(Hypertension. 1998;31:608-614.)
© 1998 American Heart Association, Inc.
Candidate Genes in the Regulation of Na+ Transport by Inner Medullary Collecting Duct Cells From Dahl Rats
Russell F. Husted;
John P. Rapp;
; John B. Stokes
From the Laboratory of Epithelial Transport, Department of Internal
Medicine, University of Iowa, Iowa City (J.B.S., R.F.H.); the Department of
Veterans Affairs Medical Center, Iowa City, IA (J.B.S.); and Department of
Physiology, Medical College of Ohio, Toledo (J.P.R.).
Correspondence to John B. Stokes, Department of Internal Medicine, University of Iowa College of Medicine, Iowa City, IA 52242.
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Abstract
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Abstract—Recently, we reported that
primary cultures of inner medullary collecting duct cells from Dahl
salt-sensitive (S) rats absorb more Na+ than do cells
cultured from Dahl salt-resistant (R) rats. To begin to
evaluate the molecular basis for this difference, we selected four
candidate gene products that on the basis of their physiology and
genetics could participate in regulation of Na+ transport
by these cells. During 24-hour exposure, inhibitors of the
cytochrome P450 enzymes had no effect on Na+ transport by
either S or R monolayers. Twenty-four-hour exposure to
NG-monomethyl-L-arginine
(0.5 mmol/L), a nonspecific inhibitor of NO synthase,
also had no effect on Na+ transport by either S or R
monolayers. Neither atrial natriuretic peptide 1–28 (100
nmol/L) nor 8-Br-cyclic GMP (100 µmol/L) had any short-term
effect on Na+ transport by either S or R monolayers.
18-Hydroxy-11-deoxycorticosterone (100 nmol/L), an adrenocorticoid
hormone that is produced in greater amounts in S rats, stimulated
Na+ transport by both S and R monolayers via the
mineralocorticoid receptor; however, its effect was less potent than
aldosterone. Congenic rats in which the R isoform of the
11ß-hydroxylase gene was bred onto the S background had monolayers
that transported Na+ at a rate similar to the S rats. These
results demonstrate that neither cytochrome P450 genes, NO synthase
genes, the atrial natriuretic peptide receptor gene, nor
the 11ß-hydroxylase gene is a likely candidate to explain the
difference in Na+ transport between S and R inner medullary
collecting duct monolayers in primary culture.
Key Words: atrial natriuretic peptides • nitric oxide synthase • cyclic GMP • 11ß-hydroxylase • rats, congenic • cell culture • electrophysiology
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Introduction
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A major underlying
reason for the hypertension exhibited by the S strain of rat involves
the inability of their kidneys to excrete an appropriate amount of
NaCl.1 Many mechanisms have been proposed to
explain the difference between the S strain and its R counterpart.
Recently, we reported that IMCD cells cultured from S rats transport
considerably more Na+ than do IMCD cells cultured
from R rats.2 This cell culture model provides a
convenient system for examining specific hypotheses regarding the
mechanisms responsible for differences in Na+
transport. Identification of factors responsible for this difference
might provide insight into the pathophysiology of salt-sensitive
hypertension and thereby suggest new therapeutic strategies. In the
present study, we considered four gene products for which there
was substantial evidence to implicate their involvement in hypertension
in the S rat.
