Assessment of the acute cardiotoxic potential of sodium nitroprusside infusions in Beagle dogs

TOXlCOLOGYANDAPPLIEDRHARhfACOLOGY46,55-64(1978)

Assessment of the Acute Cardiotoxic Potential of Sodium Nitroprusside Infusions in Beagle Dogs1p2 EUGENE HERMAN, TIBOR BALAZS, ROBERT YOUNG, AND Division

of Drug

Biology,

Food and Drug Administration, Welfare, Washington,

Received

October

261977;

U.S. Department D.C. 20204

accepted February

STEPHEN

of Health,

KROP Education,

and

13.1978

Assessment of the Acute Cardiotoxic Potential of Sodium Nitroprusside Infusions in Beagle HERMAN,E., BALAZS,T., YOUNG, R., AND KROP, S. (1978). Toxicol. AppLPharmacol. 46, 55-64. Fourteen unanesthetized dogs were infused with nitroprusside for 30 min (10 or 100 ,&kg/min), 3 hr (25 ,&kg/min), or 6 hr (12.5 or 25.0 pg/kg/min) on two consecutive days. Two animals died following the high-dose 6-hr infusion. Minimal left papillary muscle necrosis was detected in one of the four hearts taken from the dogs infused with 12.5 pg/kg/min for 6 hr. In four anesthetized dogs the 12.5. and 25.0.#g/kg/min doses depressed blood pressure by an average of 20--279/o and 45-4896, respectively. Both doses increased heart rate approximately 20%. The effect of 60-min nitroprusside infusion of 3.5-28.0 pg/kg/min on a variety of cardiovascular functions was assessed in 10 dogs. The most consistently observed changes were a dose-related decrease in blood pressure (19-50%) and an increase in coronary blood flow (30300%). In spite of the hypotensive response, only minor changes in heart rate, cardiac output, and ventricular force of contraction occurred. The results indicate that the secondary cardiovascular actions of nitroprusside are not as prominent as with other vasodilators. Consequently nitroprusside appears to possess less acute cardiotoxic potential than other agents such as minoxidil, diazoxide, or hydralazine. Dogs.

Sodium nitroprusside is a hydrated nitrosylpentacyanoferrate compound (Fig. 1) which rapidly lowers arterial blood pressure. Its action was initially thought to be due to a depression of vasomotor centers (Davidsohn, 1887, cited in Johnson, 1929). Subsequently, the agent was reported to relax isolated smooth muscles, which led to the

2 Na+

--CNCN I , CN-d--CN NO’

1 CN

FIG.

1. Structure of sodium nitroprusside.

recognition that the hypotensive effect was independent of the autonomic nervous system and due to a direct action on peripheral vascular beds (Johnson, 1929; Page et al., 1955). By dilating both arteriolar and venous vessels, nitroprusside reduces vascular ’ This paper was presented in part at the annual spring meeting of the Federation of American Societies for Experimental Biology held in Chicago, Illinois, April, 1977. *Address reprint requests to: Eugene Herman, Food and Drug Administration, Bureau of Drugs (HFD-413),200 C Street, S.W., Washington, DC. 20204. 0041-008X/78/0461-0055$02.00/0 55 Copyright @ 1978 by Academic Press. Inc. All rights of reproduction in any form reserved. Printed in Great Britain

HERMAN ET AL.

56

resistance and venous return; this reduction causes alterations in ventricular afterload and preload and in myocardial oxygen consumption (Schlant et al., 1962; Bhatia and Frolich, 1973). These actions are probably responsible for the renewed interest in the use of nitroprusside for short-term management of hypertensive emergencies (KochWeser, 1974a). In general, cardiovascular responses to nitroprusside are reported to be less severe than with other vasodilating antihypertensive agents (Tuzel, 1974). On the other hand, deaths from the use of nitroprusside for the induction of hypotension during surgery have been reported (Tinker and Michenfelder, 1976). In spite of increasing use of nitroprusside in many clinical situations, the possible acute cardiotoxicity of the agent has been the subject of relatively few experimental investigations. The present studies were initiated to examine certain aspects of the pharmacological action of nitroprusside which might lead to potential adverse cardiac effects. METHODS

