Companrson of Cardiac Output Responses to 2,4-Dinitrophenol- Induced Hypermetabolism and Muscular Work CHANG-SENG LIANG and WILLiAM B. HOOD, JR. From the Division of Medicine, Boston University School of Medicine, the Cardiology Division, Boston University Medical Service, Boston City Hospital, and the Departments of Medicine and Clinical Research, University Hospital, Boston, Massachusetts 02118 A B S TR A C T Both electrically induced exercise and infusion of 2,4-dinitrophenol (DNP) increased oxygen consumption and tissue metabolism in chloralose-anes- thetized dogs. Cardiac output increased with oxygen consumption at the same rate in both experimental con- ditions. The increase in cardiac output induced by exer- cise was, as expected, accompanied by increases in both lactate-to-pyruvate ratio and "excess lactate" in arterial blood. However, these parameters did not increase after DNP infusion until the rate of oxygen consumption had increased four- to fivefold, perhaps due to facilitation of mitochondrial electron transport by DNP. Anaerobic tissue metabolism therefore probably did not contribute significantly to increased cardiac output during the mild- to-moderate tissue hypermetabolism induced by DNP. The increased cardiac output may have been the result of metabolic changes common to both exercise and DNP infusion; muscular activity alone may not have been the primary determinant of the cardiac output response during exercise. INTRODUCTION The principal factor initiating increased cardiac output during exercise, the "work stimulus," still is not fully understood (1). Both mechanical and metabolic factors have been identified with the "work stimulus." The former have been related to the stimulation of mechani- cal sensory receptors inside the muscles or tendons, and This work was presented in part at the Annual Meeting of the American Federation for Clinical Research in At- lantic City, N. J., 29 April 1973. Dr. Liang was a recipient of a National Institutes of Health Special Research Fellowship (1 F3 HL 52986). Received for publication 18 January 1973 and in revised form 5 April 1973. the latter to metabolic changes that occur in exercising muscles. In this study, dogs were infused with 2,4-dinitrophenol (DNP)1 to produce tissue hypermetabolic changes fundamentally like those that occur in exercise but not associated with muscular movement. The cardiac output responses to DNP infusion and exercise were com- pared to assess the relative contributions of mechanical and metabolic factors in the regulation of cardiac output. The rate of oxygen consumption was taken as the index of tissue metabolism. In addition, concentrations of lactate and pyruvate in arterial blood were measured. A change in the lactate- to-pyruvate ratio indicates the presence or absence of tissue hypoxia (2), and its correlation with cardiac output provides an estimate of the role of anaerobic tissue metabolism in the cardiac output response to tis- sue hypermetabolism. METHODS Studies were performed on 22 healthy dogs of both sexes weighing between 14 and 33 kg. Anesthesia was induced with vaporized methoxyflurane, (Penthrane, Abbott Labora- tories, North Chicago, Ill.) followed by intravenous chlora- lose (60 mg/kg). The trachea was cannulated with a T tube connected to a Benedict-Roth metabolic spirometer to record the rate of oxygen consumption. A femoral artery and the right ventricle were cannulated and connected to pres- sure transducers and a Sanborn 7700 recorder (Hewlett- Packard Co., Waltham Div., Waltham, Mass.) to measure blood pressures and heart rate. Another catheter was in- serted into a carotid artery and connected to a Gilford Col- son 103 densitometer (Gilford Instrument Laboratories, Inc., Oberlin, Ohio) to determine cardiac output (3) with indo- cyanine green (Cardio-Green, kindly supplied by Hynson, 1Abbreviations used in this paper: DNP, 2,4-dinitro- phenol; LIP, lactate to pyrqva;e; PC, phosphorylcreatine; XL, excess lactate. The Journal of Clinical Investigation Volume 52 September 1973*2283-2292 2283
Transcript
Companrson of Cardiac Output Responses to 2,4-Dinitrophenol-
Induced Hypermetabolism and Muscular Work
CHANG-SENGLIANG and WILLiAM B. HOOD,JR.
