Effect of Li and of other ions on Na transport in epithelial cells of frog skin

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Biochemical Pharmacology, Vol. 29, pp. 2265.2268. 0 Pergamon Press Ltd. 1980. Printed in Great Britain.

EFFECT OF Li AND OF OTHER

IONS ON Na TRANSPORT

Thomas Department

of Physiology,

12 April

We have used the isolated investigate

the effect

We found that addition

whereas

addition

trol conditions,

damage

chloride

surements, monatomic

bathing

the Naf transport

This allowed

Except

for the brief

across

the epithelial voltage

cells

setup has the advantages to active

the current

measured

perimental

direct determination

design

the absorptive

allowed

proceeds

using standard The change

direction

techniques

across

involving

This suggested

due to alterations

that the changes

in the properties

potential

was

of the apical 2265

cells could be

short-circuit across

current

the cells

and that (IO), is

(2).

Na+ fluxes,

of Na+ (Na efflux)

spectrometry

reversible

namely

which

with 22Na by

(I,3). after the introduction upon return

to control

of Na+ (i.e. Na influx)

cell membrane,

The ex-

side of the cells

The Na+ fluxes were measured

in absorption

for Na+ re-

were not present)

to the basolateral

readily

by applying

This experimental

the epithelial

in this study began immediately

bath and

of a

the potential

the experiment

of the two unidirectional

liquid scintillation

in Na+ flux observed

of the test salt into the external conditions.

of Na+ across

the cells.

amount

of a given ion changed

the experiment.

bath to "blood side" bath) and the backflux

in opposite

by an equimolar

the electrochemical

from the apical

100 mM

of the control mea-

for bath changes,

no other ion is transported

Na+ flux (Na influx)

(i.e. from external

needed

the so-called

provided

Under con-

15 mM on both sides of the epithelial

(other forces for net Na + movement

equal to net Na+ movement

(1).

15 mfl NaCl,

the addition

cells during

under these conditions,

is specifically

side) and the external

at zero throughout

Therefore,

that any net movement

transport

in Na+ absorption

completion

was replaced

remained

(PD) was maintained device.

in epithelial

to the chamber

containing

After

us to examine whether

equal on both sides of the epithelial

assigned

it is attached

(~5 set) interruptions

clamping

a decrease

to

in Na+ absorption.

cells were identical,

when the Na+ concentration

cells.

mained

ions on Na+ transport

(chCl), 2.5 mM KHC03 and 1 mM CaC12.

test salt.

as a model membrane

the frog skin in a chamber which

solution

23298

1980)

the blood side (basolateral

the 100 mM chC1 in the external

an automatic

2 May

Virginia

of -Rana pipiens

in an increase

to the tissue where

the solutions

Richmond,

of Li+, K+, Rb+ and Cs+causes

side (apical side) of the epithelial choline

1980; accepted

were done by mounting

to prevent

of Virginia,

frog skin preparation

of Br- and I- results

The experiments

College

of Li+ and of other monatomic

cells.

designed

U.L. Biber and Terry L. Mullen

Medical

(Received

CELLS OF FROG SKIN

IN EPITHELIAL

particularly

could be

since, under

2266

Preliminary

many conditions, limiting

the entry of Nat across

step for absorption

The relative order,

ability

except

(see Figure

in presence

the entire epithelium

of the substituent

that the stimulation

(the changes

the apical cell membrane

of Na across

Li+>Kt>Rbt>Csf>cholinef

der, I->Br->Cl-

Communications

and the relative

ability

(Figure port.

of changes 1).

of Na efflux was larger in presence

major

(64 observations

in Na influx

Lit is transported

side) across steps:

purpose

tested,

Lit produced

whereas

of changes

the strongest

in the inward direction

to basolateral

involving

at least two

the apical cell membrane

:.: ::: :_ I Previous

experiments

of Lit, the

of Nat trans-

cell membrane

It has been known for some time that Na and Li uptake across related.

(Li uptake)

on the other the apical

have shown that the Na uptake ex-

n Na INFLUX q CURRENT +a0

fg 1

plJ

q NO EFFLUX

Cl

KCI

RbCl

CSCI

ChBr

un-

in IO and in PD

(i.e. from apical

the basolateral

tested,

the Na influx was

inhibition

cells of the frog skin (6) by a process

of I-

For example,

With the exception

to the sequence

of Lit from the cell across

are closely

or-

equal to the Na

of the Na influx.

for each average).

(1) the entry of Li ' into the cells across

side of the cell. cell membrane

Under the conditions

was for all practical

is identical

actively

the epithelial

and (2) the extrusion

of Br- than in presence

the Na efflux was 0.023 i 0.002 peq/cm'hr,

Of all the cations

was, in decreasing

In general, changes in Na efflux followed the same pattern

1).

of Rb+ and Cst were not significant).

