Effect of Hypoadrenalism and Excessive
Doses of Desoxycorticosterone Acetate
upon Response of the Rat to Ouabain
By
DAVID UNTERMAN, M.D., ARTHUR C. D E G R A F F ,
AND HERBERT S. KTJPPERMAN, M.D., P H . D .
M.D.,
The influence of alterations of adrenal function upon the responsiveness of the rat to intravenously
administered ouabain was studied electrocardiographically by the string galvanometer. Conduction
disturbances followed by cardiac arrest were the criteria for a definitive response. Adrenalectomized
rats showed a significant increase in sensitivity to ouabain as compared with normal controls.
Normal animals implanted with pellets of desoxycorticosterone acetate and continuously maintained ou saline drinking water did not show a significant alteration in their responsiveness to
ouabain.
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hyperpotassemia and increased intracellular
potassium.10'16 Changes in electrolyte metabolism occurring in such states might be expected to modify the responsiveness of t.hn
animal to digitalis administration.
The relationship between digitalis effect and
potassium metabolism has received considerable attention. The effect of digitalis upon the
potassium content of the myocardium has
been found to be variable. In general, therapeutic doses of digitalis appear to be associated with an increase and toxic doses with a
decrease of potassium in heart muscle.1"8 Exceptions have been noted, however, and recent
studies in dogs have shown no significant
change in the potassium content of cardiac
muscle after lethal doses of lanatoside C.7
The question of altered sensitivity to digitalis in states of potassium depletion lias also
been studied and with more uniform results.
Both experimentally and clinically, an inverse
relationship has been noted. An increased
sensitivity to digitalis has been observed during
potassium depletion and an increased protection against toxic effects during potassium
replacement. 4 ' 7i 8' 9
It was considered desirable to study the
effect of altered adrenal function upon the.
sensitivity of the experimental animal's myocardium to ouabain. It has been demonstrated
that the hypoadrenal state induced by adrenalectomy is associated with marked changes in
electrolyte metabolism characterized by a
METHOD
Albino rats weighing from 100 to 150 grams were
anesthetizel with sodium pentobarbital administered intraperitoneally. The average dose for intact
rats was 3.6 mg. per 100 gms. of body weight. Adrenalectomized rats received approximately twothirds of this dose. Supplementary ether anesthesia
was used if necessary in the course of the experiment.
A string galvanometer (Cambridge) was used for
recording electrocardiograms and photographic
paper was run at an accelerated rate of speed.
Electrodes of 30 gauge copper wire were employed.
Contact was made by winding the wires around the
shaven extremities using a standard electrode jelly
supplemented by saline-moistened gauze pads to
minimize resistance at the skin electrode surface. For
convenience, a board was constructed with two triple
pole double throw switches to permit the rapid successive recording of electrocardiograms in four animals. Control tracings were recorded in the three
standard leads. (Figure 1) Additional tracings using
lead II were taken at 10-minute intervals for a
period of two houis following administration of the
test substance. All injections were given intravenously via the tail vein. Crystalline ouabain was
weighed and freshly dissolved in normal saline immediately before each experiment. The solution was
made up so that the desired dose (from 0.25 to 1.0
mg.) was contained in 1 cc. of the diluent.
From the Department of Therapeutics, New York
University-Bellevue Medical Center College of Medicine.
Received for Publication: December 10, 1954.
The following groups of rats were studied: (1) 4
intact rats which received 1 cc. of normal saline
intravenously; (2) 21 intact rats which received
280
Circulation RcKarci, Volumt III, Ma, IMS
281
OUABAIN AFTER ADRENALECTOMY AND DCA
LEAD 2
LEAD I
LEAD 3
Fit;. 1. F,|pptrnrardiogr;ini of normal rut.
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ouabain intravenously. (4 were given 1.0 mg., 14
were given 0.5 mg. and 3 received 0.25 mg.) (3)
19 udrenulectomized rats which received 0.5 mg. of
ouabain intravenously; (4) Eight animals were implanted subcutaneously with pellets of desoxycorticosterone acetate weighing from 70 to 125 mg.
A total of 16 trials was conducted in this group.
Eight trials were made with 0.5 mg.; four trials with
0.25 mg. and four with 1.0 mg. of ouabain. (5) 17
adrenaleetomized nits received 1 cc. of normal saline
as a control for the adrenalectomized animals receiving ouabain. This was considered desirable since it
is well known that the adrenalectomized animal
shows sensitivity to various dings including anesthetics."
All animals were fed a standard rat diet consisting
of Purina chow checkem and were given tap water
except for the animals implanted with desoxycorticosterone acetate which were maintained on
normal saline.