Cytochrome P-450 Enzymes
Evidence implicating one or more of these gene products can be
summarized as follows.3 The loop of Henle of S
rats absorbs more NaCl than does the loop of R
rats4 5 ; 20-HETE, a P-450 metabolite of
arachidonic acid, inhibits Cl-
transport by the loop of Henle5 ; and the renal
medullae from S rats produce less 20-HETE than do the medullae from R
rats.6 Manipulation of the P450 enzyme system
produces changes in NaCl transport consistent with the effects
of changes in 20-HETE: inhibitors increase NaCl
absorption.5 7 An inhibitor of the
P450 system (clotrimazol) exacerbates NaCl-induced hypertension in
normal rats,8 and an inducer (clofibrate)
mitigates salt-induced hypertension in S rats.9
In addition to this physiological evidence of an
involvement of the P450 system in salt-sensitive hypertension, genetic
evidence supports a role for these enzymes. A region on chromosome 1
contains a cluster of P450 genes that cosegregates with hypertension in
F2 offspring of (SxLewis)
matings.10 Also, a locus on chromosome 5 near the
P4504A2 gene cosegregates with blood pressure in this same
cross.11 These studies provide functional and
genetic evidence that P450 metabolites such as 20-HETE might be
involved in resistance or susceptibility to salt-induced
hypertension.
NO Synthase
On the basis of several observations,12 a
deficiency of NO has been implicated in salt-sensitive hypertension.
The activity of neural NO synthase is lower in the kidneys of S rats
than in R rats13 ; infusion of
L-arginine, the precursor of NO, mitigates hypertension in
S rats14 ; and pressure natriuresis, a phenomenon
dependent on NO,15 is defective in S
rats.16 17 NO can inhibit
Na+ transport by collecting duct
cells18 ; the inducible form of NO synthase is
present in IMCD cells19 and has been
implicated in the defective NO synthesis in S
rats20 ; and a marker for the inducible NO
synthase gene (on chromosome 10) cosegregates with blood pressure in
the F2 offspring of matings of S rats with two
other strains.21 NO synthase is also an
attractive candidate because an analysis of the segmental
effect of L-arginine infusion on natriuresis shows that the
site of action in S rats is not along the loop of Henle but probably
along the collecting duct.22
ANP and cGMP
The first evidence implicating ANP in the pathophysiology of
hypertension in S rats came from experiments demonstrating that
circulating ANP levels are not diminished, but kidneys of S rats are
less responsive to infused peptide.23 24 The
ANP/guanylate cyclase receptor is liberally expressed on
IMCD cells25 26 ; ANP and cGMP have been reported
to inhibit Na+ transport by IMCD
cells27 ; and cultured IMCD cells from S rats
produce less cAMP in response to ANP than do IMCD cells from R
rats.28 Further evidence implicating the
ANP/guanylate cyclase receptor comes from the observation
that mice lacking the receptor become
hypertensive,29 30 and genetic evidence
implicates this receptor (on chromosome 2) in F2
offspring of matings of S rats with two other
strains.31
18-OH-DOC
One of the earliest differences noted between the S and R strains
was the amount of circulating 18-OH DOC.32 The 11
ß-hydroxylase gene of the S rat is different from that of the R rat
and produces more 18-OH-DOC33 34 ; cosegregation
studies implicate this gene (on chromosome 7) in
hypertension.35 These data, together with the
fact that 18-OH-DOC can act as a mineralocorticoid
hormone36 37 38 and that IMCD cells have
mineralocorticoid receptors,39 40 raise the
possibility that the difference in the 11ß-hydroxylase gene
product produces an effect on IMCD cells that contributes to the
rate of Na+ transport in primary culture.
These observations provide the rationale for examining the hypothesis
that one or more of these gene products might contribute to the
greater rate of Na+ transport by IMCD cells from
S rats.
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Methods
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Inbred strains of Dahl salt-sensitive (Dahl SS/Jr),
salt-resistant (Dahl SR/Jr), and congenic rats were bred at the
University of Iowa. We will refer to the salt-sensitive and
salt-resistant strains as S and R, respectively. All three
colonies were originally obtained from breeders at the Medical College
of Ohio.2 41 All rats were weaned at 4 weeks and
killed within a few days. The blood pressures of these S and R rats
were within the normal range.2
Preparation of Monolayers
Primary cultures of IMCD cells from rats of either sex were
prepared by use of the hypotonic lysis isolation method as previously
described for this laboratory.40 42 43 The inner
medullae were dissected, minced, and incubated in an isotonic solution
containing 0.1% collagenase for 2 to 3 hours. The solution
was made hypotonic by addition of 2 vol of distilled water containing
10 µg/mL DNAse, and cells were recovered after two
centrifugation steps. This isolation procedure yielded
20 to 40 monolayers from six kidneys. Yields from the S, R, and
congenic strains were similar.
