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The lithium–pilocarpine model mimics most features of human temporal lobe epilepsy. Following our prior studies of cerebral metabolic changes, here we explored the expression of transporters for glucose (GLUT1 and GLUT3) and monocarboxylates (MCT1 and MCT2) during and after status epilepticus (SE) induced by lithium–pilocarpine in PN10, PN21, and adult rats. In situ hybridization was used to study the expression of transporter mRNAs during the acute phase (1, 4, 12 and 24 h of SE), the latent phase, and the early and late chronic phases. During SE, GLUT1 expression was increased throughout the brain between 1 and 12 h of SE, more strongly in adult rats; GLUT3 increased only transiently, at 1 and 4 h of SE and mainly in PN10 rats; MCT1 was increased at all ages but 5-10-fold more in adult than in immature rats; MCT2 expression increased mainly in adult rats. At all ages, MCT1 and MCT2 up-regulation was limited to the circuit of seizures while GLUT1 and GLUT3 changes were more widespread. During the latent and chronic phases, the expression of nutrient transporters was normal in PN10 rats. In PN21 rats, GLUT1 was up-regulated in all brain regions. In contrast, in adult rats GLUT1 expression was down-regulated in the piriform cortex, hilus and CA1 as a result of extensive neuronal death. The changes in nutrient transporter expression reported here further support previous findings in other experimental models demonstrating rapid transcriptional responses to marked changes in cerebral energetic/glucose demand.

It is well known that the uncoupling between local cerebral glucose utilization (LCGU) and local cerebral blood flow (LCBF), i.e. decrease in LCBF rates with high LCGU, is frequently associated with seizure-induced neuronal damage. This study was performed to assess if the neuroprotective effect of the adenosinergic A1 receptor agonist R-N-phenylisopropyladenosine (R-Pia) injected prior to pilocarpine is able to reduce the uncoupling between LCGU and LCBF during status epilepticus (SE). Four groups of rats were studied: Saline, Pilo, R-Pia+Saline and R-Pia+Pilo. For LCGU and LCBF studies, rats were subjected to autoradiography using [14C]-2-deoxyglucose and [14C]-iodoantypirine, respectively. Radioligands were injected 4 h after SE onset. Neuronal loss was evaluated by Fluorojade-B (FJB) at two time points after SE onset (24 h and 7 days). The results showed a significant increase in LCGU in almost all brain regions studied in the Pilo and R-Pia+Pilo groups compared to controls. However, in R-Pia pretreated rats, the uncoupling between LCGU and LCBF was moderated in a limited number of structures as substantia nigra pars reticulata and hippocampal formation rather in favor of hyperperfusion. Significant increases in LCBF were observed in the entorhinal cortex, thalamic nuclei, mammillary body, red nucleus, zona incerta, pontine nucleus and visual cortex. The neuroprotective effect of R-Pia assessed by FJB showed a lower density of degenerating cells in the hippocampal formation, piriform cortex and basolateral amygdala. In conclusion our data shows that the neuroprotective effect of R-Pia was accompanied by a compensatory metabolic input in brain areas involved with seizures generation.

The susceptibility of rats with genetically inherited epilepsy to the genesis and consequences of secondary temporal lobe epilepsy is unknown. Here, we induced lithium-pilocarpine status epilepticus (SE) in Genetic Absence Epilepsy Rats from Strasbourg (GAERS) or in Wistar audiogenic sensitive (AS) rats. Wistar AS needed less pilocarpine than GAERS and Non-Epileptic Rats (NERs) to develop SE. Sixty six, 40 and 5% of Wistar AS, GAERS and NERs, respectively, died within 24 h after SE. In GAERS, SE prevented the occurrence of absence seizures for 5 days. Thereafter a limited number of absence seizures with low amplitude and short duration were recorded. Wistar AS developed limbic epilepsy within 9 days after SE while GAERS and NERs needed 36–39 days to develop spontaneous motor seizures. Neuronal loss consecutive to SE was similar in the three strains and particularly marked in limbic forebrain and parahippocampal cortices. In conclusion, the development of focal limbic epilepsy in GAERS largely impairs the expression of absence seizures. The genetic background underlying the expression of audiogenic seizures sensitizes strongly the rats to a further insult and compromises their survival.