From the Division of Medicine, Boston University School of Medicine, theCardiology Division, Boston University Medical Service, Boston City Hospital,and the Departments of Medicine and Clinical Research, University Hospital,Boston, Massachusetts 02118
A B S T R A C T Both electrically induced exercise andinfusion of 2,4-dinitrophenol (DNP) increased oxygenconsumption and tissue metabolism in chloralose-anes-thetized dogs. Cardiac output increased with oxygenconsumption at the same rate in both experimental con-ditions. The increase in cardiac output induced by exer-cise was, as expected, accompanied by increases in bothlactate-to-pyruvate ratio and "excess lactate" in arterialblood. However, these parameters did not increase afterDNPinfusion until the rate of oxygen consumption hadincreased four- to fivefold, perhaps due to facilitationof mitochondrial electron transport by DNP. Anaerobictissue metabolism therefore probably did not contributesignificantly to increased cardiac output during the mild-to-moderate tissue hypermetabolism induced by DNP.The increased cardiac output may have been the resultof metabolic changes common to both exercise and DNPinfusion; muscular activity alone may not have beenthe primary determinant of the cardiac output responseduring exercise.
INTRODUCTIONThe principal factor initiating increased cardiac outputduring exercise, the "work stimulus," still is not fullyunderstood (1). Both mechanical and metabolic factorshave been identified with the "work stimulus." Theformer have been related to the stimulation of mechani-cal sensory receptors inside the muscles or tendons, and
This work was presented in part at the Annual Meetingof the American Federation for Clinical Research in At-lantic City, N. J., 29 April 1973.
Dr. Liang was a recipient of a National Institutes ofHealth Special Research Fellowship (1 F3 HL 52986).
Received for publication 18 January 1973 and in revisedform 5 April 1973.
the latter to metabolic changes that occur in exercisingmuscles.
In this study, dogs were infused with 2,4-dinitrophenol(DNP)1 to produce tissue hypermetabolic changesfundamentally like those that occur in exercise but notassociated with muscular movement. The cardiac outputresponses to DNP infusion and exercise were com-pared to assess the relative contributions of mechanicaland metabolic factors in the regulation of cardiac output.The rate of oxygen consumption was taken as the indexof tissue metabolism.
In addition, concentrations of lactate and pyruvate inarterial blood were measured. A change in the lactate-to-pyruvate ratio indicates the presence or absence oftissue hypoxia (2), and its correlation with cardiacoutput provides an estimate of the role of anaerobictissue metabolism in the cardiac output response to tis-sue hypermetabolism.
METHODSStudies were performed on 22 healthy dogs of both sexesweighing between 14 and 33 kg. Anesthesia was inducedwith vaporized methoxyflurane, (Penthrane, Abbott Labora-tories, North Chicago, Ill.) followed by intravenous chlora-lose (60 mg/kg). The trachea was cannulated with a T tubeconnected to a Benedict-Roth metabolic spirometer to recordthe rate of oxygen consumption. A femoral artery andthe right ventricle were cannulated and connected to pres-sure transducers and a Sanborn 7700 recorder (Hewlett-Packard Co., Waltham Div., Waltham, Mass.) to measureblood pressures and heart rate. Another catheter was in-serted into a carotid artery and connected to a Gilford Col-son 103 densitometer (Gilford Instrument Laboratories, Inc.,Oberlin, Ohio) to determine cardiac output (3) with indo-cyanine green (Cardio-Green, kindly supplied by Hynson,
1Abbreviations used in this paper: DNP, 2,4-dinitro-phenol; LIP, lactate to pyrqva;e; PC, phosphorylcreatine;XL, excess lactate.
The Journal of Clinical Investigation Volume 52 September 1973*2283-2292 2283
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2284 C. Liang and W. B. Hood, Jr.
Westcott & Dunning, Inc., Baltimore, Md.). The dye curves,registered on a linear Hewlett-Packard 1701 BM strip chartrecorder (Hewlett-Packard Co., Palo Alto, Calif.), werecorrected for recirculation of dye, and the area measuredplanimetrically. Mean arterial blood pressure was dividedby cardiac output to calculate total peripheral vascularresistance.
