close

Вход

Забыли?

вход по аккаунту

?

Cerebal norepinephrine depletion enhances recovery after brain ischemia.

код для вставкиСкачать
Cerebral Norepinephne Depletion
Enhances Recovery after Brain Ischemia
Raul Busto, BS, Sami I. Harik, MD, Shinichi Yoshida, MD, Peritz Scheinberg, MD,
and Myron D. Ginsberg, M D
Monoamine neurotransmitters, especially norepinephrine (NE),may have an important role in the pathophysiological
aspects of postischemic cerebral dysfunction. In previous studies of post-decapitation-induced ischemia, we found that
NE depletion caused a delay in glycogen breakdown but did not influence any of the other known biochemical
abnormalities that accompany brain ischemia. In this study, we have turned to a model of transient incomplete and
diffuse forebrain ischemia in the rat to examine the effects of cerebral NE depletion on the recovery after brain
ischemia of levels of high-energy phosphate compounds, products of intermediary oxidative metabolism, and free fatty
acids. We found that a unilateral lesion of the locus ceruleus and the resultant depletion of NE in the ipsilateral
cerebral cortex had no effect on sham-operated controls nor on rats subjected to ischemia alone. However, in rats
subjected to ischemia followed by 15 minutes of recirculation, the NE-depleted cerebral cortex had significantly
higher phosphocreatine and adenosine triphosphate levels and energy charge, and lower adenosine monophosphate
and docosahexaenoic acid concentrations. With longer periods of recirculation, these side-to-side differences were not
apparent. These results suggest that activity of the central NE systems during transient brain ischemia has deleterious
effects on the biochemical recovery of the cerebral cortex from severe ischemic insults.
Busto R, Harik SI, Yoshida S, Scheinberg P, Ginsberg MD: Cerebral norepinephrine depletion enhances
recovery after brain ischemia. Ann Neurol 18:329-336, 1985
During the past decade, there has been a surge of
interest in the possible relationship between brain
monoamines and ischemic brain insults [ S , 6, 12, 18,
20, 23, 241. One reason for this increased interest is
the possibility that putative monoamine neurotransmitters may have an important role in the pathophysiological aspects of postischemic cerebral dysfunction.
Another reason is the possibility that endogenous
monoamines, through their influence on cerebral
blood vessels [9, 311, may regulate cerebral blood
reflow and perfusion after the ischemic insult and,
thereby, influence the recovery of the brain from ischemia. If these hypotheses are correct, pharmacological
manipulations of central monoaminergic activity may
enhance recovery in animal models of brain ischemia
and eventually may have a role in the treatment of
human cerebrovascular disease.
In previous work from this laboratory, we investigated the effect of unilateral locus ceruleus (LC) lesions and the resultant depletion of cerebral norepinephrine (NE) on high-energy phosphate compounds
and products of intermediary metabolism in the rat
cerebral cortex after post-decapitation-produced ischemia [14}. A unilateral LC lesion causes profound de-
pletion of NE in the ipsilateral cerebral cortex, while
leaving the contralateral cortex with its normal NE
content as an internal control. We found that NE depletion of the cerebral cortex delayed the breakdown
of glycogen, probably due to impaired cyclic adenosine
monophosphate (AMP) generation in the NE-depleted cortex; however, NE depletion had no effect on
high-energy phosphate compound levels nor on the
products of intermediary oxidative metabolism [ 141.
In another study, we examined the effect of a unilateral LC lesion on the accumulation of free fatty acids
(FFAs) in the cerebral cortex after post-decapitationinduced ischemia [381. No significant effect of NE
depletion on the accumulation of FFAs or on energy
metabolites of the brain after total ischemia was noted.
Having failed to observe an influence of N E depletion
on total irreversible brain ischemia, we examined the
effects of NE depletion on the biochemical recovery of
the cerebral cortex in a rat model of incomplete and
transient forebrain ischemia. In this paper we report
our findings that, two weeks after unilateral LC lesion,
cerebral NE depletion is associated with an earlier recovery of phosphocreatine (PCr), adenosine triphosphate ( A m ) , AMP, and FFA levels, as well as energy
From the Cerebral Vascular Disease Research Center and Neurol-
om DeDartment, Universitv of Miami School of Medicine. Miami.
Received Jan 4, 1985, and in revised form Feb 20. Accepted for
Dubfication Feb 20. 1985.
'"0"Western
and the
and Pha"aco'ogy'
Case
Reserve UniversityOfSchool of Medicine,
Cleveland,
OH 44106.
Address reprint requests to Mr Busto, Department of Neurology,
Box 016960,
D4-5, University of Miami School of Medicine,
Miami, FL.33101.
329
charge. This enhanced recovery was documented at 15
minutes of recirculation after 45 minutes of ischemia.
