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Increased long-term potentiation in the surround of experimentally induced focal cortical infarction.

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Increased Long-Term
Potentiation in the Surround
of Experimentally Induced
Focal -Cortical Infarction
Georg Hagemann, MD, Christoph Redecker, MD,
Tobias Neumann-Haefelin, MD,
Hans-Joachim Freund, MD, and Otto W. Witte, M D
in remote areas including the contralateral hemiW i t h i n these brain areas, y-aminobutyric
acid (GABA)ergic inhibition is decreased, neuronal activity enhanced,'.' and GABA-receptor activity downregulated with a change of GABA-receptor subunit
c o m p o ~ i t i o n .Taken
together, these observations indicate that cortical lesions induce a reactive plasticity in
morphologically undamaged brain regions, thereby facilitating cortical reorganization. ,3,4,10
To add support to this hypothesis, we investigated
whether the induction of long-term potentiation
(LTP), which has been associated with neuronal plasticity, learning, and memory processes,'
is facilitated i n the surround of a n d contralateral to ischemic
cortical lesions.
Functional recovery after stroke is partly due to cortical
reorganization on a structural as well as a functional
level. Recent investigations have shown that the excitability of brain areas surrounding cortical ischemic lesions is increased, probably due to a down-regulation of
y-aminobutyric acid-receptor activity. There is some
evidence that these changes might increase the susceptibility of the lesioned brain for adaptive changes and recovery. Here, we investigated the propensity for the induction of long-term potentiation (LTP) in the surround
of experimentally induced focal cortical infarcts in rat somatosensory cortex in vitro. By using standard paradigms, LTP induction was found to be facilitated ipsilaterally in slices of lesioned animals 1 week after lesion
induction. In homotopic contralateral areas, LTP was not
different from control values. As LTP is commonly associated with plasticity and learning, the results provide
further evidence for the lesion-induced amplification of
network plasticity, as it is required for the reshaping of
cortical circuits by timely training procedures.
Hagemann G, Redecker C, Neumann-Haefelin T ,
Freund H-J, Witte OW. Increased long-term
potentiation in the surround of experimentally
induced focal cortical infarction.
Ann Neurol 1998;44:255-258
Focal acute brain damage is often followed by functional cortical reorganization associated with behavioral
recovery.' This process can be substantially modified
by rehabilitative training and involves both cortex adjacent to t h e lesion as well as contralateral areas.'-' A t
the structural level, training after focal brain damage
induces pronounced synaptogenesis a n d sprouting of
neuronal dendrites ipsilateral as well as contralateral to
the lesion.*-* Functionally, a n increase in excitability
was found in brain regions close to the lesion a n d also
From the Department of Neurology, Heinrich-Heine-University,
Diisseldorf, Germany.
Received Dec 2, 1997, and in revised form Feb 20, 1998. Accepted
for publication Feb 21, 1998.
Address correspondence to D r Witte, Department of Neurology,
Heinrich-Heine-University, Moorenstr 5 , 40225 Duesseldorf,
Subjects and Methods
The experiments were performed by using adult male Wistar
SPF rats (290-310 g, n = 32). In 18 of these rats, a cortical
photothrombotic lesion was induced. l 3 Under enflurane anesthesia (1.5-2.5% in N,O/O,), a catheter was inserted into
the femoral vein, the skull was exposed, and a fiber-optic
bundle (aperture, 1.5 mm) was positioned 4 mm posterior to
bregma and 4 mm lateral to midline. For 20 minutes, the
brain was illuminated through the skull with a cold light
source. During the first minute, rose bengal (1.3 mg/100 g
of body weight) was injected. Rectal temperature was kept
constant at 37.0 2 0.2"C. After illumination, the wounds
were sutured and the animals were allowed to recover in a
warm environment with free access to food and water.
Electrophysiological recordings were performed by using
brain slices from unlesioned controls and from lesioned animals between 7 and 10 days after surgery. This time point
was chosen because of the most prominent changes of other
electrophysiological parameters as described earlier.' By using
conventional techniques, coronal neocortical slices (400 p m )
were obtained at the site of the lesion and at corresponding
levels in control animals. The slices were maintained in an
interface recording chamber at 33°C and superfused with artificial cerebrospinal fluid (ACSF; composition in mM: NaCl
124, NaHCO, 26, KCI 5, CaCI, 2, MgS04 2, N a H P 0 4
1.25, and glucose 10, equilibrated with carbogen to pH 7.4).
