Renal Na/H exchanger NHE-3 and Na-PO4 cotransporter NaPi-2 protein expression in glucocorticoid excess and deficient states

J Am Soc Nephrol

Renal

Na/H

NaP1-2

Exchanger

Protein

Deficient

Expression

Departments

of

Pediatrics

Administration

Current

data

mediate

significant

fractions

affects

NHE-3

and

rendered

and

NaP1-2

NaP1-2

and

modest

NaPr2

effects

increased

rats

was

doses

NHE-3

and

enhanced

Na/H

respectively

at

least (9-12).

and

the

abundance but

to rats

in part by Glucocorticoid

and due

to

phos-

co-

that proximal tubule Na-P04 cotransport

apiare

Received

September

University yard.

1 I, 1997. Accepted

to Dr. of Texas

Dallas.

TX

Orson

Southwestern

W.

March

proteins, renal con-

Glucocorticoid

Department

Medical

Center.

of 5323

75235-8856.

l560$03.O()/() of the American Society of Nephrology Copyright U 1998 by the American Society of Nephrology l046-6673/()9()9-

Journal

Internal Harry

Medicine, Hines

in-

proximal

tubule,

present

cotransport

by

NaP1-2,

respectively.

sup-

in both

adre-

No significant

recip-

was

in the

staining

NaP1

decreases

cortex

(14),

NaP1-2

has

Boule-

NaP1-2

but the not

or deficient

study

seen

shows

apical changes

that glu-

membrane

Na/H

in protein

abun-

the

renal

glucocorticoid

of excess

because

plasma

to be a mechanism abundance (15,16). abundance

and

of In

of these

two

in the glucocorticoid immunoblot.

increase NHE-3 and in the proximal tubule,

and

whole

distribution

relevant

variations

in the thick

in

status

We

showed

decrease NaP1-2 proand increase NHE-3

ascending

limb.

Methods

Models

Male

Sprague

Dawley

bilateral

for

at least Both

g body daily

doses), and experimental animals

rats weighing

adrenalectomy

3 d before ADX

and

wt subcutaneous

or vehicle

in each

group

were

group)

subjected operations

or sham

free access to water and rat chow. with 0.9% saline in their drinking administration

of

SHM

were

animals

injections

(phosphate-buffered

a fifth dose was given groups (ADX, SHM,

2(X) to 300 g were

(ADX

and were allowed were supplemented

glucocorticoid.

four

cellular

shown NaP-2

examined

with

expression

Materials

protein

and

is particularly

by immunohistochemistry

protein

and in either

has been membrane we

proteins

that glucocorticoids tein expression

twice

This

study,

transporter

j.g/l00

examined

insertion of apical

the present

mRNA

intranephron

been

states.

membrane regulation

water

13. 1998.

Moe,

the

animals.

tubule

and

ascending

dramatically

NaP-2

rats. The

thick

membrane

proximal

excess

(SHM group), ADX animals Correspondence

subapical

protein

Adrenalectomy

administration

on the apical

regulate and

and

in

or

NHE-3

limb.

and sham-operated

of NHE-3

to either

expression.

tubule

abundance

protein in

adrenalectomized

increased

descending

dexamethasone-treated

deficiency

protein

to either

proximal

protein

increase

Animal

NHE-3

thin

nalectomized

dance

Na-PO4

NHE-3 and NaP1-2 excess increases

the

the

dexamethasone

exchange

their effects on by stimulating

inhibiting

both

NaP2

tical NHE-3 mRNA in the rabbit (13), but its effect on NHE-3 protein in various nephron segments in the intact animal has not been examined. It is also unclear whether glucocorticoid affects

Affairs

but

H secretion increased

in

cocorticoids

deficiency The increased

excretion

whereas

in rats

data suggest exchange and

of

‘Veterans

and

significantly

not NaP1-2

rocal

( I ,7,8).

