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Cardiovascular, metabolic and neurologic effects of carbon monoxide and cyanide in the rat
Toxicology 0
Letters,
TOXLET
243
6 1 ( 1992) 243-254
1992 Elsevier Science Publishers
B.V. All rights reserved
0378-4274/92/$5.00
02738
Cardiovascular, metabolic and neurologic effects of carbon monoxide and cyanide in the rat
Robert G. Dodds, David G. Penney and Bharat B. Sutariya Department
of Phvsiology,
(Received (Accepted
17 December 19 February
Key words; Blood
Wayne State University
ofMedicine,Detroit,
School
MI (USA)
1991) 1992)
pressure:
Carbon
monoxide;
Cyanide;
Glucose;
Hypothermia;
Lactate;
Neurologic
index
SUMMARY Levine-prepared, (CO) and cyanide and neurologic
female Sprague-Dawley
rats were used to investigate
(CN) on heart rate, blood pressure,
function.
hematocrit,
the effects of carbon
monoxide
blood glucose,
lactate,
Rats were exposed to either 2400 ppm CO, 1500 ppm CO, 4 mg/kg NaCN.
or both
1500 ppm CO and 4 mg/kg NaCN for 90 min. followed to 2400 ppm CO, rats exhibited a significant bradycardia
body temperature,
by 4 h of room air recovery. Following exposure which normalized by 2 h of recovery. All groups
exhibited an initial hypotension which was either maintained rats exposed to CN, and which returned to pre-exposure
or exaggerated during exposure in all but the values by 90 min. All groups experienced a
significant hypothermia during the exposure period. with those in the 1500 ppm CO or the CN returning to initial values over the recovery period. The only significant change in hematocrit was due to 2400 ppm CO (4.
I% increase).
which
was maintained
resulted produced
During
exposure,
from the combination no significant
was significantly concentrations
all groups
in all but rats exposed
of CO and CN, whereas
rise in lactate
elevated during and neurologic
experienced
concentration.
the exposure
an initial
surge in glucose
to 2400 ppm CO. The greatest
period,
deficit in rats exposed
2400 ppm CO produced
However, returning
lactate
concentration
hyperglycemic
response
the smallest.
CN alone
concentration
in all other groups
to initial values by 4 h of recovery.
to 1500 ppm CO. when added
Lactate
to those rats treated
with CN. closely approximated the lactate and neurologic deficit of the combination treatment. Neurologic deficit was greatest in rats exposed to 2400 ppm CO. While in most cases the responses of the rats to CO and CN differed whether lationship is not suggested.
C~~rrr.vpon&~e Wayne
the substances were administered An additive or less than additive
lo: D.G. Penney. Department
State University
School of Medicine,
of Physiology 540 E. Canfield,
alone or in combination, relationship is more likely.
and Occupational Detroit.
a synergistic
and Environmental
MI 48201. USA.
re-
Health.
244
INTRODUCTION
Cyanide (CN) and carbon monoxide (CO) are two of the major combustion products of plastics and other synthetic materials [I] and are frequently found to be elevated in the blood of fire victims [2,3]. CO produces its morbid and lethal effects primarily as the result of binding to hemoglobin; thus, lowering blood oxygen carrying capacity and increasing the oxygen affinity of the remaining functional hemoglobin. In contrast, CN’s effects are primarily a result of binding to the mitochondrial cytochromes, preventing oxidative phosphorylation and ATP generation 141. The binding of CN to hemoglobin is of lesser significance. Thus, the action of CN is primarily histo- or cytotoxic. As such, CN and CO are responsible for the majority of firerelated deaths [3,5,6]. This is a particular problem in air crashes and fires [7], since so many synthetic materials are used in airplane construction. While a number of studies have examined the individual effects of CO and of CN [X- IO], information about their combined effects has been studied in only a few cases [I 11. Some studies suggest that they act synergistically [lO~~l2] in producing morbidity and mortality; however, the mechanism by which this occurs remains speculative. Hypotension, hypothermia, altered blood glucose, and increased neurologic deficit are the usual responses to acute severe CO intoxication [8,13]. At 2400 ppm CO and higher, glucose has been observed to increase during early CO exposure and then to fall back to or below normal values late during CO exposure. Higher CO concentrations exaggerate the late glucose decline. Penney and his collaborators [9,13,14] found that mortality and post-CO neurologic deficit are increased in rats incurring more extreme cases of hyper- or hypoglycemia during CO exposure. In addition, post-CO hyperglycemia and mortality are increased with increasing CO concentration. Thus. glucose level changes in a complex manner during and after CO poisoning, and appears to be an important determinant of cerebra1 viability and overall survival. Whether this is related to lactate production and the degree of acidosis achieved, remains controversial. Like CO, CN would be expected to produce hypothermia, hypotension, coma, neurologic deficit, and death at a sufficiently high dose level by interfering with oxidative metabolism. Few studies have examined the effect of CN on blood glucose, lactate, or on neurologic outcome in a well characterized and controlled animal model. Although a recent study examined several blood metabolites in mice poisoned with CO and CN for up to 12 min [I I], no one has examined the possible relationships between glucose, lactate, blood pressure. body temperature. and neurologic deficit resulting from CN intoxication. The intent of this study was to explore the cardiovascular, metabolic. and neurologic responses to CO and CN poisoning, separately and in combination. We hypothcsized that CN and CO exposure would act in an additive or synergistic manner in producing hypotension. hypothermia. in exaggerating the changes in blood glucose and lactate. and in increasing neurologic deficit. In this study, levels of CN and CO were chosen well below each substance’s individual LD,,, (CN = 7 8 mg/kg: CO =
245
3500 ppm) [12], such that most of the animals survived the combined treatment. addition, we examined the responses to two different concentrations of CO.
In
MATERIALS AND METHODS
Female Sprague-Dawley rats, 90.-140 days of age, obtained from Charles River Breeding Labs (Wilmington, MA) were used. Two to three days prior to exposure, a modified Levine ]I 5] preparation was produced as previously described under ketamine (IO7 mg/kg)/Rompun (6.5 mgikg) anesthesia 181. In it, catheters made of PIE-50 polyethylene tubing were inserted into both the jugular vein and the common carotid artery toward the heart. The catheters were threaded under the skin to the nape of the neck and tied in place. The external lengths of the catheters were plugged with Amphenol pins (#220-PO2-100) and coiled up with masking tape when not in use. This procedure effectively occludes the major blood vessels to one side of the brain, placing it at increased hypoxic/ischemic risk, while providing ports for blood withdrawal, injection, and the monitoring of vital functions. The rats were allowed to recover under close observation; they were not behaviorally different from unoperated controls. The rats were confined in plastic restrainers and exposed to 2400 ppm CO (High CO) and 1500 ppm CO (Low CO) for 90 min inside a large transparent plastic bag. Two other groups were infused with NaCN solution (4 mgikg, approximately 0.5 ml total volume) through the jugular catheter using a Harvard Apparatus Co. infusion/ withdrawal pump (model 940). Two-thirds was given over the first 20 min. and onethird over the next 70 min. One group received CN only (CN), while the other group received CN and 1500 ppm CO (CO+CN). In most cases two rats from different groups were treated at the same time. The CO-exposed rats were then transferred to room air for recovery. Meuswements
Rectal temperature was monitored with a Yellow Springs Instrument Co. telethermometer (model 43TD), using YSI 400 series probes. Blood pressure was monitored with a Statham P23id pressure transducer and recorded on a Gould model 2400 chart recorder. Heart rate was derived from the blood pressure record. Blood drawn from the carotid catheter was assayed for glucose and lactate using a Yellow Springs Instrument Co. 2300 STAT. glucose/i_-lactate analyzer. Hematocrit was determined in duplicate by the mi~rohematocrit method. Measurements were made at 0, 45, 90, 2 10 and 330 min after the start of CO and/or CN treatment. Measurements were also made at 24 and 48 h after treatment in all but the high CO group. Neurologic index was assessed before treatment, and at 330 min on a scale from 6 to 30 [ 161. It was also assessed at 24 and 48 h in all but the High CO group. A normal rat scores 6. whereas a rat with severe abnormalities in terms of appearance, posture, and motor ability scores up to 30.
