INDOMETHACIN AND RENAL HYPERTENSION/flomero and Strong
31. Sybers HD, Ingwall JS, DeLuca MA, Nimmo L: Fetal mouse heart in
organ culture; ultrastructure. Lab Invest 32: 713-719, 1975
32. Ingwall JS, DeLuca M, Sybers HD, Wildenthal K: Fetal mouse hearts;
a model for studying ischemia. Proc Natl Acad Sci USA 72: 2809-
2813, 1975
33. Bird JWC: Skeletal muscle lysosomes. In Lysosomes in Biology and
Pathology, vol 4, edited by JT Dingle, RT Dean. Amsterdam, North
The Effect of Indomethacin Blockade of
Prostaglandin Synthesis on Blood Pressure of
Normal Rabbits and Rabbits with Renovascular
Hypertension
J. CARLOS ROMERO AND CAMERON G. STRONG
With the technical assistance of Sharon M. Schryver, V. Ray Walker, and David C. Manahan
SUMMARY Indomethacin inhibits the synthesis of prosta-
glandin and the release of renin. These effects were studied in
normal rabbits and rabbits with two-kidney Goldblatt hyperten-
sion (2KGH) and one-kidney Goldblatt hypertension (1KGH) by
giving daily intravenous injections of indomethacin (3 mg/kg after
two initial doses of 9 mg/kg), and in appropriate control rabbits
given diluent phosphate buffer without indomethacin. In normal
rabbits, indomethacin significantly decreased immunoreactive
plasma prostaglandin E-like substance (IPGE) and plasma renin
activity (PRA). Indomethacin did not change plasma creatinine
(PCr) or mean blood pressure but it decreased renal blood flow
(RBF) and glomerular filtration rate (GFR). In 2KGH rabbits,
RECENT evidence suggests that the vasodilator and natri-
uretic actions of prostaglandins and their inhibitory effect
on adrenergic activity' may play important roles in the
homeostatic regulation of renal blood flow and arterial
blood pressure. In agreement with this line of thought, a
hypothesis has been advanced suggesting that renal is-
chemia subsequent to a deficient synthesis of prostaglan-
din in the kidney may induce hypertension secondary to
salt retention and stimulation of renal pressor principles.2
This concept has been supported by preliminary reports
showing that the blockade of prostaglandin synthesis by
indomethacin results in a significant increase in blood
pressure in normal rabbits3 and aggravates renovascular
hypertension in rats.4 In these reported studies, changes in
plasma renin activity and renal function were not mea-
sured. Such measurements might have allowed a better
understanding of the mechanism underlying the elevation
in blood pressure.
From the Mayo Clinic and Mayo Foundation, Rochester, Minnesota.
Supported in part by Research Grant HL-16496 of the National Insti-
tutes of Health, Public Health Service.
Dr. J.C. Romero is an Established Investigator of the American Heart
Association.
Address for reprints: Dr. J.C. Romero, Mayo Clinic, 200 First Street,
S.W., Rochester, Minnesota 55901.
Received April 2, 1976; accepted for publication August 18, 1976.
35
Holland, 1975, pp 75-109
34. Ericsson JLE: Mechanism of cellular autophagy. In Lysosomes in
Biology and Pathology, vol 2, edited by JT Dingle, HB Fell. Amster-
dam, North Holland. 1969, pp 345-394
35. Wildenthal K: Lysosomes and lysosomal enzymes in the heart. In
Lysosomes in Biology and Pathology, vol 4, edited by JT Dingle, RT
Dean. Amsterdam, North Holland, 1975, pp 167-190
responses depended on the level of renal function and, to a lesser
extent, on the level of PRA. In six of 10 2KGH rabbits in which
hypertension developed without significant changes in PRA,
IPGE, PCr, RBF, and GFR, indomethacin produced changes
similar to those seen in normals. In the other four rabbits, devel-
opment of 2KGH was accompanied by increased PRA, increased
IPGE, and decreased RBF and GFR, and indomethacin pro-
duced renal failure, oliguria, malignant hypertension, and death
within 5 days. In 1KGH rabbits, indomethacin decreased IPGE,
PRA, and renal function but increased mean blood pressure.
These observations suggest that prostaglandins exert a protective
effect on renal function in renovascular hypertension.
The present study was designed to examine the effect of
the administration of indomethacin (3 mg/kg) on blood
pressure, plasma renin activity (PRA), plasma levels of
immunoreactive prostaglandin E (IPGE), plasma creati-
nine (PCr), renal blood flow (RBF), and glomerular filtra-
tion rate (GFR) of normal rabbits and of rabbits rendered
hypertensive by constriction of one renal artery with [two-
kidney Goldblatt hypertension (2KGH)] and without
[one-kidney Goldblatt hypertension (1KGH)] the contra-
lateral kidney remaining in situ.