These results suggest that activity of central noradrenergic mechanisms during transient cerebral ischemia has a deleterious effect on the early metabolic
recovery of the cerebral cortex after severe ischemic
insult.
45' ISCHEMIA
RH
Materials and Methods
Animals
Male Wistar rats (weight, 2 5 0 to 300 gm) were used in all
experiments. The rats were kept under diurnal light conditions with free access to laboratory chow and water for one
week before LC lesion. Unilateral LC lesion was made under
chloral hydrate anesthesia (400 mgikg, intraperitoneally) and
pargyline pretreatment ( 5 0 mg/kg, intraperitoneally) by local
infusion into the LC of 5 pg of 6-.hydroxydopamine base
dissolved in 2.5 p1 of solution. The details of the lesion
procedure have been reported previously [l5, 191. Approximately equal numbers of right and left lesions were created.
The rats were returned to their cages when awake and allowed to recover for two weeks with free access to food and
water until they were used for further experiments, to be
described below.
RAT MODEL FOR INCOMPLETE, DIFFUSE, AND REVERSIBLE
FOREBRAIN ISCHEMIA. Diffuse and incomplete forebrain
ischemia was produced in the rats under nitrous oxide anesthesia by a procedure combining bilateral carotid artery occlusion, elevation of intracranial pressure to between 10 and
16 mm H g by infusion of mock cerebrospinal fluid (CSF),
and induction of moderate systemic hypotension to between
55 and 60 mm Hg by halothane inhalation.
Rats were initially anesthetized with 3% halothane, paralyzed with curare ( 5 mg/kg, intraperitoneally), tracheostomized, and ventilated with a mixture of 1% halothane,
30% oxygen, and the balance of nitrous oxide. A femoral
artery was cannulated for continuous monitoring of the mean
arterial blood pressure (MABP) and for obtaining samples of
arterial blood for oxygen pressure (PaOZ), carbon dioxide
pressure (PaC02),pH, plasma glucose levels, and hematocrit
determinations. Both common carotid arteries were isolated
and encircled with PE-10 polyethylene tubes that were then
passed through double-lumen Silastk tubes. The rat was secured in a headholder in the prone position, and the scalp
and periosteum were incised to accommodate a plastic funnel
over the calvarium for later in situ freezing of the brain. Four
electrodes were placed on the frontal and occipital bones,
bilaterally, to record bipolar electroencephalographic (EEG)
activity. The atlanto-occipital membrane was then exposed in
the midline, and a double-barrel needle was inserted into the
cisterna magna and secured with glue to the adjacent muscles. One barrel of this needle assembly was connected to a
pump for the infusion of mock CSF and the other barrel was
connected to a strain-gauge transducer for monitoring of intracranial pressure. Mock CSF was prepared as described by
Ljunggren and associates 12 11. After completion of the surgical procedure, halothane was discontinued and 4 5 minutes
were allowed for stabilization. During this period, the respiratory rate and tidal volume were adjusted to obtain P a 0 2
LH
----
40' RECIRCULATION
RH
-_
-
__
-
--
-A+----
Fig 1 . Example of the electroencephalographicactivity in the
various stages of the rat model of incomplete reversibleforebrain
ischemia. The control tracing was obtained at the end of the
stabilization period before ischemia was induced. The isoelectric
ischemic tracing was obtained at the end of 45 minutes of ischemia. The lower two tracings were obtained at 40 and 180 minutes of recirculation. (RH = right hemisphere; LH = lt$t hemisphere.)
values between 100 and 150 mm Hg and PaCOz values
between 35 and 4 5 mm Hg. Temperature (measured rectally) was maintained close to 37°C by an infrared lamp.
After the stabilization period, ischemia was induced by
inhalation of 5.0 to 5.5% halothane to produce systemic
hypotension. Within 1 to 2 minutes, MABP declined to 60
mm Hg, and at that time, the carotid arteries were occluded
bilaterally and infusion of mock CSF was started. Usually 4
minutes were required to stabilize the MABP between 55 to
60 mm Hg and the cisternal CSF pressure between 10 to 16
mm Hg. Ischemia was maintained for 4 5 minutes. The EEG
became isoelectric within 1 minute following carotid occlusion and remained so for the duration of ischemia (Fig 1).
During ischemia, the respiratory volume was reduced to
maintain PaC02 in the range of 35 to 4 5 mm Hg, and halothane concentration in the inspired gas was gradually decreased to about 2.5 to 3.0%.