Adrenalectomies were performed by the lumbar
route under light ether anesthesia. Experiments
were performed from five to seven days following
adrenalectomy. Only animals showing adrenal insufficiency as manifested by significant loss of body
weight were used.
All animals implanted with desoxycortioostarone
acetats gained large amounts of weight,—sometime
to more than twice the control weight. Studies with
ouabain were conducted from 25 to 77 days following the implantation of the D.C.A. pellets.
RESULTS
(Results are summarized in Table 1).
One of the four intact rats of group I that
received normal saline intravenously showed
considerable slowing of the heart from a
control rate of 326 to 160 per minute during
the period of observation. For this reason,
slowing of the rate was not of itself regarded as
a digitalis effect for the purposes of this
experiment. Only definite conduction disturbances such as complete auriculoventricular
dissociation or intraventricular conduction
defects followed by cardiac standstill and death
were accepted as clearcut positive effects.
(Figure 2) Sudden death during the experiment
without antecedent conduction disturbances
TABLE 1.—The Effect of Altered Adrenal Cortical Function upon the Electrocardiographic Response
of the Rat to Ouabain
Animal
Control
Adrenalectomized
Desoxj'corticosterone
Acetate Implants
Preparation
Normal Saline
Ouabain
Normal Saline
Ouabain
Ouabain
of Trials
Dose
4
21
1.0 cc.
1.0 mg. (4 trials)
0.5 mg. (14 trials)
0.25 mg (3 trials)
1.0 cc.
0.5 mg.
1.0 mg. (4 trials)
0.5 mg. (4 trials)
0.25 mg (8 trials)
17
19
16
Average Dose
mg./lOO Cm.
body weight
0.66
0.38 (0.40)*
0.19
—
0.41
0.40
0.26 (0.27)*
0.13
* Overall average in mg. per 100 Gm. of body weight for the 3 dose levels.
Positive Response
Number
0
0
1
1
3
11
1
2
2
Total %
0
(4-8%)
(4.8%)
(17.6%)
(58%)
(6.3%)
(12.5%)
(12.5%)
9.5
17.6
5S
31.3
282
UNTERMAX, D E G R A F F AND KUPPERMAN
CONTROL
IVR-375)
5 MIN
(VR-K6I
15 MIN
IVR.3O0)
HHHf
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Fia. 2.—Electrocardiograms (Lead 2) of adrenalectomized rats, receiving 0.5 mgm Ouabain intravenously, showing positive effects. A. Note progressive slowing of ventricular rate with intraventricular conduction disturbance. B. Note progressive slowing of ventricular rate with appearance of
auriculo-veutricular and intraventricular block. Time in minutes indicates interval following administration of ouabain. Arrows indicate P waves.
was not considered as a positive effect. This
occurred in a few instances before or after the
intravenous injection of either ouabain or
saline and is probably attributable to the
anesthesia. These animals were not included in
the final analysis.
In the group of 21 intacts rats receiving
ouabain, a positive effect was noted in 2
(9.5 per cent). On the other hand, in the group
of 19 adrenalectomized rats receiving ouabain,
11 (58 per cent) had a positive effect. There was
no appreciable difference in ouabain dosage per
unit of body weight between the two groups.
The average dose for the intact rats was 0.40
mg. per 100 gm. of body weight and that for the
adrenalectomized rats was 0.41 mg. per 100
gm. of body weight. There was also no significant difference in dosage in either group
between the animals showing a positive effect
and those showing a negative one.
In the group of 17 adrenalectomized rats
which were given 1 cc. of normal saline intravenously a positive effect was noted in three
(17.6 per cent). As previously mentioned, all
positive effects were characterized by electrocardiographic evidence of marked conduction disturbances followed by cardiac arrest.
In 16 trials conducted in rats implanted with
desoxycorticosterone acetate pellets, ouabain
produced a positive effect in five (31 per cent).
The average dose of ouabain in this group was
0.27 mg. per 100 gm. of body weight and there
was no appreciable dosage difference between
the positive and negative trials (the average
dose was 0.26 mg. per 100 gm. of body weight
for the positive trials and 0.27 mg. per 100 gm.
of body weight for the negative trials).