Cells were seeded onto collagen-coated 13-mm polycarbonate filters
(Poretics) glued to plastic cylinders (ADAPS) or onto similarly treated
12-mm Millicell PCF filter cups (Millipore Corp) as described
previously.40 42 Seeding density was 20 µg
DNA/filter (
350 000 cells/cm2). Cells were
grown for 3 days in a medium composed of a 1:1 mixture of DMEM and
Ham's F-12 supplemented with 50 µg/mL gentamicin, 20 µg/mL
norfloxacin, 5 pmol/L triiodothyronine, 50 nmol/L cortisol, 5 µg/mL
transferrin, 5 µg/mL bovine insulin, 10 nmol/L
Na+ selenite, and 1% wt/vol bovine
albumin. On the third day, the medium was changed to one that
had no cortisol, norfloxacin, or albumin. After 24 hours in the
steroid-free medium, RT and
Isc were measured. Each of the R, S, and
congenic groups was randomized to treatment groups by a Latin square
procedure based on the Isc.
After randomization, monolayers were exposed to 100 nmol/L
aldosterone or vehicle (ethanol; control) for 24 hours.
Some monolayers (where indicated) were exposed to 100 nmol/L each of
aldosterone and dexamethasone to ensure a
maximal steroid response. Although in general there is no additive
effect of aldosterone and
dexamethasone,43 the addition of both
steroids at times will induce a slightly greater response than one
steroid alone. Monolayers with RT <100
/cm2 were discarded (15%) because such low
resistance indicates nonconfluence.42
Electrical Measurements
Initial measurements of RT and
Isc were made under sterile conditions in
modified Ussing chambers (Jim's Instruments). Measurements were made
in media without additives at 37°C by use of a University of Iowa
voltage clamp.40 42 43 A positive
Isc indicates a flow of positive charges
from the apical to the basal surface. When measurement of electrical
properties required >3 minutes, we mounted filter cups in nonsterile
chambers that were designed to accommodate Millicell PCF
filters.44 The solution used to bathe these
monolayers consisted of a HEPES-buffered PBS solution. For every
condition reported in the present study, the
Isc was sensitive to 10 µmol/L
benzamil, reflecting the fact that the Isc
represents Na+ transport.
Construction of the 11ß-Hydroxylase Congenic Strain
This strain was constructed by breeding the steroid
11ß-hydroxylase allele from R rats onto the S background. S and R
rats were crossed, and the resulting F1 rats were
backcrossed to S. Rats heterozygous for genetic markers at the
11ß-hydroxylase locus35 were selected and
backcrossed again to S. This procedure was repeated for a total of
eight backcrosses. At the eighth backcross, two heterozygotes were
bred, and rats homozygous for the R 11ß-hydroxylase allele were
selected and bred to fix the R allele onto the S background. The
resulting congenic strain is designated S.R-Cyp11b and is maintained by
brother-sister mating.
The properties of S.R-Cyp11b are described in detail
elsewhere.45 Briefly, the congenic strain has an
R segment of chromosome 7 22 centimorgans in length that includes an
introgression of the R 11ß-hydroxylase gene into the S background.
The congenic strain has a lower blood pressure than do S rats receiving
a variety of increased salt intakes. For example, the systolic
blood pressure of male rats receiving a 4% NaCl diet for 24 days was
264±5.5 mm Hg for S and 201±8.2 mm Hg for S.R-Cyp11b
(P<.0001). Mean survival time of males fed a 4% NaCl diet
from weaning was 40±3 days for S and 112±24 days for S.RCyp11b rats
(P<.0001).