Recirculation was instituted by simultaneously stopping
halothane inhalation and mock CSF infusion, and by releasing the carotid ligatures. MABP recovered to above 100 mm
Hg within 15 minutes. Slow waves began to appear sporadically on the EEG recording at 40 to 60 minutes of reflow,
but the EEG pattern remained abnormal at the end of 180
minutes of reflow (see Fig 1). The brains were frozen in situ
330 Annals of Neurology Vol 18 No 3 September 1985
F i g 2. Schematic representation of the anatomical regzons of the
cerebral cortex sampledfor the various biochemical analyses: A
for free fatty acids; B for energy metabolites and glycolytic intermediates;and C for norepinephrine.A, B, and C samples from
each side weighed approximately 100, 5 , and 13 mg, respective&. (See Materials and Methods rection for details.)
at the end of 45 minutes of ischemia, or after 15, 30, or 180
minutes of recirculation.
There were two control groups for this experiment. Rats
in both control groups underwent the entire surgical procedure, but the common carotid arteries were not occluded,
mock CSF was not infused, and hypotension was not induced. In one control group (5 rats), the brain was frozen in
situ after the stabilization period. In the other control group
(4 rats), the brains were frozen 225 minutes after the end of
the stabilization period to ascertain whether the duration of
the procedure affected the results.
Transcalvarial freezing of the brain was performed by
pouring liquid nitrogen into the cranial funnel while the rats
were being ventilated [29). The frozen heads were sectioned
coronally at 2, 7, and 11 mm anterior to the lambda, using a
vibrating Dremel band saw, with the saw blade continuously
irrigated with liquid nitrogen. Brain blocks were stored in
liquid nitrogen until analysis. Samples of each cerebral cortex
were obtained from the coronal blocks as detailed in Figure
2. A 15 mg sample of tissue obtained from the frontal cerebral cortex was used for NE assay. Another sample ( 5 mg)
from the superior part of the frontal cortex was used for the
assay of high-energy phosphate compounds and glycolytic
intermediates. The rest of the frontal and parietal cortex
(100 mg) was used for FFA determinations. All samples
were dissected in a refrigerated glove box maintained at
- 30°C and weighed on a Cahn balance.
Assay Procedures
ATP, adenosine diphosphate (ADP), AMP, glycogen, glucose, lactate, PCr, and creatine levels were all determined in
one tissue sample weighing about 5 mg.Tissue was hornogenized in 40 pl of 0.1 N hydrochloric acid and 99% methanol
at - 30°C in a glass-glass homogenizer. Deproteinization was
then carried out with perchloric acid and the supernatant was
neutralized. ATP, ADP, AMP, glucose, lactate, PCr, and
creatine were assayed by the direct fluorometric assay procedures of Lowry and Passonneau 122). Glycogen content was
measured according to the method of Passonneau and
Lauderdale [28). Total adenylate concentrations and the energy charge were calculated according to the method of Atkinson {I).
FFAs were assayed by gas-liquid chromatography after
conversion into methyl esters by diazomethane [38, 40).
Heneicosanoic acid (C 2 1:O) was used as an internal standard.
Palmitic (C 16:0), stearic (C 18:0), oleic (C 18:1), arachidonic
(C 20:4), and docosahexaenoic (C 22:6) acid concentrations
were estimated from their peak areas.
NE was assayed in 0.1 N perchloric acid extracts of tissue
samples by high-performance liquid chromatography with
electrochemical detection (HPLC-EC) 171. In brief, tissue
was weighed in the frozen condition and homogenized in
twenty volumes of ice-cold 0.1 N perchloric acid. The
homogenates were centrifuged at 20,000 g for 15 minutes
and aliquots of the supernatant were injected directly into
the HPLC-EC system (model LC-304 CA; Bioanalytical Systems). This system uses a biophase (octadecyl silane, 5 pm)
column, 25 cm in length and 5 mm in diameter. The potential at the glassy-carbon electrode was 0.8 V against a silversilver chloride reference electrode. The mobile phase consisted of 10% methanol in water; 1% acetic acid; 1 mM
EDTA; and 2 m~ I-heptane sulfonic acid. The p H was adjusted to 3.8. The flow rate of the mobile phase was 1.8 mY
min.
Analysis of Data
The effects of ischemia and ischemia followed by various
periods of recirculation in the contralateral and ipsilateral
cerebral cortices of rats with unilateral LC lesions were estimated by one-way analysis of variance (ANOVA). The effects of NE depletion were estimated by the paired Student's
t test (two-tailed), which compared the results obtained from
the ipsilateral and contralateral cerebral cortex of each rat
used in the analysis of data. Significance for both of these
statistical methods was considered at <0.05.
The success of the LC lesion was assessed from NE data.
An LC lesion was considered successful when the NE level
in the ipsilateral hemisphere was reduced to less than onethird of the NE content of the contralateral cortex of control
rats. Data from rats not satisfying this criterion were not
considered in data analysis.
There were no significant differences in any of the variables between the two groups of control rats. For this reason,
the results obtained from the two control groups were combined (9 rats). Five rats in the ischemiagroup, and 6, 7, and 8
rats in the groups undergoing recirculation for 15, 30, and
180 minutes, respectively, were included in the analysis of
data.