246
Data analysis and graphic display were carried out on a Macintosh microcomputer. Most values are means k SEM. Student’s t-test was used for statistical analysis. Differences that resulted in P-values of 0.05 or smaller were considered significant. RESULTS
Acute CO exposure at both CO concentrations and in the presence of CN resulted in hypotension after 90 min as compared to initial values. The CN rats did not show significant hypotension after 90 min, however mean blood pressure was significantly depressed at 45 min (Fig. 1). The greatest decrease in blood pressure occurred in the High CO rats (119 to 75 mmHg), with blood pressure decreasing nearly as much in the Low CO rats (119 to 83 mmHg). Hypotension was less severe in the CO+CN rats (123 to 9 1 mmHg). Blood pressure increased after 4 h of recovery (330 min), returning to initial values in all but the High CO group. Blood pressure in that group was significantly lower than all other groups after 2 and 4 h of recovery. No significant changes in heart rate were seen in the Low CO, CN and CO+CN rats as compared to initial values (data not shown). In contrast. the High CO rats showed a significant bradycardia at 90 min (478-395 bpm); heart rate had returned to normal after 2 h of recovery. All groups displayed a significant hypothermia after 90 min as compared to initial values (Fig. 2). The High CO rats had the greatest body temperature drop of 4.4”C. with the Low CO rats decreasing half as much, 2.2”C. The CN rats dropped I .2” C. while the CO+CN rats fell 2.8”C. The High CO group was significantly more hypothermic than all others at both 45 and 90 min, whereas the CN group was significantly less hypothermic than the High CO. Low CO and CO+CN rats after 90 min. Only the Low CO and CN rats regained their initial body temperatures after 4 h of recovery.
Fig.
I A comparison
of lhe blood
pressure in rata exposed to either 2400 ppm CO. I500 ppm CO. 3 mgkg
NaCN. or a combination of 1500 ppm and 4 mgikg CN for 90 min with subsequent 4 h recovery period. Values are means and the vertical bars equal 2 SEM. Symbols: +P>O.O5. ++P~O.Ol. +++P~O.OOOl as compared to initial values, (+) P~0.05. (++) P~0.01. and (+++) P~O.0001 as compared to 1500 ppm CO. The numbers in parentheses represent the quantity of animals in each group.
247
32 L--
Fig. 2. A comparison of the body temperature in rats exposed to either 2400 ppm CO, 1500 ppm CO, 4 mg/kg NaCN, or a combination of 1500 ppm CO and 4 mg/kg CN for 90 min with subsequent 4 h recovery period.
Values are means and the vertical
bars equal 2 SEM. Symbols
are the same as in Figure
I.
The High CO group was significantly different from the CN and CO+CN groups after 2 h of recovery and the CO+CN group after 4 h of recovery. The High CO rats showed a significant increase in hematocrit after 45 and 90 min (4.1%) as compared to the initial values, while the other groups showed no significant changes (Fig. 3). The hematocrit of the High CO rats returned to the initial value after 4 h of recovery. The Low CO rats had a slightly decreased hematocrit following 2 h of recovery, which became significant after 4 h of recovery. Inexplicably the initial values of the High CO group were significantly different from the Low CO and CO+CN groups. For this reason, comparisons between groups were based upon differences from the initial, not the absolute values. All groups exhibited a significant hyperglycemia after 45 min as compared to the initial values (Fig. 4). The Low CO, CN and CO+CN rats maintained a significant
364
,
,
o~co,c.’--ATime
Fig. 3. A comparison
of the change
in hematocrit
,
,
I
ir 17ecover145 (Hours)
in rats exposed
to either 2400 ppm CO, 1500 ppm CO, 4
mg/kg NaCN, or a combination of 1500 ppm CO and 4 mg/kg CN for 90 min with subsequent 4 h recovery period. Values are means and the vertical bars equal 2 SEM. Symbols are the same as in Figure I.
24X
hyperglycemia after 90 min, hut glucose level in the High CO rats had decreased to the initial value by that time. Blood glucose concentration increased the most in the CO+CN rats (A = 13I mg/dl and 113 mgidl at 45 and 90 min. respectively), and the CN rats slightly less (A = 106 mg/dl and 71 mg/dl at 45 and 90 min, respectively). The Low CO group was significantly less hyperglycemic than the CN and CO+CN groups after 45 min (A = 52 mgidl), but its glucose level continued to rise at 90 min (A = 83 mgidl). The High CO rats displayed the smallest hyperglycemi~~ at 4.5 min (A = 36 mg/dl). The blood glucose concentration of the High CO group was slightly but significantly elevated after 2 h of recovery. but had returned to initial values by 4 h. The other three groups exhibited somewhat decreased blood glucose after 4 h of recovery. The High CO, Low CO and CO+CN rats displayed signi~cant increases in lactate after 90 min as compared to the initial values (Fig. 5). Lactate did not change significantly in the CN group. The lactate concentration in the High CO rats increased 135 mgidl, which was significantly greater than all other groups. Lactate in the CO+CN group increased 105 mg/dl after 45 min, then declined somewhat at 90 min. Lactate in the Low CO group increased by 67 mg/dl after 90 min. The CO+CN rats had a sign~~cantly higher lactate than the Low CO and CN groups after 45 min. but they were significantly higher than only the CN group after 90 min. Lactate in the Low CO and CO+CN rats returned to initial levels after 2 h of recovery. while in the High CO rats this occurred after 4 h of recovery. All experimental groups displayed a significant increase in neurologic index after 4 h of recovery, indicating the development of neurologic deficit (Fig. 6). The neurologic index in the High CO rats increased the most (A = 7.6). with the COKN rats increasing somewhat less (d = 4.6). The increases in the Low CO and the CN groups were similar (A = 1.8 and 2.2, respectively), and were significantly less than in the High CO group. The neurologic index of the CO-tCN group was significantly higher
249
200 -
‘I?+
,:::,
B
3 e E -
&T.;,,::;, ;r’
loo-
B z
/ _! /
8 -I i
*,
p” (:I:,
++
(22)
j .: . . . . . +......&.. 0
Fig. 5. A comparison period.
(ii
;,,
of the lactate concentration
ppm CO. 4 mg/kg NaCN, 4 h recovery
“’
. . . . . . . . . . . . . . . . . . . . . . . . . . ‘+:!;eu;uiiiii~K~
or a combination
Values are means
in the blood of rats exposed to either 2400 ppm CO, 1500
of 1500 ppm CO and 4 mg/kg CN for 90 min with subsequent
and the vertical Figure
bars equal 2 SEM. Symbols
are the same as in
I.
than the Low CO group following 4 h of recovery. The neurologic index of the Low CO group, when added to that of the CN group, closely approximated the neurologic index of the CO+CN group. There were no significant differences from initial values in blood pressure, heart rate, hematocrit, body temperature, or glucose at 24 and 48 h. Measurements were not made at these time points in the High CO group. DISCUSSION
The hypotension, hypothermia, and neurologic deficit observed in the present study are the usual responses to acute severe CO intoxication in animals [8,13], as is
(24EO"pppm)
co
CN
(1500 wml
14mgW
CO + CN (1500 ppm. 4 mgikg,
Fig. 6. A comparison of the neurologic index in rats previous to exposure in tither 2400 ppm CO. IS00 ppm CO. 4 m&/kg NaCN, or a combination of 1500 ppm CO and 4 m&/kg CN for 90 min and following the subsequent 4 h recovery period. A healthy rat will score a 6 while a severely impaired rat scores a 30. Values are means and the vertical
bars equal I SEM. Symbols
arc the same as in Figure
I.