Adult beagle dogs of both sexes weighing from 7.8 to 10.2 kg were used in all experiments. Infusions in UnanesthetizedDogs

The right saphenous vein in each of 14 dogs was cannulated with polyethylene tubing by means of a previously inserted thin-wall, 18-gauge stainless-steel needle. The sodium nitroprusside3 was dissolved, just before use, in a volume of 0.9% saline such that a constant flow infusion pump delivered the required dose at a rate of 0.123 ml/min. For 6-hr infusions, fresh nitroprusside was prepared after 3 hr. Before and during the infusions the lead II electrocardiogram (ECG) and heart rate were obtained from subcutaneous electrodes and recorded on a multichannel polygraph. Dogs received sodium nitroprusside with minimal restraint at the following rates and times of infusion on 2 consecutive days: A. 30 min, two dogs at the rate of 10 pg/kg/min, and two dogs at the rate of 100 pg/kg/min; B. 3 hr, two dogs at the rate of 25 ,ug/kg/min; C. 6 hr, four dogs at the rate of 12.5 pg/kg/min, and four dogs at the rate of 25.0 hg/kg/min. In each of the experiments, dogs were killed by electrocution 24 hr after the last infusion of nitroprusside, and necropsy was performed immediately. The hearts were examined grossly, and sections were prepared from the left ventricular anterior and posterior papillary muscles, right ventricular anterior, middle, and posterior papillary muscles, and left ventricular free wall ,@d left ventricular septal wall. The sections were stained with hematoxylin and eosin and examined under the light microscope. Infusions in Anesthetized Dogs Three-hour infusion. Four dogs were anesthetized with 30 mglkg of pentobarbital sodium. Cannulas were placed in the right femoral artery and vein for monitoring arterial blood pressure and drug injection, respectively. Subcutaneous electrodes were inserted in the appropriate limbs for monitoring the lead II ECG and heart rate. 3 Nipride, Ayerst Laboratories, New York, N.Y.

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Nitroprusside was infused for 3 hr in two dogs at a dose of 12.5 puglkglmin and in two dogs at 25.0 pg/kg/min. Each dose of nitroprusside was freshly dissolved in 0.9% saline at a concentration to allow the pump infusion rate to be set at 0.123 ml/min. All physiological parameters were recorded continuously on a Hewlett-Packard polygraph before, during, and up to 30 min after cessation of the nitroprusside infusion. Cardiovascular function experiments. Ten dogs were anesthetized with sodium pentobarbital (30 mg/kg, iv), and cannulas were inserted into the right femoral vein and artery and the trachea. The tracheal cannula was connected to a positive-pressure respirator; the heart was then exposed by means of a thoracotomy at the left fifth intercostal space. A 6- to 8-mm section of the left circumtlex coronary artery was cleared and a 2.0-mm Biotronix electromagnetic flow probe was inserted around the artery. Similarly, a section of the pulmonary artery was cleared of surrounding tissue, and a 12.0-mm Biotronix electromagnetic flow probe was inserted around this artery. A Walton-Brodie strain gauge was sutured to the left ventricular myocardium (Boniface et al., 1953). The chest was sutured closed, and the animal was allowed to resume spontaneous respiration. Systolic, diastolic, and mean arterial pressure were monitored from a pressure transducer attached to the femoral artery cannula. The lead II ECG and heart rate were recorded from subcutaneous needle-tipped electrodes, and heart rate was recorded by a cardiotachometer triggered by the R wave of the ECG. Changes in coronary blood flow and cardiac output were obtained from the left circumflex coronary artery and pulmonary artery probes attached to an electromagnetic flowmeter. All parameters were recorded on a multichannel polygraph. Once spontaneous respiration resumed, the animals were allowed to stabilize 30 min before the drug was administered. Sodium nitroprusside was infused by means of a constant flow infusion pump into the femoral vein. Two doses of nitroprusside were given during each experiment. The first dose, either 3.5 or 7.0 pg/kg/min, was infused for 60 min. After a 30-min recovery period (sufficient for blood pressure and heart rate to return to control value) a second dose of 14.0 (following 3.5) or 28.0 (following 7.0) ,ug/kglmin was infused for 60 min. Each pair of doses was given to five dogs. The nitroprusside was dissolved in 0.9% saline in a concentration such that the infusion rate was 0.123 ml/min (3.5- and 7.0;uglkglmin dose) or 0.25 ml/min (14.0- and 28.0~pg/kg/min dose). All experiments were terminated 30 min after the second infusion. The mean percentage change from control +_SE in mean arterial pressure, heart rate, force of contraction, coronary blood flow, and cardiac output were calculated for each of the four infusion doses.