Immunohistochemical cal membrane Na/H mediated respectively

and abundance

Adrenalec-

and

administration rats

pressed

abundance,

buffer

excretion,

exchange

Departments

Center,

sham-operated

respectively. excess or defi-

of dexamethasone

(3-6). Glucocorticoids exert HCO1 and P04 absorption

membrane

transport,

and

Switzerland; Medical

dexamethasone

limb,

adrenalectomy,

tubular

urinary

and

Zurich,

Southwestern

creased

compared

protein

increased

ammoniagenesis

phate excretion proximal tubule apical

involves

of Zurich,

Texas

mem-

Protein

Glucocorticoid excess increases and glucocorticoid decreases net acid excretion by the kidney (2,3). net acid excretion HCO1 absorption

apical

immunohistochemistry.

on

proteins

abundance

by bilateral

pharmacologic

immunoblots

had

protein

in control

of glucocor-

tubule

transporters.

glucocorticoid-deficient

using

and

of proximal

of these

and to rats receiving tomy

NHE-3

and Na/P04 cotransport, whether glucocorticoid

distribution

of

tubule apical membrane cotransport activity (I).

Na/PO4

the

University

University

doses

proximal

that

brane Na/H exchange This study examines

of NHE-3

Excess

Texas.

of pharmacologic

suggest

Physiologv,

Medicine,

Dallas,

increases renal exchange and decreases

intrarenal

Cotransporter

in Glucocorticoid

and

‘InternaI

Center,

in vivo

ciency

Na-P04

MARIUS LOTSCHER,* BRIGITTE KAISSLING,* JURG BIBER,t MOUIN SEIKALY, ROBERT J. ALPERN,* MOSHE LEVI,*i and ORSON W. MOE11

*A,zato,n.

citid

Medical

ticoid Na/H

and

1998

States JAN LOFFING,* HEINI MURER,t MICHEL BAUM,*

Abstract.

NHE-3

9: 1560-1567,

either given

or

either

60

of dexamethasone saline

[PBSI)

2 h before sacrifice. ADX + DEX, SHM used

vehicle

for NHE-3

and

(DEX) for

2 d (four

Initially, +

NaP-2

four

DEX)

with

staining

J

Am

Soc

Nephrol

9:

l56O-l567.

by immunohistochemistry membranes. Subsequently, formed

DEX,

+

and immunoblots of cortical an additional set of experiments

the same

using

ADX

four

conditions

= 6: SHM

a

expression in both cortical were sacrificed and kidneys preparation

Renal

or for

NHE-3

munoblots istry

Glucocorticoid

1998

‘1

protein

ADX,

and

after

expression groups

ADX

+

5: SHM.

=

a

4:

perfusion

for

staining

with

PBS,

(Dakopatts)

Expression

Sigma,

sections

by im-

of actin

of

were

were

St. Louis,

studied

Reichert-Jung.

by

Vienna.

and

NHE-3

Finally.

applied

2.5% MO)

NaP-2

l561

phalloidin ( 1/1(X) diluadded to the secondary

was

filaments.

coverslips

containing

(DABCO:

fixation.

was quantified

antibody rinsed

NHE-3

membranes. The animals for either renal membrane

immunohistochemistry

and NaP-2

a

Renal

Glostrup, Denmark). Rhodamine-conjugated tion: Molecular Probes. Eugene, OR)

and apical was per-

4) to examine

=

and medullary were harvested

in all four experimental

in SHM,

(ADX.

DEX.

+

and

the

with

sections

1 ,4-diazahicyclo-(2.2.2)-octane as a fading retardant.

epifluorescence

were

DAKO-Glycergel

microscopy

and the (Polyvar.

Austria).

and by immunohistochem-

DEX

animals.

Statistical

Analysis

Quantitative

differences

were

assessed

by ANOVA.