2.50
the rise in blood glucose we report [8,14,17]. The increase in blood lactate we observed presumably results from the CO-stimulation of glycolysis. Similar increases in lactate have been noted by others in the CO-exposed rat (Sutariya et al. and Jalukar et al., unpublished data) [ 171. Increasing levels of CO produce increasing morbidity and lethality [lo]. In the present study, the higher CO concentration resulted in greater hypothermia and lactate concentration, The initial increase in glucose at the higher CO concentration was attenuated, whereas the glucose surge was maintained at the lower CO level during CO exposure. This is consistent with our earlier studies in which even higher CO levels (e.g. 2700 and 3000 ppm) were found to profoundly depress glucose after 90 min of CO exposure [14]. With regard to heart rate, bradycardia was seen only at the higher CO concentration, suggesting that significant hypoxic depression of pacemaker activity and/or the conduction system was present only in this case. CO exposure at 2400 ppm produces greater morbidity during recovery than 1500 ppm CO with or without CN, or CN alone. based upon the recovery of normal blood pressure and lactate concentration. In terms of the neurologic index, the morbidity produced by 2400 ppm CO was greater than that at 1500 ppm CO or CN alone. but not significantly greater than with 1500 ppm CO and CN in combination. Body temperature recovery of the 2400 ppm CO-exposed rats was also impaired and may have contributed to elevation of the neurologic index. This indicates that within the CO concentration range used. with less than a doubling, there is a striking increase in morbidity; however, it is no greater than that resulting from the co-administration of 4 mg/kg CN. The response to CN differed in several important respects from that of CO. i.e.. lack of hypotension and elevation of blood lactate concentration, less severe hypothermia, and an exaggerated hyperglycemic response. Nevertheless, the neurologic deficit was similar to that of 1500 ppm CO, following 4 h of recovery. The hypotension induced by carboxyhemoglobinemia is thought to involve peripheral vasodilation mediated by local peripheral control mechanisms. The less severe hypotension observed with CN suggests that this mechanism ~ontroIling vascular resistance may not be operating with CN. Although CN is known to have major effects on the heart [Is]. the level of CN used ~lpp~ir~ntly did not markedly compromise cardiac pump function since the blood pressure was more or less maintained. This contrasts with tht‘ report that heart cytochrome oxidase is 42% inhibited after 2 h in mice treated with 4 mgikg CN [19]. The smaller hypothermia with CN may also be reflective of less peripheral vasodilation. and thus heat loss; and/or the lesser depression of metabolic heat production by CN relative to CO. Others. however, have observed profound hypotension in rats given sodium CN i.v. 1201. It is unclear whether increased glucose production or decreased glucose utilization was responsible for the rapid increase in blood glucose we noted with CN alone. Based on the lack ofchangc in blood lactate in these rats, it is clear that large amounts of glucose were not being metabolized to lactate. This suggests that glucose consumption was decreased. it is reported from in vitro ~~pcriments that the catalytic cl’-
251
ficiency of rat liver phosphorylase u is elevated 2-fold in the presence of CN [21]. The increased release of glucose from tissue glycogen could be another mechanism underlying the large increase in blood glucose in the CN-treated rats. Although CN continued to be infused into our animals over 90 min, the maintenance of adequate blood CN concentration was a concern. In the dog, the half-life of CN was found to be approximately 23 min, however elimination of CN was biphasic with an initial rapid loss followed by loss at a slower rate [22]. Because CN was not measured, we cannot state the actual blood CN concentrations achieved. Based upon the present data, the effects of simultaneous exposure to CO and CN appear to be largely additive or less than additive, rather than synergistic. In terms of body temperature and blood pressure, the responses were identical to that of CO alone during both treatment and recovery. The early glucose surge was the same as that with CN alone, while the lactate response was inexplicably greater than with CO alone. The latter suggests a synergistic response for lactate. In contrast, the neurologic index for the combined treatment equalled the sum of the indices for CO and CN alone, strongly suggesting additive effects of CO and CN. In the dog, both CO and CN are reported to increase cerebral blood flow, in at least an additive fashion, although the effects of these two agents on metabolism may be synergistic [23]. In mice the presence of CO does not affect the CN blood levels achieved, but prior CN treatment decreases the uptake of CO [l 11. The latter would slow the onset of CO’s effects and may be a contributing factor to our observations of an additive, or less than additive, relationship between CO and CN. In this regard, the depression of blood pH by CO and CN was found to be additive when the two substances were co-administered [ 1 I]. Recently, Moore et al. [l l] treated mice with 5 mg/kg CN, and/or 3500 ppm CO, over 11.5 min. Unlike the present study, they found an elevation of blood lactate in all three cases, with the greatest lactate increase with the combined COiCN treatment. The difference in response with CN alone, could have been due to differences in the species, study duration. or route of administration. In a clinical report, lactate increased only 4-fold in a man who ingested 600 mg of potassium CN [24]. a dose (i.e. mg/kg body wt.) well above that given to our rats. The fact that CN was administered all at once in the Moore et al. study [l 11, rather than gradually in the way that fire victims usually take up gaseous CN, limits the relevance of their study. Their protocol more closely simulates the effects of CN poisoning resulting from ingestion. With combined CO/CN treatment, the lactate level they observed was greater than 150 mg/dl [I 11, similar to values obtained in the present study. They stated that such a lactate level is prognostic of decreased survival, but did not examine survival; the virtually complete survival of the rats in the present study does not support this notion. Moore et al. [l l] found that blood pH fell to below 6.8 after 1 1.5 min in mice given the combined CO/CN treatment. This presumably hinged on the greatly elevated blood lactate concentration in those rats. Although we did not measure blood pH, similar levels of acidosis may have been achieved in the present study. In mice given
CO and CN alone. pH values of 7.1 and 7.3, respectively, were observed. Sokal [17] reported a blood pH value of 6.85 in rats exposed to 4000 ppm CO for 40 min. Moreover, increases in brain lactate of over 12-fold have been observed in anesthetized rats inhaling 20 000 ppm CO for 30 min [25]. It is believed by many investigators that hypoxiciischemic brain damage is the direct result of excessive lactate production and the attendant acidosis [26], and that brain lactate can be used to calculate brain pH. Recent magnetic resonance studies of brain lactate and pH, however. suggest that the two become dissociated during hypoxia!ischemia [27.28]. Considerable hemoconcentration occurred during CO exposure at 2400 ppm CO. but not at 1500 ppm CO exposure, alone or with CN, or with CN alone. This has been observed before [9], and is likely due to a redistribution of body fluid. In one animal study [29]. blood volume fell 20% after 30 min at 60% COHb, resulting mainly from a sharp reduction in plasma volume caused by increased vascular permeability. The substantial post-CO decline in hematocrit WC observed at 2400 ppm CO may have come about through fluid conservation and/or a reversal of the fluid redistribution process that occurred during CO exposure. Increased hematocrit and lowered body temperature act to increase blood viscosity. This. along with depressed cardiac function, compromise cerebral perfusion. In the Levine-prepared rat, the oligemia is more extreme during CO exposure on the operated side of the brain, leading to edema and behavioral evidence of unilateral brain dysfunction [8]. In summary, the present study demonstrated the following characteristics of acute CO poisoning. alone and in combination with CN, and of CN poisoning alone. (1) The increasingly severe hypotension observed with CO at increasing concentrations. does not occur with CN. The addition of CN does not modify this response to CO. Restitution of normal blood pressure during recovery is impeded by increased CO concentration. (2) 2400 ppm CO results in approximately twice as much hypothermia as 1500 ppm CO, whereas CN produces minimal hypothermia. The addition of CN does not modify this hypothermic response to CO. Restitution of normal body tcmperature during recovery is impeded by an elevated CO concentration or by the addition of CN. (3) The initial surge in blood glucose observed with CO exposure is maintained at 1500 ppm CO, but declines at 2400 ppm CO. CN alone, or in combination with CO at the lower concentration exaggerates and maintains the initial glucose surge. (4) 2400 ppm CO produces approximately twice as large an increase in blood lactate as 1500 ppm CO. The lactate elevation is increased by the addition of CN. while CN alone produces no increase in lactate. (5) 2400 ppm CO produces a much greater neurologic deficit than does 1500 ppm CO. The neurologic deficit produced by 1500 ppm CO when added to that produced by CN. closely approximates the deficit produced by the combined treatment. suggesting an additive effect on brain dysfunction. ACKNOWLEDGEMENTS
This work was supported
by an American
Heart Association
of Michigan,
Summer
253
Research Fellowship to R.G.D. We wish to thank Vishram Jalukar for technical assistance. REFERENCES I Loke, J., Matthay,
R.A. and Walker
Smith, G.J. (1988) The toxic environment
tions with special emphasis
on smoke inhalation.