Cardiac output was also measured by the Fick prin-ciple, and these data were used exclusively for statisticalcomparisons (see Results). Outputs vere obtained by draw-ing simultaneous 4-ml samples of right ventricular and ar-terial blood over a period of 2 min and determining therate of oxygen consumption during the same interval. How-ever, during exercise, the blood was sampled at doublespeed. The blood samples were analyzed promptly for oxy-gen content using a Beckman GC-2A gas chromatograph(Beckman Instruments, Fullerton, Calif.) by the method ofRamsey (4); in our laboratory, this method yields resultsconsistent with those of the method described by Van Slykeand Neill (5).
The oxygen capacity of blood was determined by a cyan-methemoglobin method (6). Oxygen saturation was calcu-lated by dividing blood oxygen content by oxygen capacity.Blood pH and gas tensions were measured on a RadiometerPHM71 Acid Base Analyzer (The London Company, West-lake, Ohio).
To determine lactate and pyruvate concenitrations, arterialblood was allowed to flow freely into a tube of ice-cooledtrichloroacetic acid. The filtrate was analyzed for lactateby the enzymatic method of Friedland and Dietrich (7),and for pyruvate by the method of Friedemann and Haugen,as modified by Huckabee (8). Blood water content wasdetermined by the reduction in weight after drying. Con-centrations of "excess lactate" (XL) were calculated fromchanges in pyruvate and lactate concentrations by the fol-lowing equation (2):
XL = (Ln-L.)- (P- -P.) (L.IPo)where L. and L. and P0 and P. are lactate and pyruvateconcentrations in arterial blood water in control and experi-mental conditions.
The experimental animals were divided into two groups.In one group of eight dogs, tissue hypermetabolism wasinduced by three or four successive infusions of a 200 mg/100 ml DNP aqueous neutral solution, spaced at 15-minintervals. The dose of DNPfor each infusion was 2 mg/kggiven into the femoral artery catheter over a 2-min period.In another group of 14 dogs, muscular work was inducedby direct stimulation of limb muscles with electrical pulsesof 3/s frequency, 5 ms duration, and 80 V intensity de-livered from a Grass SD 5 stimulator (Grass InstrumentCo., Quincy, Mass.) over a period of 10 min. Each dogwas stimulated once.
Circulatory and metabolic measurements were made atfrequent intervals before and after both kinds of experi-mental intervention. Preliminary experiments using the in-dicator dilution method showed that cardiac output rose toa steady state within 3 min after onset of exercise andwithin 5 min after each infusion of DNP. Therefore it wasdecided to obtain Fick cardiac outputs between 6 and 9 minafter the start of exercise and DNPinfusion.
In each group of experiments, the significance of differ-ences between control and experimental values was deter-mined using t tests for paired comparisons. Correlationsand regression coefficients were computed for both groups ofanimals, and the slopes of the regression lines for thesetwo groups were compared (9).
RESULTS
Experimental results obtained in animals that receivedDNP infusions are shown in Table I. Table II providesa statistical summary of both exercise and DNP infu-sion experiments. With the exception of stroke volume,there were not significant differences between the controlvalues for the two groups of animals, as determined bytunpaired t tests.
Oxygen consumption and cardiac output. The rate ofoxygen consumption increased 2-10-fold after DNP in-fusion and exercise, and was accompanied by an in-crease in cardiac output. Fig. 1 illustrates the stepwiseincrease of both oxygen consumption and cardiac outputaftet successive doses of DNP. Fig. 2 shows that cardiacoutput increased with oxygen consumption at the samerate in both groups of dogs. The regression coefficientand its standard error were 5.460 and 0.462 for theDNP-treated dogs, and 5.235 and 0.655 for the dogssubjected to electrically induced exercise. There wasno difference between the regression coefficients of thesetwo groups (t = 0.27, P > 0.5).
The slope of the regression line in Fig. 2, which is theratio of increment in cardiac output to increment in oxy-gen consumption (AQ/AVo2), is called the "exercisefactor" when employed in studies of exercise (10). Ithas been shown repeatedly (11-16) that cardiac outputincreases linearly with oxygen consumption during ex-ercise, to rates of oxygen consumption up to 10-15times the normal resting value. The calculated AQ/AVo2is constant over this range of oxygen consumption. Thisratio was calculated for individual animals with bothexercise and DNP infusion; their means in these twogroups did not differ significantly (Table II).
