(Stroke. 1998;29:1229-1239.)
© 1998 American Heart Association, Inc.
A Reproducible Model of Circulatory Arrest and Remote Resuscitation in Rats for NMR Investigation
Serguei Liachenko, PhD;
Pei Tang, PhD;
Ronald L. Hamilton, MD;
Yan Xu, PhD
From the Departments of Anesthesiology and Critical Care Medicine (S.L.,
P.T., Y.X.), Pathology (R.L.H.), and Pharmacology (Y.X.), University of
Pittsburgh (Pa).
Correspondence to Yan Xu, PhD, W-1358 Biomedical Science Tower, University of Pittsburgh, Pittsburgh, PA 15261. E-mail xu{at}smtp.anes.upmc.edu
 |
Abstract
|
|---|
Background and Purpose—Because
noninvasive physiological monitoring of cerebral
blood flow, metabolic integrity, and brain ion and water
homeostasis can now be accomplished with new, state-of-the-art MR
spectroscopy and imaging techniques, it is appropriate to develop
controllable and reproducible animal models that permit prolonged
circulatory arrest and resuscitation in the magnet and also allow for
studies of long-term survival and outcome. We have developed such a
model in rats that involves minimal surgical preparations and can
achieve resuscitation remotely within precisely controlled
time.
Methods—Cardiac arrest was induced by asphyxiation, the duration
of which ranged from 8 to 24 minutes. Resuscitation was achieved
remotely by a slow, intra-aortic infusion of oxygenated
blood (withdrawn either from the same rat before asphyxia or from a
healthy donor rat) along with a resuscitation cocktail containing
heparin (50 U/100 g), sodium bicarbonate (0.1 mEq/100 g), and
epinephrine (4 µg/100 g). The body temperature was measured
by a tympanic thermocouple probe and was controlled either by a heating
pad (constant tympanic temperature=37°C) or by warm ambient air
(constant air temperature=37°C). Interleaved
31P/1H nuclear magnetic resonance (NMR)
spectroscopy was used in a selected group of rats to measure the
cerebral metabolism before and during approximately 20
minutes of circulatory arrest and after resuscitation.
Results—The overall success rate of resuscitation, irrespective
of the duration of cardiac arrest, was 82% (51 of 62). With a
programmed infusion pump, the success rate was even higher (95%). The
survival time for rats subjected to 15 and 19 minutes of asphyxia with
core temperature tightly controlled was significantly lower than that
with ambient temperature control (P<0.001 and
P<0.04, respectively). High-quality NMR spectra can be
obtained continuously without interference from the resuscitation
effort. Final histological examinations taken 5 days
after resuscitation showed typical neuronal damages, similar to those
found in other global ischemia models.
Conclusions—Because the no-flow time and resuscitation time can
be precisely controlled, this outcome model is ideally suited for
studies of ischemic and reperfusion injuries in the brain and
possibly in other critical organs, permitting continuous assessment of
long-term recovery and follow-up in the same animals.
Key Words: heart arrest • ischemia • nuclear magnetic resonance • reperfusion • resuscitation • rats
|
Introduction
|
|---|
Heart disease, the
primary cause of death in the United States,1
leads to cardiac arrest in many patients. The high mortality rate of
cardiac arrest is due in part to our current inability to prevent and
reverse tissue damage in several vital organs, particularly the brain
and the heart, after a short period of ischemia. Of the 70 000
patients per year in the United States who are successfully rescued by
CPR to ROSC, 60% subsequently die in the hospital; only 3% to 10%
have a chance to resume their former life
activities.2
To date, investigations of mechanisms of tissue damage after global
ischemia have largely focused on models with reduced
complexity, such as cell cultures,3 perfused
organs,4 5 and occlusion of major
vessels.6 7 8 9 Although such models can provide
valuable tools to define the cascade of cellular events and to identify
the important components of injury process after ischemia, they
often do not permit assessment of postresuscitation
syndrome10 11 12 13 and long-term outcome.
Be- cause it has become clear that in the brain, and possibly in other
organs, the progression and maturation of injuries after circulatory
arrest occur well after the point of ROSC and are often reperfusion
dependent,14 development of intact animal models
that precisely mimic the clinical state of cardiac arrest and
resuscitation is of crucial importance for evaluation of effective
postresuscitation therapy and outcome.
We report here a new cardiac arrest and resuscitation model in rats.
The model involves minimal surgical preparation. Cardiac arrest and
resuscitation can be controlled remotely and with high efficacy.
Although the model can potentially be used in various experimental
settings, it is particularly useful for state-of-the-art NMR
measurements because once the animals are positioned in the magnet,
there is no interference to the magnetic field homogeneity from the
resuscitation efforts. High-quality imaging and spectroscopic
measurements are thus possible throughout the crucial periods of
cardiac arrest, resuscitation, and subsequent recovery.
|
Materials and Methods
|
|---|
The experimental protocol was approved by the Institutional
Animal Care and Use Committee at the University of Pittsburgh. Male
Sprague-Dawley rats (Harlan Sprague-Dawley, Indianapolis, Ind),
weighing 164±33 g, were used. Younger rats were chosen for their
relevance to asphyxial cardiac arrest in infants and children.
Sixty-six rats were randomized into 10 groups (Table 1
). In group 1 (sham operation), rats
were surgically prepared, as described below, but were not subjected to
asphyxia and cardiac arrest. In groups 2 through 7, rats were
respectively subjected to 8, 8, 10, 12, 15, and 19 minutes of asphyxia.
In these groups, the rat body temperature, measured by a tympanic
temperature probe, was precisely controlled to 37.0±0.5°C throughout
the experiment, with the use of a feedback-controlled heating pad
(temperature control method I). Groups 8, 9, and 10 underwent 15, 19,
and 24 minutes of asphyxia, respectively, with body temperature
controlled by a flow of 37.0°C ambient air (temperature control
method II). This second method of temperature control was tested
because of its clinical relevance, despite the fact that it was
insufficient to maintain a constant tympanic temperature during
prolonged circulatory arrest.
Animal Preparation
In a typical experiment, rats were anesthetized with 4%
isoflurane in an O2 (30%) and
N2O (70%) mixture and intubated orotracheally.