IFfllflUt?OblOtS

Renal

or the inner

corteX

and homogenized

(buffer

stripe

of the outer

containing

in mM:

medulla 300

pH 7.50. and 5 ethyleneglycol-bis(f3-aminoethyl tic

acid:

in ig/ml:

aprotmnin,

100

Results

dissected 20

fluoride,

A: Bninkman

polytron),

of Glucocorticoids

Effect

Hepes.

ether)N,N’-tetra-ace-

phenylmethylsulkniyl

and 2 pepstatin

was

mannitol.

Figure

2 leupeptin,

2

1 A and ,

amining

NHE-3

on NHE-3

B, shows

protein

representative

abundance

in

immunoblots cortical

cx-

apical

mem-

and the membrane

fraction was obtained by centnifugation (Beckman J2-2l M. JA-20 rotor. 20.0(X) rpm, 40 mm, 4#{176}C).Apical membrane vesicles were prepared from branes to three final

apical

the cortical consecutive

homogenate precipitations

membrane-enriched

pernatant Twenty

(Beckman micrograms

branes. dodecyl

or cortical apical sulfate-polyacrylamide

vesicles

were

membranes. I 568 against

lution)

and

cortical memMgCl,, and the

pelleted

J2-21 M, JA-20 rotor. 20.000 of either cortical membranes.

nitrocellulose serum (no. (9)

by exposing in 15 mM

from

the so-

rpm. 40 mm. 4#{176}C). medullary mem-

membranes was fractionated by sodium gel electrophoresis and transferred to For epitope

anti-NaP-2

imniunoblotting. DSFLQADGPEEQLQ

antiserum

(against

epitope

Anti-f3-actin

antibody

(1:1000

dilution)

hanced

cheniiluniinescence

(Amersham,

quantified by densitometry. NHE-3 ized to -actin signals and expressed

+

DEX

-j

-

86 kD

was

used

Heights,

IL) and

Cortex

A

and NaP-2 signals were normalas a percentage of the SHM group.

perfusion

abundance

through

(9.12.15).

dehyde/0.05%

(pH 7.4,

of NHE-3

and

the

Briefly.

kidneys

protein

in three

cooled

sections

(3 to 4 jtm thick)

by liquid

gelatin-coated was performed

sections

were

NaP-2,

with

3%

Sections

were

then

N,.

and stored

were

cut in a cryostat

with

pretreated

with

milk

10% normal

powder

incubated

and with

1/8000

LALPAHHNATRL.

(against

den) overnight binding

epitope

in a humidified of

and placed

the

primary

swine-anti-rabbit

goat

0.3%

in PBS/BSA)

serum

DEX

DEX -

_______

previ-

-86kD

( I :40

NHE-3 and For NHE-3.

antiserum

and

for

PBS/BSA:

Figure

1. Immunoblot

Mobility

1566

creased

anpowrinsing FITC-

Dakopatts.

ofrenal

in kilodaltons

(no.

with

-.

51 kD

,-

Medulla

membranes:

repeated

-.--.---

B

PBS.

in PBS/milk

detected

-_--

in

or with anti-NaP-2

was

-acfln

Serial

on chrome

X-l00

at 4#{176}C. After

antibodies

use.

in PBS,

Triton

I :500

chamber lgG

for (9.12.15).

anti-NHE-3

tiserum

conjugated

NHE-3

+

3% paraformal-

-80#{176}C until

at

slides. Inimunohistostaining as described previously

YSRHELTPNEDEKQ,

PBS.