Neuronal–astrocytic interactions in 1-month-old Genetic Absence Epilepsy Rats from Strasbourg (GAERS) before the occurrence of seizures are compared to those in non-epileptic rats (NERs) and in adult GAERS expressing epilepsy. Animals received [1-13C]glucose and [1,2-13C]acetate, preferential substrates of neurons and astrocytes, respectively, and extracts from cerebral cortex, subcortex and cerebellum were analyzed by NMR spectroscopy. Increased mitochondrial metabolism took place in the cortical neurons of immature and adult GAERS and therefore does not seem to be a consequence of the occurrence of absence seizures. Glutamine supply to GABAergic neurons was reduced in cortex and subcortex in young GAERS, as reflected by increased glutamine content and decreased 13C-labeling of GABA. In the brain of immature GAERS, interactions between glutamatergic neurons and astrocytes appeared normal whereas increased astrocytic metabolism took place in adult GAERS, suggesting that astrocytic alterations could possibly be the cause of seizures.

The role of endothelial nitric oxide (NO) in the cerebrovascular response to partial seizures was investigated in mice deleted for the endothelial NO synthase gene (eNOS−/−) and in their paired wild-type (WT) congeners. Local cerebral blood flow (LCBF, quantitative [14C]iodoantipyrine method) was measured 3–6 h after unilateral kainate (KA) injection in the dorsal hippocampus; controls received saline. In WT mice, KA seizures induced a 22 to 50% LCBF increase restricted to the ipsilateral hippocampus, while significant LCBF decreases (15–33%) were noticed in 22% of the contralateral areas, i.e., the parietal cortex, amygdala and three basal ganglia areas, compared to saline-injected WT mice. In eNOS−/− mice, no LCBF increases were recorded within the epileptic focus and generalized contralateral LCBF decreases (22–46%) were noticed in 2/3 of the brain areas, compared to saline-injected eNOS−/− mice. Thus, endothelial NO is the mediator of the cerebrovascular response within the epileptic focus and participates in the maintenance of LCBF in distant areas.

The role of neuronal nitric oxide (NO) in the cerebrovascular response to partial seizures induced by intrahippocampal injection of kainate (KA) was investigated in mice deleted for the neuronal NO synthase gene (nNOS−/−) and in wild-type controls (WT). A second group of WT mice received the nNOS inhibitor, 7-nitroindazole (WT-7NI). Local cerebral blood flow (LCBF) was measured using the quantitative 14C-iodoantipyrine method. Within the epileptic focus, all three groups of seizing mice (WT, WT-7NI, and nNOS−/−) showed significant 26–88% LCBF increases in ipsilateral hippocampus, compared to saline-injected mice. Contralaterally to the epileptic focus, KA seizures induced a 21–47% LCBF decreases in hippocampus and limbic cortex of WT mice and in most contralateral brain structures of nNOS−/− mice, while WT-7NI mice showed no contralateral CBF change. Neuronal NO appears to be not involved in the cerebrovascular response within the epileptic focus, but may rather have a role in the maintenance of distant LCBF regulation during seizures.

To better understand the role of inflammatory responses in temporal lobe epilepsy, we characterized Interleukin1-β (IL1-β), Nuclear Factor-κB (NF-κB), and Cyclooxygenase-2 (COX-2) expression together with neurodegeneration in the rat lithium–pilocarpine model. The immunohistochemical expression of IL1-β, NF-κB, and COX-2 started by 12 h post-injection, persisted for 24 h (status epilepticus period), and returned to basal levels by 3 and 6 days (latent period). The regional distribution of IL1-β, NF-κB, and COX-2 occurred mainly in structures prone to develop neuronal damage. Using double-staining protocols, we detected IL1-β expression in glial cells, COX-2 expression in neurons, and NF-κB in both cell types. The presence of Fluoro-Jade-B-positive degenerating neurons was associated with IL1-β, NF-κB, and COX-2 proteins expression during status epilepticus but not during the latent period while neurons were still degenerating. These data suggest that seizure-related IL1-β, NF-κB, and COX-2 expression may contribute to the pathophysiology of epilepsy by inducing neuronal death and astrocytic activation.