Slices were left undisturbed for at least 1 hour before starting
any recording.
Extracellular recordings of field potentials were performed
with glass micropipettes filled with ACSF, which were placed
in layer II/III of the sensory cortex (granular primary somatosensory cortex ~ a r l , ' * )approximately I to 2 mm lateral
to the lesion, in the homotopic contralateral area, or in corresponding control areas. In separate experiments, 5 mmollL
bicuculline methiodide (Sigma Chemicals, St Louis, MO)
was added to the solution in the recording pipette, to suppress GABA, response^.'^ Synaptic responses were evoked
with 50-psec pulses delivered through a concentric bipolar
stimulation electrode positioned in layer IV. LTP induction
was only started after a stable baseline, with field responses 2
to 4 mV in amplitude for longer than 20 minutes, was
achieved. The induction protocol for LTP consisted of a sequence of six theta bursts delivered at 0.1 Hz. Each burst
Copyright 0 1998 by the American Neurological Association
consisted of 10 sequences delivered at 5 Hz, and one sequence included five 200-psec pulses at 100 H Z . ' ~Data
were digitized, stored to disk, and analyzed off-line. Results
are given as mean % SEM of each time point before and
after theta-burst application. Statistical analysis was performed by using the Wilcoxon rank test, with p < 0.01 considered to be significant.
All animals that underwent surgery showed a clearly
demarcated cortical infarct that was easily localized in
the recording chamber. There was no overall difference
between field potentials of different groups of animals.
After pooling and normalizing results to baseline
level in slices of control animals, amplitude and slope
were potentiated by theta-burst application to 102.8 2
2.3% and 106.8 % 4.3%, respectively (9 animals, 18
slices) (Fig 1). Application of a theta burst ipsilaterd to
the lesion increased field potential amplitude to
109.3 t 1.8% and slope to 130.1 -t- 4.8% (10 animals, 14 slices; difference significant for lesioned vs
control), In the homotopic contralateral area, the same
protocol only increased field potential amplitude to
102.4 2 3.4% and slope to 113.7 2 5.4% (8 animals,
Fig 1. (4) Schematic diagram of a coronal neocortical slice
preparation. The granular cortex is represented by gray stippling in layer I K The lesion is indicated by an indentation.
The recording electrode (FP) was positioned in layer II/III and
the stimulating electrode (stim) was placed in layer I K For
clarity, both pairs of electrodes are depicted. (B) Average o f
Jive Jield potentials obtained in control and lesioned animals.
Field potentials before and a$er application of a theta burst
are superimposed.
5 ms
Annals of Neurology
Vol 44
No 2
August 1998
16 slices; difference in slope but not amplitude significant for lesioned vs control). The quantitative differences are depicted in Figure 2A. In control animals,
the first response after application of the theta burst
was diminished in slope as well as in amplitude; in
slices of lesioned animals, this decline could not be observed (see Fig 2).
As it is known that GABAergic inhibition is diminished in the surround of the lesion, we investigated
whether pharmacological reduction of GABAergic inhibitionI5 gives results similar to those of cortical lesioning. With this approach, amplitude was potentiated to 107.0 2 2.3% and slope to 118.7 t 8.5% (7
animals, 9 slices; significant differences for slope and
amplitude vs both, control and lesioned).
This study demonstrates that the induction of LTP is
facilitated in the surround of focal cortical infarcts in
vitro. LTP is a transmitter and second messengerdependent phenomenon that is believed to underly
complex processes such as learning and memory.