Results
I n this rat model of incomplete bilateral forebrain ischemia, cerebral blood flow is reduced to 3 to 5% of
Busto et al: NE Depletion and Brain Ischemia
331
PbyJiological Variablesa
Control
Variable
45 Minutes of Ischemia followed by Recirculation for
Ischernia
(45 min)
start
End
start
144.0
148.9
t
t
12.7
16.7
149.0
+
10.2
123.6
141.3
22.3
133.6
+11.5
40.3
t
15 rnin
180 min
start
End
Start
End
start
End
56.0
151.4
145.7
144.0
132.0
151.0
127.9
?
f
k
5
f
t
5
4.2
16.5
11.3
19.2
12.5
12.3
19.7
"
128.3
135.7
131.6
144.7
156.5
141.4
t
5
t
%
11.3
20.5
11.3
38.9
40.5
38.3
38.6
37.8
f
5
t
t
3.6
2.2
1.7
1.6
7.419
7.411
7.416
f
2
0.045
0.032
c
0.022
204
-c
19
286
+
66
225
+
52
44.3
45.0
f
t
2.6
3.1
t
End
30 min
18.2
19.6
37.8
39.8
41.1
t
t
t
t
0.7
6.9
38.6
-c
1.2
136.7
r
12.5
2.0
2.1
2.0
1.8
7.391
7.408
7.375
7.427
7.389
t
c
2
t
c
0.027
0.045
0.040
0.053
0.056
7.402
+
0.656
7.406
+
0.041
258
161
*
t
12.0
211
+
21
192
+
41
279
267
210
f
t
t
f
64
83
47.7
43.8
43.0
42.0
23
44.8
31
44.2
c
63
39.8
f
t
t
f
t
k
t
t
2.9
2.9
2.5
3.9
7.2
6.2
3.5
4.0
41.6
'Results are expressed as means f SD of the physiological variables of each group of rats measured at the start (i.e,, at the end of the period of
stabilization after surgical procedures) and at the end (just before transcalvarial freezing) of each experiment.
MABP = mean arterial blood pressure; PaO, = arterial oxygen tension; PaC02 = arterial carbon dioxide tension; Hct
control values in the frontal and parietal cerebral cortex, and returns to preischemic levels within 30 minutes of recirculation (Harik and colleagues, unpublished observations, 1985). The determinations of
physiological variables obtained at the start of the experiments and immediately before sacrifice for each of
the experimental groups are presented in the Table.
At the end of 45 minutes of ischemia, PCr and ATP
levels were nearly depleted, AMP and creatine concentrations were increased, and the energy charge and
total adenylate values were decreased (Figs 3, 4). Also,
cerebral glycogen and glucose levels were almost totally depleted (Fig 5). These results were expected after severe cerebral ischemia. Lactate accumulated but
not as much as was expected in a model of incomplete
ischemia (Fig 5).
When recirculation was established, the following
pattern was observed: the PCr level gradually recovered and attained values that were higher than control
values in the NE-depleted cerebral cortex at 30 minutes of recirculation, and in both sides at 180 minutes
of recirculation (p < 0.01; Fig 3). Creatine concentration also decreased and became significantly lower than
control values after 180 minutes of recirculation (p <
0.01; Fig 3). The ATP level gradually increased with
time but never reached control values, even at 180
minutes of recirculation (p <: 0.01; Fig 4). ADP levels
=
hematocrit.
did not change significantly throughout the period of
ischemia and recirculation, and AMP concentration
decreased during recirculation, attaining low control
values afer 30 minutes (Fig 4). Total adenylate values
steadily increased but never reached control values.
Energy charge in the contralateral cortex was significantly lower than control at 15 minutes of recirculation (p < 0.01), whereas it reached normal values in
the NE-depleted cortex. At 30 minutes of recirculation, both sides attained control energy charge values
that were maintained for the duration of the experiment (Fig 4).
The glycogen value recovered slowly, probably because of the relatively low activity of brain glycogen
synthetase [13, 25, 271, and attained values that were
slightly higher than control by 180 minutes of recirculation (Fig 5). The glucose value, on the other hand,
recovered quickly, reaching control values by 15 minutes of recirculation and exceeding control values at 30
minutes (p < 0.01; Fig 5). The glucose content then
declined slightly and was still significantly higher than
control in the ipsilateral cortex by 180 minutes of recirculation (p < 0.05; Fig 5). These results suggest that
the capability of the postischemic cerebral cortex to
use glucose is impaired. Lactate gradually decreased to
control values by 180 minutes of recirculation (Fig 5).