Materials
Unless otherwise specified, chemicals were purchased from
Sigma Chemical Co. Media and gentamicin were obtained from the
University of Iowa Diabetes and Endocrinology Research Center. ODYA,
20-HETE, and ethoxyresorufin were purchased from Biomol. ANP 1–28 was
purchased from Nova Biochem. Collagenase was purchased from
Boehringer Mannheim. Bovine albumin (Bovuminar) was
purchased from Bayer. RU-38486 was a kind gift from Roussel-UCLAF
(Romainville, France).
Statistics
Comparisons between groups were made by ANOVA. If the data
exhibited significant heterogeneity of variance
(Bartlett's test), statistical analysis was performed on
log-transformed data. Subsequent analysis comparing specific
groups was made by Newman-Keuls or Bonferroni test as appropriate.
Significance was concluded at the level of P<.05.
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Results
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Inhibitors of Cytochrome P450 Enzymes
We reasoned that if the difference in Na+
transport by cultured IMCD cells from Dahl S and R rats was caused by a
difference in the activity of one or more of the cytochrome P450
enzymes, we would be able to mitigate its effect with
inhibitors. We tested the effects of two agents known to
inhibit P450 monooxygenase enzymes involved in regulating
ion transport by renal epithelial cells.5 46 47
ODYA inhibits the conversion of arachidonic acid to
20-HETE by P4504A,5 7 an action that is of
potential importance in the pathophysiology of hypertension in the Dahl
S rat.5 6 48 Ethoxyresorufin, also an
inhibitor of the P450 monooxygenase system, has
been shown to alter Na/K ATPase activity in renal epithelial
cells.47
Fig 1
shows a single experiment testing
the effect of these inhibitors on the magnitude of
Na+ transport by IMCD monolayers from S and R
rats. The Table summarizes the results of
several experiments. These inhibitors had no effect on the
magnitude of Na+ transport by either S or R
monolayers regardless of whether Na+ transport
was increased by steroid treatment. We also tested the effect of 1
µmol/L 20-HETE on Na+ transport (as assessed by
Isc) by S and R monolayers. There was no
effect over a 24-hour period of observation (data not shown).
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Figure 1. Effect of P450 inhibitors ODYA (1
µmol/L) or ethoxyresorufin (EtR; 1 µmol/L) on Na+
transport by IMCD monolayers from S or R rats.
Representative experiment with six monolayers in each
group. Open bars represent monolayers from R rats; hatched
bars, S rats. Monolayers were exposed to steroids (Ster; 100 nmol/L
dexamethasone and 100 nmol/L aldosterone)
and/or P450 inhibitors for 24 hours before measurement of
Isc. Virtually all of the
Isc was sensitive to 10 µmol/L
benzamil.
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Table 1. Effect of Inhibitors of Cytochrome P450
Monooxygenase Enzymes on Isc in
IMCD Monolayers From S and R Rats
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Inhibition of NO Synthase
We next tested the hypothesis that a difference in
endogenous production of NO might account for the
difference in Na+ transport by IMCD cells. We
used a nonspecific inhibitor because we were interested in
testing all NO synthase enzymes, not just the inducible
form.21 Fig 2 shows
the effect of 0.5 mmol/L L-NMMA on Na+
transport. L-NMMA had no effect on Isc in
either S or R monolayers whether or not they had been treated with
steroids.
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Figure 2. Effect of inhibition of NO synthase on
Na+ transport by IMCD monolayers from S or R rats.
Isc is shown for 8 to 11 monolayers from two
isolations. Shaded bars represent monolayers exposed to L-NMMA
(0.5 mmol/L). All monolayers were exposed to steroids. *S
monolayers have greater Isc than R
monolayers (P<.02). There was no effect of L-NMMA on
Isc.
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Effects of ANP and cGMP
One of the circulating peptides thought to regulate
Na+ transport by the IMCD is
ANP.27 Because of the possibility that the
coupling between circulating ANP levels and intracellular cGMP levels
might be different in S and R IMCD cells,24 28 31
we conducted a series of experiments measuring the short-term effect of
100 nmol/L ANP and, subsequently, 100 µmol/L 8-Br-cGMP on
Isc.