inhalation
Dekker,
2 Clark,
Injurtes.
Marcel
C.J.. Campbell,
survivors.
and its medical implica-
In: J. Loke (Eds.). Pathophysiology
and Treatment
of
Inc., New York, pp. 453~-504.
D. and Reid. W.H. (1981) Blood carboxyhemoglobin
and cyanide
levels in ftre
Lance1 ii. 1333 1335.
3 Anderson,
R.A. and Harland.
cyanide.
W.A. (1982) Fire deaths
in the Glasgow
area: II. The role of hydrogen
Med. Sci. Law 72. 3540.
4 Piantadosi,
C.A. and Sylvia, A.L. (1984) Cerebral
rats. Toxicology 5 Kindwall.
cytochrome
a,a3 inhibition
by cyanide
in bloodless
33, 67-79.
E.P. (1980) Carbon
monoxide
poisoning
and cyanide
poisoning.
Hyperbar.
Oxygen
1.
Rev.
115-122. 6 Jones, G.R.N. (198X) Cyanide and tire victims. Lancet ii, 457. 7 Mohler,
S.R. (lY75) Air crash survival:
Med. 46, X6 X8. 8 Penney. D.G., Verma. acute carbon
ln,juries and evacuation
toxic hazards.
K. and Hull, J.A. (1989) Cardiovascular,
monoxide
poisoning
in the rat. Toxicol.
metabolic
Aviat. Space Environ.
and neurologtc
effects of
Lett. 45, ?O7- 213.
9 Penney, D.G.. Helfman. C.C.. Hull. J.A.. Dunbar. J.C. and Verma. K. (1990) Elevated blood glucose is associated with poor outcome in the carbon monoxide-p~~isone~~ rat. Toxicol. Lett. 54. 2X7-298. 10 Maujak-Schaper, oxygen.
M. and Alarie.
J. Cornbust.
Toxicol.
Y.
( 1982)Toxicity of carbon
monoxide.
hydrogen
cyanide
and low
9. 21 61.
1 I Moore. S.J., Ho. I.K. and Hume. .A.S (1991) Severe hypoxia produced by concomitant intoxication with sublethal doses of carbon monoxide and cyanide. Tox~ol. Appt. Pharmacol. 109.412 420. 12 Norris, carbon
J.C., Moore, monoxide
13 Penney,
D.G.,
poisoning
S.J. and Hume.
and cyanide.
Helfman.
A.S.
Toxicology
C.C., Dunbar,
t lY86) Synergistic 40, 13I l2Y.
J.C. and McCoy,
in the rat: effects of hyperglycemia
lethality
L.E.
and hypoglycemia
induced
by the combination
f 1991)Acute severe carbon on mortality.
recovery
deficit. Can. J. Physiol. Pharmacol. 69. I168 1177. I4 Penney. D.G., Sharma, P., Sutariya, B.B. and Nallamothu, B.G. (1990) Development is associated with death during carbon monoxide poisoning, J. Crit. Care 5, I -7. 15 Lcvinc. S. (1960) Anoxic I6 Penney.
D.G.
ischemic
f 1991) Modifyillg
encephalopathy role of plasma
in rats. Am. J. Pathol. glucose
in acute carbon
of
monoxide
and neurologic of hypoglycemia
?6, I-17. monoxide
pojsoning.
Arch.
Toxicol. (Recent Dev. Toxicoi.: Trends Methods Prom.) Suppl. 14, 23Y 244. 17 Sokal. J.A. ( lY75) Lack of the correlation between biochemical effects on rats and blood carboxyhemoglobin concentrations
in various
conditions
of single acute exposure
to carbon
monoxide.
Arch. Toxi-
col. 34. 331-336.
of
I8 Baskin. S.1. (19Yl) The cardiac effects cyanide.In: S.1. Baskin (Ed.), Principles ogy. CRC Press, Boca Raton. FL, pp. 41’3-430. I9 Buzateh.
A.M.,
Vazques,
ES. and Bathe.
A.M.D.C.
(1990) An experinlental
ofCardiacToxicol-
assessment
of cyanide
challenge dose-response, time-course and recovery of indicative toxicity parameters, Gen. Pharmacol. 21. x39 x43. 20 Brierley. J.B.. Brown. A.W. and Calverly, J. (1976) Cyanide intoxication in the rat: physiological and neuropathological aspects. J. Neurol. Neurosurg. Psychiatr. 39, 1299140. 21 Vandebroeck, A.. Uyttenhove, K.. Bollen. M. and Stalmans, W. (1988) The hepatic glycogenolysis induced by reversible ischaemia or KCN is exclusively catalyscd by phosphorylase a. Biochem. J. 256. 685 -68X.
254
22 Bright, J.E. and Marrs,
T.C.
(1988)Pharmacokinetics
of intravenous
potassium
cyanide.
Hum. Toxicol.
7. 183-186. 23 Pitt. B.R.. Radford, E.P., Gurtner. and cyanide on cerebral circulation
G.H. and Traystman, R.J. (1979) Interaction of carbon and metabolism. Arch. Environ. Hlth 34, 354-359.
24 Graham, D.L., Laman, D., Theodore. J. and Robin, E.D. (1977) Acute cyanide by lactic acidosis and pulmonary edema. Arch. Intern. Med. 137, 1051 1055. 25 MacMillan, V. (1977) Cerebral Brain Res. 121. 271--286. 26 Rehncrona.
S., Rosen.
of neuronal 27 Chopp.
damage
M.. Welch,
intracellular 28 Lotito. Benabid,
metabolism
K.M.A..
Acta Physiol.
Tidwell,
hyperglycemia
P., Francois.
A.L. (1989) Correlation
Stand.
M.. Shimazaki.
experimental
carbon
acute carbon
monoxide
acidosis:
An important
C.D. and Helpern.
A., von Kienlin. between
S.. Tanaka.
monoxide
J.A. (1988) Global in cats. Stroke
M.. Remy.
intracellular
Indust.
intoxication. mechanism
a combined
“P-‘H
cerebral
ischemia
J.P.,
Decorps.
M. and
levels in the rat brain
in vivo nuclear
magnetic
T. (1974) Blood volume
Hlth 12. 12I- 125.
and
19, 1383 1387.