Methods
ANIMAL PROTOCOL
The study involved 42 male New Zealand rabbits weigh-
ing 2.5-3.0 kg. After a control period of 6 days, during
which basal values were obtained for PRA, IPGE, PCr,
RBF, and GFR, the left renal artery was constricted in 28
rabbits. The remaining 14 rabbits did not undergo surgery
and were used as the normal group. At the end of the 2nd
week the contralateral kidney was removed from 14 of the
28 clipped rabbits. For this procedure the rabbits were not
randomly selected; the contralateral kidney was removed
from the rabbits that exhibited the lesser increase or no
change in blood pressure 1 week after clipping. Thereaf-
ter, all the rabbits were housed in metabolic cages and fed
36 CIRCULATION RESEARCH
regular rabbit pellets (Purina Rabbit Chow) and water ad
libitum for 35 days. During this time, hypertension devel-
oped in all the clipped and in the clipped-unilaterally
nephrectomized rabbits. At the end of the 35th day treat-
ment with indomethacin was begun with two injections of
9 mg of indomethacin each, 2 hours apart, followed by 3
mg/kg per day for 10 days administered in two doses at
12:00 a.m. and 12:00 p.m. by the same route. As in
previous studies,5'6 these doses of indomethacin were sus-
pended in 0.5 ml of phosphate buffer (pH 8.4). This
treatment was given to 10 of the 14 normal rabbits, 10 of
the 14 rabbits with 2KGH, and 10 of the 14 rabbits with
1KGH. The remaining four rabbits in each group (four
normals, four 2K.GH, and four 1KGH) were treated with
an identical amount of phosphate buffer that did not
contain indomethacin.
BLOOD SAMPLING AND PREPARATION OF PLASMA
FOR DETERMINATION OF PRA, IPGE, AND PCr.
During the 6-day control period and during the 35 days
that followed the surgical procedure, 6-ml blood samples
for determination of PRA, IPGE, and PCr were obtained
every week. Blood samples also were drawn 9 hours after
treatment with indomethacin began and daily or every
other day during the 10 days of treatment. The samples
were drawn between 8:00 a.m. and 9:00 a.m. from a 21-
gauge needle inserted in the central artery of the ear. The
first 2 ml were collected in a separate tube for determina-
tion of PRA, whereas the remaining 4 ml were collected in
different tubes for determination of IPGE and PCr. This
was done to avoid an artifactual increase in PRA induced
by blood sampling. Blood samples for determination of
PRA were centrifuged, and plasma was separated and
kept frozen until determination by radioimmunoassay.7 A
similar procedure was followed for PCr. Plasma samples
for determinations of IPGE were separated after refriger-
ated centrifugation and were frozen immediately at 0°C
until extraction. Plasma (1 ml) was acidified to a pH
between 3 and 4 with 1 N HC1; 4 ml of redistilled ethyl
acetate (Fisher Chemical) were added to plasma and vor-
texed vigorously for 30 seconds. After centrifugation
(2,000 rpm, 5 minutes) the ethyl acetate phase was sepa-
rated, and the residue was extracted again with an equal
amount of ethyl acetate. Both extractions were then com-
bined and flash evaporated (Buchler-Rotary Evapormix).
Column chromatography was performed according to
the procedure recommended by Caldwell et al.8 After
separation of IPGE and conversion to prostaglandin B
(PGB) by alkaline treatment,5 PGB was radioimmunoas-
sayed using labeled PGB, and anti-PGB2 antibodies (Clin-
ical Assays). Free-labeled PGB, was separated from PGB,
bound to antibodies with charcoal.8 Final values are ex-
pressed in nanograms per milliliter of plasma. Recoveries
from extraction and column separation obtained with 3H-
labeled prostaglandin E, (3H-PGE,) (New England Nu-
clear) in 30 samples from normals, 30 samples from
2KGH, and 30 samples from 1KGH ranged from 50% to
70%. Conversion of PGE to PGB in these samples ranged
from 90% to 95%.
Renal clearances of p-aminohippurate (PAH) and inulin
VOL. 40, No. 1, JANUARY 1977
(In) were performed on the 6th day of the control period,
on the 35th day after the development of hypertension,
and on the 5th and 10th days of treatment with indometh-
acin. This was done by giving the priming and mainte-
nance dose of In and PAH through the marginal vein of
the right ear. Blood samples (3 ml) for determination of In
and PAH were obtained through a 21-gauge needle in-
serted in the central artery of the ear. In general, the
procedure was identical to that used in previous studies,9
with the exception that urine samples were collected
through a 7-Fr. cannula (Foley) inserted into the bladder
through the urethra rather than by catheterizing the ure-
ters.
During the experiment, blood pressure was recorded
with a modification of the Grant-Rothschild capsule10 ex-
cept during the 9 hours that followed the first two injec-
tions of indomethacin when blood pressure was recorded
on a polygraph (Grass model 7) connected to a transducer
(Statham DB23) that sensed pressure through a 21-gauge
butterfly needle placed in the central artery of the ear.
This was done to detect acute changes in blood pressure.
PCr levels were estimated by a modified picric acid
method6 used in previous studies."
PRA was measured by radioimmunoassay according to
the procedure of Haber et al.7 Anti-angiotensin I antibod-
ies were obtained from New Zealand rabbits treated with
serum albumin-angiotensin 1 complex prepared according
to the procedure of Goodfriend et al.12 Angiotensin I
labeled with 125I and inhibitors of the converting enzyme
angiotensinase were purchased from Squibb. The repro-
ducibility of this method has been published elsewhere.13
STATISTICAL ANALYSIS
The significance of differences in changes between the
groups or occurring within one group at different times
was analyzed by the unpaired and paired /-test. When the
variances between the two groups were unequal, treat-
ment differences were analyzed by a rank-sum test.
