in the presence of calcium or magne-sium chloride， both with and without the d田 orptiontreatment of plant r∞ts at 250 and 20C.

2) A t a higher concentration (1.0 mM) of monovalent cations， the sodium uptake decreased at 250 and 20C， and the rubidium uptake increased at 250C in the pre田 nceof calcium or magnesium chloride with or without the desorption treatment. However， at 20C， calcium and magnesium chlorid田 d配 rea田 dthe rubidium uptake without the desorption treatment of plant roots， but increased it with the desorption treatロlent.

Stimulation of the rubidium uptake was greater in the pre舘 nceaf calcium than in the pre田 nceof magn白 ium.

3) The stimulating effect of calcium was more marked when sodium was pr田 entin the absorption solution.

4) 1t has been posited here， as noted in the previous paper， that the calcium effect on the rubidium up回kewas dependent on the meta-bolic process of ion uptake. Moreover， the results support the contention that sodium plays an important role in the mechanism(s) of calcium d配t.

REFERENCES

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2. Epstein. E. and Leggett. J. E. 1954. The ab回 rptionof a1kaline earth cations by bar1ey r∞ts: Kinetics and mechanism. Am. J. Botany 41: 785-791.

3. Gauch. H. G. 1957. Minera1 nutrition of p1ants. Ann. Rev. P1ant Physio1. 8: 31-64.

28 T. Kawasaki & S. Hori

4. Esptein， E. 1955. Passive permeation and active transport of ions in plant r∞ts. Plant Physiol. 30: 529-日5.

5. Kahn， J. S. and Han田 n，J. B. 1957. The effect of calcium on potassium accumula-tion in corn and釦 ybeanr∞ts. Plant Physiol. 32: 312-316.

6. Han回 n，J. B. and Kahn， J. S. 1957. The kinetics of potassium accumulation by ∞rn r，∞ts as a function of cell maturity. Plant Physiol. 32: 497-498.

7. Tanada， T. 1962. Localization and mechanism of calcium stimulation of rubidium absorption in the mung bean r∞t. Am. J. Botany 49: 10伺ー1072.

8. Jacobson， L.， M∞re， D. P. and Hannapel， R. J. 1960. Role of calcium in 油田rptionof monovalent cations. Plant Physio1. 35: 352-3田.

9. ]a∞bson， L.， Hannapel， R. ].， M∞re， D. P. and Schaedle， M. 1961. Inf1uence of 伺 lciumon selectivity of ion absorption pr∞ess. Plant Physio1. 36: 58-61.

10. Waisel， Y. 1962. The effect of Ca on the uptake of monovalent ions by excised bar1ey r∞ts. Physio1. Plantarum 15: 709-724.

11. Sutcliffe，]. F. 1957. The selective uptake of alkali cations by red beet r'∞t tissue. J. Exp. Botany 8: 36-49.

12. Epstein， E. and Hagen， C. E. 1952. A kinetic study of the absorption of alkali cations by barley r∞ts. Plant Physiol. 27: 457-474.

13. Bange， G. G.]. 1959. Interactions in the potassium and sodium ab田 rptionby intact maize鑓 edlings. Plant Soil 11: 17-29.

14. Epstein. E. 1961. The essential role of calcium in sel即日vecation transport by plant cells. Plant Physiol. 36: 437-444.

15. Handley， R.， Metwally， A. and Overstr，田t，R. 1965. Divalent cations and the per-m国民lityto Na of the r∞t meristem of Zea mays. Plant Soil 22: 200-206.

16. Handley， R.， Metwally， A. and Over百紅白t，R. 1965. Effects of Ca upon metabolic and nonmetabolic uptake of Na and Rb by r∞t segments of Zea muys. Plant Physiol. 40: 513-520.