C.. Albrand.
pH and lactate
N., Ukai. T. and Sugimoto,
poisoning.
complicated
110, 435- 437.
and hypoglycemia
potassium cyanide induced metabolism blockade: copy study. Neurosci. Lett. 97. 91- 96. 29 Ogawa.
during
I. and Siesjo. B.K. (1980) Excessive cellular
in the brain.
pH during
S., Blondet,
carbohydrate
poisoning
monoxide
during
spectroschanges
in
Letters,
TOXLET
243
6 1 ( 1992) 243-254
1992 Elsevier Science Publishers
B.V. All rights reserved
0378-4274/92/$5.00
02738
Cardiovascular, metabolic and neurologic effects of carbon monoxide and cyanide in the rat
Robert G. Dodds, David G. Penney and Bharat B. Sutariya Department
of Phvsiology,
(Received (Accepted
17 December 19 February
Key words; Blood
Wayne State University
ofMedicine,Detroit,
School
MI (USA)
1991) 1992)
pressure:
Carbon
monoxide;
Cyanide;
Glucose;
Hypothermia;
Lactate;
Neurologic
index
SUMMARY Levine-prepared, (CO) and cyanide and neurologic
female Sprague-Dawley
rats were used to investigate
(CN) on heart rate, blood pressure,
function.
hematocrit,
the effects of carbon
monoxide
blood glucose,
lactate,
Rats were exposed to either 2400 ppm CO, 1500 ppm CO, 4 mg/kg NaCN.
or both
1500 ppm CO and 4 mg/kg NaCN for 90 min. followed to 2400 ppm CO, rats exhibited a significant bradycardia
body temperature,
by 4 h of room air recovery. Following exposure which normalized by 2 h of recovery. All groups
exhibited an initial hypotension which was either maintained rats exposed to CN, and which returned to pre-exposure
or exaggerated during exposure in all but the values by 90 min. All groups experienced a
significant hypothermia during the exposure period. with those in the 1500 ppm CO or the CN returning to initial values over the recovery period. The only significant change in hematocrit was due to 2400 ppm CO (4.
I% increase).
which
was maintained
resulted produced
During
exposure,
from the combination no significant
was significantly concentrations
all groups
in all but rats exposed
of CO and CN, whereas
rise in lactate
elevated during and neurologic
experienced
concentration.
the exposure
an initial
surge in glucose
to 2400 ppm CO. The greatest
period,
deficit in rats exposed
2400 ppm CO produced
However, returning
lactate
concentration
hyperglycemic
response
the smallest.
CN alone
concentration
in all other groups
to initial values by 4 h of recovery.
to 1500 ppm CO. when added
Lactate
to those rats treated
with CN. closely approximated the lactate and neurologic deficit of the combination treatment. Neurologic deficit was greatest in rats exposed to 2400 ppm CO. While in most cases the responses of the rats to CO and CN differed whether lationship is not suggested.
C~~rrr.vpon&~e Wayne
the substances were administered An additive or less than additive
lo: D.G. Penney. Department
State University
School of Medicine,
of Physiology 540 E. Canfield,
alone or in combination, relationship is more likely.
and Occupational Detroit.
a synergistic
and Environmental
MI 48201. USA.
re-
Health.
244
INTRODUCTION
Cyanide (CN) and carbon monoxide (CO) are two of the major combustion products of plastics and other synthetic materials [I] and are frequently found to be elevated in the blood of fire victims [2,3]. CO produces its morbid and lethal effects primarily as the result of binding to hemoglobin; thus, lowering blood oxygen carrying capacity and increasing the oxygen affinity of the remaining functional hemoglobin. In contrast, CN’s effects are primarily a result of binding to the mitochondrial cytochromes, preventing oxidative phosphorylation and ATP generation 141. The binding of CN to hemoglobin is of lesser significance. Thus, the action of CN is primarily histo- or cytotoxic. As such, CN and CO are responsible for the majority of firerelated deaths [3,5,6]. This is a particular problem in air crashes and fires [7], since so many synthetic materials are used in airplane construction. While a number of studies have examined the individual effects of CO and of CN [X- IO], information about their combined effects has been studied in only a few cases [I 11. Some studies suggest that they act synergistically [lO~~l2] in producing morbidity and mortality; however, the mechanism by which this occurs remains speculative. Hypotension, hypothermia, altered blood glucose, and increased neurologic deficit are the usual responses to acute severe CO intoxication [8,13]. At 2400 ppm CO and higher, glucose has been observed to increase during early CO exposure and then to fall back to or below normal values late during CO exposure. Higher CO concentrations exaggerate the late glucose decline. Penney and his collaborators [9,13,14] found that mortality and post-CO neurologic deficit are increased in rats incurring more extreme cases of hyper- or hypoglycemia during CO exposure. In addition, post-CO hyperglycemia and mortality are increased with increasing CO concentration. Thus. glucose level changes in a complex manner during and after CO poisoning, and appears to be an important determinant of cerebra1 viability and overall survival. Whether this is related to lactate production and the degree of acidosis achieved, remains controversial. Like CO, CN would be expected to produce hypothermia, hypotension, coma, neurologic deficit, and death at a sufficiently high dose level by interfering with oxidative metabolism. Few studies have examined the effect of CN on blood glucose, lactate, or on neurologic outcome in a well characterized and controlled animal model. Although a recent study examined several blood metabolites in mice poisoned with CO and CN for up to 12 min [I I], no one has examined the possible relationships between glucose, lactate, blood pressure. body temperature. and neurologic deficit resulting from CN intoxication. The intent of this study was to explore the cardiovascular, metabolic. and neurologic responses to CO and CN poisoning, separately and in combination. We hypothcsized that CN and CO exposure would act in an additive or synergistic manner in producing hypotension. hypothermia. in exaggerating the changes in blood glucose and lactate. and in increasing neurologic deficit. In this study, levels of CN and CO were chosen well below each substance’s individual LD,,, (CN = 7 8 mg/kg: CO =
245
3500 ppm) [12], such that most of the animals survived the combined treatment. addition, we examined the responses to two different concentrations of CO.
In
MATERIALS AND METHODS
Female Sprague-Dawley rats, 90.-140 days of age, obtained from Charles River Breeding Labs (Wilmington, MA) were used. Two to three days prior to exposure, a modified Levine ]I 5] preparation was produced as previously described under ketamine (IO7 mg/kg)/Rompun (6.5 mgikg) anesthesia 181. In it, catheters made of PIE-50 polyethylene tubing were inserted into both the jugular vein and the common carotid artery toward the heart. The catheters were threaded under the skin to the nape of the neck and tied in place. The external lengths of the catheters were plugged with Amphenol pins (#220-PO2-100) and coiled up with masking tape when not in use. This procedure effectively occludes the major blood vessels to one side of the brain, placing it at increased hypoxic/ischemic risk, while providing ports for blood withdrawal, injection, and the monitoring of vital functions. The rats were allowed to recover under close observation; they were not behaviorally different from unoperated controls. The rats were confined in plastic restrainers and exposed to 2400 ppm CO (High CO) and 1500 ppm CO (Low CO) for 90 min inside a large transparent plastic bag. Two other groups were infused with NaCN solution (4 mgikg, approximately 0.5 ml total volume) through the jugular catheter using a Harvard Apparatus Co. infusion/ withdrawal pump (model 940). Two-thirds was given over the first 20 min. and onethird over the next 70 min. One group received CN only (CN), while the other group received CN and 1500 ppm CO (CO+CN). In most cases two rats from different groups were treated at the same time. The CO-exposed rats were then transferred to room air for recovery. Meuswements
Rectal temperature was monitored with a Yellow Springs Instrument Co. telethermometer (model 43TD), using YSI 400 series probes. Blood pressure was monitored with a Statham P23id pressure transducer and recorded on a Gould model 2400 chart recorder. Heart rate was derived from the blood pressure record. Blood drawn from the carotid catheter was assayed for glucose and lactate using a Yellow Springs Instrument Co. 2300 STAT. glucose/i_-lactate analyzer. Hematocrit was determined in duplicate by the mi~rohematocrit method. Measurements were made at 0, 45, 90, 2 10 and 330 min after the start of CO and/or CN treatment. Measurements were also made at 24 and 48 h after treatment in all but the high CO group. Neurologic index was assessed before treatment, and at 330 min on a scale from 6 to 30 [ 161. It was also assessed at 24 and 48 h in all but the High CO group. A normal rat scores 6. whereas a rat with severe abnormalities in terms of appearance, posture, and motor ability scores up to 30.