FFA levels in control rats were in agreement with
332 Annals of Neurology Vol 18 No 3 September 1985
pmol/g
N O R E PINE PH R I N E
ATP
3
300
2
200
t
++
++
I
100
0
0
prnol/g
AOP
I
PHOSPHOCREATINE
02
0
++
I
CREATINE
Z AOENYLATES
4 t
2
CONTROL
45'
ISCHEMIA
15'
30'
180'
0
RECIRCULATION
Fig 3. Mean values (SEM) of observationsfrom the contralateral
(open columns) and ipsilateral (shaded columns) cerebral cortex of
9 rats in the control group; 5 rats in the ischemia group; and 6 ,
7, and 8 rats in the groups in which ischemia was followed by
recirculationfor 15, 30, and 180 minutes, respectively. Norepinephrine values are expressed as nanograms per gram of tissue
while phosphocreatine and creatine values are expressed as micromolesper gram of tissue. ior t wer one or both columns
indicates that results of one or both hemispheres are significanti)
d;ffent from the control group ( + at p < 0.05 or t t at p <
0.01 by ANOVA followed by Dunni- multiple comparison test).
* or ** indicates significant differences between ipsilateral and
contralateral results within the same group (" d t p < 0.05 and
** at p < 0.01 by the paired Student's t test, two-tailed).
+
values reported in the literature for rat cerebral cortex
not exposed to ischemia [lo, 32,401. During ischemia,
all FFAs that we assayed increased markedly in concentration, with a preferential increase in stearic and
arachidonic acids as originally found by Bazan 121 (Fig
6). At the end of ischemia, total FFA concentrations
increased by 13-fold over control values, but invariably
returned to control values by 180 minutes of recirculation (Figs 6, 7). Total FFA values were generally
higher in the contralateral cerebral cortex during ischemia and recirculation, with the difference between
the two sides reaching maximum at 45 minutes of ischemia followed by 15 minutes of recirculation (Fig 7).
Consistent with previous results, the polyunsaturated
FFAs, arachidonic and docosahexaenoic acids, returned to control levels more quickly than other FFAs
C37, 39, 401.
A unilateral LC lesion produced severe loss of NE
E N E R G Y CHARGE
CONTROL
45'
ISCHEMIA
IS'
30'
180'
RECIRCULATION
Fig 4. Results of adenosine triphosphate (ATP), adenosine
diphosphate (ADP), and adenosine monophosphate (AMP), total adenylates, and energy charge measurements expressed as micromoles per gram of tissue. (See legend for Figure 3 for remaining expkznation.)
in the ipsilateral cerebral cortex, similar to previous
results from our laboratory [14-16, 19, 401. The severe loss of NE was noted in control rats and after
ischemia, with and without recirculation (Fig 3). NE
levels in the contralateral cerebral cortex decreased
significantly during ischemia and declined further after
15 and 30 minutes of recirculation. NE content increased slightly at 180 minutes of tecirculation, but
never fully recovered to control levels (p < 0.01).
These results showing decreased NE levels in the contralateral cerebral cortex after ischemia and recirculation are compatible with previous reports [5, 61.
The important consideration in these data is the effect of NE depletion on brain ischemia and the recovery from ischemia that could be gleaned by assessing
the differences between the NE-depleted ipsilateral
cerebral cortex and the contralateral cerebral cortex.
With the exception of NE, there were no significant
Busto et al: NE Depletion and Brain Ischemia
333
pmol/g
nrnor/g w e t weight
G L Y C 0G E N
3t
500
c
++
16:O
T
T
250
0
18:O
G LUC 0 S E
++
r+
++
18:l
++
500
+
250
0
++
200
100
++
I
LACTATE
0
500
++
/-
++
20:4
250
"
45'
CONTROL
ISCHEMIA
15'
180'
30'
0
RECIRCULATION
++
22:6
45'
IS'
Fag 5 . Results of glycogen, glucose, and lactate measurements expressed as micromoles per gram oftissue. (See legend for Figure 3
for remaining explanation.)
differences in any of the variables measured between
the two hemispheres of control rats. After ischemia,
both cerebral hemispheres also reacted in a similar
fashion, thus indicating that NB depletion has little or
no role in the reaction of the cerebral cortex to incomplete diffuse forebrain ischemia. However, early after
recirculation (i.e., 45 minutes of ischemia followed by
15 minutes of recirculation), there were significant differences between the two sides in PCr, ATP, and
AMP values, and energy charge (Figs 3, 4). Similarly,
FFA levels decreased more rapidly after recirculation
in the NE-depleted ipsilateral cerebral cortex, with the
differences between the two sides reaching statistical
significance for docosahexaenorc acid (Fig 6). All sideto-side differences attributed to NE depletion faded
with longer periods of recirculation. It should be noted
that all differences between the two hemispheres were
in the direction of quicker recovery from the ischemic
insult in the NE-depleted ipsilateral cerebral cortex.
Discussion
Two major aspects of the results warrant discussion.