RESULTS

Unanesthetized Dogs Thirty-minute infusions. The infusion of nitroprusside produced dose-related pharmacological effects. The two dogs receiving the low dose (10 pglkglmin) appeared essentially normal with no electrocardiographic changes noted during the infusion period.

HERMAN

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ET AL.

The two dogs receiving the high dose (100 ~glkglmin) vomited several times shortly after the infusion started. Later, both dogs defecated and urinated a little; both appeared to become disoriented at times with vocalization, struggling, and difficulty in standing toward the end of the infusion. The respiratory rate increased noticeably in both dogs. One dog developed ventricular ectopic beats during the last 5 min of the infusion. Both dogs appeared to recover somewhat from the effects of this dose of nitroprusside by 1 hr after termination of the infusion. These two dogs showed similar effects (vomiting, defecation, and urination) on the second day of infusion but with less frequency and intensity. The results of gross and histopathologic examination of hearts from these dogs and others infused with nitroprusside are summarized in Table 1. TABLE SUMMARY OF UNANESTHETIZED

1

GROSS AND HISTOPATHOLOGICAL MYOCAR~IAL ALTERATIONS BEAGLE DOGS AFTER Two CONSECUTIVE DAILY INFUSIONS NITROPRUSSIDE

Gross observations Nitroprusside dose rate (ug/kg/min)

Duration (mid

No. of animals

Myocardial hemorrhage

10 100 25 12.5 25

30 30 180 360 360

2 2 2

None None t a None

Myocardial necrosis None None None None None

SEEN IN OF SODIUM

Microscopic observations Myocardial hemorrhage None None None None None

Myocardial necrosis None None None

Minimal (4) None

0 Two died after first day.

No treatment-related gross or microscopic alterations were seen in any of these four hearts. Three-hour infusions. One of the dogs infused with 25 ,ug/kg/min vomited on both days. Both dogs were restless at the beginning of the infusion but were relatively calm when the infusion was terminated 3 hr later. The heart rate increased in both dogs, but no abnormal ECG patterns were observed during the experimental period. Myocardial necrosis was not evident in any of these hearts. In one of the four hearts, fresh petechial epicardial hemorrhages were noted in the adipose tissue adjacent to some coronary vessels. These hemorrhages have also been noted occasionally in control dogs and are probably due to electrocution. Hemorrhage was not detected in the histopathologic examination of the subendocardium. Six-hour infusions. At a dose of 12.5 ,ug/kg/min, nitroprusside induced one or two episodes of vomiting in each of four dogs. A slight to moderate tachycardia occurred in all dogs but was readily reversible once the infusion was stopped (Fig. 2). On gross observation no evidence of myocardial necrosis was seen in these hearts, but a very minimal focal area of left ventricular papillary muscle necrosis was found in one heart under microscopic examination. Fresh petechial hemorrhages were seen in one heart, as described above. The dogs infused with 25.0 pg/kg/min of nitroprusside vomited, defecated, and

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CARDIOTOXICITY

urinated at various times after the infusion began. During the initial 30 min the animals were restless, but they became more subdued and began to salivate and pant as the infusion progressed. On the first day, two dogs developed tonic-clonic convulsions during the fourth hour of infusion. At this point the infusion was terminated in one dog, but the dog died 1.5 hr later; the second dog died overnight following the first day’s infusion. This dog was depressed at the end of the infusion and was the only one to develop ECG abnormalities such as premature ventricular beats during the infusion. In