Abstract: It is well known that the uncoupling between local cerebral glucose utilization (LCGU) and local cerebral blood flow (LCBF), i.e. decrease in LCBF rates with high LCGU, is frequently associated with seizure-induced neuronal damage. This study was performed to assess if the neuroprotective effect of the adenosinergic A1 receptor agonist R-N-phenylisopropyladenosine (R-Pia) injected prior to pilocarpine is able to reduce the uncoupling between LCGU and LCBF during status epilepticus (SE). Four groups of rats were studied: Saline, Pilo, R-Pia+Saline and R-Pia+Pilo. For LCGU and LCBF studies, rats were subjected to autoradiography using [14C]-2-deoxyglucose and [14C]-iodoantypirine, respectively. Radioligands were injected 4h after SE onset. Neuronal loss was evaluated by Fluorojade-B (FJB) at two time points after SE onset (24h and 7days). The results showed a significant increase in LCGU in almost all brain regions studied in the Pilo and R-Pia+Pilo groups compared to controls. However, in R-Pia pretreated rats, the uncoupling between LCGU and LCBF was moderated in a limited number of structures as substantia nigra pars reticulata and hippocampal formation rather in favor of hyperperfusion. Significant increases in LCBF were observed in the entorhinal cortex, thalamic nuclei, mammillary body, red nucleus, zona incerta, pontine nucleus and visual cortex. The neuroprotective effect of R-Pia assessed by FJB showed a lower density of degenerating cells in the hippocampal formation, piriform cortex and basolateral amygdala. In conclusion our data shows that the neuroprotective effect of R-Pia was accompanied by a compensatory metabolic input in brain areas involved with seizures generation. [Copyright &y& Elsevier]

Neuronal-astrocytic interactions in 1-month-old Genetic Absence Epilepsy Rats from Strasbourg (GAERS) before the occurrence of seizures are compared to those in non-epileptic rats (NERs) and in adult GAERS expressing epilepsy. Animals received [1-13C]glucose and [1,2-13C]acetate, preferential substrates of neurons and astrocytes, respectively, and extracts from cerebral cortex, subcortex and cerebellum were analyzed by NMR spectroscopy. Increased mitochondrial metabolism took place in the cortical neurons of immature and adult GAERS and therefore does not seem to be a consequence of the occurrence of absence seizures. Glutamine supply to GABAergic neurons was reduced in cortex and subcortex in young GAERS, as reflected by increased glutamine content and decreased 13C-labeling of GABA. In the brain of immature GAERS, interactions between glutamatergic neurons and astrocytes appeared normal whereas increased astrocytic metabolism took place in adult GAERS, suggesting that astrocytic alterations could possibly be the cause of seizures.

The role of neuronal nitric oxide (NO) in the cerebrovascular response to partial seizures induced by intrahippocampal injection of kainate (KA) was investigated in mice deleted for the neuronal NO synthase gene (nNOS-/-) and in wild-type controls (WT). A second group of WT mice received the nNOS inhibitor, 7-nitroindazole (WT-7NI). Local cerebral blood flow (LCBF) was measured using the quantitative (14)C-iodoantipyrine method. Within the epileptic focus, all three groups of seizing mice (WT, WT-7NI, and nNOS-/-) showed significant 26-88% LCBF increases in ipsilateral hippocampus, compared to saline-injected mice. Contralaterally to the epileptic focus, KA seizures induced a 21-47% LCBF decreases in hippocampus and limbic cortex of WT mice and in most contralateral brain structures of nNOS-/- mice, while WT-7NI mice showed no contralateral CBF change. Neuronal NO appears to be not involved in the cerebrovascular response within the epileptic focus, but may rather have a role in the maintenance of distant LCBF regulation during seizures.