After intubation, anesthesia was reduced to 1.25%
isoflurane in an O2 (30%) and air (70%)
mixture, and the rats were paralyzed with pancuronium bromide (0.02
mg/100 g) and mechanically ventilated with a tidal volume of 1 mL/100
g, 40 breaths per minute, and a positive end-expiratory pressure of 5.5
cm H2O. Baseline arterial pH was
maintained at 
7.4, PaCO2 at
30 mm Hg, and PaO2
>170 mm Hg. Both femoral arteries and the left femoral vein were
cannulated. One of the arterial catheters was attached to a
pressure transducer (Baxter Edwards, model 53-DTS-260), which was
interfaced through a Grass 7D Polygraph recorder (Grass Instrument
Co) to a PowerMac 8100 computer (Apple Computer, Inc) running the
LabVIEW program (National Instruments) for continuous display and
recording of arterial blood pressure,
arterial pulse pressure, and HR. The other
arterial catheter was slowly advanced through the abdominal
aorta to the thoracic aorta for later resuscitation use (see below) and
for arterial blood sampling. The arterial blood
gases and pH were determined every 60 minutes with a Corning 178
pH/blood gas analyzer (Corning Medical and Scientific). A
conventional six-lead ECG was recorded with the Grass 7D Polygraph
recorder. The tympanic membrane temperature was measured by a
flexible thin thermocouple inserted deeply into the left auditory
canal, which was then closed with cotton.
Cardiac Arrest and Resuscitation
Five minutes before induction of asphyxial cardiac arrest, rats
received a dose of the short-acting muscle relaxant vecuronium bromide
(0.01 mg/100 g IV, plus 50 U/100 g heparin) to prevent spontaneous
breathing during asphyxia. A minute before asphyxia, a small amount of
oxygenated arterial blood (1 mL/100 g body wt)
was collected into a syringe (groups 1 and 3 to 10). Group 2 differed
from others in that no blood was withdrawn from the rat under study but
rather from a healthy donor, of the same litter, 1 minute before the
planned resuscitation. The syringe collecting the blood contained a
mixture of heparin (50 U/100 g), sodium bicarbonate (0.1 mEq/100 g),
and epinephrine (4 µg/100 g) in a total volume of 0.25 mL/100
g body wt. Asphyxiation was caused by disconnecting the ventilation
tubes from the ventilator and clamping them. This led to
electromechanical dissociation, as indicated by a decrease in MABP to
<10 mm Hg and an arterial pulse pressure to <5
mm Hg, and subsequent pulseless electric activity. Resuscitation began
with 100% O2 ventilation and a slow infusion of
the oxygenated arterial blood into the thoracic
aorta. In earlier experiments the infusion was done manually, but for
the 19 rats in the most recent experiments it was accomplished
automatically at an infusion rate of 0.45 mL/100 g per minute with the
use of a programmable syringe pump (KDS-210, KD Scientific Inc). For
group 2, the same volume of venous blood was collected during infusion
to maintain the hemodynamic balance.
With this intra-aortic infusion method, resuscitation to ROSC could be
achieved without electric defibrillation or chest compression. ROSC was
determined as the point at which MABP reached a value >60 mm Hg
with a supraventricular cardiac rhythm. In group 2, donor
blood infusion was discontinued at the point of ROSC. In all groups, if
ROSC was impossible after all withdrawn blood had been returned, no
further resuscitation was attempted. The resuscitated rats were
continually ventilated for 
2 hours with a mixture of
O2 (30%) and air (70%). Anesthesia
was reinstated with 1% isoflurane if signs of awakening were observed
or if HR increased to 300 beats per minute.
At least 2 hours after resuscitation and under isoflurane general
anesthesia, the arterial and venous catheters
were surgically removed and the wound was closed. Rats were then
mechanically ventilated with room air and allowed to extubate
themselves. After extubation, they were returned to their cages and
monitored closely for long-term outcome evaluation. Whenever needed,
rats were given fluids subcutaneously or fed with ground chew with
water. Rats that did not extubate themselves, or later became morbid or
could not attend to their physiological needs, were
killed and prepared for histological examination.
Outcome Evaluation
Animals were closely observed for 3 to 5 days after
resuscitation. In animals that survived for >3 days, the neurological
deficit scores (NDS) were evaluated by the same investigator using the
criteria proposed by Neumar and coworkers.15
After final NDS evaluation, animals were reanesthetized (1.5%
isoflurane through a nose cone mask). A thoracotomy was performed, and
a 14-gauge catheter was advanced into the aortic arch through the apex
of the left ventricle. The descending aorta was then clamped. Neutrally
buffered 3% paraformaldehyde was infused through the
catheter under a pressure of 100 cm H2O. The
right atrium was incised, and perfusion was continued until the fluid
draining from the right atrium was clear (approximately 60 mL). The rat
was then decapitated, and the head was stored in 3%
paraformaldehyde for 24 hours. The brain was then
removed from the skull and stored in the fixative. Paraffin-embedded
coronal sections, 6 µm thick, were made through the level of
hippocampus and stained with hematoxylin-eosin.
Interleaved 31P/1H NMR
Spectroscopy
To demonstrate the compatibility of the model with NMR
investigation, 6 rats from group 9 were subjected to continuous
interleaved 31P/1H NMR
spectroscopic measurements before, during, and after circulatory arrest
and resuscitation. Experiments were performed with an Otsuka
CMXW-400SLI spectrometer, equipped with a 9.4-T, 111-mm bore magnet.
Because of the vertical orientation of the magnet, rats were snugly
positioned, head up, in a specially designed cradle. A double-resonance
surface-coil probe was used, consisting of an 11x13-mm elliptical
surface coil tuned to 162.367 and 401.102 MHz for
31P and 1H resonance
frequencies, respectively. The pulse sequences for interleaved
acquisition consisted of a conventional one-pulse sequence for
31P and a spin-echo sequence with gaussian water
suppression for 1H. The pulse widths for
31P and 1H were 32 and 40
microseconds, respectively, which produced nominal 90° flip at a
position 7 mm perpendicular from the center of the surface coil.
Spin-echo time for 1H was set to 136 milliseconds
to maximize water suppression and lactate
detection.16 Data were acquired in 4096 complex
points with a spectral width of 10 kHz for a pair of interleaved
31P and 1H spectra.
Interleaving was achieved by acquiring data from one nucleus during the
mandatory relaxation period of the other. Additional relaxation time
was added to ensure a repetition delay of 1.8 seconds for
31P and 1.6 seconds for 1H.
A total of 152 scans were summed and zero-filled once before Fourier
transform. The temporal resolution for a pair of interleaved spectra
was 5 minutes.