Results
EFFECT OF INDOMETHACIN ON BLOOD PRESSURE,
PRA, IPGE, AND RENAL FUNCTION OF NORMAL
RABBITS
During the control period, as well as during the 35 days
preceding treatment with indomethacin, blood pressure,
PRA, RBF, PCr, and IPGE remained constant in normal
rabbits (Fig. 1). The administration of indomethacin was
followed during the next 24 hours by a significant (P >
0.01) decrease in the mean \alues of IPGE. On the 10th
day of treatment, these mean values returned to approxi-
mately 50% of the control values.
Changes in IPGE were accompanied by a significant
(85%) (P < 0.01) decrease in the mean value of PRA,
and this decrease remained during the 10 days of treat-
ment. On the contrary, no significant changes were de-
tected in blood pressure during the first 5 days of treat-
ment; later, the blood pressure decreased slightly. The
total decrease from the 5th day to the 10th day was 12 mm
Hg(P < 0.05).
On the 10th day, indomethacin induced moderate but
significant (P < 0.05) decreases in RBF (from 40.4 ±3.3
 1NDOMETHACIN AND RENAL HYPERTENSION/Komero and Strong 37

120 36 ± 3 mm Hg but no significant change in PRA, RBF,

r—Indomethocin treotment-i
1 1 Jdfinrl f l f l l U r i
t
n fin
F .ft. . . . n - , . . ft
Control0 1 2 3 4
5 3 6 9 2 3 4 5 6 7 8 9 10
Weeks Hours Doys
FIGURE 1 Effect of indomelhacin treatment on normal rabbits.
Plasma renin activity (PRA) immunoreactive plasma prostaglan-
din E-like substance (IPGE) were reduced, but renal blood flow
(RBF) and plasma creatinine did not change. Mean blood pressure
did not change significantly. Hatched area represents range of
individual values.
to 33.9 ± 1.3 ml/min) and GFR (from 8.2 ± 0.6 to 5.1 ±
0.5 ml/min) (Fig. 1). PCr levels did not change signifi-
cantly.
The administration of phosphate buffer without indo-
methacin to four normal rabbits did not result in any
significant change in any of the parameters studied (Table
1).
Histological examination of the kidneys removed from
the normal rabbits that were treated showed localized
areas of interstitial and periglomerular fibrosis infiltrated
with inflammatory cells. These lesions were absent in the
nontreated rabbits.
EFFECT OF INDOMETHACIN ON BLOOD PRESSURE,
PRA, IPGE, AND RENAL FUNCTION ON
2KGH RABBITS
In six rabbits the clipping of one renal artery caused a
significant (P < 0.01) increase in mean arterial pressure of
IPGE, or PCr (Fig. 2). In four other rabbits the renal
arterial constriction elicited a much greater increase in
mean arterial pressure. The increase in blood pressure in
these four rabbits was accompanied by a significant in-
crease (P < 0.05) in PRA and IPGE. These four rabbits
also showed significant reductions (P < 0.01) in RBF.
The response to indomethacin was different in these two
groups of rabbits. In the group of six rabbits in which
L hypertension evolved without changes in RBF, indometh-
acin treatment was followed by a series of changes that
resembled those noted in normal rabbits, in that the initial
decrease in circulating prostaglandin was accompanied by
a marked decrease in PRA with no increment in blood
pressure. These rabbits also exhibited a transient decrease
in RBF (from 40.0 ± 1.4 to 26.8 ± 1.8 ml/min) and GFR
1 ("i (from 8.5 ± 0.4 to 5.4 ± 0.1 ml/min) on the 5th day of
treatment. However, PCr remained unaltered during the
treatment.
In the four 2KGH rabbits that had decreased RBF in
response to clipping, treatment with indomethacin caused
renal failure with a progressive increase in PCr, oliguria,
and malignant hypertension and a progressive increase in
PRA. These rabbits did not survive for more than 5 days.
All these described changes induced by indomethacin in
2KGH rabbits were not observed in the control 2KGH
untreated group in which the administration of phosphate
buffer without indomethacin did not result in any signifi-
cant change in BP, PRA, IPGE, or PCr (Table 1).
The histological appearance of kidneys from six 2KGH
rabbits treated with indomethacin revealed the existence
of interstitial and periglomerular fibrosis and infiltration
with chronic inflammatory cells similar to those found in
the normal group that was treated. These lesions were
more frequent in the contralateral than in the clipped
kidney, and grossly the involvement of the renal paren-
chyma was greater than that seen in the normal group that
was treated. On the contrary, the occurrence of interstitial
and periglomerular fibrosis was rare in the group of un-
treated 2KGH rabbits.