SHM

picnic acid in a 6:4 mixture of 0. 1 M cacodylate buffer adjusted to 300 mosmol/kg water with sucrose) and 10%

propane

in

SHM

were fixed by

as described

aorta,

perfused

in the

experimental

The kidneys

abdominal were

NaP1-2

hydroxyethyl starch. After 5 mm of fixation. the fixative was washed out by perfusion with 0. 1 M cacodylate/sucrose buffer. Coronal slices of the fixed kidneys were mounted on cork disks, frozen in liquid

alum NaP-2

ADX

+

were studied by immunohistochemistry (SHAM, ADX, and ADX + DEX).

vascular ously

and

kD

to ensure

peroxidase-coupled were detected by en-

Arlington

-51

-acfin

ADX Localization

SHM

DEX

Imnmnimno/iistochemmstr’ kidney groups

SHM

LALPAHH-

,

equal loading. After incubation with horseradish mouse anti-rabbit secondary antibody. signals

NHE-3

ADX

+

anti-NHE-3 antiat I :200 di-

NATRL at I : l()00 dilution) ( I I I 2) were used as primary antisera. The specificity of both antisera have been characterized previously (9,12,17).

ADX

NHE-3

ADX

slightly

expression.

NHE-3

is indicated decreased, Number

and f3-actin on the

right.

whereas

protein (A)

DEX

of experiments:

abundance.

Cortical

apical

significantly ADX,

in-

9: SHM.

+ DEX. 10: SHM + DEX. 8. (B) Medullary membranes: ADX decreased, whereas DEX increased NHE-3 expression. Number ofexpeniments: ADX, 5: SHM. 4: ADX + DEX. 6: SHM + DEX. 4.

8: ADX

DEX. operated.

dexamethasone-treated:

ADX.

adrenalectomi

zed : SHM.

sham-

1562

Journal

branes and experimental

of the American

medullary groups.

not significantly was

in both

(P

which

served

Summary

relative

NHE-3

shown).

in the ADX

the ADX

of four signals:

DEX, 245 of cortical

+

Figure

and

DEX

+

from the abundance

SHM

±

SHM, 7%;

100

ADX

membranes 1B

shows

showed

+ DEX,

280

similar

a representative

animals

significantly

16%.

dance

compared

with

SHM

ANOVA). Summary NHE-3 signals: SHM,

NHE-3

(not

immunoblot

of

and ADX

(P

0.05

<

(Figure the

Figure

2B).

brush

of

NHE-3 border.

was localized

of

compartments

border (B): F-actin

tubules

(revealed

NaP,-2

in

the

was

(A),

and over

costaining

NHE-3

(B),

the entire

height

by rhodamine-conjugated

and

of

of

F-actin

(C)

of the brush phalloidin)

an axial

decrease

In S3

segments

+

was

the

3, A through immunoblots

depletion consistently,

way.

and

SHM

similar

above.

The

effect

of

seemed

to affect

The

S 1 and

described

effects

(Figure was were

seen

in

This

evident

in the immunoblots

proximal

border is present

(A): over

variable

of

the

the entire

on immuno-

rats,

whereas

TAL

NHE-3

labyrinth.

is restricted of the brush-border

expression

NaP-2

larger

of the

is visible

microvilli

also

membrane was

to the base

ADX

was

on medullary

NHE-3

NHE-3

much

animals

on TAL

performed

cortical

lengths

expression In SHM

animals,

of ADX

TAL

immunostaining

B).

the individual

effect

However,

tubules

NHE-3

all

among

groups.

above.

on NHE-3

4, A and

similar

differences

S2

above magni-

modest

slight

pattern

to both

groups. (TAL),

had

with

abundance

ADX are

or excess

in a similar

in

findings

shown

in

sham-operated

NHE-3 both

These

evident

expression

with

animals,

C).

not

in NHE-3

to

for

to cortex and was border (Figure 2A).

compared DEX

of

immunoreactivity

staining

compared

of

in the

shown

in

no

decrease

3B)

In ADX

tubules

variations

F-actin

stripe,

small

increased

histochemistry

labyrinth

was

segments.