Data Analysis
Statistical analysis was performed with the use of the
SPSS program (SPSS Inc). Simple factorial two-way ANOVA was used to
determine the effects of asphyxia time, temperature control method, and
rat orientation on long-term survival, NDS, and HR. Repeated-measures
ANOVA was used to compare the physiological
variables among groups at three time points: before cardiac arrest
and 1 and 2 hours after resuscitation. If a significant change was
indicated for a given variable, then Student-Newman-Keuls multiple
comparisons were made to determine the differences between time points
within groups or between groups at a given time point. Differences in
NDS among groups were compared with the nonparametric
Kruskal-Wallis test. Survival of rats was analyzed with
Kaplan-Meier Survival Analysis with the log-rank test for
between-group comparisons. The mean survival times were also compared
among all groups with the Duncan multiple comparisons test. All data
are expressed as mean±SD.
|
Results
|
|---|
Figure 1 shows the
representative traces of arterial blood
pressure and HR before, during, and after cardiac arrest and
resuscitation from a rat in group 9 (19 minutes of asphyxia). The
arterial blood pressures in the shaded time interval in
Figure 1 are expanded in Figure 2 to
indicate several key events, labeled I through V. The withdrawal of
arterial blood (interval I-II) led to a slight (<4%)
decrease in MABP. The time from the onset of asphyxia to
electromechanical dissociation (interval II-III) was 2.88±0.95
minutes, averaged among 57 rats in groups 3 to 10, and was 1.33±0.31
minutes for group 2 (no preasphyxia blood withdrawal). Event III marks
the beginning of circulatory arrest by definition. The infusion of
oxygenated blood and reventilation (event IV) resulted in
an MABP increase, the initial rate of which was controlled mainly by
the rate of infusion. A rapid elevation in MABP, leading to ROSC (event
V by definition), occurred either during or immediately after
completion of the infusion in 51 of 62 rats (a success rate of 82%).
The success rate with pump resuscitation was even higher and seemed
independent of the no-flow time. Of the 19 rats resuscitated by the
pump, only one did not show ROSC after all shed blood was returned.
Table 1
summarizes the number of resuscitated rats of the total rats in
each group, the intervals between the major events, the mean survival
time, and the NDS measured 3 days after resuscitation. Note that the
duration of asphyxia and that of circulatory arrest (no-flow time) are
indicated in Table 1 by II-IV and III-V, respectively.
View larger version (26K):
[in this window]
[in a new window]
|
Figure 1. Representative
cardiovascular response to 19 minutes of asphyxiation
(from group 9). Traces of systolic, mean, and
diastolic arterial blood pressure (A) and HR
(B) before, during, and after cardiac arrest and resuscitation are
displayed as a function of time. Time 0 is arbitrarily assigned to the
moment immediately before the arterial blood
withdrawal.
|
|
View larger version (22K):
[in this window]
[in a new window]
|
Figure 2. Detailed changes in arterial blood
pressure during the shaded interval in Figure 1A. Important events are
as follows: I, beginning of blood withdrawal; II, onset of asphyxia;
III, beginning of no-flow by definition; IV, beginning of
resuscitation; and V, ROSC by definition. The duration of asphyxia and
that of circulatory arrest are defined by the intervals II-IV and
III-V, respectively.
|
|
The MABP and HR immediately before arterial blood
withdrawal or 1 minute before asphyxia for group 2 (arbitrarily
assigned as time 0) and 1, 2, and 3 hours after ROSC are listed in
Table 2. Corresponding data from group 1,
with time 0 assigned to the point half an hour after surgical
preparation and before blood withdrawal, are also included. MABP showed
a similar decrease after cardiac arrest in all animals. The HR was
significantly lower in groups 9 and 10 at 2 hours after ROSC compared
with the sham-operated rats (group 1).
Other important physiological variables are
listed in Table 3 for all groups. After
24-minute asphyxiation (groups 7 and 10), any attempts to take an
arterial blood sample after ROSC caused a significant
decrease in MABP. Therefore, no blood sample was taken for these two
groups after ROSC. In other groups, arterial blood pH
showed a tendency to recover soon after ROSC. Only group 9 showed a
significantly lower value at 1 hour after ROSC compared with the sham
operation. No significant changes in
PaCO2 and
PaO2 were found.
Representative ECG tracings (lead II) for groups 3
through 10 are displayed in Figure 3.
Electric silence was not observed after the mechanical asystole. In
groups with prolonged asphyxia (19 and 24 minutes), however,
ventricular complexes disappeared 3 to 7 minutes before
resuscitation. In almost all rats, the electric activity of the heart
reappeared immediately after the onset of resuscitation procedure and
intensified before ROSC. In groups 3 and 8, ECG returned to normal 2
hours after ROSC and remained normal 3 to 5 days after resuscitation.
All other groups showed different extents of ECG abnormalities,
indicating certain degree of ischemic and hypoxic damage to the
myocardium.17
View larger version (29K):
[in this window]
[in a new window]
|
Figure 3. Representative ECG tracing in rats
before, during, 2 hours, and 3 days after cardiac arrest. There was no
immediate electric silence in ECG after mechanical asystole, indicating
electric-mechanical dissociation. In most rats in the prolonged
asphyxiation groups (19 minutes of asphyxia), ventricular
complexes disappeared before the resuscitation procedure had begun.
ROSC was preceded by restoration of normal supraventricular
cardiac rhythm. ECG tend to recover after mild ischemic insult
(groups 3 and 8) but showed hypoxic/ischemic myocardial damage
in other groups.
|
|
All sham-operated rats (group 1) and all resuscitated rats in group 8
were alive 5 days after the episode. When the tympanic membrane
temperature was precisely controlled to 37°C throughout the
experiments, the percentages of 3-day survival of the successfully
resuscitated rats were 60% for group 2 (3 of 5), 50% for group 3 (3
of 6) and group 4 (2 of 4), and 0% for groups 5, 6, and 7 (0 of 4, 6,
and 2, respectively). In contrast, when the body temperature was
controlled with 37°C ambient air (the tympanic temperature was
gradually decreased to 34°C to 35°C during cardiac arrest), the
long-term survival was significantly better. The percentage of 3-day
survival was 100% for group 8 (5 of 5) and 67% for group 9 (8 of 12).
However, after prolonged 24-minute asphyxiation (group 10), the mean
survival time was significantly lowered.
For rats that remained alive 3 days after the surgery (group 1) or
ROSC (groups 2 to 4 and 8 to 10), there was no significant difference
in NDS (P>0.63). Rats that died <3 days after
resuscitation were excluded from the NDS evaluation. The reduction in
the NDS primarily reflected impairment to the hind limbs. It was not
determined whether this impairment was due to neurological damage or to
surgical procedures.
Because of the orientation of our magnet, rats had to be positioned
vertically for NMR measurements. To determine the effects of
orientation, we assigned 41 rats to be positioned vertically in the NMR
probe and 25 horizontally on the bench top. Two-way ANOVA with respect
to asphyxia time, temperature control method, and rat orientation
showed that the orientation had no significant effects on long-term
survival (P>0.79), NDS (P>0.82), and HR before
(P>0.17) and after (P>0.07) cardiac arrest.
Figure 4A shows
representative interleaved
31P/1H NMR spectra before,
during, and after 20 minutes of circulatory arrest. The peaks assigned
are ATP, phosphocreatine, inorganic phosphate,
phosphomonoester, and MDPA in the
31P spectra, and NAA, creatine, choline,
glutamate/glutamine, and lactate in the 1H
spectra. pHi can be calculated on the basis of
chemical shifts in the 31P
spectra.18 The MDPA peak was from a sealed
external reference, whose position relative to the NMR coil was fixed.