Heart rate and stroke volume. Fig. 3 shows that theincreases in cardiac output were associated with in-creases in both heart rate and stroke volume in bothgroups of animals.
Arterial and right ventricular pressures and totalperipheral vascular resistance. Mean arterial bloodpressure did not change significantly in dogs after DNPinfusion (Tables I and II), whereas it fell immediately20-25% below control after initiation of electrically in-duced exercise. As exercise continued, it rose and stabi-lized at about 15% below the control (Table II). Rightventricular end-diastolic pressure did not change sig-nificantly after DNP infusion. Right ventricular pres-sure tracings for dogs undergoing exercise were dis-torted and are not reported. The calculated total periph-eral vascular resistance was reduced both with exerciseand DNP infusion, and its changes were inversely re-lated to changes in cardiac output.
Blood oxygen saturation and blood gases. Arterialblood was fully saturated with oxygen in dogs at rest,
Cardiac Output and Tissue Hypermetabolism 2285
TABLE I IStatistical Summary of
Experi-mentalcondi-
Wt tions Vo2 Q HR sv P TPVR
kg ml/kg/min mlikglmin beats/min ml/kg mmHg mmHg,'liier/min
2, 4-Dinitrophenol19.640.2 A 4.92±0.37 153.8±14.2 111.9+7.2 1.37±0.08 143.0±5.9 50.2±4.6
A, control state; B, C, D, anid E, after the first, second, third, and fourth infusions of DNP (2 mg/kg); Wt, body weight;HR, heart rate; [L], Lactate concentration in arterial blood water; Pa, mean arterial blood pressure; [P], pyruvate con-centration in arterial blood water; Paco2, arterial blood carbon dioxide tension; SV, stroke voluime; SV,o2, right ventricularblood oxygen saturation; TPVR, total peripheral vascular resistance.
after DNP infusion, and during exercise. On the otherhand, right ventricular blood oxygen saturation fell asoxygen consumption increased in both groups of animals(Fig. 4). Neither DNP infusion nor exercise producedsignificant changes in arterial pH or Pco2 (Table II).
Arterial blood lactate and pyruvate concentrations.Arterial blood concentrations of lactate and pyruvate fell
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progressively reachinig their lowest values of 40-50% ofthe control after the second infusion. With additional(loses of DNP, both lactate and pyruvate increased. Inthree experiments, lactate concentrations even rose abovethe control values after the third infusion (Table I). Onthe other hand, exercise was consistently accompanied byincreased concentrations of lactate and pyruvate; at no
10DNP DNP r
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Oxygen Consumption
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CONrROL M/NUTES
FIGURE 1 A typical experiment showing the changes in oxygen consumption, cardiac outputand L/P ratio in a dog after four successive infusions of 2,4-DNP. Straight lines depictingoxygen consumption were calculated from the slopes of spirometer tracings taken at thosetime intervals.
2286 C. Liang and W. B. Hood, Jr.
Experimental Results
Syo2 P.cO2 pH AQ/[Vo, tL] [P] L/P XL
% mmHg mmoles mmoles mmolesper liter per liter per liter
time did either of them fall below control values (Table no XL was formed after the first two infusions of DNP;II). but after the third infusion, L/P ratio rose slightly and
Lactate to pyruvate ratio and "excess lactate." Lac- XL appeared (Fig. 1, Table I). On the other hand, ex-tate to pyruvate (L/P) ratio was reduced slightly and ercise was always accompanied by increases in both
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OXYGENCO,,'SUPiii7O0,V (ml, STPO/min)FIGURE 2 A graphic representation of the relationship between cardiac output and oxygen
consumption in dogs both during exercise and after DNPinfusions.
Cardiac Output and Tissue Hypermetabolism 2287
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FiGURE 3 Relationships of heart rate and stroke volume to cardiac output ininfusions (A), and during exercise (B).
L,/P ratio and XL. The mieans of these two parametersduring exercise were significantly higher than those ob-served after DNP infusion (P < 0.01).