Transmitter release is potentiated and the postsynaptic
The increased synaptic efficacy
accounts for associativity and specificity, which are
hallmarks of LTP, resulting in neuronal plasticity." It
is therefore reasonable to assume that the facilitation
of LTP-induction after cortical lesioning reflects an
underlying mechanism of cortical reorganization and
The increased propensity for LTP in the surround of
ischemic cortical lesions is well compatible with the hyperexcitability and decrease of GABA-receptor activity
observed in widespread brain areas in the surround and
contralateral to such lesion^.^.^'^^' This explanation
gains support from experiments with pharmacological
reduction of GABAergic inhibition, which also facilitated LTP although to a somewhat lesser degree than
cortical lesioning. Part of the hyperexcitability is probably due to deafferentati~n.'~
Thus, if deep cortical
layers, which give rise to extended intracortical collaterals, are damaged, it affects widespread and remote
brain areas. Although in our model deep cortical layers
were affected in all animals, there was increased LTP
contralateral to the lesion concerning slope but not
Receptor autoradiographic studies have shown that,
in addition to the GABAergic system, other transmitter
systems are also altered in the surround of cortical les i o n ~ . ' ~It' ~is therefore possible that in addition to the
reduction of GABAergic inhibition, other factors also
contribute to the enhancement of LTP after cortical ischemia. An increment in N-methyl-D-aspartate
(NMDA)-receptor activity may have a direct impact on
LTP induction. Furthermore, the neuronal membrane
potential is less negative in these brain areas,8 causing a
ipsilateral to lesion
40 min
time after theta-burst
contralateralto lesion
40 min
time after theta-burst
Fig 2. Long-term efects of theta-burst stimulation on j e l d potential peak amplitudes (A,, B,) and slopes (A2, Bz). Graphs plot the
mean result -t SEM of all experiments. (A) Data obtained in slices of control animals (0)and ipsilateral to the lesion (a).
(B) Data of controls (0)and contralateral to the lesion (a). After 20 minutes of baseline recording, a theta burst was applied at
time point zero and data normalized to baseline level. The stippled line indicates 100%. In terms of slope as well as amplitude,
long-term potentiation was clearly facilitated ipsilateral to the lesion. There is short-term potentiation in slices contralateral to
the lesion (B2).
further increase of NMDA-dependent transmission as
the Mg2+ block is released.
The facilitation of LTP induction is likely to be an
essential mechanism of lesion-induced plasticity and
may be a prerequisite for functional re~titution.'~'~
Furthermore, this capacity may prove useful for additional restorative measures. Morphological changes associated with behaviorally beneficial outcome are most
pronounced2 weeks after lesioning. In how far such experimental data may clarify the presence of an optimal
Brief Communication: Hagemann et al: Lesion-Induced Plasticity
time window for therapeutic measures, such as input
manipulation and training, is presently unclear. However, early overuse of a lesion-affected extremity can
worsen functional outcome3,* and increase lesion size,
possibly due to excessive energy demands causing adFurther analysis of these inditional neuronal
teractions is necessary to provide better insights into
the implementation of therapeutic strategies in patients
with acute focal brain damage.
The investigations were supported by SFB 194 B2.
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Mitochondrial DNA
in Focal Dystonia:
A Cybrid Analysis
S. J. Tabrizi, MD, ChB,* J. M. Cooper, PhD,'
and A. H. V. Schapira, DSc, MD, FRCP*t
The cause and pathophysiology of dystonia remain unknown. The recent identification of mitochondrial complex I deficiency in platelets from patients with sporadic
focal dystonia suggests that a defect of energy metabolism
may be relevant in a proportion of patients. We have addressed the possible contribution of mitochondrial DNA
(mtDNA) to the complex I deficiency in dystonia by the
use of genome transfer technology. Platelets from patients deficient for complex I were fused with A549 p"
(mtDNA-less) cells to form cybrids comprising the A549
nucleus and dystonia mtDNA. Mixed cybrid cell lines
were analyzed for 9 controls and 9 dystonia patients,
and clonal cybrid lines were generated for 2 control
and 2 dystonia patients. Subsequent biochemical analysis
showed that the dystonia complex I defect was complemented in both the mixed and the clonal cybrid lines.
These results contrast with similar studies in mitochondrial myopathy and Parkinson's disease patients, in
which the mitochondrial defect was maintained in at
least a proportion of A549 cybrids, and suggest that the
complex I defect in dystonia is not caused by an mtDNA
Tabrizi SJ, Cooper JM, Schapira AHV.
Mitochondrial DNA in focal dystonia: a cybrid
analysis. Ann Neurol 1998;44:258-261
Idiopathic torsion dystonia (ITD) is the most common
form of primary dystonia and can be classified according to anatomical distribution into focal, segmental,
From the *University Department of Clinical Neurosciences, Royal
Free Hospital School of Medicine, and ?University Department of
Clinical Neurology, Institute of Neurology, London, UK.
Received Aug 22, 1997, and in revised form Feb 2, 1998. Accepted
for publication Feb 6, 1998.
Address correspondence to Prof Schapira, Clinical Neurosciences,
Royal Free Hospital School of Medicine, Rowland Hill Street, London N W 3 2PF, UK.
Copyright 0 1998 by the American Neurological Association
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