Intracellular

(Figure

3A).

found

there S2

1998

tude of the changes in all animals of the corresponding In the apical membrane of the thick ascending limb ADX

of S I and

outer

a very

(Figure

proximal

by

in the base

by

profoundly

expression

cortical

predominantly

as demonstrated

2. Localization

intracellular m.

proximal

(Figure

was

of

the

Soc Nephrol 9: 1560-1567,

NaP1 staining was restricted the entire length of the brush

glucocorticoid

abun-

for both

rats

those

the

observed.

tubules

animals

of all animals showed the following 100 ± 25%; ADX, 54 ± 51%; SHM +

DEX, 345 ± 23%; ADX + DEX, 367 ± 26%. NHE-3 protein was expressed in the brush border segments

Immu-

rays,

along and

caused

proximal

results

protein

was

ADX

20%;

±

In medullary rays

NHE-3

membranes. The effect variable than in cortex,

increased

expression

following

103

±

2C).

NHE-3

these sections. evident along

ADX suppressed NHE-3 expression in the medulla by ANOVA). DEX administration to either SHM or

ADX

S2

DEX

+

(Figure medullary

in the four the

showed

but

of -actin,

ADX,

5l%

±

four was

animals,

and the SHM

experiments

NHE-3 protein abundance in medullary of ADX on NHE-3 abundance is more but overall (P < 0.05

J Am

0.05 for both by ANOVA). Labeling as a loading control, was not different

<

groups. SHM noblots

of Ncphrology

membrane from animals In renal cortex, NHE-3

different

increased

animals

Society

dramat-

in

small

of the brush (C).

Bar. 20

J Am Soc Nephrol

Figure

9: 1560-1567.

3. Detection

rat (C).

by immunofluorescence

Immunostaining

dexamethasone for each

ically

is seen

significantly

increased

increases

in

with

the

ADX

SHM in the thin descending limb affected by ADX or ADX

either

Figure abundance

NHE-3

+

ADX

out of four

Renal

of a sham-operated

tubules.

in the brush

DEX

animals

quantifying from the four

NaP-2

immunoexpression

of animals,

suppressed

with

NaP-2

no change

expression

A summary of all four sets relative NaP1-2 levels: SHM,

217

SHM

35%: <

0.05

for

+

(Figure

immunoblot vesicles

increased

sets

dramatically

2% (P

cortex

of proximal

abundance

SHM animals. the following ±

in rat renal

border

4C)

staining was not

on NaP1-2

5 is a representative in apical membrane groups.

three DEX

brush

or ADX animals. NHE-3 (TDL), on the other hand, + DEX.

(?t Glucocorticoid.s

mental

of NHE-3

in the

and

rat (A),

Adrenalectomy

border

of proximal

Expression

an ADX

minimally tubules.

Bar.

of NHE-3

rat (B),

reduces 100 m.

and

NaP-2

1563

and an ADX

whereas Number

+

DEX

administration

of

of experiments:

four

group.

compared

Effect

Glucocorticoid

1998

DEX.

15

all comparisons

± 45%:

except

in set.

in all ADX

or

of animals showed 100 ± 23%; ADX,

ADX

+

between

DEX,

18

SHM

On kidney munoexpression

sections (Figure 6). ADX increased in the brush border of all ncphrons

compared

with

in ADX

animals

SHM was

(Figure

6, A and

slightly

more

B).

NaP-2 imexamined

Intracellular

distinct,

but

cortical

did

staining not

appear

to be more abundant than in SHM controls (Figure 6, A and B). ADX + DEX (Figure 6C) drastically reduced NaP1-2 expres-

NaP1-2 experi-

in one

DEX and ADX + DEX: ANOVA). Immunoblots with membranes showed a similar pattern (data not shown).

sion

in the

for NaP-2 than

brush

was

in ADX

border

more

or SHM

of

all

diffuse

nephrons.

Intracellular

and

abundant

in ADX

apical

membrane

Na/H

staining +

DEX

animals.