DISCUSSIONIn the l)resent experiments, oxygen consumption was in-
creased in dogs by both DNP infusion and electricallvinduced exercise. DNP uncouples oxidation and phos-phorylationi in the mitochondrial cvtochromes (17).resulting in a net increase in ADP an(l a (lecrease inATP (18, 19). ATP hydrolysis also occurs int muscles
during contraction (20, 21). The increasedl availabilityof tissue ADP in turn increases tissue metabolism and
oxygen consumption (22, 23). Thus, the hvpermetabo-lism induced by DNP is a result of mechaniismis basicallvsimilar to those occurring during exercise.
The validity of the assumption that electrically in-
duced work simulates spontaneous exercise has beenpreviously established (24, 25). Fig. 2 shows thatcardiac output increased with oxygen consumptionduring electrically induced exercise. The regression lineis similar to those obtained in unaniesthetized dogs run-
ning on a treadmill (11, 12).Both heart rate and stroke voluiie were augnmeinted
during electrically induced exercise, but systemic ar-
terial blood pressure was reduced (Fig. 3, Table II).A similar change in arterial blood pressure was observedearlier by Euler and Liljestrand (26) using the same
method of muscle stimulation in chloralose-anesthetized
dogs after DNP
animals, but it did not occuir when muscular work was in-duced by ventral root stimulation (27). Therefore, itappears likely that the hypotensive response was peculiarto the site of electrical stimulation employed in our
exp)eriments.DNP-indltced hypermietabolisnm was accompanied by
an increase in cardiac output comparable to that whichocculrredl during exercise (Fig. 2). Furthermore, as
occurred during exercise, the increase in cardiac outputafter DNP infusion was brought about by increases in
both heart rate and stroke volume (Fig. 3). DNPinfu-sion has been reported to increase cardiac output in in-tact dogs (28, 29). However, these reports did not pro-
vide sufficient data to allow quantitative comparison withour experimental results. Before we can attribute thisincreased cardiac output to the metabolic effects of DNPon peripheral tissues, the effects of DNPon the heart, ar-
terial chenmoreceptors, and central nervous systenm mustbe considered. Fawaz anid Tutunji (30) found that DNPdidl not change left ventricular output in a heart-lungpreparation within 1 h, even at concentrations more
than 10 times those achieved in the present experiments;left ventricular output fell after more prolonged DNPexposure. DNPis capable of stimulating carotid chemo-receptors (31, 32). The immediate and transient byper-ventilation and hypertension which occur when DNP isinjected into the carotid artery are abolished by thedenervation of carotid chemoreceptors. This indicatesthat these changes are not caused by direct effects of
2288 C. Liang and W. B. Hood, Jr.
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A. DNP (n -32)r 0.582
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B. EXERCISE (n =26)r* 0.641
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Decreasing -
80 60 40 20 0 80 60 40 20 0
RIGHT VENTRICULAR BLOOD OXYGENSATURATION (%)
FIGURE 4 Relationship of cardiac output to right ventricular blood oxygen saturation indogs after DNP infusions (A), and during exercise (B).
DNP on the central nervous system. Relatively largeconcentrations of DNP are required to stimulate caro-
tid chemoreceptors (31, 32). Similarly high concentra-tions probably were not attained in the present experi-ments because the observed ventilatory and circulatoryresponses were gradual in onset and in 5 min reacheda steady state well adjusted to the tissue metabolic ratewith no change in arterial blood Pco2. Carbon dioxidetension fell only during severe hypermetabolism. Norin all likelihood was the central nervous svstem im-portant in the regulation of the cardiac output response
to DNP infusion because. as shown by Banet andGuyton (29). cardiac output increased normally afterDNP infusion in decapitated (logs as long as arterialblood pressure was maintained. Furthernmore, using a
cross-perfusion technique in which the head of one dog(recipient) was perfused by a second dog (donor),Levine and Huckabee (33) found that DNP. when givento the head of the recipient dog, did not produce any
change in the recipient dog's ventilation, whereas thedonor dog's ventilatory rate increased markedly. On thebasis of these results from other investigators, it seems
likely that the hemodynamic and respiratorx changes ob-served in intact dogs after DNP infusion were relatedto the hypermetabolic state created by DNP in periph-eral tissues.