As shown in Fig 3, neither ANP nor cGMP
had an effect on either S or R monolayers. Fig 3 shows the results of a
set of experiments testing sequential addition of these agents. In
other experiments (not shown), neither ANP nor cGMP had an effect by
itself when the rats were followed up for longer periods of time. These
data are consistent with our previous report of a lack of
effect of ANP on Wistar rat IMCD monolayers over a 24-hour
period.49 In addition, we have tried to
demonstrate a short-term effect of ANP and cGMP on
Isc using Wistar monolayers but have never
been able to do so (unpublished data). Of course, we cannot be certain
that a lack of effect of cGMP (or any agent) in cultured IMCD cells
precludes an effect on these cells in vivo. We also cannot be certain
that other culture conditions might uncover an effect of these agents.
Nevertheless, we conclude that the difference in
Na+ transport between S and R IMCD monolayers is
not the result of a difference in responsiveness to ANP or cGMP.
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Figure 3. Effect of sequential addition of 100 nmol/L ANP
and 100 µmol/L 8-Br-cGMP on Na+ transport by IMCD
monolayers from S and R rats. Open circles, R; closed circles, S
monolayers. Three sets each of S and R monolayers were normalized to
the change in Isc with time control
(untreated) monolayers. Neither ANP nor cGMP had an effect on
Isc.
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Role of 18-OH-DOC
Of the genes contributing to the blood pressure difference between
S and R rats, one of the best understood is
11ß-hydroxylase.34 35 As outlined above, the
different activity of this enzyme causes a higher production of
18-OH-DOC in S rats and contributes to their salt-sensitive
hypertension. To explore the possibility that 18-OH-DOC might play a
role in the different rates of Na+ transport in S
and R IMCD monolayers, we first questioned whether 18-OH-DOC could
stimulate Isc in either S or R monolayers.
Fig 4 shows that it can, but at equal
concentrations (100 nmol/L), 18-OH-DOC is a weaker agonist than
aldosterone. The concentration of 18-OH-DOC used in these
experiments is within the physiological range for
Wistar rats.37 Thus, despite the fact that
18-OH-DOC is a less potent mineralocorticoid agonist than
aldosterone, it seems possible that 18-OH-DOC could exert a
mineralocorticoid effect on IMCD cells in vivo.
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Figure 4. Effect of 18-OH-DOC on Na+ transport
by S and R IMCD monolayers. n=37 to 113 monolayers from eight
isolations. Both 18-OH-DOC and aldosterone (Aldo) were
added to 100 nmol/L final concentration, and monolayers were incubated
for 24 hours. *S monolayers had greater Isc
than R monolayers (P<.01); #aldosterone
stimulated Isc in S and R monolayers
(P<.01);  18-OH-DOC stimulated
Isc over control untreated monolayers but
had a lesser effect than aldosterone
(P<.01).
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We next addressed the question of which steroid receptor was primarily
responsible for the 18-OH-DOC effect. Fig 5 shows that the mineralocorticoid
receptor antagonist spironolactone inhibited the
stimulation produced by 18-OH-DOC, whereas the glucocorticoid receptor
antagonist RU-38486 produced a significantly smaller
inhibition. These results are consistent with the idea that
18-OH-DOC produces its stimulatory effect on Na+
transport primarily via the mineralocorticoid receptor.
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Figure 5. Effect of steroid receptor antagonists
on the ability of 18-OH-DOC to stimulate Na+ transport in S
and R monolayers. All monolayers were treated with 100 nmol/L 18-OH-DOC
for 24 hours. *S monolayers had a greater
Isc than R monolayers
(P<.01). #Spironolactone (Spiro; 10 µmol/L;
solid bars), an antagonist of the mineralocorticoid
receptor, inhibited the effect of 18-OH-DOC (P<.01).