Discussion ± +

Increased activity

proximal

contributes

tubule to the

increased

NaHCO3

exchanger

absorption

and

I 564

Journal

of the American

4. Detection

Figure

Society

of Nephrology

by immunofluorescence

of NHE-3

in the inner

stripe

of the rat renal

medulla

of a sham-operated

and an ADX + DEX rat (C). Immunostaining is seen in the thick ascending limbs and the thin descending decreases whereas administration of dexarnethasone significantly increases NHF.-3 abundance in the lurninal cells. The weak 100 p.m. Nuniher

of the

imniunostaining of

experiments:

ADX DEX

lurninal membrane for each group.

four

ADX

SHM

of thin limbs

SHM DEX

+

is not

affected

and

immunohistochernistry

studies

+

by

ectomy

adrenalectomy

apical

Ofl

per

protein

‘

:

-

86 kD

.4.-

decrease

did

se

-

:i:

proximal

detectable

by

increase

in NHE-3 was

:i

-51

kD

profound.

/3-actin

and

the right. Number

of

protein ADX

renal

the

expected

because

animals

abundance.

increased

of experiments:

apical

Mobility

whereas

DEX

four

each

fir

niembranes

in kilodaltons decreased

for

NaP-2

is indicated NaP-2

study

cant Ofl

expression.

of

increased I ,3.4).

nalectomy whereas the Na/H

NH4 Kinsella

secretion

In the

group.

in states

and coworkers

per ce did not lower apical dexamethasone administration

exchanger

by 40 to 90%

for Na or H ( I .7). This increase ground of both adrenalectomized (

I

).

of

have

Our findings

of changes

glucocorticoid demonstrated

excess that adre-

membrane Na/H increased the

medulla,

without

altering

V1.5

of

its affinity

was observed in the hackor sham-operated animals

in NHE-3

antigen

by immunoblot

a small not

both

and

indepetident This

Na/H

and

of

finding

of

activ-

abundance

NHE-3 apical

is

dose

exchange

NHE-3

that

tubule

The

immunoblots

was

between

be

by dexametha-

a pharmacologic

was

demonstrable to

with

TAL

on

more

in the

mediates

a signifi-

cx-

Na/H

membrane

even

in

immunoblots

in TAI

The

increased

..

originating

NHE-3

large

from

TAL

was

NHE-3

all

cxand

Immunoblots

could

TDL

in the

or TAL

clearly

The lack of response

variation compared

in immunoblots

immunohistochemistry medullary

the

of immunoblots

unequivocal

a variable

The suppression

despite

nature

was

to have

expression.

inimtinohistochemistry.

changes

whereas

increased

pression

anii’nals

seemed

NHE-3

quantitative

in DEX

distinguish

medulla. the

the

on

immunohistochemistry.

evident

not

adrenalectomy

effect

due

pression

activity.

that may

change.

suppressive

(

by

( I .7)

confirms

proximal

NHE-3 cortical

induced

represents

stu(Iies

further

portion

on

on total

expression

increase

agreement

previous

effect

adrenalectomized.

jig/kg

60

previous

immunohistochemistry.

assayed

were

The

the

in

present

cortical

and

Bar.

Adrenal-

it is possible

NHE-3

abundance

as

with activity.

performed

This

whether

ity

agreement

exchanger

although

protein

rat (B),

treatment.

a significant

apical

inii’nunohistochemistry.

Cortex 5. Irntiiunohlot

have

immunoblot

an ADX

rat (A).

in

on immunoblots

glticocorticoid.

figllr(’

not

1998

dexamethasone

Na/H

membranes.

in

sone

3-acfln

are

abundance

9: l560-1567.

limbs. Adrenalectomy minimally membrane of thick ascending limb or by

membrane

or brush-border

NaPi-2

Soc Nephrol

J Am

due

showed

to increased

in the TDL

suggests

that cx-

that

J Am Soc Neplirol

Figure rat

9:

560-1567.

