Summary: The aim of this study was to examine the development of cortical epileptic afterdischarges (ADs) in genetic absence epilepsy WAG/Rij rats, and to compare them with two strains with minimal incidence of spike-and-wave (SW) episodes (ACI and Wistar). Epileptic ADs were elicited by stimulation of sensorimotor cortex in 12-, 18-, and 25-day-old rats of the three strains. The threshold current intensities were established for movements accompanying stimulation, for ADs of the SW type and accompanying clonic seizures and for transition into limbic type of ADs (characterized by behavioral automatisms). Individual groups were formed by 7-12 rats. There were no differences among the three strains in the thresholds for elicitation of stimulation-bound movements. In contrast, WAG/Rij and ACI rats exhibited easier elicitation of SW ADs than Wistar rats at the age of 18 and 25 days. There was no difference among the three strains in transition into the limbic type of ADs in 18-and 25-day-old rats. Lower thresholds for SW ADs in 18-and 25-day-old WAG/Rij and ACI rats in comparison with Wistar rats are in agreement with our data from adult animals as well as with development of pharmacologically induced models of absence seizures. The failure to find a specific difference between WAG/Rij rats and the other two strains might indicate a difference in generation of SW episodes and SW cortical AD. Key Words: Absence seizures-Epileptic afterdischargesCerebral cortex-Electrical stimulation-Ontogeny-Rat. Human absences represent an age-bound type of seizures characterized by short loss of consciousness and spike-and-wave (SW) rhythm in the EEG; they appear in late preschool-and school-age children (Stefan and Snead, 1997). There are two genetic models of absence epilepsy in rats characterized by spontaneous appearance of episodes of SW rhythm-GAERS and WAG/Rij (for revie