Unfortunately, the sudden death of 2KGH rabbits pre-
cluded a systematic histological evaluation, but the kid-
neys obtained from two of these rabbits, 24 and 36 hours
after the development of malignant hypertension, showed
microscopic lesions that resembled those seen during the
development of experimental vasomotor acute renal fail-

TABLE 1 Normal and Hypertensive Rabbits Treated With Phosphate Buffers

Normal control rabbits (n=4) 2KGH rabbits (n = 4) 1KGH rabbits (n = 4)

After clip- PO4-buffer After clip- PCVbuffer After clip- PO^buffer

Control, 6th ping, 35th treatment, Control , 6th ping, 35th treatment, Control , 6th ping, 35th treatment,
Parameter day day 10th day day day 10th day day day 10th day

BP (mm Hg) 72 ± 3 74 ± 2 76 ± 3 73 ± 3 110 ± 4 109 ± 3 75 ± 1 110 ± 4 106 ± 3

PRA (ng/ml/hr) 16 ± 4 20 ± 3 18 ± 5 20 ± 1 20 ± 2 19 ± 1 18 ± 3 19 ± 4 21 ± 2
PGE (ng/ml) 1.3 ± 0.1 1.5 ± 0.2 1.2 ± 0.2 1.4 ± 0 2 1.5 ± 0.4 1.6 ± 0.3 1.2 ± 02 1.5 ± 0.2 1.4 ± 0.1
RBF (ml/min) 40 ± 2.7 38 ± 3.1 39.1 ± 1.0 37.1 ± 1.4 26.3 ± 1.3 27.3 ± 1.2
GFR (ml/min) 7.4 ± 0.4 7.8 ± 0.4 8.5 ± 0.4 8.6 ± 0.3 5.4 ± 0.4 5.3 ± 0.3
PCr (mg/dl) 0.9 ± 0.1 1.0 ± 0.1 1.1 ± 0.1 0.9 ± 0 1 0.9 ± 0.1 1.1 ± 0.1 0.9 ± 01 0.9 ± 0.2 0.7 ± 0.1
Results are expressed as mean ± SE.
2KGH = two-kidney Goldblatt hypertension; 1KGH = one-kidney Goldblatt hypertension; BP = blood pressure; PRA = plasma renin activity; PGE •
prostaglandin E; RBF = renal blood flow; PCr = plasma creatinine.
38 CIRCULATION RESEARCH VOL. 40, No. 1, JANUARY 1977

teo rlndomethocm progressive and significant (P < 0.01) increment, which

treatment was paralleled by a significant (P < 0.01) increase in
blood pressure. On the 10th day of treatment the mean
values for blood pressure increased significantly (P <
0.05) over those recorded before treatment began. The
administration of indomethacin to 1KGH rabbits resulted
in a significant (P < 0.01) decrease in PRA and IPGE. As
in previous treated groups, the decrease in IPGE was not
sustained and the values measured on the 8th and 10th
days of treatment had returned to 50% of the pretreat-
ment levels. None of these changes was observed in the
group of 1KGH control rabbits that were not treated with
indomethacin (Table 1).
The histological appearance of these kidneys was similar
to that found in the 2KGH rabbits that did not develop
malignant hypertension. The histological alterations of
renal parenchyma produced by the interstitial and periglo-
merular fibrosis were the same. However, four of these
rabbits had renal cortical infarction that was not seen in
any other group, with the exception of one of the four
rabbits that developed malignant hypertension.
Discussion
The major finding of this study is that the blockade of
prostaglandin synthesis by the administration of indo-
methacin to normal rabbits and to 2KGH and 1KGH
tf, ^L J1! J1 Vt — A I I 1
rabbits is not accompanied by any significant increase in
'Control0 ' 2 3 4
5 3 6 9 1 2 3 4 5 6 78 9 1 0 blood pressure unless the treatment induced a significant
Hours Days reduction in the renal function. Our data also showed that
FIGURE 2 Effect of indomethacin on 2 KGH rabbits. Six rabbits renal failure did not occur in a consistent fashion, occur-
(closed circles, solid bars) showed changes similar to those seen in ,so
normal rabbits. Four rabbits (open circles, open bars) showed plndomethacin treotment-x
«;
increased PRA and IPGE with decreased RBF during development 160 •
ment
of 2 KGH; in these rabbits, indomethacin produced renal failure, j£ 1 4 0 ' clip T
malignant hypertension, and death within 5 days. 2KGH = two- Right
kidney Goldblatt hypertension; other abbreviations as in Figure 1. Nx
100 ; \

ure." The renal tubules were filled with hemoglobin casts, 80 t—<
and there was evidence of epithelial and glomerular is- 60
chemic lesions. These histological disturbances were simi- Jg 20
lar in severity in the contralateral and the clipped kidney. S co
In one of these rabbits, in the contralateral kidney there
§• o
were numerous renal infarctions. L

EFFECT OF INDOMETHACIN ON BLOOD PRESSURE,

PRA, AND RENAL FUNCTION IN 1KGH RABBITS
Changes in blood pressure, PRA, RBF, PCr, and IPGE
occurred during the development of 1 KGH and during the
subsequent treatment with indomethacin (Fig. 3).
i x 20
e IO
0

;
I I
The development of one-kidney hypertension in this
group was not accompanied by any significant changes in
PRA, PCr, or IPGE. However, there was a significant (F
< 0.05) reduction in RBF (from 38.5 ± 2 to 27.3 ± 1.46
ml/min) and GFR (from 7.4 ± 0.3 to 5.45 ± 0.26 ml/ C o n t r o t0 1 2 3 4
j tt.h
5 3 6 9 1 2 3 4 5 6 7 8 9 10
min). The decrease in renal flow was accentuated during Weeks Hours Days
the treatment with indomethacin. On the 5th and 10th FIGURE 3 Effect of indomethacin on I KGH rabbits. PRA and
days of treatment, the mean values of RBF were de- IPGE were reduced, hypertension was aggravated, RBF was de-
creased by 20% from those recorded immediately before creased, and plasma creatinine was increased by indomethacin
treatment. During the indomethacin treatment the rabbits treatment. 1 KGH = one-kidney Goldblatt hypertension; other
developed a progressive oliguria. The PCr exhibited a abbreviations as in Figure I.