The first documents the efficacy of this rat model of
diffuse forebrain ischemia in producing the various
biochemical changes of ischemic brain insult, and provides evidence that recovery of these biochemical parameters takes place during recirculation. The mecha334
Annals of Neurology
Vol 18 No 3
CONTROL
ISCHEMIA
180'
30'
RECIRCULATION
Fig 6. Results of free fatty acid determinations:palnzrtic (C
16:0), stearic (C 18:0), oleic (C 18:1), arachidonic (C 20:4),
and docosahexaenoic (C 226) acids, expressed as nanomoles per
gram of tissue. (See legend for Figure 3 for remaining explanation.)
nmol/g
++
2000
LT
++
i
1500
1000
500
0
m
45'
CONTROL
CHEMIA
15'
30'
180'
RECIRCULATION
Fig 7. Results of totalfree fatty acid (FFA) determinations derived by the addition of the five individual FFAs we assayed,
expressed as nanomoles per gram of tissue (see Fig 6j. (See legend
for Figure 3 j & remaining explanation.)
September 1985
nisms underlying the changes in levels of energy
metabolites and glycolytic intermediates after ischemia
and recirculation have been the subject of several reports that employed different animal models of diffuse
ischemia C17, 30, 33, 361. Our results are consistent
with the general patterns reported by others. Another
parameter employed in our experiments was brain
FFA concentrations, which increase during ischemia
12) and return to normal after recirculation ~ 3 2 37,
,
39,403. Accumulation of brain FFAs in ischemia is the
result of both activation of phospholipases [8) and inhibition of FFA activation to their acyl-coenzyme As
C3, 35, 38). During recirculation, a major portion of
the accumulated FFAs probably becomes reacylated
and subsequently incorporated into phospholipids as
the energy state of the brain recovers [34]. A small
fraction of free arachidonic acid undergoes enzymatic
peroxidation to yield various prostaglandins I1 11 and
leukotrienes C261. Faster incorporation of polyunsaturated FFAs into phospholipids, as compared with
saturated FFAs, is probably responsible for the rapid
preferential decrease of polyunsaturated FFAs that we
have noted during recirculation (Fig 6 ) [341.
The second and more important aspect of these results is the effect of NE depletion on the reaction of
the brain to ischemia and recovery from ischemic insult. Monoamines, and NE in particular, have been
suspected of having a role in the reaction of the brain
to ischemia 123). Most of the evidence for this is indirect. For example, increased central monoaminergic
activity has been postulated during ischemia and recirculation because of decreased concentrations of NE
and serotonin and increased levels of their metabolites
in ischemic brain and after recirculation. However, to
our knowledge, this is the first time that changes in
central noradrenergic transmission have been documented to have an effect on the biochemical accompaniments of recovery of the brain from ischemic insult. Our success in demonstrating the effects of NE
depletion is probably the result of the particular suitability of our experimental model in which we compare, within the same animal with a unilateral LC
lesion, the results obtained from the NE-depleted ipsilateral cerebral cortex with those obtained from the
contralateral cerebral cortex with its normal content of
NE. This side-to-side comparison is resistant to interanimal variations that exist in most biological experiments of this nature. These results strongly suggest
that the cerebral cortex which has been depleted of its
endogenous NE is capable of faster recovery from
ischemia than control cortex. The absence of differences in this experimental model between the two
hemispheres after ischemia without recirculation is
compatible with our previous results obtained after decapitation-induced ischemia C14, 40).
There are several possible explanations for the salu-
tary effect of endogenous NE depletion on the process
of recovery from ischemia. One explanation invokes
the possible effects of endogenous NE on the cerebral
circulation 19, 311. If endogenous NE mediates vasoconstriction and decreased cerebral reflow, then these
results may be explained by the better perfusion of the
NE-depleted ipsilateral cerebral cortex during recirculation. This explanation is unlikely in view of the recent work of Blomqvist and colleagues C41, who
showed that endogenous NE depletion by the LC lesion did not affect postischemic brain hypoperfusion in
a rat model of incomplete and reversible forebrain
ischemia similar to the model used in our experiments.
However, because these investigators examined cerebral blood flow at 60 minutes of recirculation after 15
minutes of ischemia, a slight possibility that endogenous NE may have a vascular effect during the earlier
period of recirculation remains. Another possible
mechanism accounting for how NE depletion is associated with enhanced recovery from ischemia invokes a
direct metabolic effect of NE on the cerebral cortex.
We have previously presented evidence of interaction
between endogenous NE depletion and cerebral oxidative metabolism measured in situ by reflection spectrophotometry and microfluorometry C16, 191. In
these experiments, NE depletion 2 weeks after an LC
lesion was associated with a slower metabolic response
to sudden increases in energy demand induced by direct cortical electrical stimulation.