Detection

6.

by immunotluorescence

(C). Immunostaining

immunoreactivity

reduces experiments:

there

is seen

the

in

NaP-2

Glticocorticoid

1998

brush

abundance four

are nephron

brush

border

with

in the

for each

border

little border

brush

in rat renal of

proximal

change of

in the

proximal

on NHE-3

of gene

coid-induced

factors

mediating

the effect

by

an

increase

in steady-state

oftranscription

increases

in NHE-3

and

transepithelial

maturing apical tion by

was

decreased

adrenalectomy

the

(2 1 .22).

expression finding

by 40% and

gene

at

pattern. the

restored

that

he accounted levels (18-20).

The

decrease

adrenalectomized

in

rat

TAL

in an in i’ii’o to

(

normal

NaHCO3

microperfusion

by

low-dose

(25.26) sone.

port

the glucocorti-

transcript

NHE-3

neonate

NHE-3 with

can

with

for due

to

Parallel

protein. Na/H exchange activity. flux have been described in the

mRNA, of the

membrane

is compatible

ofthe

cells.

activity NHE-3

NaHCO3

nephron

staining increases

of NHE-3

an ADX

compartments.

23

Na-K-ATPase

In OKP

in NHE-3

activation

and

Expression

rat (A).

in intracellular

methasone

expression.

transcription.

increase

of a sham-operated and

intracellular tubules

The increased NHE-3 protein abundance is compatible the current model of activatioti of NHE-3 by glucocorticoids the level

cortex tubules

Renal

and

NaP-2

and an ADX

rat (B).

Adrenalectomy

Administration

intracellular

I 565

NaP1-2

of dexamethasone

staining.

Bar.

1(8)

DEX

+

increases

drastically

jtm.

Number

of

group.

segment-specific

of glucocorticoid

of NaP-2

in the

and

in TAL rats absorp-

The

).

(24)

can

adrenalectomyand

be restored

by

It is conceivable

by adrenal

cocorticoid

effect

on

of

that

steroids the

and activity has

been

be

on in

NaCl but

not

mediated

tubule

NHE-3

in chronic

study

mineral and

dexamethatrans-

effects rather

of

glu-

than

the

Na-K-ATPase. NHE-3

mRNA metabolic

A protein

and

protein

acidosis

in

(27,28).

The effect of glucocorticoids on phosphate plex and inclrtdes effects on gastrointestinal

dexa-

by

by

NHE-3

proximal

i n TAL absorption

NaHCO

hasolateral

and TAL

described

decrease

of TAL

membrane

increase

abundance

aldosterone

may

apical

abundance

induced

transepithelial

regulation

mineralocorticoids

commensurate

rats

TAL

balance. hypophosphatemia

and

renal have

phosphate been

handling. described

balance absorption,

is comhone

Phosphatunia in patients

with

1566

Journal

and

in animal

principal

of the American

models

regulatory

of earlier

asone

reduced

of renal

mately

30%,

membrane definite mized

adrenalectomy Na-P1

try.

increase

NaP1-2 Levi

had

protein

study

( 14) is far greater

decrease in apical Na-P04 ence between the transport

cotransport data and

likely

due

to the fact

Na-PO4

more

than

one

tnibutions

that

form

of Na-P1

of NaP2

isoforms

to the

little

and

Na-P04

cotransport

on by

in the

report

the observed

protein

activity

Na-PO4

the decrease

sone-treated tion

in apical

animals

and/or

suggests

increased

account

the decrease

in apical

apical ticking

membrane

4.

5.

could

well

cortical

be

on

protein

of

playing

NaP1-2.

membranes

a role Because

also

by which administration in mediating

with

7.

the

to

8.

traf-

10.

of

status,

13.

glucocorticoids may sequent intracellular

abundance.

first induce degradation

endocytosis of NaP-2.

In summary, the present study in Na/H exchange and suppressed

demonstrates Na-PO4

previously

described

due to changes respectively. apical

In

in the TDL

tubule

followed

of the status

is not

TAL

of the

regulated

is inversely

animal,

whereas

that

Science

Grant

3100-

membrane

proportional NHE-3

border

MK,

the phos-

membrane

vesicles.