 INDOMETHACIN AND RENAL HYPERTENSION/flomero and Strong
ring only in rabbits that exhibited an RBF below normal at
the time treatment with indomethacin began. These find-
ings strongly suggest that the elevation of blood pressure
produced by indomethacin is not a phenomenon strictly
related to the blockade of prostaglandin synthesis but one
indirectly mediated by a decrease in renal function.
Therefore, it is relevant to analyze why indomethacin
induced renal failure only in rabbits that already had a
significant decrease in RBF and to evaluate whether there
is an identifiable factor common to the production of renal
failure and the aggravation of hypertension.
Previous studies have shown that the reduction of RBF
induced by mechanical occlusion of the renal artery14 or by
infusion of angiotensin II15 elicits the release of prosta-
glandin E. These findings suggested that the vasodilator
action of renal prostaglandin E could be important as a
local regulator in minimizing renal ischemia.14 This con-
cept has received further support from other investigators
who have shown that prostaglandin may be responsible for
the reactive hyperemia that occurs after the temporary
occlusion of the blood supply to the kidney1" and to other
tissues such as skeletal17 and cardiac18 muscle. If the en-
hanced synthesis of prostaglandin which follows renal is-
chemia is chronically operative,19 then it could be assumed
that the remaining RBF in kidneys in which the circulation
was severely impaired by the narrowing of the renal artery
(such as the clipped kidney of 2KGH and 1KGH rabbits)
or by the vasoconstrictor action of excessive amounts of
circulating angiotensin (such as the contralateral kidney of
rabbits with 2KGH and high levels of PRA) is primarily
maintained by an enhanced synthesis of prostaglandin.
Hence, blockade of prostaglandin synthesis in these kid-
neys would lead to a significant reduction in RBF and
renal failure. In contrast, such effects are not seen when
the renal circulation is not severely impaired (as in normal
rabbits or in 2KGH rabbits that have normal RBF). The
validity of these assumptions is further supported by the
finding in previous studies20 that the decrease in RBF
produced by indomethacin is proportional to the previous
decrement in RBF and indicates that the importance of
renal prostaglandins in maintaining RBF increases with
the degree of renal ischemia. Furthermore, the concept
that the enhanced synthesis of renal prostaglandin plays an
important role in the maintenance of the remaining RBF
when renal circulation is compromised also comes from
the previous finding" that the blockade of prostaglandin
synthesis with indomethacin aggravates the course of glyc-
erol-induced (vasomotor type) renal insufficiency in which
the major initial pathological event is believed to be an
exaggerated constriction of the glomerular afferent arteri-
oles.21 In contrast, indomethacin does not aggravate the
course of the nephrotoxic type of acute renal failure in-
duced by mercuric chloride11 in which the primary event is
tubular damage.22 Thus, the histological appearance of
renal tubular and glomerular ischemic lesions and renal
infarction seen in kidneys from rabbits with severe 2KGH
and 1 KGH might be caused by the cessation of the vasodi-
lator protective effect of prostaglandins.
In analyzing the consequences derived from the block-
ade of prostaglandin synthesis, one should consider the
39
difficulties in ruling out the possibility that such effects
were produced directly by indomethacin. These draw-
backs have been partially overcome by other investigators
who determined the reproducibility of the observed effect
when the blockade of prostaglandin synthesis was induced
by other aspirin-like substances such as meclofenamate.23
These uncertainties cannot be resolved by the results of
this study, which was designed to define the mechanisms
underlying the reported elevation in blood pressure when
the synthesis of prostaglandin is blockaded with indometh-
acin.3'4 Within this context, renal insufficiency followed
the treatment with indomethacin and was observed only in
rabbits that already had a significant decrease in RBF.
Because blood pressure increased only in rabbits that
developed renal insufficiency, blood pressure should be
analyzed further to determine to what extent further in-
creases were conditioned by failure in renal function. If
the blood pressure of 1KGH rabbits were "volume-de-
pendent"24 and they developed progressive renal failure
with a urinary output significantly lower than that found in
1KGH untreated control rabbits, it would be logical to
assume that the extracellular fluid volume of these rabbits
was expanded during the treatment with indomethacin and
that this expansion contributed to the aggravation of hy-
pertension.
The aggravation of hypertension induced by indometha-
cin in 1 KGH rabbits was also accompanied by an upward
trend in PRA (Fig. 3). However, several factors tend to
abrogate its importance in the aggravation of hyperten-
sion: first, the maintenance of one-kidney hypertension is
relatively independent of changes in PRA;25'26 second, in
these same rabbits, one-kidney hypertension occurred
without significant change in PRA; and third, the signifi-
cant decrease in PRA which followed the treatment with
indomethacin was not accompanied by any change in
blood pressure.