180

NITROPRUSSIDE INFUSION IpgIKgIMINI

a0 -

-

125

-

12.5 DAY2

-

250

RATE

DAY 1

DAY 1

60 I 01530

M I

P-INFUSION

I 60

25.0+ DAY 2 (AVERAGE I 120

2 ANIMALS1 I

180

ON

I 240

I 300

1 I 360 15 J,INFUSION ’ OFF

TIME (MINI

2. Changes in heart rate during 6-hr iv infusion of either 12.5 or 25.0 pg/kg/min of sodium nitroprusside in unanesthesized beagle dogs. Values obtained were converted to percentages of control. Each point of 12.5 pg/kg/min on Days 1 and 2 and 25.0 &kg/min on Day 1 represents the mean f SE of four separate determinations. The points on Day 2,25.0 pg/kg/min, represent the average of two animals. FIG.

general heart rates increased 20-50% shortly after the infusion began (Fig. 2) and varied a great deal during the first day, but increases were more consistent in the two surviving dogs on the second day. Gross and microscopic examination revealed no abnormalities in the hearts from these two animals. Anesthetized Dogs Three-hour infusions. In these experiments

,ug/kg/min

two dogs were infused with 12.5 and two dogs with 25.0 .ug/kg/min of nitroprusside for 3 hr. The effect of

HERMAN

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ET AL.

these infusions on mean arterial blood pressure and heart rate are summarized in Fig. 3. The low dose lowered blood pressure 20-27% compared to 45-48% after the high dose. Simultaneously both doses of nitroprusside increased heart rate; this increase, although variable, averaged approximately 20% or more at all time intervals. Changes in both

NlTROPRUSSlOE INFUSION IpgIKgIMIN) 150

M

I

RATE

25.0

I

I

I

I

I

I

15

30

60

120

180

15

I--INFUSION

0Nd

OFF

TIME (MINI

FIG. 3. Effect of 3-hr infusion of sodium nitroprusside on mean arterial pressure and heart rate in four anesthetized dogs given doses of 12.5 pg/kg/min (2) or 25.0 @g/kg/min. Values obtained were converted to percentages of control. Each point represents the mean of two separate determinations.

parameters occurred almost immediately after the infusion was started and were completely reversed within 15 to 30 min after the infusion ended. Cardiovascularfunction studies.The effect of 3.5, 7.0, 14.0, or 28.0;ug/kg/min nitroprusside infusions on a variety of cardiovascular parameters is summarized in Fig. 4. The most consistently observed changes were in mean arterial pressure and coronary blood flow. Mean arterial blood pressure was depressed approximately 19% by the

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lowest dose (3.5 ,&kg/mm) to 56% by the highest dose (28.0 &kg/mm). Coronary blood flow increased in a dose-related manner from 33% after the low dose to 300% after the high dose. In spite of the hypotensive response, only minor increases in heart

NITROPRUSSIDE INFUSION WKgIMINl

35” 300

RATE

M

3.5

-

14.0

-

7.0

-

28.0

120 LL

100

I 1

TIME OF INFUSION

IMINI

FIG. 4. Effect of 60min infusions of 3.5, 7.0, 14.0, or 28.0 pg/kg/min of sodium nitroprusside on a variety of cardiovascular functions in the anesthesized beagle dog. Values obtained were converted to percentages of control. Each point represents the mean + SE of five separate determinations.

rate occurred during any of the infusions, perhaps because control rates in these dogs averaged about 150 beats/min compared to 100 beats/min in the unanesthetized dogs. Cardiac output fell 12-17% during the 3.5- and 7.0~pg/kg/min infusions while left ventricular force of contraction increased approximately 20% after the high dose.

62

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ETAL.

Changesin all parametersbegan shortly after the infusion was begun and returned to control levels within 15min onceinfusion was ended. DISCUSSION