RU-38486 (486; 10 µmol/L; hatched bars), an
antagonist of the glucocorticoid receptor, had no effect
(by ANOVA). n=11 to 12 monolayers from two isolations.
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Congenic Rats
To test the idea that the gene encoding 11ß-hydroxylase
(P45011ß) could contribute to the difference in
Na+ transport between S and R monolayers, we
developed congenic rats. These rats were bred to have the section of
chromosome 7 containing P45011ß from R rats superimposed
onto the S genetic background. Monolayers of IMCD cells from S, R, and
congenic rats were prepared simultaneously, and
Isc was measured. As shown in Fig 6, monolayers from the congenic rats had
Isc values similar to S rats, and both had
values greater than for R monolayers. This conclusion was valid whether
or not the monolayers were treated with aldosterone. These
results indicate that the gene(s) responsible for the difference in
Na+ transport by S and R IMCD monolayers is (are)
not located in or around the P45011ß gene.
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Figure 6. Comparison of IMCD monolayers from S, R, and the
congenic strain S.R-Cyp11b, in which the P45011ß gene
from R rats was imposed on the S genetic background. n=13 to 18
monolayers from three isolations. *P<.01 vs R rats
(open bars). #All aldosterone-treated monolayers had
greater ISC than comparable control groups
(P<.01). There was no difference in
Isc values between S rats (solid bars) and
the congenic strain (hatched bars).
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Discussion
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The present results demonstrate that the actions of four
candidate gene products probably do not explain the different rates
of Na+ transport by IMCD cells cultured from Dahl
S and R rats. Our results also demonstrate that 18-OH-DOC can stimulate
Na+ transport by these cells and that the
majority of its action appears to be mediated via the mineralocorticoid
receptor rather than the glucocorticoid receptor.
Despite the attractiveness of attributing the different
Na+ transport rates in cultured S and R IMCD
cells to differences in cytochrome P-450 enzymes and NO synthase
activities, we must conclude that they probably do not contribute to
these different rates. In fact, because manipulation of these
activities did not produce any effect on monolayers from either strain,
we question whether they play any significant role in regulating
Na+ transport in this system.
We also conclude that ANP and cGMP do not influence electrogenic
Na+ transport in S and R IMCD monolayers. This
conclusion may be surprising to some investigators because there is
evidence that these agents do influence Na+
transport by IMCD cells, at least under some conditions. The most
convincing evidence for an effect of ANP on Na+
transport comes from rabbit IMCD cells in
suspension.50 51 Additional evidence has been
presented by investigators using measurements of single ion
channels of cultured rat IMCD cells.52 When this
technique is used, the type of channel that has been reported to be
inhibited by cGMP is a nonselective cation channel. In intact, cultured
rat IMCD cells grown on a permeable support, the principal (if not the
only) route of entry for Na+ across the apical
membrane is the highly selective epithelial Na+
channel. The nonselective cation channel is seen only in excised
patches.53 On the basis of its regulatory
characteristics, the nonselective cation channel is not likely to be
active in a normal, intact IMCD cell.53 54 55
The lack of effect of ANP on Na+ transport by
these cultured IMCD cells is similar to the lack of effect of ANP on
isolated, perfused rat CCDs,56 57 although there
is not unanimity on this point.58 The reasons for
the differences in the reported effect of ANP on
Na+ transport by the collecting duct probably
relate to species differences and types of preparation used. However,
we must conclude that a simple cascade of events, which starts with an
interaction of ANP with its receptor on the collecting duct, which
causes an increase in intracellular cGMP, which in turn reduces the
activity of the apical membrane Na+ channel, is
probably too simplistic. In any event, the hypothesis that a difference
in responsiveness to ANP might explain the different rates of
Na+ transport by IMCD cells from S and R rats
cannot be supported by these data.