Detailed changes in ATP, phosphocreatine, inorganic phosphate,
pHi, NAA-choline ratio, and lactate, averaged for
six rats in group 9, are depicted in Figure 4B. As can be seen,
immediately after cardiac arrest phosphocreatine was completely
depleted; ATP decreased to <20% of control; pHi
was lowered to 6.2; the NAA-choline ratio decreased gradually during
the 20 minutes of circulatory arrest and continued to decline after
ROSC for 30 minutes to 70% of the control; inorganic phosphate
increased by >500%; and lactate increased by 340%. The residual ATP
during the cardiac arrest is believed to arise from the slightest
signal contamination from the extracranial muscle. Because surgery was
minimized in this outcome model, the extracranial muscle was not
extracted.
After resuscitation, inorganic phosphate and phosphocreatine recovered
to the control level within 30 and 80 minutes after ROSC, respectively.
ATP, however, did not fully recover until 2 hours after ROSC. The
NAA-choline ratio recovered slowly over a course of 3 hours but only to
90% of the control level, suggesting a certain degree of neuronal
damage. Recovery of pHi was biphasic in this
particular group of rats; there was a rapid return of
pHi to 7.1, which correlated in time with the
clearance of lactate, and a slow normalization over a period of 2 to 3
hours.
Figure 5A depicts a typical section of
hippocampus from a rat 5 days after an 8-minute cardiac arrest (from
group 3). The neurons of the CA1 region showed typical
hypoxic/ischemic changes, similar to those found in other
global ischemia models. These changes included vacuolization
and hypereosinophilia of the neuronal cytoplasm, nuclear pyknosis, and
karyorrhexis. Injuries were only rarely present in the cortical
neurons and were not present in other portions of the hippocampal
formation. In rats killed 3 days after cardiac arrest, only rare
neurons in this area showed eosinophilia, but there was considerable
vacuolization in some cases (Figure 5B). None of these changes were
seen in rats killed within 24 hours of cardiac arrest (Figure 5C). In
animals in which method II of temperature control was used, only
vacuolization of cytoplasm could be seen in the CA1 region 3 days after
ROSC.
View larger version (85K):
[in this window]
[in a new window]
|
Figure 5. A, Section of hippocampus from a rat 5 days after
8-minute cardiac arrest. The neurons of the CA1 region show
hypoxic/ischemic changes, including hypereosinophilia and
vacuolization of the neuronal cytoplasm, nuclear pyknosis, and
karyorrhexis (arrows). These changes are only rarely present in
cortical neurons and are not present in other portions of the
hippocampal formation. B, In rats killed 3 days after cardiac arrest,
some show vacuolization in the CA1 region but only rare eosinophilic
neurons. C, None of the changes can be found in rats killed within 24
hours of cardiac arrest. Hematoxylin-eosin–stained sections of
paraffin-embedded tissue. Bar=10 µm in A and C; bar=20 µm
in B.
|
|
|
Discussion
|
|---|
A highly reproducible cardiac arrest and resuscitation model in
rats is presented. The model is developed specifically for use
with noninvasive MR spectroscopic and imaging techniques, but clearly
it can be used for other studies in which remotely controlled
resuscitation and minimal animal movement during data acquisition are
required. In addition to the ease of control of the no-flow and
resuscitation times, a particular advantage of the present model
for MR investigation is its minimal (essentially none) perturbation to
the magnetic field homogeneity during resuscitation. Unlike
conventional CPR or electric defibrillation in which unpredictable
animal movements due to CPR or the noise and secondary field caused by
the electric current can interfere with magnetic field homogeneity, the
present resuscitation method permits uninterrupted MR measurements
during the most crucial period of circulatory arrest and reperfusion.
Moreover, because of its clinical relevance, the model is ideally
suited for investigation of the mechanisms of postresuscitation
syndrome,10 such as perfusion failure in the
brain and possibly other organs, reoxygenation injury
leading to cell necrosis, extracerebral organ derangement, and blood
derangement. The feasibility of long-term survival in this model also
allows for assessment of possible postarrest pharmacological
interventions and follow-up studies during the crucial period of
recovery after prolonged circulatory arrest.
Although asphyxia differs from ventricular fibrillation as
the primary cause for circulatory arrest, the postarrest resuscitation
and the evaluation of reperfusion injury and delayed neuronal death are
similar. Moreover, the asphyxia-induced cardiac arrest is, in its own
right, of great clinical importance: coma-induced hypoventilation is
the second most common indication for CPR and cerebral resuscitation,
and in children the predominant cause of cardiac arrest is
asphyxia.19
The resuscitation regimen used in the present model (ie, 2- to
3-minute infusion of oxygenated arterial blood
directly into the thoracic aorta) resembles closely the procedures for
cardiopulmonary bypass, implicating the potential clinical
importance of the model. Partial cardiopulmonary bypass (or
partial cardiopulmonary support20) is
routinely used in clinical practice for extracorporeal life support in
humans and sometimes also for experimental
resuscitation.21 22 23 24 25 Although
intra-arterial administration of medication (eg,
epinephrine) should be practiced with caution, the low doses
used in the model can be further reduced when donor blood is used (such
as in group 2), because the withdrawn donor blood and predetermined
dose of medication need not be infused after ROSC is observed. As an
experimental approach, our intra-aortic blood infusion proves to be a
very reliable method for remotely controlled resuscitation after
prolonged cardiac arrest.
Except for group 2, the amount of heparin administered was high.
Although we observed no evidence of hemorrhage after
resuscitation, heparin may alter hemodynamic,
metabolic, and immunologic responses after resuscitation
from cardiac arrest. In our recent modification of the method using
donor rats, we tested the use of heparin-coated extracorporeal
devices26 without administrating free heparin.
Excellent resuscitation was obtained (data not shown), and no blood
coagulation or microclots were found in the coated devices. Numerous
studies27 28 29 in which the same type of coating
in cardiopulmonary bypass was used have shown greatly improved
outcome. Release of heparin from the coating was virtually
undetectable.26
Successful resuscitation after prolonged cardiac arrest of up to 32
minutes has been demonstrated previously with the use of extracorporeal
circulation,30 two-stage
resuscitation,31 or internal cardiac
massage.32 Not all of these, however, permit
evaluation of animal survival and neurological outcome. In one study in
dogs, after 20 minutes of ventricular fibrillation followed
by resuscitation with cardiopulmonary
bypass,23 nine of 10 dogs were alive 72 hours
after arrest, but none regained normal neurological function at 96
hours after arrest.