M.D Pavel Mareš

Pavel Mareš

†elena Tolmacheva

CiteSeerX

Epilepsia, 48(Suppl. 5):164–167, 2007
Blackwell Publishing, Inc.
C© International League Against Epilepsy
Comparison of Cortical Epileptic Afterdischarges in Immature
Genetic Absence Epilepsy WAG/Rij Rats with Those in Two Other
Strains (ACI and Wistar)
∗Pavel Mareš and ∗†Elena Tolmacheva
∗Institute of Physiology, Academy of Sciences, Prague, Czech Republic; and †Biological Faculty, Lomonosov State University,
Moscow, Russia
Summary: The aim of this study was to examine the devel-
opment of cortical epileptic afterdischarges (ADs) in genetic
absence epilepsy WAG/Rij rats, and to compare them with two
strains with minimal incidence of spike-and-wave (SW) episodes
(ACI and Wistar). Epileptic ADs were elicited by stimulation
of sensorimotor cortex in 12-, 18-, and 25-day-old rats of the
three strains. The threshold current intensities were established
for movements accompanying stimulation, for ADs of the SW
type and accompanying clonic seizures and for transition into
limbic type of ADs (characterized by behavioral automatisms).
Individual groups were formed by 7–12 rats. There were no dif-
ferences among the three strains in the thresholds for elicitation
of stimulation-bound movements. In contrast, WAG/Rij and ACI
rats exhibited easier elicitation of SW ADs than Wistar rats at
the age of 18 and 25 days. There was no difference among the
three strains in transition into the limbic type of ADs in 18-
and 25-day-old rats. Lower thresholds for SW ADs in 18- and
25-day-old WAG/Rij and ACI rats in comparison with Wistar
rats are in agreement with our data from adult animals as well
as with development of pharmacologically induced models of
absence seizures. The failure to find a specific difference be-
tween WAG/Rij rats and the other two strains might indicate a
difference in generation of SW episodes and SW cortical AD.
Key Words: Absence seizures—Epileptic afterdischarges—
Cerebral cortex—Electrical stimulation—Ontogeny—Rat.
Human absences represent an age-bound type of
seizures characterized by short loss of consciousness and
spike-and-wave (SW) rhythm in the EEG; they appear
in late preschool- and school-age children (Stefan and
Snead, 1997). There are two genetic models of absence
epilepsy in rats characterized by spontaneous appearance
of episodes of SW rhythm—GAERS and WAG/Rij (for
review Depaulis and van Luijtelaar, 2006). It was found
that these episodes start in restricted part of the cere-
bral cortex of WAG/Rij rats (Meeren et al., 2002) sup-
porting thus a primary role of neocortex in generation of
the SW rhythm (van Luijtelaar and Sitnikova, 2006). In
agreement with this finding is easier formation of GABA-
withdrawal cortical focus in GAERS than in outbred Wis-
tar rats (Brailowski et al., 1999) as well as a recent demon-
stration of higher excitability of slices of somatosensory
cortex from WAG/Rij rats in comparison with cortex of
Wistar animals (D’Arcangelo et al., 2006). SW episodes
are found not only in the two strains mentioned above,
Address correspondence and reprint requests to Pavel Mareš, M.D.,
D.Sc., Dept. Developmental Epileptology, Institute of Physiology,
Academy of Sciences of the Czech Republic, Videnska 1083, CZ-14220
Prague 4, Czech Republic. E-mail: maresp@biomed.cas.cz
doi: 10.1111/j.1528-1167.2007.01305.x
but also in other inbred (including ACI rats commonly
used as controls in studies of WAG/Rij rats—Inoue et al.,
1990) and outbred (Wistar) strain (Chocholová, 1983),
the differences are only quantitative. Using cortically in-
duced epileptic afterdischarges (AD) we found an easier
induction of SW (ADs), that is, an increased excitability
of sensorimotor cortex not only in adult WAG/Rij but also
in ACI strain in comparison with Wistar rats (Tolmacheva
et al., 2004). On the other hand WAG/Rij rats needed lower
stimulation current intensities than the other two strains
to spread the epileptic activity into the limbic structures
(Tolmacheva et al., 2004).
There is a contradiction between the ontogeny of SW
episodes (the only developmental study performed in
GAERS was able to register SW episodes since the age of
40 days (Vergnes et al., 1986) and pharmacological mod-
els of absence seizures demonstrating SW rhythm since
the third postnatal week in Wistar rats (Schickerová et al.,
1984; Snead, 1994). Therefore we decided to study de-
velopment of cortical excitability of the three rat strains
(WAG/Rij, ACI, and Wistar) by means of epileptic ADs