In analyzing the factors responsible for blood pressure
elevation in 2KGH rabbits that developed malignant hy-
pertension during indomethacin treatment, one should
consider that this type of hypertension is highly dependent
on the levels of PRA. 2526 In fact, the simultaneous
changes in blood pressure and PRA exhibited by the four
rabbits from the beginning of the experiment support this
concept. The development of two-kidney hypertension
was paralleled by an increase in PRA; the decrease in
PRA induced by indomethacin was accompanied by a
transient decrease in blood pressure, and with the devel-
opment of the malignant phase, both blood pressure and
PRA increase. In these rabbits there also was a positive
relationship between the increments in PCr and in blood
pressure, and during this period they experienced signifi-
cant oliguria. Therefore, expansion of the extracellular
fluid volume could have potentiated the pressor effect of
renin.27 An interesting observation made on the four rab-
bits with severe 2KGH was that malignant hypertension
developed in the presence of levels of renin which were
not higher than those recorded before treatment began. If
one considers that other investigators have shown that
indomethacin potentiates the vasopressor responses to
catecholamines and angiotensin,28 one might assume that
40 CIRCULATION RESEARCH
in these four rabbits the vasopressor action of angiotensin
could have been considerably potentiated by the blockade
of prostaglandin. The sudden increase in blood pressure
observed in the four rabbits with decreased RBF seems
unlikely to be explained by the natural evolution of the
disease because such an outcome has never been observed
in 2KGH rabbits with similar blood pressure levels kept
alive for 80 days.29
A highly reproducible change that was observed in nor-
mal, 1KGH, and 2KGH rabbits treated with indometha-
cin was a significant decrease in PRA. This confirms the
previous observations of our group.9-30 From these experi-
mental studies designed to elucidate the mechanism of the
lowering effect of indomethacin on PRA, we concluded
that such a phenomenon is not due to the interference by
this drug with the renin-angiotensinogen reaction but
rather to the blockade of renin release.31 If a decrease in
renin release, and thereby in PRA, is due entirely to
indomethacin and is not related to the blockade of prosta-
glandin synthesis, then it could be theorized that indo-
methacin is not the drug of choice to investigate the effect
of prostaglandin withdrawal on blood pressure. The eleva-
tion in blood pressure expected after the disappearance of
the vasodilator action of prostaglandin would be blunted
by the depressive side effect of this drug on renin release.
However, if the decrease in renin release is inherent in the
blockade of prostaglandin synthesis and the withdrawal of
prostaglandin is physiologically accompanied by a reduc-
tion of PRA, then the role of prostaglandin in the regula-
tion of blood pressure will have to be considered within
the context of the parallel changes elicited in the renal
pressor system. The latter speculation may be correct,
since attempts to separate the effects of blockade of pros-
taglandin synthesis from those exerted on renin release by
the use of other blockers of prostaglandin synthesis with a
molecular configuration different from that of indometha-
cin (such as meclofenamate and aspirin) have been nega-
tive so far.32 All these considerations are important be-
cause they restrict our experimental design in examining
the cause of indomethacin enhancement of blood pressure
reported by other investigators to experimental circum-
stances in which the blockade of prostaglandin will pre-
dictably be accompanied by a decrease in PRA.
In summary, the data obtained from this study show that
the effect on blood pressure of the blockade of prostaglan-
din synthesis produced by indomethacin in normal rabbits
and in renovascular hypertensive rabbits is conditioned by
renal function. In 2KGH and 1KGH rabbits with severe
impairment in renal circulation, the administration of in-
domethacin caused renal insufficiency and increased hy-
pertension and this could be attributed largely to either
volume expansion or a further increase in PRA.
In renal vascular hypertensive rabbits with normal renal
circulation and in normal rabbits, the blockade of prosta-
glandin synthesis produced by indomethacin is followed by
a significant and sustained decrease in PRA and by no
major changes in blood pressure.
Acknowledgments
Indomethacin was generously donated by Merck Sharp & Dohme, West
Point, Pennsylvania.
VOL. 40, No. 1, JANUARY 1977
References
1. Hedqvist P: Effect of prostaglandins and prostaglandin synthesis inhib-
itors on norepinephrine release from vascular tissue. In Prostaglandin
Synthetase Inhibitors: Their Effects on Physiological Functions and
Pathological States, edited by H Robinson, JR Vane. New York,
Raven Press, 1974, pp 303-309
2. Lee JB: Prostaglandins and the renal antihypertensive and natriuretic
endocrine function. Recent Prog Horm Res 30: 481-522, 1974
3. Colina-Chourio J, McGiff JC, Nasjletti A: Development of high
blood-pressure following inhibition of prostaglandin synthesis (abstr).