Both isocapnic (hyperpnea) and hypocapnic hyper-
ventilation occurred in dogs after DNP infusion. Rich-ardson, Kontos, Raper, and Patterson (34) found thathyperpnea had no effects on cardiac output, whereas
hypocapnic hyperventilation produced a transient in-crease in cardiac output which disappeared in 4 min.These observations make it unlikely that the sustainedincrease in cardiac output observed in the present ex-
periments after DNP infusion was caused by either a
decreased arterial Pco2 or an increased respiratory move-
ment of the chest.It should be noted that, except for the increases in
respiratory movement, there was no visible muscularhyperactivity in dogs that received DNP infusions, even
when the rate of oxygen consumption increased eight tonine times above control values. Since cardiac outputincreased to the same extent in response to tissue hy-permetabolism, whether associated with muscular move-
ment or not, it appears that mechanical movement of theworking muscles and the stimulation of mechanore-ceptors did not play an important role in the regulationof cardiac output. Instead, the increased cardiac outputwas closely related to the metabolic changes common toboth exercise and DNP infusion experiments, sug-
gesting that the "work stimulus" is a metabolic one.
This is certainly true during DNP infusion, in whichmuscular movement is absent. However, it cannot betotally excluded that exercise may be associated witheffects upon either cardiac output or oxygen consump-
tion, both of which are related to mechanoreceptor stim-ulation. Furthermore, because we achieved only a
doubling of cardiac output with DNP-induced hyper-metabolism, these experiments do not exclude the pos-
sibility that the stimulation of mechanoreceptors may
Cardiac Output and Tissue Hypermetabolim 2289
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contribute to the increased cardiac output during severeexercise, which can increase cardiac output as mucl-as 500r, above its resting value.
The conclusion that the "work stimulults" is primarilymetabolic in origin is supportedl by the recently pub-lished electrophysiological studies of Perez-GonzAlezand Coote (35). They studied the activity of afferentnerve fibers originating from muiiscle spindles and tendonsduring tetanic muscular contraction, and found that af-ferent nerve activities did not correlate with either theintensity of mulscular contractioni or the aniticipated pres-sor responses. They, too, concluded that mechanorecep-tors do not play a role iIn the circulatory response toexercise.
The concept that the "work stimulus" is metabolicallylinked was first postulated by Alam anid Snmirk (36),who found that the pressor response to exercise in manwas greatly exaggerated when the circulation of theworking muscles was occluded, and that the blood pres-sure remained elevated, even after cessation of rnuscu-lar activity, as long as the circulation through the previ-ously working muscles was arrested. Similar resultswere reported more recentlv by Asmiiussen and Nielsen(37).
The exact metabolic nature of the 'work stimulus"has never been ascertained. Since the breakdown of ATPto ADP is common and fundamental to both muscularexercise and the uncoupling effect of DNP, the hy-drolysis of ATP probably is linke(d to the "workstimulus."
In intact tissues, as soon as ATP is hydrolvzed toADP, ATP is resynthesized via oxidative phosphoryla-tion, anaerobic glycolysis. or breakdown of phosphoryl-creatine (PC) to creatine and inorganic phosphate.There usually is little or no change in tissue ATP con-centrations. Fawaz, Hawa, and Tutunji (38) found thatDNP, when infused into a canine heart-lung preparation,significantly lowered the PC content with no appreciablechange in the ATP content. Similarly, Hultman, Berg-str6m, and McLennan Anderson (39) found in man thatPC in working muscles decreased markedlv. while thechange in tissue ATP concentration was relatively small.They also showed that the PC concentration in themuscles during exercise was inversely related to thework load but directly related to the pulse rate. Wemay speculate that the changes in high energy phos-phate and the intimately associated metabolic changesmay cause the increase in cardiac output that occursduring exercise and after DNP infusion. However, di-rect evidence that would either substantiate this hy-pothesis or implicate any of the substances involved inthese metabolic processes as the "work stimulus" isstill not available.
Another feature common to tissue hypermetabolisminduce(d by both DNP infusion and exercise is the re-duction in tissue oxygen tension as oxygen consumptionincreases. In our experiments, no direct measurementsof tissue oxygen tension were nmade, but the right ven-tricular blood oxygen saturation, which represents thesummation of tissue oxygen saturation in all parts ofthe bod-, wN-as determinie(l. Fig. 4 slhow s that cardiac out-put increased as right ventricular blood oxygen satu-ration fell in dogs both during exercise and after DNPinfusioni. Althouglh there is no direct evidence thatlow-ering of tissue oxygen tension itself can increasecardliac output, such a possibility is not excluded andwarrants further studies.