1.004 2 0.048 Peq/cm2hr sequence

(4,5).

to stimulate

influx since the Na efflux was only a very small fraction conditions

is the rate-

ions to inhibit Na influx was, in decreasing

the net Na flux (Na influx minus Na efflux)

der control

(Na uptake)

Chl

Figure 1. Average percentage changes ? SEM after substitution of 100 mM chC1 by different Number of observations for Na influx, Na efflux, PD and short-circuit current test salts. measurements was 8, 16, 24 and 24, respectively.

Preliminary

hibits

saturation

kinetics

and can be described

Na uptake = I (Jm where Jm is the maximal ability

Naext)/(KNa + Na,,,)}

pared to the saturating

describes

component

take is exclusively tenable magnitude

gradient

is substantially

the external

Figure

bath (9,lO).

showed

1 indicates

transport

that Li+ proceeds

of Li

elude that Li+ absorption conditions

increased

their observation

from the elec-

potential

bath (8).

of Li+ across

the basolateral

proceeds

against

In

cell

an elec-

bladder

gradient

as Na (11).

in more proximal

Na transport

inhibition

the apical

The nature

at the apical

cell membrane

of decreasing

current

work which

(6) one must COW

with Li+

from the Na influx.

that Lit cannot

and Kashgarian

be transported

capability

against

for reabsorption

(12) as possible

no Lit is reabsorbed electrochemical

in pro-

rate than the Na influx under

This limited

explanation

for

in the distal parts of the

gradient,

whereas

75% of the fil-

parts of the nephron.

cell membrane.

of Na uptake

rate when

From this and from earlier

differed

indicate

that, in the rat kidney,

at a considerable

be noted here that the experiments

current

in which Lit faces a large adverse

in which

across

(8) is not

along a favorable

by 20% instead

higher

This study shows that K+ causes a 45% inhibition

ments

path-

of Li+ to ten times the level in

by an equivalent

It should

has been used by Hayslett

tered Lit is reabsorbed

values.

at a slightly

short-circuit

done on toad urinary

a gradient

inhibits

potential

in the external

in an inward direction

is accompanied

(0.6 ueq/cm2hr).

as great an electrochemical

nephron

+.

proceeded

were the only ones in which

against

con-

of the Na up-

the intracellular

since this transfer

bath, since the current

that net movement

Studies

proceeds

preparation,

accumulation

to the Na influx to 15% of the control

control

Li+ uptake

On the other hand the extrusion to active

that inhibition

from the one calculated

of high Li'+ levels

for the observed

with an inhibitor

shift in cell membrane

shift.

that when Li+

that Na+ and Li+ share a common

explanation,

to be

gradient.

added to the external Portion

potential

In this

Jm and KNa were determined

is vastly different

even in presence

account

must be assigned

trochemical

Another

in the short-circuited

since,

negative,

fact such potentials

membrane

of inhibition

bath (7).

is very small when com-

competitively

This suggests

the result of a Li+-induced

of the cell membrane

trochemical

is inhibited

into the cells.

since the degree

which

CY is a perme-

the same study it was discovered

In

bath, Na uptake

constant",

in the external

(first term in equation).

.+ stant for Li , KLi, of about 25 mM (7). way or site for entrance

Michaelis

a linear component

and 14.3 mM, respectively.

is added to the external

+ caNaext)

KNa is an "apparent

Na uptake,

the last term, aNaext

4.0 peq/cm'hr

as:

and Naext is the Na concentration

coefficient

equation,

Communications

of Na influx,

presumably

This view is supported

by Kt was observed

because

it

by other experi-

in direct measurements

of Na entry

(13).

of the interaction

of ions other

than Li+ with Na uptake

remains

to be de-

2268

Preliminary

dermined.

It is interesting

the cation

series as well as the increase

are correlated ure 2).

to note that both the decrease in stimulation

the interaction

that dimensional

with Na transport.

properties

Further

in inhibition

of Na influx in

of Na influx in the anion series

with the size of the radius of the crystal

This suggests

interaction

Communications

of the different

ions (see Fig-

of these ions are an important

studies

factor

are under way to try to characterize

in this

more fully.

-100

!

!

0

I

2 SOUARE

Figure 2. Changes ion radius.

3

OF CRYSTAL

in Na influx shown in Figure

ION

RADIUS

I

I

4

5

(A’,

1 plotted

against

the square of the crystal

RtFERENCES

1.

T.U.L.

Biber and T.L. Mullen,

2.

H.H. Ussing

3.

T.U.L.

Biber and P.F. Curran,

4.

T.U.L.

Biber and L.J. Cruz, Am. J. Physiol.

5.

L.J. Cruz and T.U.L.

6.

K. Zerahn, Acta Physiol.

7.

T.U.L.

8.

W. Nagel,

9.

H. Hvid Hansen and K. Zerahn,

and K. Zerahn,

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Stand.

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Acta Physiol.

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G. Leblanc,

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F.C. Herrera,

R. Egea and A.M. Herrera,

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J.P. Hayslett

and M. Kashgarian,

13.

C.A. Rotunno,

F.A. Villalonga,

(1970).

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231, 995 (1976).

Stand.

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