elicited by stimulation of the sensorimotor cortex and
compare the results with data from adult animals as well
as with data on development of SW activity. We expected
164
CORTICAL SEIZURES IN IMMATURE RATS 165
FIG. 1. Original recordings of afterdischarges in 18-day-old rats from Wistar, WAG/Rij and ACI strains (from left to right). Individual leads
in each section demonstrate activity of the left frontal area. Recordings demonstrate the last second of stimulation (an arrow marks the
end of stimulation) and 39 s after stimulation. Individual rows show increasing stimulation intensities—from top to bottom 0.2, 0.4, 0.6,
0.8, 1, 1.4, 1.8, 2.2, 2.6, 3, 3.5, 4, 5, 6, and 8 mA. Time mark 5 s, amplitude calibration 5 mV.
a marked increase of cortical excitability between the 12th
and 18th postnatal day in agreement with the appearance
of spike-and wave EEG rhythm elicited by convulsant
drugs (Schickerová et al., 1984; Snead, 1994; Zouhar et al.,
1989) or by thalamic stimulation (Mareš et al., 1982). This
expected result could indicate different mechanisms gen-
erating SW rhythm in acute models and in genetic models
of absence seizures.
METHODS
The experiments were approved by the Animal Care
and Use Committee of the Institute of Physiology to be in
agreement with European Community Council directives
86/609/EEC.
Three age groups (12, 18, and 25 days old) of the
three rat strains (WAG/Rij, ACI and Wistar) were studied.
Flat cortical silver electrodes (<1×1 mm) were implanted
epidurally under ether anesthesia. Stimulation electrodes
were localized over right sensorimotor area (AP −1 and
+1, L 2 mm), registration electrodes over left sensorimo-
tor, parietal, and visual areas and over right visual area.
Reference electrode was in nasal bone, ground in the oc-
cipital bone. Electrodes were fixed with a fast curing dental
acrylic. Surgical preparation lasted 10–12 min, then ether
anesthesia was interrupted and the animals were allowed
to recover for at least 1 h. Their body temperature was
maintained by means of a pad heated to 34◦C, that is, to
the temperature in the nest.
EEG activity was digitalized at a rate of 500 Hz and
saved on the hard disc (Kaminskij Biomedical Research
Systems, Prague). Behavior of animals was coded directly
into the recording. Epileptic ADs were elicited by stim-
ulation with 15-s series of 1-ms biphasic pulses at 8-Hz
frequency. Interval between the subsequent stimulations
was at least 10 min, current intensity was increased from
0.2 to 15 mA. Individual age groups were formed by 7–12
rats with equal number of male and female rat pups (with
the exception of one group with N = 7). No significant
differences between the two sexes were found in any strain
and age groups, therefore the results were put together.
Threshold intensities of stimulation current were esti-
mated for movements directly elicited by stimulation, for
ADs of the SW type and their behavioral correlate (clonic
seizures) and for transition into the second, limbic type
of ADs accompanied by behavioral automatisms (mixed
type, for details (Mareš et al., 2002; Mareš and Kubová,
2006). Duration of all ADs was measured and average val-
ues for individual stimulation intensities were calculated.
ANOVA with subsequent pairwise comparison by Tukey’s
test (SigmaStat, SPSS Inc., Chicago, IL) was used for sta-
tistical evaluation. The level of statistical significance was
set at 5%.
RESULTS
Movements bound to stimulation, SW ADs, and clonic
seizures were elicited in all animals. Transition into the
limbic type of ADs was observed in majority but not in all
rats. There were no differences in EEG pattern of ADs as
well as in frequency of sharp graphoelements among the
three strains (Fig. 1); it was described in detail in Wistar
rats (Mareš et al., 2002).
The three strains exhibited different developmental pro-
files. Developmental differences in Wistar rats did not
reach the level of statistical significance whereas 12-day-
old WAG/Rij pups exhibited significantly lower threshold
for elicitation of stimulation-bound movements than the
Epilepsia, Vol. 48, Suppl. 5, 2007
166 P. MAREŠ AND E. TOLMACHEVA
two older groups. Limbic type of ADs was more easily
elicited in 25-day-old than in 18-day-old rat pups in this
strain. Movements bound to stimulation were most easily
elicited in 18-day-old ACI animals whereas SW type of
ADs as well as clonic seizures needed higher stimulation
intensities in 12-day-old than in both 18- and 25-day-old