The reduction in total peripheral vascular resistanceprobably was caused in part by the reduced tissue oxygentensioin or the metabolic changes produced by DNP.This change, however, did not (lecrease systemic arterialbloo1( pressure or increase right ventricular end-diastolicpressure (Tables I and II ). Vasodilation in exercisingmuscles prob.bly is not responsible for the increasedcardiac output, because cardiac output remainis elevatedev en wheni the circulation through exercising limbs isseparated from the general circulation by occlusion ofblood flow to the limiibs (37). Furthermore. Banlet andGuvton (29) showed that when DNPwas infused intodogs in which the central nervous system had beendestroyed, total periplleral vascular resistance declinedas imiuchl as it (lid in intact dogs, but cardiac outputincreased only slightly.
Relative tissue hypoxia mav occur during tissue hy-permetabolism. Tissue lhypoxia impairs cellular oxida-tion and phosphorylation, resulting in an increase inmitoclhonidrial NADH/NADratio. Based on the follow-ing relationship:
Lactic dehydrogenase[Pyruvate] + [.NADH]
[Lactate] + [NAD],
[Lactate] . .X
[NADH][Pyruvate] Equilbrum constant X D]
Huckabee (2, 40) suggested the use of L/P ratio anldXL as measures of tissue anaerobic metabolism. BothL/P ratio and XL increased during exercise (Table II).However, when tissue hypermetabolism was induced byDNP, the increase in cardiac output was accompaniedinitially by a decreased L/P ratio and a negative XL.The L/P ratio and XL did not increase until oxygenconsumption had exceeded four to five times its controlvalue (Tables I and II). Therefore, anaerobic metabo-lism probably was not linked to the "work stimulus"that caused the increased cardiac output during the mild
2290 C. Liang and W. B. Hood, Jr.
to moderate tissue hypermetabolism, but the presentstudy does not preclude the possibility that anaerobicmetabolism might contribute to increased cardiac outputwhen it occurs during severe tissue liypermetabolism.In fact, since the circulatory occlusion of exercisingmuscles exaggerated the hemodynamic responses duringsteady-state exercise (36, 37), it appears likely thatanoxia could potentiate either the production of the"work stimulus" or the responsiveness of receptor organsto the "work stimulus."
Presumably glycolysis is accelerated after DNP in-fusion because phosphofructokinase activity is enhancedby the decrease in ATP and the increases in ANIP,ADP, and inorganic phosphate (41). However, de-spite this increased rate of glycolysis, blood lactate andpyruvate concentrations and the L/P ratio actually fellafter the two lowest doses of DNP in these experiments.This may have resulted from concurrent facilitation ofelectron transport by DNP (19) of a magnitude thatexceeded its effects on glycolysis. This facilitation ofelectron transport is, however, counteracted by the fall-ing tissue oxygen tension. A critical oxygen tensionmay exist below which the NADH/NADor L/P ratiosrise, in spite of maximum facilitation of electron trans-port. Oxygen consumption probably increases to roughlythe same extent in all parts of the body after DNP in-fusion, although it would be expected to increase chieflyin the working muscles during exercise. In the lattersituation, muscle oxygen tension might decrease to thecritical level, causing anaerobic metabolism with a rela-tively small increase in total body oxygen consumption.A comparable DNP-induced increase in oxygen con-sumption might result in only a slight reduction in tissueoxygen tension in all parts of the body. The L/P ratiodid not increase until oxygen consumption had in-creased more than fivefold.
It should be noted that cardiac output increased inproportion to the increase in oxygen consumption nomatter where oxygen was consumed. It appears that the"'work stimulus" for the increase in cardiac output couldhave been produced not only in working muscles but alsoin other parts of the body.
ACKNOWLEDGMENTSThe authors thank Miss Adele Rymut and Mrs. BarbaraKozol for expert technical assistance and Miss Betty Gott-fried for secretarial help.
The work was supported by grants from the USPHS(HE 07299 and HL 14646) and from the American HeartAssociation (71 1016).
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