A synaptosomal mechanism involving the release of
NE has been proposed for the elevation of brain FFAs
in pathological conditions 131. Although we failed to
obtain evidence for this hypothesis during ischemia,
NE depletion had an effect on cerebral FFAs during
recirculation. Thus, the level of free docosahexaenoic
acid at 15 minutes of reflow was lower in the NEdepleted cortex than in the contralateral cortex. The
faster recovery of the energy state in the NE-depleted
cerebral cortex could explain this result, because acylation of FFAs, which is an important initial step for
phospholipid synthesis, is an energy-requiring reaction
134). Alternately, NE might inhibit the activity of acyl
transferases for polyunsaturated acyl groups.
Whatever the mechanism, or mechanisms, by which
the effects of NE depletion are mediated, the results
presented in this manuscript suggest that activity of the
cerebral noradrenergic system that originates in the LC
has a deleterious effect on the recovery of the cerebral
cortex from incomplete diffuse forebrain ischemia.
These results should stimulate research to determine
the exact mechanism by which these effects are produced and to study the effects of specific adrenoceptor
blockade on the recovery of the brain from ischemia.
Such research may provide a rational scientific basis for
therapeutic strategies in the treatment of human cerebrovascular diseases.
Busto et
al:
NE Depletion and Brain Ischemia 335
Supported in part by United States Public Health Service grant NS05820 and by a grant from the Cleveland Foundation.
We thank Mss I. Valdez, 0. Alonso, E Martinez, M. Santiso, and L.
Iacofmo for inspired technical assistance.
References
1. Atkinson DE: The energy charge of the adenylate pool as a
regulatory parameter: interaction with feedback modifiers.
Biochemistry 7:4030-4034, 1968
2. Bazan NG: Effects of ischemia and electroconvulsive shock on
free fatty acid pool in the brain. Biochim Biophys Acta 218:l10, 1970
3. Bazan NG: Free arachidonic acid and other lipids in the nervous
system during early ischemia and after electroshock. Adv Exp
Med Biol 72:317-335, 1976
4. Blomqvist P, kndvall 0, Wieloch T Delayed postischemic hypoperfusion: evidence against involvement of the noradrenergic
locus ceruleus system. J Cereb Blood Flow Metab 4:425-429,
1984
5. Brown RM, Carlson A, Ljunggren D, et al: Effect of ischemia
on monoamine metabolism in the brain. Acta Physiol Scand
90:789-791, 1974
6. Calderini G, Carlson A, Nordstrom CH: Influence of transient
ischemia on monoamine metabolism in the rat brain during
nitrous oxide and phenobarbirone anesthesia. Brain Res
157:303-310, 1978
7. Cheng CH, Wooten GF: Dopamine turnover estimated by
simultaneous LCEC assay of dopamine and dopamine metabolites. J Pharmacol Methods 8:123-133, 1982
8. Edgar AD, Strosznajder J, Horrocks LA: Activation of
ethanolamine phospholipase Al in brain during ischemia. J
Neurochem 39:1111-1116, 1982
9. Edvinsson L, Lindvall M, Nielsen K-C, Owman CH: Are brain
vessels innervated also by central (non-sympathetic) adrenergic
neurones? Brain Res 63:496-499, 1973
10. Gardiner hl, Nilsson B, Rehncrona S, Siesjo BK: Free fatty
acids in the rat brain in moderate and severe hypoxia. J
Neurochem 36:1500-1505, 1981
11. Gaudec RJ, Levine L Transient cerebral ischemia and brain
prostaglandins. Biochem Biophys Res Commun 862393-901,
1979
12. Gaudet R, Welch KMA, Chabi E, Want T-P: Effect of transient
ischemia on monoamine levels in the cerebral cortex of gerbils.
J Neurochem 30:751-757, 1978
13. Goldberg ND, OToole AG: ’The properties of glycogen synthetase and regulation of glycogen biosynthesis in rat brain. J
Biol Chem 244:3053-3061, 1’369
14. Harik SI, Busto R, Marrinez E: Norepinephrine regulation of
cerebral glycogen utilization during seizures and ischemia. J
Neurosci 2:409-414, 1982
15. Harik SI, Duckrow RB, LaMannaJC, et al: Cerebral compensation for chronic noradrenergic denervation induced by locus
ceruleus lesion: recovery of receptor binding, isoproterenolinduced adenylate cyclase activity, and oxidative metabolism. J
Neurosci 1:64 1-649, 1981
16. Harik SI, LaManna JC, Light AI, Rosenthal M: Cerebral norepinephrine: influence on cortical oxidative metabolism in situ.