In earlier models of asphyxial cardiac arrest in rats, resuscitation
was achieved by extrathoracic compression either
manually11 15 or with the use of a remotely
controlled pneumatic balloon.33 The present
model has an advantage over these earlier models in that the time from
the beginning of resuscitation to ROSC can be precisely controlled by
the rate at which the oxygenated arterial blood
is infused, thus making the no-flow time reproducible. This will ensure
a similar degree of insult to all animals subjected to the same period
of asphyxia, so that results from different animals can be
compared.
Intravenous injection of 0.5 mol/L KCl has been used
in a transient cardiac arrest model in rats,34 in
which 2% venous blood (milliliters per gram body weight) was
withdrawn after KCl injection, and rats were resuscitated 3.5 minutes
later by external cardiac compression and intravenous blood
infusion. Although KCl injection causes isoelectric ECG almost
instantly and thus offers a good control of the time to induce cardiac
arrest, resuscitation after KCl-induced cardiac arrest requires
CPR34 35 and is impossible without electric
defibrillation if no-flow time is prolonged (unpublished data, Y. Xu,
J. Melick, 1994). Any large variation in resuscitation time defeats the
purpose of having a tight control of the arrest time in the first
place. Moreover, KCl poisoning is a rather rare medical problem; the
clinical implication of KCl-induced cardiac arrest is thus not as
clearly defined as that of asphyxial cardiac arrest.
When our model is compared with other global ischemia models,
it should be noted that the present study uses relatively young
rats, chosen for their relevance to asphyxial cardiac arrest in infants
and children. Although not investigated here, the age of the animal may
affect the resuscitation and survival rates. However, in terms of
excitotoxicity, the rats used in the present study (164±33 g; 46
to 50 days old) are comparable to adult rats. Recent studies using
intracerebral microinjection of
neurotoxins36 37 have shown that in rats peak
vulnerabilities to N-methyl-D-aspartate,
-amino-3-hydroxy-5-methyl-4 isoxazole propionic acid (AMPA), and
kainate occur at postnatal days 6, 10, and 21, respectively, and
the sensitivity to excitotoxic damage flattens after the age of 21
days.
The body temperature during and after cardiac arrest and resuscitation
has a profound impact on the severity of injury and long-term
outcome.24 38 39 40 The two temperature control
methods used in this study (viz, precise control with a feedback
heating device wrapped around the rats to maintain the tympanic
membrane temperature at 37°C and the ambient air control to maintain
the ambient temperature at 37°C) resulted in significantly different
outcomes after prolonged cardiac arrest. For example, when group 6 is
compared with group 8 (both were subjected to 15 minutes of asphyxia,
resulting in 14 minutes of circulatory arrest) and group 7 is compared
with group 9 (19 and 20 minutes of circulatory arrest, respectively),
the mean survival time was significantly shorter (P<0.001
for 15 minutes and P<0.04 for 19 minutes) for rats in which
method I was used (Table 1). Although the ambient temperature was
controlled at 37°C, method II was insufficient to maintain a constant
rat body temperature. In the case of group 9 (19 minutes of asphyxia),
the rat body temperature was gradually decreased to 34°C toward the
end of circulatory arrest. Correspondingly, the
histological changes in the vulnerable CA1 region of
the hippocampus were less expressed in animals with the temperature
control of method II (Figure 5B).
The issue of temperature control in studies of cardiac arrest and
resuscitation should be viewed from two different perspectives. First,
precise temperature control is necessary for mechanistic studies,
especially when the effects of therapeutic agents are under
investigation. However, temperature control with ambient air, allowing
the body temperature to decrease naturally during circulatory arrest,
is more closely relevant to a clinical setting. Our two methods of
controlling temperature can be selected to ideally meet both needs.
No special treatment was administered after ROSC to correct the
arterial pH in the present model. The
31P NMR data showed that brain
pHi was lowered from the control level of 7.4 to
6.2 during cardiac arrest in group 9, recovered to 7.1 immediately
after ROSC, stayed at this level for 2 hours, and then gradually
returned to the control level. It has been
suggested41 that reduction of the extracellular
HCO3- concentration greatly
inhibits the rate of pHi recovery and that
correction of systemic metabolic acidosis after cardiac
arrest may improve neurological outcome. Because substantial
controversy exists regarding whether systemic treatment with sodium
bicarbonate is beneficial in cardiopulmonary resuscitation, the
current model, in conjunction with the 31P NMR
measurement of pHi, provides a means to study the
mechanism of postischemic normalization of intracellular
and extracellular pH and to evaluate the treatment along with the
long-term outcome.
The no-reflow phenomenon in the brain is believed to be one of the
limiting factors for postischemic
survival.14 42 Reflow-promoting therapy has been
suggested to prevent injury after prolonged circulatory
arrest.14 Compared with the earlier resuscitation
model with extrathoracic compression,33 the
relatively better outcome seen in the present model may result from
the initial arterial blood infusion into the thoracic
aorta; the high perfusion pressure helped to overcome the resistance in
the microvasculature due to increased blood viscosity and reduced
microvascular lumen. Indeed, the initial hyperperfusion during
blood-infusion resuscitation can be seen clearly in the studies of MR
cerebral perfusion mapping with this model. This aspect, and
particularly the MR quantification of the protracted cerebral
hypoperfusion at later stage of reperfusion, will be presented
elsewhere.
In conclusion, an outcome model of cardiac arrest and resuscitation in
rats is presented. The model is ideally suited for
investigations in which remote resuscitation with minimal
disturbance to animal position is required. Examination of
injury development and maturation, evaluation of treatment strategies,
and follow-up of long-term outcome are all possible with this
model.
|
Selected Abbreviations and Acronyms
|
|---|
| CPR |
= |
cardiopulmonary resuscitation |
| HR |
= |
heart rate |
| MABP |
= |
mean arterial blood pressure |
| MDPA |
= |
methylene diphosphonic acid |
| NAA |
= |
N-acetyl-aspartate |
| NDS |
= |
neurological deficit scores |
| NMR |
= |
nuclear magnetic resonance |
| pHi |
= |
intracellular pH |
| ROSC |
= |
restoration of spontaneous circulation |
|
|
Acknowledgments
|
|---|
This study was supported in part by grants from the National
Institute of Neurological Disorders and Stroke (1R01NS36124), National
Institutes of Health, and the University Anesthesiology and Critical
Care Medicine Foundation, University of Pittsburgh. We thank Dr
Nicholas G. Bircher for suggestions and comments, Dr Lisa Goetz for
editorial assistance, and Dr Leonard Firestone for encouragement
and support.
Received October 20, 1997;
revision received March 16, 1998;
accepted March 17, 1998.
|
References
|
|---|
1.
Becker LB. The
epidemiology of sudden death. In: Paradis NA,
Halperin HR, Nowak RM, eds. Cardiac Arrest: The Science and
Practice of Resuscitation Medicine. Baltimore, Md: Williams &
Wilkins; 1996:28–47.
2.