Fed Proc34: 368, 1975
4. Pugsley DJ, Beilin LJ, Peto R: Renal prostaglandin synthesis in the
Goldblatt hypertensive rat. Circ Res 36 (suppl I): 81-88, 1975
5. Levine L, Gutierrez Cernosek RM, Van Vunakis H: Specificities of
prostaglandins B,, F,a, and F2o antigen-antibody reactions. J Biol
Chem 246:6782-6785, 1971
6. Levinson SA, MacFate RP: Clinical Laboratory Diagnosis, ed. 7.
Philadelphia, Lea & Febiger, 1969, p 413
7. Haber E, Koerner T, Page LB, Kliman B, Purnode A: Application of
a radioimmunoassay for angiotensin I to the physiologic measurements
of plasma renin activity in normal human subjects. J Clin Endocrinol
Metab 29: 1349-1355, 1969
8. Caldwell BV, Burstein S, Brock WA, Speroff L: Radioimmunoassay
of the F prostaglandins. J Clin Endocrinol Metab 33: 171-175, 1971
9. Romero JC, Ott CE, Aguilo JJ, Torres VE, Strong CG: Role of
prostaglandins in the reversal of one-kidney hypertension in the rab-
bit. Circ Res 37: 683-689, 1975
10. Wilson DM, Romero JC, Strong CG, Lee KE, Schryver SM: Indirect
blood pressure measurements in the rabbit; correlations with direct
aortic and ear pressures. J Lab Clin Med 86: 1032-1039, 1975
11. Torres VE, Strong CG, Romero JC, Wilson DM: Indomethacin en-
hancement of glycerol-induced acute renal failure in rabbits. Kidney
Int 7: 170-178, 1975
12. Goodfriend TL, Levine L, Fasman GD: Antibodies to bradykinin and
angiotensin; a use of carbodiimides in immunology. Science 144:
1344-1346, 1964
13. Mancia G, Romero JC, Shepherd JT: Continuous inhibition of renin
release in dogs by vagally innervated receptors in the cardiopulmonary
region. Circ Res 36: 529-535, 1975
14. McGiff JC, Crowshaw K, Terragno NA, Lonigro AJ, Strand JC,
Williamson MA, Lee JB, Ng KKF: Prostaglandin-like substances
appearing in canine renal venous blood during renal ischemia; their
partial characterization by pharmacologic and chromatographic proce-
dures. Circ Res 27: 765-782, 1970
15. McGiff JC, Crowshaw K, Terragno NA, Lonigro AJ: Release of a
prostaglandin-like substance into renal venous blood in response to
angiotensin II. Circ Res 27 (suppl I): 121-130, 1970
16. Herbaczynska-Cedro K, Vane JR: Prostaglandins as mediators of
reactive hyperaemia in kidney. Nature 247: 492, 1974
17. Herbaczynska-Cedro K, Staszewska-Barczak J, Janczewska H: The
release of prostaglandin-like substances during reactive and functional
hyperemia in the hind leg of the dog. Pol J Pharmacol Pharm 26: 167-
170, 1974
18. Alexander RW, Kent KM, Pisano JJ, Keiser HR, Cooper T: Regula-
tion of postocclusive hyperemia by endogenously synthesized prosta-
glandins in the dog heart. J Clin Invest 55: 1174-1181, 1975
19. Jaffe BM, Parker CW, Marshall GR, Needleman P: Renal concentra-
tions of prostaglandin E in acute and chronic renal ischemia. Biochem
Biophys Res Commun 49: 799-805, 1972
20. Aguilo JJ, Romero JC, Ott CE, Strong CG, Bernatz PE: Prostaglan-
din synthesis blockade in release of renal artery constriction. Surg
Forum 26: 570-572, 1975
21. Oken DE, Arce ML, Wilson DR: Glycerol-induced hemoglobinuric
acute renal failure in the rat. I. Micropuncture study of the develop-
ment of oliguria. J Clin Invest 45: 724-735, 1966
22. Flanigan WJ. Oken DE: Renal micropuncture study of the develop-
ment of anuria in the rat with mercury-induced acute renal failure. J
Clin Invest 44: 449-457, 1965
23. Lonigro AJ, Itskovitz HD, Crowshaw K, McGiff JC: Dependency of
renal blood flow on prostaglandin synthesis in the dog. Circ Res 32:
712-717, 1973
24. Swales JD, Thurston H, Oueiroz FP, Medina A, Holland J: Dual
mechanism for experimental hypertension. Lancet 2: 1181-1184,
1971
25. Romero JC, Hoobler SW, Kozak TJ, Warzynski RJ: Effect of anti-
renin on blood pressure of rabbits with experimental renal hyperten-
sion. Am J Physiol 225: 810-817, 1973
26. Romero JC, Mak SW, Hoobler SW: Effect of blockade of angiotensin
I-converting enzyme on the blood pressure of renal hypertensive
rabbits. Cardiovasc Res 8: 681-687, 1974
27. Ames RP, Borkowski AJ, Sicinski AM, Laragh JH: Prolonged infu-
sions of angiotensin II and norepinephrine and blood pressure, elec-
 RECUPERATIVE POTENTIAL IN HYPERTROPHY AND FAILURE/Cou/son et al.