rats. Surprisingly, transition to limbic type of ADs ap-
peared in 12-day-old ACI rats at significantly lower inten-
sities than in the two older groups but this type of seizures
was elicited only in four animals.
If the three strains were compared (Fig. 2) 12-day-old
rats exhibited a significant difference only in thresholds
for limbic type of ADs; it was lower in ACI rats that in
the two other strains. Eighteen- and 25-day-old WAG/Rij
and ACI rats needed lower intensities to elicit SW type of
ADs and accompanying clonic seizures than Wistar rats.
Twelve-day-old animals of all three strains generated
longer ADs than older rats. The difference reached the
level of statistical significance only with medium stimu-
FIG. 2. Threshold intensities (mean + S.E.M.) for elicitation of
stimulation-bound movements, spike-and-wave afterdischarges,
clonic seizures, and limbic (mixed) type of afterdischarges (see
inset) in 12-, 18-, and 25-day-old rats (from top to bottom). Ab-
scissae from left to right: Wistar, WAG/Rij and ACI rats; ordinates:
intensity of stimulation current in mA. Asterisks denote statisti-
cally significant difference in comparison with corresponding phe-
nomenon in the other two strains.
FIG. 3. Duration of afterdischarges (mean + S.E.M.) in 12-, 18-,
and 25-day-old rats (from top to bottom). Individual columns—
afterdischarges elicited by stimulation intensity of 1, 2, 3, 4, or
5 mA (see inset). Abscissae from left to right: Wistar, WAG/Rij,
and ACI rats; ordinates: duration of afterdischarges in seconds.
Circles mark a significant difference between Wistar and WAG/Rij
rats, asterisks denote a difference between WAG/Rij and ACI rats,
double crosses a difference between ACI and Wistar rats.
lation intensities in Wistar rats whereas it was significant
with nearly all current intensities in the two inbred strains
(Fig. 3). An enormous increase of duration of ADs with in-
creasing intensities of stimulation observed in 12-day-old
ACI rats (Fig. 3, uppermost part) was due to a transition
to the limbic type of ADs at relatively low current inten-
sities. Systematic significant differences between 18- and
25-day-old rats were found only in ACI rats—ADs in the
younger group were longer than those in 25-day-old rats.
Comparison of ADs duration among the three strains
did not show consistent changes. It demonstrated longer
ADs in 12-day-old ACI rats than in the two other strains,
some shorter ADs in 18-day-old WAG/Rij rats in com-
parison with ACI and Wistar rats and a few differences
between 25-day-old Wistar and ACI rats (Fig. 3).
DISCUSSION
The difference in thresholds for SW and clonic seizures
at the age 18 and 25 days was the same as in adult
Epilepsia, Vol. 48, Suppl. 5, 2007
CORTICAL SEIZURES IN IMMATURE RATS 167
animals—Wistar rats needed higher stimulation current
intensities than WAG/Rij and ACI rats. The difference in
cortical excitability between WAG/Rij and Wistar rats is
in agreement also with data of D’Arcangelo et al. (2006).
Unfortunately, these authors did not study ACI rats. No
similar differences among the three strains were observed
in 12-day-old pups—at the age when SW EEG rhythm
cannot be elicited (Mareš et al., 1982; Schickerová et al.,
1984; Snead, 1994). The only difference between adult
WAG/Rij rats and two other strains (an easier transition to
the limbic type of ADs) did not appear in our age groups.
This transition matures during the fourth postnatal week
in Wistar rats (Mareš et al., 1982) and therefore the dif-
ference may appear later in life.
Failure of changes specific for WAG/Rij rats could
be due to the fact that SW episodes are present in all
three strains and they differ only quantitatively (Inoue
et al., 1990; Chocholová, 1983). Localization of stimu-
lation electrodes might also play a role—they were local-
ized more medially than the locus found by Meeren et al.
(2002), but we used relatively large electrodes (surface in
contact with dura mater was nearly 1×1 mm) so that the
stimulated field is broad and at least at high stimulation
intensities involves the locus of origin of SW episodes.
The possible effect of stimulated area could be excluded
only with cortical microstimulation. Nevertheless our re-
sults raise again an old question if an increased cortical ex-
citability is the main change in generation of SW episodes.
Higher excitability of cerebral cortex found in 18- and
25-day-old WAG/Rij and ACI pups in comparison with