Krause GS, Kumar K, White BC, Aust SD, Wiegenstein JG.
Ischemia, resuscitation, and reperfusion: mechanisms of tissue
injury and prospects for protection. Am Heart J. 1986;111:768–780.[Medline]
[Order article via Infotrieve]
3.
Abu RS, Trembovler V, Shohami E, Lazarovici P. A
tissue culture ischemic device to study eicosanoid release by
pheochromocytoma PC12 cultures. J Neurosci Methods. 1993;50:197–203.[Medline]
[Order article via Infotrieve]
4.
Espanol MT, Xu Y, Litt L, Yang GY, Chang LH, James TL,
Weinstein P, Chan PH. Modulation of glutamate-induced intracellular
energy failure in neonatal cerebral cortical slices by kynurenic acid,
dizocilpine, and NBQX. J Cereb Blood Flow Metab. 1994;14:269–278.[Medline]
[Order article via Infotrieve]
5.
Yanagida S, Luo CS, Doyle M, Pohost GM, Pike MM.
Nuclear magnetic resonance studies of cationic and energetic
alterations with oxidant stress in the perfused heart: modulation with
pyruvate and lactate. Circ Res. 1995;77:773–783.[Abstract/Free Full Text]
6.
Kohno K, Back T, Hoehn-Berlage M, Hossmann KA. A
modified rat model of middle cerebral artery thread occlusion under
electrophysiological control for magnetic
resonance investigations. Magn Reson Imaging. 1995;13:65–71.[Medline]
[Order article via Infotrieve]
7.
Gonce ME, Brackett DJ, Squires RA, Gibson DD,
Kajdacsy-Balla A, Lerner MR, McCay PB, Pennington LR. A new model for
inducing total hepatic ischemia while preventing circulatory
collapse secondary to splanchnic vascular congestion. Shock. 1995;3:440–446.[Medline]
[Order article via Infotrieve]
8.
Forsting M, Reith W, Schabitz WR, Heiland S, von KR,
Hacke W, Sartor K. Decompressive craniectomy for cerebral infarction:
an experimental study in rats. Stroke. 1995;26:259–264.[Abstract/Free Full Text]
9.
Hughes JD, Mattar SG, Chen C, Someren A, Noe B, Suwyn
CR, Lumsden AB. Renal artery perfusion modifies
ischemia/reperfusion injury. J Surg Res. 1996;60:321–326.[Medline]
[Order article via Infotrieve]
10.
Safar P. Prevention and therapy of postresuscitation
neurologic dysfunction and injury. In: Paradis NA, Halperin HR, Nowak
RM, eds. Cardiac Arrest: The Science and Practice of
Resuscitation Medicine. Baltimore, Md: Williams & Wilkins;
1996:859–887.
11.
Katz L, Ebmeyer U, Safar P, Radovsky A, Neumar R.
Outcome model of asphyxial cardiac arrest in rats. J Cereb
Blood Flow Metab. 1995;15:1032–1039.[Medline]
[Order article via Infotrieve]
12.
Safar P. Cerebral resuscitation after cardiac arrest: a
review. Circulation. 1986;74(suppl IV):IV-138-IV-153.
13.
Safar P. Effects of the postresuscitation syndrome on
cerebral recovery from cardiac arrest. Crit Care Med. 1985;13:932–935.[Medline]
[Order article via Infotrieve]
14.
Hossmann KA. Ischemia-mediated neuronal injury.
Resuscitation. 1993;26:225–235.[Medline]
[Order article via Infotrieve]
15.
Neumar RW, Bircher NG, Sim KM, Xiao F, Zadach KS,
Radovsky A, Katz L, Ebmeyer E, Safar P. Epinephrine and sodium
bicarbonate during CPR following asphyxial cardiac arrest in rats.
Resuscitation. 1995;29:249–263.[Medline]
[Order article via Infotrieve]
16.
Xu Y, Cohen Y, Litt L, Chang LH, James TL. Tolerance of
low cerebral intracellular pH in rats during hyperbaric hypercapnia.
Stroke. 1991;22:1303–1308.[Abstract/Free Full Text]
17.
Bestetti RB, Oliveira JS. The surface
electrocardiogram: a simple and reliable method for
detecting overt and latent heart disease in rats. Braz J Med
Biol Res. 1990;23:1213–1222.[Medline]
[Order article via Infotrieve]
18.
Petroff OA, Prichard JW, Behar KL, Alger JR, den HJA,
Shulman RG. Cerebral intracellular pH by 31P
nuclear magnetic resonance spectroscopy. Neurology. 1985;35:781–788.[Abstract/Free Full Text]
19.
Safar P, Paradis NA. Asphyxial cardiac arrest. In:
Paradis NA, Halperin HR, Nowak RM, eds. Cardiac Arrest: The
Science and Practice of Resuscitation Medicine. Baltimore, Md:
Williams & Wilkins; 1996:702–726.
20.
Dluzneski J, Casthely PA, Niedzinski J. Partial
cardiopulmonary support. In: Casthely PA, Bregman D, eds.
Cardiopulmonary Bypass: Physiology, Related
Complications, and Pharmacology. Mount Kisco, NY: Futura
Publishing Co Inc; 1991:179–189.
21.
Iijima T, Back T, Hossmann KA. Effect of extracorporeal
life support on cerebral blood flow, metabolism and
electrophysiology in normothermic cats. Intensive
Care Med. 1995;21:38–44.[Medline]
[Order article via Infotrieve]
22.
Kano T, Hashiguchi A, Sadanaga M, Ashimura K, Sakamoto
M, Morioka T. Cardiopulmonary-cerebral resuscitation by using
cardiopulmonary bypass through the femoral vein and artery in
dogs. Resuscitation. 1993;25:265–281.[Medline]
[Order article via Infotrieve]
23.
Safar P, Abramson NS, Angelos M, Cantadore R, Leonov Y,
Levine R, Pretto E, Reich H, Sterz F, Stezoski SW, Tisherman S.
Emergency cardiopulmonary bypass for resuscitation from
prolonged cardiac arrest. Am J Emerg Med. 1990;8:55–67.[Medline]
[Order article via Infotrieve]
24.
Safar P, Xiao F, Radovsky A, Tanigawa K, Ebmeyer U,
Bircher N, Alexander H, Stezoski SW. Improved cerebral resuscitation
from cardiac arrest in dogs with mild hypothermia plus blood flow
promotion. Stroke. 1996;27:105–113.[Abstract/Free Full Text]
25.
Xu X, Zhou Y, Ma Q, Zhu J, Ma D. Establishing a
resuscitation model in rabbits with closed-thoracic
cardiopulmonary by-pass. Resuscitation. 1994;27:61–66.[Medline]
[Order article via Infotrieve]
26.