246
Data analysis and graphic display were carried out on a Macintosh microcomputer. Most values are means k SEM. Student’s t-test was used for statistical analysis. Differences that resulted in P-values of 0.05 or smaller were considered significant. RESULTS
Acute CO exposure at both CO concentrations and in the presence of CN resulted in hypotension after 90 min as compared to initial values. The CN rats did not show significant hypotension after 90 min, however mean blood pressure was significantly depressed at 45 min (Fig. 1). The greatest decrease in blood pressure occurred in the High CO rats (119 to 75 mmHg), with blood pressure decreasing nearly as much in the Low CO rats (119 to 83 mmHg). Hypotension was less severe in the CO+CN rats (123 to 9 1 mmHg). Blood pressure increased after 4 h of recovery (330 min), returning to initial values in all but the High CO group. Blood pressure in that group was significantly lower than all other groups after 2 and 4 h of recovery. No significant changes in heart rate were seen in the Low CO, CN and CO+CN rats as compared to initial values (data not shown). In contrast. the High CO rats showed a significant bradycardia at 90 min (478-395 bpm); heart rate had returned to normal after 2 h of recovery. All groups displayed a significant hypothermia after 90 min as compared to initial values (Fig. 2). The High CO rats had the greatest body temperature drop of 4.4”C. with the Low CO rats decreasing half as much, 2.2”C. The CN rats dropped I .2” C. while the CO+CN rats fell 2.8”C. The High CO group was significantly more hypothermic than all others at both 45 and 90 min, whereas the CN group was significantly less hypothermic than the High CO. Low CO and CO+CN rats after 90 min. Only the Low CO and CN rats regained their initial body temperatures after 4 h of recovery.
Fig.
I A comparison
of lhe blood
pressure in rata exposed to either 2400 ppm CO. I500 ppm CO. 3 mgkg
NaCN. or a combination of 1500 ppm and 4 mgikg CN for 90 min with subsequent 4 h recovery period. Values are means and the vertical bars equal 2 SEM. Symbols: +P>O.O5. ++P~O.Ol. +++P~O.OOOl as compared to initial values, (+) P~0.05. (++) P~0.01. and (+++) P~O.0001 as compared to 1500 ppm CO. The numbers in parentheses represent the quantity of animals in each group.
247
32 L--
Fig. 2. A comparison of the body temperature in rats exposed to either 2400 ppm CO, 1500 ppm CO, 4 mg/kg NaCN, or a combination of 1500 ppm CO and 4 mg/kg CN for 90 min with subsequent 4 h recovery period.
Values are means and the vertical
bars equal 2 SEM. Symbols
are the same as in Figure
I.
The High CO group was significantly different from the CN and CO+CN groups after 2 h of recovery and the CO+CN group after 4 h of recovery. The High CO rats showed a significant increase in hematocrit after 45 and 90 min (4.1%) as compared to the initial values, while the other groups showed no significant changes (Fig. 3). The hematocrit of the High CO rats returned to the initial value after 4 h of recovery. The Low CO rats had a slightly decreased hematocrit following 2 h of recovery, which became significant after 4 h of recovery. Inexplicably the initial values of the High CO group were significantly different from the Low CO and CO+CN groups. For this reason, comparisons between groups were based upon differences from the initial, not the absolute values. All groups exhibited a significant hyperglycemia after 45 min as compared to the initial values (Fig. 4). The Low CO, CN and CO+CN rats maintained a significant
364
,
,
o~co,c.’--ATime
Fig. 3. A comparison
of the change
in hematocrit
,
,
I
ir 17ecover145 (Hours)
in rats exposed
to either 2400 ppm CO, 1500 ppm CO, 4
mg/kg NaCN, or a combination of 1500 ppm CO and 4 mg/kg CN for 90 min with subsequent 4 h recovery period. Values are means and the vertical bars equal 2 SEM. Symbols are the same as in Figure I.
24X
hyperglycemia after 90 min, hut glucose level in the High CO rats had decreased to the initial value by that time. Blood glucose concentration increased the most in the CO+CN rats (A = 13I mg/dl and 113 mgidl at 45 and 90 min. respectively), and the CN rats slightly less (A = 106 mg/dl and 71 mg/dl at 45 and 90 min, respectively). The Low CO group was significantly less hyperglycemic than the CN and CO+CN groups after 45 min (A = 52 mgidl), but its glucose level continued to rise at 90 min (A = 83 mgidl). The High CO rats displayed the smallest hyperglycemi~~ at 4.5 min (A = 36 mg/dl). The blood glucose concentration of the High CO group was slightly but significantly elevated after 2 h of recovery. but had returned to initial values by 4 h. The other three groups exhibited somewhat decreased blood glucose after 4 h of recovery. The High CO, Low CO and CO+CN rats displayed signi~cant increases in lactate after 90 min as compared to the initial values (Fig. 5). Lactate did not change significantly in the CN group. The lactate concentration in the High CO rats increased 135 mgidl, which was significantly greater than all other groups. Lactate in the CO+CN group increased 105 mg/dl after 45 min, then declined somewhat at 90 min. Lactate in the Low CO group increased by 67 mg/dl after 90 min. The CO+CN rats had a sign~~cantly higher lactate than the Low CO and CN groups after 45 min. but they were significantly higher than only the CN group after 90 min. Lactate in the Low CO and CO+CN rats returned to initial levels after 2 h of recovery. while in the High CO rats this occurred after 4 h of recovery. All experimental groups displayed a significant increase in neurologic index after 4 h of recovery, indicating the development of neurologic deficit (Fig. 6). The neurologic index in the High CO rats increased the most (A = 7.6). with the COKN rats increasing somewhat less (d = 4.6). The increases in the Low CO and the CN groups were similar (A = 1.8 and 2.2, respectively), and were significantly less than in the High CO group. The neurologic index of the CO-tCN group was significantly higher
249
200 -
‘I?+
,:::,
B
3 e E -
&T.;,,::;, ;r’
loo-
B z
/ _! /
8 -I i
*,
p” (:I:,
++
(22)
j .: . . . . . +......&.. 0
Fig. 5. A comparison period.
(ii
;,,
of the lactate concentration
ppm CO. 4 mg/kg NaCN, 4 h recovery
“’
. . . . . . . . . . . . . . . . . . . . . . . . . . ‘+:!;eu;uiiiii~K~
or a combination
Values are means
in the blood of rats exposed to either 2400 ppm CO, 1500
of 1500 ppm CO and 4 mg/kg CN for 90 min with subsequent
and the vertical Figure
bars equal 2 SEM. Symbols
are the same as in
I.
than the Low CO group following 4 h of recovery. The neurologic index of the Low CO group, when added to that of the CN group, closely approximated the neurologic index of the CO+CN group. There were no significant differences from initial values in blood pressure, heart rate, hematocrit, body temperature, or glucose at 24 and 48 h. Measurements were not made at these time points in the High CO group. DISCUSSION
The hypotension, hypothermia, and neurologic deficit observed in the present study are the usual responses to acute severe CO intoxication in animals [8,13], as is
(24EO"pppm)
co
CN
(1500 wml
14mgW
CO + CN (1500 ppm. 4 mgikg,
Fig. 6. A comparison of the neurologic index in rats previous to exposure in tither 2400 ppm CO. IS00 ppm CO. 4 m&/kg NaCN, or a combination of 1500 ppm CO and 4 m&/kg CN for 90 min and following the subsequent 4 h recovery period. A healthy rat will score a 6 while a severely impaired rat scores a 30. Values are means and the vertical
bars equal I SEM. Symbols
arc the same as in Figure
I.