All doses of sodium nitroprusside produced a rapid fall in arterial pressure on iv administration. The hypotension seenin anesthetizeddogs was accompaniedby doserelated increasesin coronary blood flow with minimal elevations in heart rate and other indices of cardiac function. Rowe and Henderson (1974), giving a similar nitroprusside dose to anesthetizeddogs, found a 30% increase in cardiac output together with declinesin total peripheral resistance,pulmonary vascular resistance,and right arterial pressure. They found that nitroprusside induced less hypotension than that which occurred in our studies, but in both instances coronary blood flow increased significantly. The reduction in arterial pressure and the associateddecline in total peripheral resistancehas beendocumentedin virtually every study of nitroprusside and forms the basisfor its use in treatment of various types of heart failure. Besidesnitroprusside, other vasodilators such as minoxidil, diazoxide, and hydralazine decrease peripheral resistance and arterial blood pressure. The hypotension associated with these three agents leads in certain instances to clinical signs of palpitation, angina pectoris, and rarely, myocardial necrosis (Hamby et al., 1968; Koch-Weser, 1974b; Moyer, 1953; Mellen, 1970; Pettinger and Mitchell, 1973). Infusions of high dosesof nitroprusside for up to 180 min have failed to produce any evidence of myocardial cellular damage. When the infusion was extendedto 360 min (12.5 or 25.0 pg/kg/min), a time frame in other experimentsin which minoxidil has produced lesions,minimal necrosiswas detectedin only one of six hearts.It should be noted that the 25.0~pg/kg/min dose was essentiallyan LDSOin the present studies,and yet the myocardia from the two surviving animals were normal. The lack of significant acute cardiac toxicity implies important differences in the effects of nitroprusside and the other vasodilating drugs such as minoxidil or diazoxide on the heart. Clinically, changes in pulse rates accompanying nitroprusside hypotension have been reported (Page et al., 1955; Maraca et al., 1962). In one clinical study both nitroprusside and diazoxide significantly reduced arterial pressure and reflexly increasedheart rate (Maraca et al., 1962).The increasein rate was significantly greater with diazoxide. In addition, diazoxide increased cardiac output and left ventricular ejectionrate. Reflex tachycardia in the presence of hypotension, as a consequenceof the pharmacological effects of minoxidil, diazoxide, or hydralazine, appears to be an important factor in maintaining the functional integrity of the myocardium. Experimental acute myocardial necrosiscan be detectedafter short-term treatment with thesethree agents(Balazsand Payne, 1971; Balazset al., 1975; Herman et al., 1975).Tachycardia itself may not produce ischemia; however, the concurrent hypotension with its effects on coronary circulation and ultimately capillary perfusion pressure impairs the blood supply to the myocardium. In separatestudies(Herman, E., Balazs,T., Young, R., Earl, F., and Krop, S., unpublished observations) where arterial blood pressure was decreasednearly 50%, minoxidil (3 mg/kg) increased coronary blood flow approximately 25% while under similar hypotensive conditions in the present experimentsthe

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increase after nitroprusside was over 330%. Thus the impairment in coronary blood flow due to low perfusion would appear to be more serious with minoxidil than with nitroprusside. Coronary blood flow is further hindered by reflex tachycardia because oxygen demand increases and the diastolic period, when most perfusion occurs, is reduced. Experimentally, increases in heart rate of 100% or more have been observed in beagle dogs after certain doses of minoxidil (Herman et al., 1975) or diazoxide (Balazs er al., 1975). In contrast, heart rate rarely increased above 50% during high-dose nitroprusside infusions. At present it is not known whether the apparent reduction in magnitude of tachycardia is due to a direct myocardial suppressant effect. In any event, a significant increase in coronary blood flow and a lower level of reflex tachycardia may be some of the factors which reduce the potential acute cardiac toxicity of nitroprusside. While the hemodynamic effects of nitroprusside administration do not appear as likely to produce acute cardiotoxicity as other vasodilating compounds, one additional aspect of the compound is of concern. The nitroprusside molecule contains five cyanide groups which may be released by a reaction with hemoglobin in red blood cells and then transformed at least in part to thiocyanate (Tinker and Michenfelder, 1976). In one instance, a child treated with nitroprusside developed an anginal-like syndrome which the investigators felt might be due to cyanide and thiocyanate interference with myocardial oxygen utilization (Palmer and Lasseter, 1975). This aspect of nitroprusside toxicity deserves further attention. ACKNOWLEDGMENTS

Histopathological examinations were performed by Drs. W. Busey and J. Ferrell, Experimental Pathology Laboratories, Herndon, Virginia. The authors thank Mr. Donald Gates and Mr. Arlen Sager for technical assistanceduring the experiments. REFERENCES

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