trolyte balance, and aldosterone and cortisol secretion in normal man
and in cirrhosis with ascites. J Clin Invest 44: 1171-1186, 1965
28. Aiken JW, Vane JR: Intrarenal prostaglandin release attenuates the
renal vasoconstrictor activity of angiotensin. J Pharmacol Exp Ther
184: 678-687, 1973
29. Hoobler SW, Romero JC: Experimental hypertension; the role of
renin in the blood pressure elevation. Excerpta Medica International
Congress Series No. 302, 1973, pp 75-81
30. Romero JC, Strong CG, Torres VE, Ott C, Knox FG: Plasma prosta-
Recuperative Potential of Cardiac Muscle following
Relief of Pressure Overload Hypertrophy and Right
Ventricular Failure in the Cat
RICHARD L. COULSON, SHAHRIAR YAZDANFAR, EMIR RUBIO, ALFRED A. BOVE,
GERALD M. LEMOLE, AND JAMES F. SPANN
SUMMARY This study examined the recuperative potential of
cat hearts subjected to experimental right ventricular pressure
overload (for a 10- to 14-day period) which provoked hypertro-
phy with and without congestive heart failure. Five groups of cats
were studied: normal controls; one group with 70% pulmonary
artery constriction which produced right ventricular hypertrophy
(RVH); one group with an 87% constriction which also produced
right ventricular hypertrophy but with congestive heart failure
(CHF); and two groups which had been similarly subjected to
pressure overload but which had been allowed a recovery period
of 30 days after relief of the pressure overload. Both the 70% and
87% pulmonic constrictions were associated with extensive right
ventricular hypertrophy, depression of myocardial contractile
function, and severe reduction of cardiac norepinephrine stores
(normal, 1.42 /xg/g: RVH, 0.11 /ig/g; CHF, 0.01 /ig/g). After a
30-day period of relief from the pulmonic constriction normal
hemodynamic function returned. In cats in which RVH had been
MYOCARDIAL contractile function and function of the
cardiac sympathetic system are impaired when ventricular
hypertrophy and congestive heart failure result from a
pressure overload on the heart.'"5 There is well established
therapy to relieve the pressure overload; for example,
systemic hypertension can be treated by pharmacological
means and aortic stenosis can be relieved by cardiac sur-
gery. However, relatively little is known of the potential
for recovery of contractile function and repletion of my-
From the Section of Cardiology and the Section of Cardiac and Thoracic
Surgery, Temple University Health Sciences Center, Philadelphia, Penn-
sylvania.
Supported by the Southeastern Pennsylvania Heart Association and by
Grants 5T01-HL 05712 and 3R01-HL 17631 from the National Institutes
of Health. This work was completed during the tenure of fellowships
granted to Dr. Coulson by the Medical Research Council of Canada and
the Canadian Heart Foundation.
Dr. Bove is an Established Investigator, American Heart Association.
Dr. Yazdanfar's present address is: Albert Einstein Medical Center,
Northern Division, Philadelphia, Pennsylvania 19141.
Received July 28, 1975; accepted for publication August 26, 1976.
41
glandins, plasma renin activity and blood pressure in normal and renal
hypertensive rabbits treated with indomethacin. In Abstracts of the
Annual Meeting of the American Society of Nephrology, 1973, p 89
31. Romero JC, Strong CG, Ott CE, Walker R, Schryver S, Manahan D:
The effect of indomethacin on the renin-angiotensin system (abstr).
Clin Res 23: 372A, 1975
32. Romero JC, Dunlap CL, Strong CG: The effect of indomethacin and
other anti-inflammatory drugs on the renin-angiotensin system. J Clin
Invest 58: 282-288, 1976
relieved, right ventricular weight and contractile function were
normal but catecholamine depletion persisted. Cats with relieved
CHF showed depressed contractile function and depleted my-
ocardial norepinephrine, and the right ventricular weight did not
return to normal. Cardiac muscle of all pressure-overloaded
nonrelieved hearts showed depressed velocity of shortening and
depressed ability to sustain load. Cats with RVH alone regained
normal muscle shortening velocity and load-bearing ability after
relief. However, cardiac muscle from the CHF-relieved group
recovered only unloaded shortening velocity while the ability to
sustain load remained depressed. We conclude that the recupera-
tive potential of myocardium damaged by pressure overload is
adequate provided congestive heart failure has not occurred.
Heart failure produces a persistent reduction in force-generating
ability of the myocardium. Hypertrophy due to pressure over-
load, with or without CHF, leads to cardiac catecholamine deple-
tion which is not readily reversed by relief of the overload.
ocardial catecholamine stores following relief of a pressure
overload, despite the clinical relevance of such informa-
tion for the correct timing of therapeutic interventions.e~a
Even less is known of factors that may determine the
potential for return of contractile function and norepi-
nephrine stores. A recent study10 has established that the
contractile defect of hypertrophy due to pressure overload
without heart failure, produced by experimental pulmonic
constriction, is totally relieved after relief for approxi-
mately 4-5 weeks from the pulmonic stenosis.
The mechanics of cat papillary muscle can be described
in a manner similar to that employed for skeletal muscle."
It has been shown that the variation in mechanical muscle
function is small within different groups of cats, thus
functional parameters from different groups can be com-
pared quantitatively.12 Since it is possible by constriction
of the pulmonary artery in the cat to produce right ventric-
ular hypertrophy with and without overt congestive heart
failure,1 a source of myocardium from hypertrophied and
failing hearts is available. In addition, surgical reversal of