Larm O, Larsson R, Olsson P. A new non-thrombogenic
surface prepared by selective covalent binding of heparin via a
modified reducing terminal residue. Biomater Med Devices Artif
Organs. 1983;11:161–173.[Medline]
[Order article via Infotrieve]
27.
Bozdayi M, Borowiec J, Nilsson L, Venge P, Thelin S,
Hansson HE. Effects of heparin coating of cardiopulmonary
bypass circuits on in vitro oxygen free radical production
during coronary bypass surgery. Artif Organs. 1996;20:1008–1016.[Medline]
[Order article via Infotrieve]
28.
Wendel HP, Heller W, Gallimore MJ, Hoffmeister HE.
Heparin-coated oxygenators significantly reduce contact system
activation in an in vitro cardiopulmonary bypass model.
Blood Coagul Fibrinolysis. 1994;5:673–678.[Medline]
[Order article via Infotrieve]
29.
Wendel HP, Heller W, Gallimore MJ. Heparin-coated
devices and high-dose aprotinin optimally inhibit contact system
activation in an in vitro cardiopulmonary bypass model.
Immunopharmacology. 1996;32:128–130.[Medline]
[Order article via Infotrieve]
30.
Iijima T, Bauer R, Hossmann KA. Brain resuscitation by
extracorporeal circulation after prolonged cardiac arrest in cats.
Intensive Care Med. 1993;19:82–88.[Medline]
[Order article via Infotrieve]
31.
Seo K, Ishimaru S, Hossmann KA. Two-stage resuscitation
of the cat brain after prolonged cardiac arrest. Intensive Care
Med. 1991;17:410–418.[Medline]
[Order article via Infotrieve]
32.
Barnett WM, Alifimoff JK, Paris PM, Stewart RD, Safar
P. Comparison of open-chest cardiac massage techniques in dogs.
Ann Emerg Med. 1986;15:408–411.[Medline]
[Order article via Infotrieve]
33.
Xu Y, Liachenko S, Tang P, Yan B, Melick JA. Cardiac
arrest and resuscitation at 9.4 T: interleaved
31P/1H NMR evaluation of
cerebral protection by fructose-1,6-bisphosphate. Proc Int Soc
Magn Reson Med. 1996;4:912. Abstract.
34.
Blomqvist P, Wieloch T. Ischemic brain damage
in rats following cardiac arrest using a long-term recovery model.
J Cereb Blood Flow Metab. 1985;5:420–431.[Medline]
[Order article via Infotrieve]
35.
Rosenthal RE, Chanderbhan R, Marshall G, Fiskum G.
Prevention of post-ischemic brain lipid conjugated diene
production and neurological injury by hydroxyethyl
starch-conjugated deferoxamine. Free Radic Biol
Med. 1992;12:29–33.[Medline]
[Order article via Infotrieve]
36.
Saransaari P, Oja SS. Dizocilpine binding to cerebral
cortical membranes from developing and ageing mice. Mech Ageing
Devel. 1995;85:171–181.[Medline]
[Order article via Infotrieve]
37.
McDonald JW, Trescher WH, Johnston MV. Susceptibility
of brain to AMPA induced excitotoxicity transiently peaks during early
postnatal development. Brain Res. 1992;583:54–70.[Medline]
[Order article via Infotrieve]
38.
Wass CT, Lanier WL. Hypothermia-associated protection
from ischemic brain injury: implications for patient
management. Int Anesthesiol Clin. 1996;34:95–111.[Medline]
[Order article via Infotrieve]
39.
Simon D, Taylor MJ, Elrifai AM, Shih SR, Bailes JE,
Davis D, Kluger Y, Diamond DL, Maroon JC. Hypothermic blood
substitution enables resuscitation after hemorrhagic shock and 2 hours
of cardiac arrest. ASAIO J. 1995;41:M297–M300.[Medline]
[Order article via Infotrieve]
40.
Leonov Y, Sterz F, Safar P, Radovsky A. Moderate
hypothermia after cardiac arrest of 17 minutes in dogs: effect on
cerebral and cardiac outcome. Stroke. 1990;21:1600–1606.[Abstract/Free Full Text]
41.
Bircher NG. Acid-base physiology during advanced
cardiac life support. Anesthesiol Clin North Am. 1995;13:799–808.
42.
Fischer M, Hossmann KA. No-reflow after cardiac arrest.
Intensive Care Med. 1995;21:132–141[Medline]
[Order article via Infotrieve]
Editorial Comment
Lawrence Litt, PhD, MD, Guest Editor
Departments
of Anesthesia and Radiology and the
Cardiovascular Research Institute University of
California at San Francisco San Francisco, California
|
Introduction
|
|---|
Complex, remote physiological manipulations of
animals in an NMR spectrometer are not new to Dr Xu, the senior author.
He previously used NMR to study brain intracellular pH in very
hypercapnic rats (PCO2
750 mm Hg) in a hyperbaric chamber.1
Although advantages of rodent models include reduced animal costs,
relevant cerebrovascularity, and a small brain size convenient for
fixation and immunohistochemistry, the impressive new
"in-the-magnet" stroke model of Liachenko et al is clearly
"technology driven." Rodents fit into high-field magnets, and the
new model opens rodent studies of complete cerebral ischemia to
sophisticated NMR methodologies currently being used for preclinical
studies of focal cerebral ischemia. The article is
appropriately brief about plans for future rodent studies. It stays
focused on details of the model. However, we must appreciate that the
authors have a high-field system (9.4-T magnet; 400-MHz proton
frequency), with custom-made radiofrequency coils and a versatile
computer console. From an NMR perspective, sophisticated NMR
spectroscopy and MRI techniques are possible, and also improved, by
having smaller animals at higher magnetic fields. For example, spin
tagging of arterial water protons in the neck should allow
for reliable perfusion MRI images (ie, images of cerebral blood flow)
in the rodents. Tagged water protons can traverse the brain more
quickly in the rat than in the human, where transit time, long enough
for relaxation effects to be important, is a limiting factor for NMR
approaches to cerebral blood flow
determinations.2 Similarly, brain imaging of
pH and lactate,3 4 as well as several other NMR
methods, should be more feasible at high magnetic fields. Ultimately,
technological NMR advances might permit in vivo
physiological monitoring to be accomplished
completely via specialized NMR imaging and spectroscopy. NMR-derived
parameters can be used to regulate and define
physiological insults, while neurological
examinations and histology/immunohistochemistry are used to quantify
outcome and injury. Such physiological monitoring
would, of course, be even more welcome for studies in mice, where it
could assist valuable testing of genetic modulations. But even if NMR
methods are not extending rapidly to mice, genetic methods are slowly
being extended to rats. Genetic engineering in rats has become a
reality, with genetic overexpression being accomplished more easily at
this time than genetic knock-out.5 One can
imagine, for example, using the authors' model to study transgenic
rats that overexpress human copper-zinc superoxide dismutase
1.5 The authors could test interesting
hypotheses about the role of oxidative stress6 in complete
cerebral ischemia.