Wistar rats is in accordance with findings in pentetrazol-
and gama-hydroxybutyrate-induced models of absences
(Schickerová et al., 1984; Snead, 1994) as well as with
electrically induced SW rhythm—it was recorded since
the third postnatal week (Mareš, et al., 1982, 2002; Mareš
and Kubová, 2006). This result is in contradiction with
finding that SW episodes in the other strain with sponta-
neous absence seizures (GAERS) were recorded since the
age of 40 days (Vergnes et al., 1986). Identical develop-
ment of SW ADs in WAG/Rij and ACI rats indicate that
generation of cortical epileptic ADs differs from gener-
ation of SW episodes. It is supported by a difference in
frequency of SW activity induced by cortical stimulation
and SW episodes (Mareš et al., 1982). Further analysis
of this possibility including detailed ontogenetic study of
spontaneous EEG activity of WAG/Rij rats is necessary.
Acknowledgment: This study was supported by a grant
No. 305/05/2581 of the Grant Agency of the Czech Republic
and by a research project AV0Z 50110509.
REFERENCES
Brailowski S, Montiel T, Boehrer A, Marescaux C, Vergnes M. (1999)
Susceptibility to focal and generalized seizures in Wistar rats with
genetic absence-like epilepsy. Neuroscience 93:1173–1177.
Chocholová L. (1983) Incidence and development of rhythmic episodic
activity in the electroencephalogram of a large rat population under
chronic conditions. Physiologia Bohemoslovaca 32:10–18.
D’Arcangelo G, D’Antuono MD, Tancredi V, Avoli M. (2006) Neocor-
tical hyperexcitability in a genetic model of absence seizures and its
reduction by levetiracetam. Epilepsia 47:1144–1152.
Depaulis A, van Luijtelaar G. (2006) Genetic models of absence epilepsy
in the rat. In Pitkänen A, Schwartkroin PA, Moshé SL (Eds) Models
of Seizures and Epilepsy. Elsevier, Amsterdam, pp. 233–248.
Inoue M, Peters BW, van Luijtelaar EL, Vossen JM, Coenen AM. (1990)
Spontaneous occurrence of spike-wave discharges in five inbred
strains of rats. Physiology and Behavior 48:199–201.
Mareš P, Kubová H. (2006) Electrical stimulation-induced models of
seizures. In Pitkänen A, Schwartkroin PA, Moshé SL (Eds) Models
of Seizures and Epilepsy. Elsevier, Amsterdam, pp. 153–159.
Mareš P, Marešová D, Trojan S, Fischer J. (1982) Ontogenetic devel-
opment of rhythmic thalamo-cortical phenomena in the rat. Brain
Research Bulletin 8:765–769.
Mareš P, Haugvicová R, Kubová H. (2002) Unequal development of
thresholds for various phenomena induced by cortical stimulation in
rats. Epilepsy Research 49:35–43.
Meeren HK, Pijn JP, van Luijtelaar EL, Coenen AM, Lopes da Silva
FH. (2002) Cortical focus drives widespread corticothalamic net-
works during spontaneous absence seizures in rats. Journal of Neu-
rosciences 22:1480–1495.
Schickerová R, Mareš P, Trojan S. (1984) Correlation between electro-
corticographic and motor phenomena induced by metrazol during
ontogenesis in rats. Experimental Neurology 84:153–164.
Snead OC III. (1994) The ontogeny of [3H]gamma-hydroxybutyrate and
[3H]GABAB binding sites: relation to the development of experi-
mental absence seizures. Brain Research 659:147–156.
Stefan H, Snead OC III. (1997) Absence seizures. In Engel J, Jr, Pedley
TA (Eds) Epilepsy. A Comprehensive Textbook. Lippincott – Raven,
Philadelphia, pp. 579–590.
Tolmacheva E, van Luijtelaar G, Chepurnov SA, Kaminskij Yu, Mareš P.
(2004) Cortical and limbic excitability in rats with absence epilepsy.
Epilepsy Research 62:189–198.
van Luijtelaar G, Sitnikova E. (2006) Global and focal aspects of ab-
sence epilepsy. The contribution of genetic models. Neuroscience
and Biobehavioral Reviews 30:983–1003.
Vergnes M, Marescaux C, Depaulis A, Micheletti G, Warter JM. (1986)
Ontogeny of spontaneous petit mal-like seizures in Wistar rats. De-
velopmental Brain Reearch 30:85–87.
Zouhar A, Mareš P, Lišková-Bernášková K, Mudrochová M. (1989)
Motor and electrocorticographic epileptic activity induced by bicu-
culline in developing rats. Epilepsia 30:501–510.
Epilepsia, Vol. 48, Suppl. 5, 2007


Comparison of Cortical Epileptic Afterdischarges in Immature Genetic Absence Epilepsy WAG/Rij Rats with Those in Two Other Strains (ACI and Wistar)

http://citeseerx.ist.psu.edu/viewdoc/download;jsessionid=B926063E17FCAD819FE81D8F77AF43DA?doi=10.1.1.1071.2055&rep=rep1&type=pdf

Comparison of Cortical Epileptic Afterdischarges in Immature Genetic Absence Epilepsy WAG/Rij Rats with Those in Two Other Strains (ACI and Wistar)

Abstract

Similar works

Full text

Available Versions

CiteSeerX