1982 Levine

DZ:

rat. Kidney

Renal

Jut

J, Eby

E, Lau

Kinsella

JL.

Na-H

net

acid

19: 516-528,

Boross

rabbit

Frieberg

for the phosphaturia

but not on diet

P04

of

or PTH.

1982 uptake

Role

JM,

of

Sacktor

in renal

B:

brush

F233-F239.

M, Kinsella

by proximal

cells

dexamethasone.

A,n

J

J, Cheng

L, Sacktor renal

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Glucocorticoid

border

An,

J Phvsiol

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250:

J, L#{246}tscherM, Kaissling

Moe OW:

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limb.

phos1986

B. Alpern

in the apical

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inorganic

F827-F833,

M, Loffing

thick

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B: Glucocorticoids

transport

Amemiya

and

activation

membrane

1985

acidosis-induced

tubule

proximal

91 : I 10-1 14, 1993

1985

exchange

calcium

A: acid-

1980

invest

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248:

Sebastian systemic

stimulate

K: Mechanism

F74-F83,

RD. and

F30-F43,

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rabbit

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239:

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249:

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on renal

J Phvsiol

acidification.

Phs’siol

EL

steroids

tubule

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RJ,

membrane of rat hit 48: 1206-

Kidney

expression

5983,

1993

Custer

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Acknowledgments

and immunohistochemistry.

Ant

Baum

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Gentry

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F437-F442,

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1994

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mRNA.

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microtubules

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99: Amemiya

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OW,

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glycosphingolipid

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renal

is

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Murer

by glucocorticoids.

The present studies are supported by the Department Affairs Research Service (Drs. Levi and Moe). National

Byers

increase

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R: Glucocorticoids

P. Vandewalle

coids 14.

that the increase cotransport activity apical

Na

brush

Bonner

Am

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membrane

glucocorticoid

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1215, 1995

changes in kinetics of insertion and internalization cannot be the sole mode of regulation of NaP1-2 in response to chronic changes in glucocorticoid excess. At present, one cannot condude on the relative roles of trafficking versus protein synthesis/degradation in mediating the effect of glucocorticoid on tubule

Guntupalli

proximal

in total

glucocorticoid

Licht

M, Quigley

phate, 9.

traf-

effect

Nair

metabolism.

Az

dietary regulates

abundance

USA

of glucocorticoid

Baum

of

inser-

(I 5, 16), and

NaP1-2

varied

acute

HN,

metabolic

in part Protein

Hulter

An,

6.

in dexametha-

protein.

abundance

to Dr. Baum,

National

B: Glucocorticoids the

in renal

Sci

AH.

NH4CI:

adminbut does

membrane

decrease

in the adrenalectomized

convoluted

is cur-

contribute

NaP1-2

mechanism hormone

NaP1-2

glucocorticoids

likely

membrane

ficking is a well described phosphate or parathyroid

staining

Acad

Dubrovsky

base

con-

for the lack

that decreased

internalization

DK-416l2

the Swiss

to Dr.

J, Sacktor

and systems

Nail

Effects

cotransporter

that dexamethasone decreases NaP1-2

membrane

and

I 981

reflects

in vesicles

JM, Kinsella exchange

excretion 3.

detectable effect of adrenalectomy on apical membrane NaP04 cotransport. The increase in intracellular staining of NaP1-2 that accompanies

2.

is

relative

Freiberg

Proc

30%

abundance

The

can also

3 1-47742.96

phate-uptake

not affect NaP-l protein abundance (14). The contribution of Na-P04 cotransporter isoforms other than NaP1-2 to apical membrane

Grant

Moe),

to Dr. Murer).

Na/H

and

flux

I.

in adrenalecto-

cotransport

rently unknown. It has been shown istration in adrenal-intact animals

a

( 1 ,3 I). The differ-

potential

(Grant

to Dr. Alpern, to Dr.

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