Science 20669-71, 1979
17. Kobayashi M, Lust WD, Passonneau JV: Concentrations of energy metabolites and cyclic nucleotides during and after bilateral
ischemia in the gerbil cortex. J Neurochem 29:53-59, 1977
18. Kogure K, Scheinberg P, Marsunloto A, et al: Catecholamines
in experimental brain ischemia. Arch Neurol 32:2 1-24, 1975
19. LaManna JC, Harik SI, Light AI, Rosenthal M: Norepinephrine
336 Annals of Neurology
Vol 18 No 3
depletion alters cerebral oxidative metabolism in the “active
state.” Brain Res 204237-101, 1981
20. Lavyne MH, Moskowitz MA, Larin F, er al: Brain H3catecholamine metabolism in experimental cerebral ischemia.
Neurology (Minneap) 25:483-485, 1975
21. Ljunggren B, Schutz H, Siesjo BK: Changes in energy state and
acid-base parameters of the rat brain during complete compression ischemia. Brain Res 73:277-289, 1974
22. Lowry OH, Passonneau JV: A Flexible System of Enzymatic
Analysis. New York, Academic, 1972
23. Lust WD, Arai H , Yasumoto Y, et al: Ischemic encephalopathy.
In McCandless DW (ed): Cerebral Energy Metabolism and Metabolic Encephalopathy. New York, Plenum, 1985, pp 79-1 17
24. Lust WD, Mrsulja BB, Mrsulja BJ, et al: Putative neurotransmitters and cyclic nucleotides in prolonged ischemia of the cerebral cortex. Brain Res 98:394-399, 1975
25. Lust WD, Passonneau JV: Cyclic nucleotides in murine brain:
effect of hypothermia on adenosine 3’,5’ monophosphate, glycogen phosphorylase, glycogen synthase and N e metabolites
following maximal electroshock or decapitation. J Neurochem
26111-16, 1976
26. Moskowitz MA, Kiwak KJ, Hekimian K, Levine L Synthesis of
compounds with properties of leukotrienes C4 and D4 in gerbil
brain after ischemia and reperfusion. Science 224:886-889,
1984
27. Passonneau JV, Brunner EA, Molstad C, Passonneau R: The
effects of altered endocrine state and of ether anaesthesia on
mouse brain. J Neurochem 18:2317-2328, 1971
28. Passonneau JV, Lauderdale VR: A comparison of three
methods of glycogen measurement in tissues. Anal Biochem
60:405-412, 1974
29. Ponten U, Ratcheson RA, Salford LG, Siesjo BK: Optimal
freezing conditions for cerebral metabolites in rats. J
Neurochem 2 1:I 127-1 138, 1973
30. Pulsinelli WA, Duffy TE: Regional energy balance in rat brain
after transient forebrain ischemia.J Neurochem 40: 1500- 1503,
1983
31. h c h l e ME, Hartman BK, Eichling JO, Sharpe LG: Central
noradrenergic regulation of cerebral blood flow and vascular
permeability. Proc Natl Acad Sci USA 72:3726-3730, 1975
32. Rehncrona S, Westerberg E, Akesson B, Siesjo BK: Brain cortical fatty acids and phospholipids during and following complete
and severe incomplete ischemia. J Neurochem 38234-93, 1982
33. Siesjo B K Brain Energy Metabolism. New York, Wiley, 1978,
pp 453-526
34. Sun GY: Metabolism of arachidonate and stearate injected simultaneously into the mouse brain. Lipids 12:661-665, 1977
35. Sun GY, Su KL, Der OM, Tang W: Enzymic regulation of
arachidonate metabolism in brain membrane phosphoglycerides. Lipids 14:229-235, 1979
36. Welsh FA, Ginsberg MD, Rieder W, Budd WW: Diffuse cerebral ischemia in the cat. 11. Regional metabolites during severe
ischemia and recirculation. Ann Neurol 3:493-501, 1978
37. Yoshida S, Abe K, Busto R, et al: Influence of transient ischemia on lipid-soluble antioxidants, free fatty acids and energy
metabolites in rat brain. Brain Res 245:307-3 16, 1982
38. Yoshida S, Harik SI, Busto R, et al: Free fatty acids and energy
metabolites in ischemic cerebral cortex with noradrenergic depletion. J Neurochem 425’1 1-717, 1984
39. Yoshida S, Inoh S, Asano T, et al: Effect of transient ischemia
on free fatty acids and phospholipids in the gerbil brain: lipid
peroxidation as a possible cause of postischemic injury. J
Neurosurg 5 3323-33 1, 1980
40. Yoshida S, Inoh S, Asano T, et al: Brain free fatty acids, edema,
and mortality in gerbils subjected to transient, bilateral ischemia, and effect of barbiturate anesthesia. J Neurochem 40:
1278-1286, 1983
September 1985
Документ
Категория
Без категории
Просмотров
0
Размер файла
853 Кб
Теги
ischemia, recovery, cerebal, enhance, norepinephrine, brain, depletion
1/--страниц
Пожаловаться на содержимое документа