But what about the fact that the authors' model employs asphyxia?
Other "in-the-magnet" cardiac arrest models with larger animals
have been used for important studies,7 8 but
with fibrillation followed by defibrillation. Such an approach more
closely models the situation for adult humans. Asphyxial cardiac arrest
is more relevant to pediatric global ischemia, a point
appreciated and discussed by the authors. If asphyxia is a disadvantage
to the model, a redeeming aspect is the consistency and
reproducibility of hemodynamic and
metabolic resuscitation. Perhaps the model's achievements
in resuscitation exploit the fact that fibrillating rodent hearts can
be defibrillated without electroshock.
When investigators undertake in-the-magnet studies, will they make this
model as popular as the "suture occlusion"
model9 10 11 has been made for NMR studies of
focal ischemia12,13? As stated in a
study and review by Laing et al,14 the
"suture occlusion" model was first published in 1986 by Koizumi et
al 9 and then
published in a variation by Longa et al.10
Important information about the evolution of T2-weighted imaging and
diffusion-weighted imaging in focal ischemia, first done in
animal studies,15 is turning out to be very
useful in MRI evaluations of human stroke,16
especially in differentiating new infarcts from old ones. In summary,
the new complete cerebral ischemia model by Liachenko et al
should permit important rodent studies of new drugs and regimens. If
protocols can be found that improve rodent neurological outcome after
cardiac arrest, one can then search for NMR/MRI parameters
that are associated with improvement and gain insight into mechanisms
and measures to be focused upon in humans. Interest in the authors'
stroke model will clearly increase after it is used for important
preclinical NMR investigations.
|
Selected Abbreviations and Acronyms
|
|---|
| CPR |
= |
cardiopulmonary resuscitation |
| HR |
= |
heart rate |
| MABP |
= |
mean arterial blood pressure |
| MDPA |
= |
methylene diphosphonic acid |
| NAA |
= |
N-acetyl-aspartate |
| NDS |
= |
neurological deficit scores |
| NMR |
= |
nuclear magnetic resonance |
| pHi |
= |
intracellular pH |
| ROSC |
= |
restoration of spontaneous circulation |
|
Received October 20, 1997;
revision received March 16, 1998;
accepted March 17, 1998.
|
References
|
|---|
1.
Xu Y, Cohen Y, Litt L, Chang L-H, Severinghaus JW,
James TL. Tolerance of low cerebral intracellular pH during severe
hypercapnia and adequate oxygenation in rats: an in
vivo NMR study in a hyperbaric chamber. Stroke. 1991;
22:1303–1308.
2.
Wong EC, Buxton RB, Frank LR. Implementation of
quantitative perfusion imaging techniques for functional brain mapping
using pulsed arterial spin labeling. NMR Biomed.. 1997;10:237–249.[Medline]
[Order article via Infotrieve]
3.
Van Zijl PC, Barker PB. Magnetic resonance spectroscopy
and spectroscopic imaging for the study of brain
metabolism. Ann N Y Acad Sci.. 1997;820:75–96.[Medline]
[Order article via Infotrieve]
4.
Ebisu T, Rooney WD, Graham SH, Mancuso A, Weiner MW,
Maudsley AA. MR spectroscopic imaging and diffusion-weighted MRI for
early detection of kainate-induced status epilepticus in the rat.
Magn Reson Med.. 1996;36:821–828.[Medline]
[Order article via Infotrieve]
5.
Chan PH. Transgenic mice, In: Welch KMA, Caplan LR, Reis
DJ, Siesj
BK, Weir B, eds. Primer on Cerebrovascular
Diseases. New York, NY: Academic Press Inc; 1997:28–31.
6.
Chan PH. Role of oxidants in ischemic brain
damage. Stroke. 1996; 27:1124–1129.
7.
Schmitz B, Fischer M, Bockhorst K, Hoehn-Berlage M,
Hossmann KA. Resuscitation from cardiac arrest in cats: influence of
epinephrine dosage on brain recovery. Resuscitation. 1995;30:251–262.[Medline]
[Order article via Infotrieve]
8.
Eleff SM, Sugimoto H, Shaffner DH, Traystman RJ, Koehler
RC. Acidemia and brain pH during prolonged cardiopulmonary
resuscitation in dogs. Stroke.1995;26:1028–1034.
9.
Koizumi J, Yoshida Y, Nakazawa T, Ooneda G. Experimental
studies of ischemic brain edema, I: a new experimental model of
cerebral embolism in rats in which recirculation can be introduced in
the ischemic area. Jpn J Stroke.. 1986;8:1–8.
10.
Longa EZ, Weinstein PR, Carlson S, Cummins R. Reversible
middle cerebral artery occlusion without craniectomy in rats.
Stroke.. 1989;20:84–91.[Abstract/Free Full Text]
11.
Yang GY, Chen SF, Kinouchi H, Chan PH, Weinstein PR.
Edema, cation content, and ATPase activity after middle cerebral artery
occlusion in rats. Stroke.. 1992;23:1331–1336.[Abstract/Free Full Text]
12.
Jiang Q, Zhang ZG, Chopp M, Helpern JA, Ordidge RJ,
Garcia JH, Marchese BA, Qing ZX, Knight RA. Temporal evolution and
spatial distribution of the diffusion constant of water in rat brain
after transient middle cerebral artery occlusion [published correction
appears in J Neurol Sci. 125:113]. J Neurol
Sci.. 1993;120:123–130.[Medline]
[Order article via Infotrieve]
13.
Minematsu K, Fisher M, Li L, Sotak CH. Diffusion and
perfusion magnetic resonance imaging studies to evaluate a
noncompetitive N-methyl-D-aspartate
antagonist and reperfusion in experimental stroke in rats.
Stroke. 1993; 24:2074–2081.
14.
Laing RJ, Jakubowski J, Laing RW. Middle cerebral artery
occlusion without craniectomy in rats: which method works best?
Stroke.. 1993;24:294–297.[Abstract/Free Full Text]
15.
Moseley ME, Cohen Y, Mintorovitch J, Chileuitt L, Shimizu
H, Kucharczyk J, Wendland MF, Weinstein PR. Early detection of regional
cerebral ischemia in cats: comparison of diffusion- and
T2-weighted MRI and spectroscopy. Magn Reson Med.. 1990;14:330–346.[Medline]
[Order article via Infotrieve]
16.
Lutsep HL, Albers GW, DeCrespigny A, Kamat GN, Marks MP,
Moseley ME. Clinical utility of diffusion-weighted magnetic resonance
imaging in the assessment of ischemic stroke. Ann
Neurol.. 1997;41:574–580.[Medline]
[Order article via Infotrieve]
Top
Abstract
Introduction
), according to
pHi=6.683+log[(