2.50
the rise in blood glucose we report [8,14,17]. The increase in blood lactate we observed presumably results from the CO-stimulation of glycolysis. Similar increases in lactate have been noted by others in the CO-exposed rat (Sutariya et al. and Jalukar et al., unpublished data) [ 171. Increasing levels of CO produce increasing morbidity and lethality [lo]. In the present study, the higher CO concentration resulted in greater hypothermia and lactate concentration, The initial increase in glucose at the higher CO concentration was attenuated, whereas the glucose surge was maintained at the lower CO level during CO exposure. This is consistent with our earlier studies in which even higher CO levels (e.g. 2700 and 3000 ppm) were found to profoundly depress glucose after 90 min of CO exposure [14]. With regard to heart rate, bradycardia was seen only at the higher CO concentration, suggesting that significant hypoxic depression of pacemaker activity and/or the conduction system was present only in this case. CO exposure at 2400 ppm produces greater morbidity during recovery than 1500 ppm CO with or without CN, or CN alone. based upon the recovery of normal blood pressure and lactate concentration. In terms of the neurologic index, the morbidity produced by 2400 ppm CO was greater than that at 1500 ppm CO or CN alone. but not significantly greater than with 1500 ppm CO and CN in combination. Body temperature recovery of the 2400 ppm CO-exposed rats was also impaired and may have contributed to elevation of the neurologic index. This indicates that within the CO concentration range used. with less than a doubling, there is a striking increase in morbidity; however, it is no greater than that resulting from the co-administration of 4 mg/kg CN. The response to CN differed in several important respects from that of CO. i.e.. lack of hypotension and elevation of blood lactate concentration, less severe hypothermia, and an exaggerated hyperglycemic response. Nevertheless, the neurologic deficit was similar to that of 1500 ppm CO, following 4 h of recovery. The hypotension induced by carboxyhemoglobinemia is thought to involve peripheral vasodilation mediated by local peripheral control mechanisms. The less severe hypotension observed with CN suggests that this mechanism ~ontroIling vascular resistance may not be operating with CN. Although CN is known to have major effects on the heart [Is]. the level of CN used ~lpp~ir~ntly did not markedly compromise cardiac pump function since the blood pressure was more or less maintained. This contrasts with tht‘ report that heart cytochrome oxidase is 42% inhibited after 2 h in mice treated with 4 mgikg CN [19]. The smaller hypothermia with CN may also be reflective of less peripheral vasodilation. and thus heat loss; and/or the lesser depression of metabolic heat production by CN relative to CO. Others. however, have observed profound hypotension in rats given sodium CN i.v. 1201. It is unclear whether increased glucose production or decreased glucose utilization was responsible for the rapid increase in blood glucose we noted with CN alone. Based on the lack ofchangc in blood lactate in these rats, it is clear that large amounts of glucose were not being metabolized to lactate. This suggests that glucose consumption was decreased. it is reported from in vitro ~~pcriments that the catalytic cl’-
251
ficiency of rat liver phosphorylase u is elevated 2-fold in the presence of CN [21]. The increased release of glucose from tissue glycogen could be another mechanism underlying the large increase in blood glucose in the CN-treated rats. Although CN continued to be infused into our animals over 90 min, the maintenance of adequate blood CN concentration was a concern. In the dog, the half-life of CN was found to be approximately 23 min, however elimination of CN was biphasic with an initial rapid loss followed by loss at a slower rate [22]. Because CN was not measured, we cannot state the actual blood CN concentrations achieved. Based upon the present data, the effects of simultaneous exposure to CO and CN appear to be largely additive or less than additive, rather than synergistic. In terms of body temperature and blood pressure, the responses were identical to that of CO alone during both treatment and recovery. The early glucose surge was the same as that with CN alone, while the lactate response was inexplicably greater than with CO alone. The latter suggests a synergistic response for lactate. In contrast, the neurologic index for the combined treatment equalled the sum of the indices for CO and CN alone, strongly suggesting additive effects of CO and CN. In the dog, both CO and CN are reported to increase cerebral blood flow, in at least an additive fashion, although the effects of these two agents on metabolism may be synergistic [23]. In mice the presence of CO does not affect the CN blood levels achieved, but prior CN treatment decreases the uptake of CO [l 11. The latter would slow the onset of CO’s effects and may be a contributing factor to our observations of an additive, or less than additive, relationship between CO and CN. In this regard, the depression of blood pH by CO and CN was found to be additive when the two substances were co-administered [ 1 I]. Recently, Moore et al. [l l] treated mice with 5 mg/kg CN, and/or 3500 ppm CO, over 11.5 min. Unlike the present study, they found an elevation of blood lactate in all three cases, with the greatest lactate increase with the combined COiCN treatment. The difference in response with CN alone, could have been due to differences in the species, study duration. or route of administration. In a clinical report, lactate increased only 4-fold in a man who ingested 600 mg of potassium CN [24]. a dose (i.e. mg/kg body wt.) well above that given to our rats. The fact that CN was administered all at once in the Moore et al. study [l 11, rather than gradually in the way that fire victims usually take up gaseous CN, limits the relevance of their study. Their protocol more closely simulates the effects of CN poisoning resulting from ingestion. With combined CO/CN treatment, the lactate level they observed was greater than 150 mg/dl [I 11, similar to values obtained in the present study. They stated that such a lactate level is prognostic of decreased survival, but did not examine survival; the virtually complete survival of the rats in the present study does not support this notion. Moore et al. [l l] found that blood pH fell to below 6.8 after 1 1.5 min in mice given the combined CO/CN treatment. This presumably hinged on the greatly elevated blood lactate concentration in those rats. Although we did not measure blood pH, similar levels of acidosis may have been achieved in the present study. In mice given
CO and CN alone. pH values of 7.1 and 7.3, respectively, were observed. Sokal [17] reported a blood pH value of 6.85 in rats exposed to 4000 ppm CO for 40 min. Moreover, increases in brain lactate of over 12-fold have been observed in anesthetized rats inhaling 20 000 ppm CO for 30 min [25]. It is believed by many investigators that hypoxiciischemic brain damage is the direct result of excessive lactate production and the attendant acidosis [26], and that brain lactate can be used to calculate brain pH. Recent magnetic resonance studies of brain lactate and pH, however. suggest that the two become dissociated during hypoxia!ischemia [27.28]. Considerable hemoconcentration occurred during CO exposure at 2400 ppm CO. but not at 1500 ppm CO exposure, alone or with CN, or with CN alone. This has been observed before [9], and is likely due to a redistribution of body fluid. In one animal study [29]. blood volume fell 20% after 30 min at 60% COHb, resulting mainly from a sharp reduction in plasma volume caused by increased vascular permeability. The substantial post-CO decline in hematocrit WC observed at 2400 ppm CO may have come about through fluid conservation and/or a reversal of the fluid redistribution process that occurred during CO exposure. Increased hematocrit and lowered body temperature act to increase blood viscosity. This. along with depressed cardiac function, compromise cerebral perfusion. In the Levine-prepared rat, the oligemia is more extreme during CO exposure on the operated side of the brain, leading to edema and behavioral evidence of unilateral brain dysfunction [8]. In summary, the present study demonstrated the following characteristics of acute CO poisoning. alone and in combination with CN, and of CN poisoning alone. (1) The increasingly severe hypotension observed with CO at increasing concentrations. does not occur with CN. The addition of CN does not modify this response to CO. Restitution of normal blood pressure during recovery is impeded by increased CO concentration. (2) 2400 ppm CO results in approximately twice as much hypothermia as 1500 ppm CO, whereas CN produces minimal hypothermia. The addition of CN does not modify this hypothermic response to CO. Restitution of normal body tcmperature during recovery is impeded by an elevated CO concentration or by the addition of CN. (3) The initial surge in blood glucose observed with CO exposure is maintained at 1500 ppm CO, but declines at 2400 ppm CO. CN alone, or in combination with CO at the lower concentration exaggerates and maintains the initial glucose surge. (4) 2400 ppm CO produces approximately twice as large an increase in blood lactate as 1500 ppm CO. The lactate elevation is increased by the addition of CN. while CN alone produces no increase in lactate. (5) 2400 ppm CO produces a much greater neurologic deficit than does 1500 ppm CO. The neurologic deficit produced by 1500 ppm CO when added to that produced by CN. closely approximates the deficit produced by the combined treatment. suggesting an additive effect on brain dysfunction. ACKNOWLEDGEMENTS
This work was supported
by an American
Heart Association
of Michigan,
Summer
253
Research Fellowship to R.G.D. We wish to thank Vishram Jalukar for technical assistance. REFERENCES I Loke, J., Matthay,
R.A. and Walker
Smith, G.J. (1988) The toxic environment
tions with special emphasis
on smoke inhalation.
