1. FEDERAL UNIVERSITY OF CEARá / BRAZIL FACULTY OF MEDICINE
DEPARTMENt of PHYSIOLOGY and pharmacology
Laboratory of Neuropsychopharmacology
INTRACEREBROVENTRICULAR OUABAIN-INDUCED SEIZURES: A MODEL OF SECONDARILY GENERALIZED STATUS EPILEPTICUS
LUSTOSA, Í R*; BORGES, L T N; BARBOSA T M; HOLANDA, F T G; DOS SANTOS, R C; DOS SANTOS JR, M A; VASCONCELOS, G S; LIMA, C N C; XIMENES, N C; MEDEIROS, I S; JUCÁ, M M; VASCONCELOS, S M**
except for two major differences: 1) ouabain P1 is consistently heralded by seizures of 1-2 Racine´s stages; 2) ouabain seizures evolves as a clonic status epilepticus (SE), not as an ictal depression with recover after 15 min as in audiogenic models.
INTRODUCTION
The 2nd antiepileptic drugs developed in the last two decades failed to modify the prevalence of pharmacoresistant epilepsies (33-40%), and there is a growing concern over models that may be useful in pharmacoresistance studies in addition to the predictive validity.
International League Against Epilepsy (ILAE) recommended establish anatomic substrates and pathophysiologic mechanisms that allow set different seizure types as discrete entities more than focuse on a purely desccriptive phenomenology.
We have induced seizures in rats through ouabain intracerebroventricular (i.c.v.) infusion and have characterized it behaviorally.
Behavior
Incidence
(n - %)
Latency (L)
Duration (D)
Time (s)
(mean ± SEM)
p (SW) P1 % L
79,33 ± 8,243 * D
14,33 ±1,818 F
106,1 ± 44,25 P2
7 - 77,77%
169,6 ± 55,38
>0,05
17,00 ± 2,545
PCS
4 - 44,44%
277,8 ± 77,56
LRR
491,2 ± 55,56
1CTGmáx
6 - 66,66%
2117 ± 166,3
5 - 55,55%
2755 ± 83,77
OBJECTIVE
To characterize behaviorally the convulsive seizures regarding the localizing semiology.
Table1. Incidence, latency (L) and duration (D) times of i.c.v. ouabain-induced behavioral seizures. 1st procursive seizure focal, hindbrain (P1); 2nd procursive seizure focal, hindbrain (P2); freezing (F); partial clonic seizure extralimbic forebrain (includes clonic seizure stage 3 of Racine); loss of righting reflex (LRR); 1st generalized clonic-tonic seizure generalized, hindbrain stage 8 of Jobe (1GCTmáx); death (D); standard error of the menan (SEM); p<0,05; Shapiro-Wilk goodness-of-fit test (SW); sample size: n=9 animals; time of analysis: 1 h.
MATERIALS AND METHOD
Surgery: adult male Wistar rats ( g of body weight, n=22) anesthetized with ketamine + xylazine ( mg/kg i.p, respectively). AP -0.9; ML -1.8; DV -2.5.
Behavior
Figure 1. Ethogram of i.c.v. ouabain-induced seizures. Non convulsoive (NC)
CONCLUSION
Adapted from: Remie, 2000
The i.c.v. ouabain-induced seizures may not serve to high throughput screening of substances with antiepileptic activity due to it is labor-intensive protocol.
Notwithstanding it may serve to study mechanisms of ictogenesis and neuronal network synchronization that in turn can lead to a severe status epilepticus
Induction: Between 5th and 7th postoperative days. Internal cannula 30 G (protruding 1 mm beyond guide cannula) loaded with ouabain (10-2 M, 5µL in artificial cerebrospinal fluid (ACSF) or ACSF 5µL alone (control group)) and fitted into the guide. Animals were posed at the center of an open filed and acclimated during 5 min. Injection performed throughout 1 min. Cannula was left in place for 1 additional min to avoid back flow. Digital video record throughout 1 h. Video analysis: offline. Latency and duration times in seconds were registered. Descriptive statistics. Ethogram.
REFERENCES
KOKARE, D. M. et al. A simple and inexpensive method to fabricate a cannula system for intracranial injections in rats and mice. Journal of Pharmacological and Toxicological Methods, v. 64, n. 3, p. 246–250, 2011.
FERRY, B.; GERVASONI, D.; VOGT, C. Stereotaxic Neurosurgery in Laboratory Rodent - Handbook on best practices. 1a ed. Paris: Springer, 2014.
PAXINOS, G.; WATSON, C. The Rat Brain in Stereotaxic Coordinates. 5a ed. San Diego: Elsevier Academic Press, 2005.
RIEGEL, R. E. et al. Animal model of mania induced by ouabain: Evidence of oxidative stress in submitochondrial particles of the rat brain. Neurochemistry International, v. 55, n. 7, p. 491–495, 2009.
EL-MALLAKH, R. S. et al. Efficacy of olanzapine and haloperidol in an animal model of mania. Progress in Neuro-Psychopharmacology and Biological Psychiatry, v. 30, n. 7, p. 1261–1264, 2006.
VINOGRADOVA, L. V; GRINENKO, O. A. Ictal electrographic pattern of focal subcortical seizures induced by sound in rats. Brain Research, v. 1635, p. 161–168, 2016.
From: Kokare et a., 2011
RESULTS AND DISCUSSION
Circa 85% of the animals receiving µg of i.c.v. ouabain exhibit a behavior consisting of one or two violent bursts of running or jumping (procursive seizures, P1, P2) intercalated by a period of freezing (F) evolving as generalized clonic SE with late tonic phenomena.
This pattern strikingly seems that of a full blown audiogenic seizure in susceptible strains (WAR, GEPR-9, DBA/2, GASH:Sal), which in turn is attributed to a focal (inferior colliculus, IC) hindbrain ictal initiation (procursive) followed by secondary generalization (clonic)
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