DISCUSSION
The results indicate a significantly greater
sensitivity of the adrenalectomized rats to the
cardiotoxic effects of ouabain as compared
with intact animals. In evaluating these results, the known sensitivity of the adrenalectomized animals to many noxious pharmacologic agents must be considered. That
conduction disturbances followed by cardiac
arrest in these experiments may not invariably
represent a specific digitalis effect is suggested
by the fact that such changes occurred in three
of 17 adrenalectomized rats receiving saline
intravenously. In addition, it should be noted
that three other adrenalectomized rats included in the above series died while under
OUABAIN AFTER ADRENALECTOMV AXD DCA
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anesthesia; one before receiving saline and the
other two at ten and 120 minutes, respectively,
following the saline injection. In none of these
was any conduction disturbance demonstrable
prior to death. However, the incidence of
clearcut conduction disturbances followed by
cardiac arrest was significantly greater among
the adrenalectomized rats receiving ouabain
than among those receiving saline and may
therefore be attributed in part to a specific
digitalis effect upon the myocardium.
The factors involved in the interrelationship
between altered adrenal-cortical function and
the responsiveness of the experimental animal
to digitalis glycosides may be considered in
terms of the influence of adrenal function on
electrolyte metabolism. Several studies have
demonstrated an increase in cellular potassium
content of skeletal muscle in adrenalectomized
animals, usually associated with an increase in
serum potassium levels.10"16 However, cardiac
muscle failed to show a significant increase in
potassium in at least one study and the same
was true of the liver and kidneys.11 It has also
been suggested that the actual concentration of
skeletal muscle potassium following adrenalectomy remains unchanged because of a
concomitant increase of intracellular water.
The increased sensitivity of the adrenalectomized rat to ouabain cannot be explained in
terms of altered potassium balance. On the contrary, the tendency toward potassium-retention and hyperkalemia in the hypoadrenal
state should lead to greater resistance to
digitalis. At present, it may only be inferred
that some other metabolic changes incident
to hypoadrenalism may account for this increased sensitivity.
Although the present study suggests an
increased sensitivity to ouabain in the rats
maintained upon desoxycorticosterone acetate
implants and saline dnnking water, this difference did not prove to be significant when
compared with the control rats receiving ouabain alone. It should be noted, however, that
the dosage of ouabain per unit of body weight
in the DCA implanted animals was only about
two-thirds that of the controls. Strictly comparable doses of ouabain ]>er unit of body weight may
have yielded a significant difference between
283
the two groups. Since desoxycorticosterone
acetate and saline have a tendency to deplete
the animal of potassium, increased sensitivity
to ouabain might be anticipated on this basis.
In recent experiments in dogs similarly depleted of potassium, increased sensitivity to
lanatoside C was demonstrated by Zeeman,
et al.7 One should note, however, that these
authors employed a continuous infusion of the
glycoside to the point of cardiac arrest rather
than the administration of a fixed dose as used
in the present study.
SUMMARY AND CONCLUSION
The responsiveness of the rat to intravenously administered ouabain was studied
electrocardiographically using conduction disturbances followed by cardiac arrest as the
criteria for positive effect. Four normal rats received saline intravenously and served as
controls. Twenty-one normal and 19 adrenalectomized rats received ouabain intravenously.
An additional 17 adrenalectomized rats received saline intravenously and served as controls for the adrenalectomized ouabain injected animals. In addition, 16 trials with
ouabain were conducted in 8 rats which had
been implanted subcutaneously with pellets of
desoxycorticosterone acetate and maintained
on normal saline solution.
Increased sensitivity to ouabain was observed in the adrenalectomized animals as
compared with the normal controls. A positive
effect characterized by conduction disturbances terminating in cardiac standstill was
noted in 11 of the 19 adrenalectomized rats and
in only 2 of the 21 normal rats. This difference
proved to be highly significant. In the group of
rats maintained on desoxycorticosterone acetate and saline drinking water and receiving
ouabain, a positive effect was noted in 5 of
16 trials. This did not prove to be significantly
different from the incidence noted in the control
rats receiving ouabain alone.
These observations are discussed in terms of
some of the available information regarding the
relationship between potassium balance and
digitalis sensitivity and the altered electrolyte
metabolism in abnormal adrenal states.
284
I'XTKUMAX,
DEGRAFF
AcKNOWLEDd EMENT
Appreciation is expressed to Dr. Norman L.
Heminway, Associate Director, Division of Clinical
Research, Schering Corpomtion for the steroid
hormone preparation used in this study.
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Effect of Hypoadrenalism and Excessive Doses of Desoxycorticosterone Acetate upon
Response of the Rat to Ouabain
DAVID UNTERMAN, ARTHUR C. DEGRAFF and HERBERT S. KUPPERMAN
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Circ Res. 1955;3:280-284
doi: 10.1161/01.RES.3.3.280
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