inhalation
Dekker,
2 Clark,
Injurtes.
Marcel
C.J.. Campbell,
survivors.
and its medical implica-
In: J. Loke (Eds.). Pathophysiology
and Treatment
of
Inc., New York, pp. 453~-504.
D. and Reid. W.H. (1981) Blood carboxyhemoglobin
and cyanide
levels in ftre
Lance1 ii. 1333 1335.
3 Anderson,
R.A. and Harland.
cyanide.
W.A. (1982) Fire deaths
in the Glasgow
area: II. The role of hydrogen
Med. Sci. Law 72. 3540.
4 Piantadosi,
C.A. and Sylvia, A.L. (1984) Cerebral
rats. Toxicology 5 Kindwall.
cytochrome
a,a3 inhibition
by cyanide
in bloodless
33, 67-79.
E.P. (1980) Carbon
monoxide
poisoning
and cyanide
poisoning.
Hyperbar.
Oxygen
1.
Rev.
115-122. 6 Jones, G.R.N. (198X) Cyanide and tire victims. Lancet ii, 457. 7 Mohler,
S.R. (lY75) Air crash survival:
Med. 46, X6 X8. 8 Penney. D.G., Verma. acute carbon
ln,juries and evacuation
toxic hazards.
K. and Hull, J.A. (1989) Cardiovascular,
monoxide
poisoning
in the rat. Toxicol.
metabolic
Aviat. Space Environ.
and neurologtc
effects of
Lett. 45, ?O7- 213.
9 Penney, D.G.. Helfman. C.C.. Hull. J.A.. Dunbar. J.C. and Verma. K. (1990) Elevated blood glucose is associated with poor outcome in the carbon monoxide-p~~isone~~ rat. Toxicol. Lett. 54. 2X7-298. 10 Maujak-Schaper, oxygen.
M. and Alarie.
J. Cornbust.
Toxicol.
Y.
( 1982)Toxicity of carbon
monoxide.
hydrogen
cyanide
and low
9. 21 61.
1 I Moore. S.J., Ho. I.K. and Hume. .A.S (1991) Severe hypoxia produced by concomitant intoxication with sublethal doses of carbon monoxide and cyanide. Tox~ol. Appt. Pharmacol. 109.412 420. 12 Norris, carbon
J.C., Moore, monoxide
13 Penney,
D.G.,
poisoning
S.J. and Hume.
and cyanide.
Helfman.
A.S.
Toxicology
C.C., Dunbar,
t lY86) Synergistic 40, 13I l2Y.
J.C. and McCoy,
in the rat: effects of hyperglycemia
lethality
L.E.
and hypoglycemia
induced
by the combination
f 1991)Acute severe carbon on mortality.
recovery
deficit. Can. J. Physiol. Pharmacol. 69. I168 1177. I4 Penney. D.G., Sharma, P., Sutariya, B.B. and Nallamothu, B.G. (1990) Development is associated with death during carbon monoxide poisoning, J. Crit. Care 5, I -7. 15 Lcvinc. S. (1960) Anoxic I6 Penney.
D.G.
ischemic
f 1991) Modifyillg
encephalopathy role of plasma
in rats. Am. J. Pathol. glucose
in acute carbon
of
monoxide
and neurologic of hypoglycemia
?6, I-17. monoxide
pojsoning.
Arch.
Toxicol. (Recent Dev. Toxicoi.: Trends Methods Prom.) Suppl. 14, 23Y 244. 17 Sokal. J.A. ( lY75) Lack of the correlation between biochemical effects on rats and blood carboxyhemoglobin concentrations
in various
conditions
of single acute exposure
to carbon
monoxide.
Arch. Toxi-
col. 34. 331-336.
of
I8 Baskin. S.1. (19Yl) The cardiac effects cyanide.In: S.1. Baskin (Ed.), Principles ogy. CRC Press, Boca Raton. FL, pp. 41’3-430. I9 Buzateh.
A.M.,
Vazques,
ES. and Bathe.
A.M.D.C.
(1990) An experinlental
ofCardiacToxicol-
assessment
of cyanide
challenge dose-response, time-course and recovery of indicative toxicity parameters, Gen. Pharmacol. 21. x39 x43. 20 Brierley. J.B.. Brown. A.W. and Calverly, J. (1976) Cyanide intoxication in the rat: physiological and neuropathological aspects. J. Neurol. Neurosurg. Psychiatr. 39, 1299140. 21 Vandebroeck, A.. Uyttenhove, K.. Bollen. M. and Stalmans, W. (1988) The hepatic glycogenolysis induced by reversible ischaemia or KCN is exclusively catalyscd by phosphorylase a. Biochem. J. 256. 685 -68X.
254
22 Bright, J.E. and Marrs,
T.C.
(1988)Pharmacokinetics
of intravenous
potassium
cyanide.
Hum. Toxicol.
7. 183-186. 23 Pitt. B.R.. Radford, E.P., Gurtner. and cyanide on cerebral circulation
G.H. and Traystman, R.J. (1979) Interaction of carbon and metabolism. Arch. Environ. Hlth 34, 354-359.
24 Graham, D.L., Laman, D., Theodore. J. and Robin, E.D. (1977) Acute cyanide by lactic acidosis and pulmonary edema. Arch. Intern. Med. 137, 1051 1055. 25 MacMillan, V. (1977) Cerebral Brain Res. 121. 271--286. 26 Rehncrona.
S., Rosen.
of neuronal 27 Chopp.
damage
M.. Welch,
intracellular 28 Lotito. Benabid,
metabolism
K.M.A..
Acta Physiol.
Tidwell,
hyperglycemia
P., Francois.
A.L. (1989) Correlation
Stand.
M.. Shimazaki.
experimental
carbon
acute carbon
monoxide
acidosis:
An important
C.D. and Helpern.
A., von Kienlin. between
S.. Tanaka.
monoxide
J.A. (1988) Global in cats. Stroke
M.. Remy.
intracellular
Indust.
intoxication. mechanism
a combined
“P-‘H
cerebral
ischemia
J.P.,
Decorps.
M. and
levels in the rat brain
in vivo nuclear
magnetic
T. (1974) Blood volume
Hlth 12. 12I- 125.
and
19, 1383 1387.
C.. Albrand.
pH and lactate
N., Ukai. T. and Sugimoto,
poisoning.
complicated
110, 435- 437.
and hypoglycemia
potassium cyanide induced metabolism blockade: copy study. Neurosci. Lett. 97. 91- 96. 29 Ogawa.
during
I. and Siesjo. B.K. (1980) Excessive cellular
in the brain.
pH during
S., Blondet,
carbohydrate
poisoning
monoxide
during
spectroschanges
in