The present demonstration that 18-OH-DOC increases
Na+ transport is consistent with the idea
that this compound produces hypertension by increasing
Na+ retention by the
kidney.32 36 37 The observation that 18-OH-DOC
increases Na+ transport by the IMCD adds some
important information to its mechanism of action. The traditional
method of demonstrating an effect of a steroid hormone on the
collecting duct in intact animals is to measure the effect on the ratio
of Na+ and K+ excretion.
This effect is produced primarily by the CCD, in which steroid action
couples Na+ absorption and
K+ secretion. However, steroid hormones stimulate
only Na+ absorption by the IMCD; they do not
stimulate K+
secretion.40 59 Agents that stimulate ion
transport only by the IMCD and not by the CCD would be expected to
increase NaCl absorption without affecting K+
balance. In this respect, the IMCD is more like steroid-responsive
amphibian tissues (ie, skin and urinary bladder) than is the CCD.
Use of Congenic Rats
These experiments illustrate the usefulness of congenic rat
strains in determining cellular mechanisms of action of gene
products. The most direct use of such strains is to test the idea
that a specific gene can have an effect on a quantitative
phenotype in an intact animal. Our experiments extend the
usefulness of these animals to include examining specific hypotheses
regarding the mechanisms of action of these gene products apart
from the in vivo setting.
An obvious use of congenic rats to examine cellular mechanisms of
action in primary cell cultures would be to test gene products
directly involved in a specific cell function. For example, if a gene
encoding one of the subunits of the epithelial
Na+ channel were implicated in hypertension,
congenic rats could be constructed to determine if that gene
product actually made a difference in blood pressure. Primary
cultures of IMCD cells from these congenic rats could then be used to
examine the extent to which the specific gene in question contributed
to a difference in Na+ transport in vitro.
The 11ß-hydroxylase gene product is not expressed in IMCD cells
but is confined to the adrenal gland. Is there a reason to suspect that
a product not intrinsic to the IMCD might influence its behavior in
primary culture? There is a precedent for such an effect. The
development of hypertension in the SHR is associated with a reduction
in the ability to synthesize endothelin by primary cultures of IMCD
cells.60 Thus, the behavior of cells in primary
culture is not necessarily related to an immutable or inevitable
phenotype. Extrinsic factors acting in vivo before cells are
cultured can alter behavior in vitro. However, such does not seem to be
the case for 18-OH-DOC and IMCD cells.
Combining the results from congenic rats with measurements of the
action of 18-OH-DOC on Na+ transport by IMCD
cells, we can conclude that (1) the IMCD from S and R rats is
responsive to 18-OH-DOC and (2) although replacing the
11ß-hydroxylase gene in S rats with the gene from R rats can reduce
blood pressure in S rats, this replacement does not affect the capacity
of IMCD cells to transport Na+. The implication
of this information is that circulating levels of 18-OH-DOC in vivo do
not alter the intrinsic capacity of the IMCD cells to transport
Na+, at least in primary culture. The difference
in Na+ transport by S and R IMCD monolayers must
be the result of other gene products.
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Selected Abbreviations and Acronyms
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|---|
| ANP |
= |
atrial natriuretic peptide |
| CCD |
= |
cortical collecting duct |
| Isc |
= |
short-circuit current |
| IMCD |
= |
inner medullary collecting duct |
| L-NMMA |
= |
NG-monomethyl-L-arginine |
| ODYA |
= |
17-octadecynoic acid |
| 18-OH-DOC |
= |
18-hydroxy-11-deoxycorticosterone |
| R |
= |
Dahl/Rapp salt-resistant |
| RT |
= |
transmonolayer electrical resistance |
| S |
= |
Dahl/Rapp salt-sensitive |
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Acknowledgments
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This research was supported in part by United States Public
Health Service grant HL-55006 and a grant from the Department of
Veterans Affairs. We appreciate the technical assistance of Dick
Shaffer for breeding the rats.
Received March 18, 1997;
first decision April 24, 1997;
accepted October 9, 1997.
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References
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Abstract
Introduction
