Nonconvulsive status epilepticus in rats leads to brain pathology.

OBJECTIVE: Status epilepticus (SE) is an abnormally prolonged epileptic seizure that if associated with convulsive motor symptoms is potentially life threatening for a patient. However, 20%-40% of patients with SE lack convulsive events and instead present with more subtle semiology such as altered consciousness and less motor activity. Today, there is no general consensus regarding to what extent nonconvulsive SE (NCSE) is harmful to the brain, which adds uncertainty to stringent treatment regimes. METHODS: Here, we evaluated brain pathology in an experimental rat and mouse model of complex partial NCSE originating in the temporal lobes with Western blot analysis, immunohistochemistry, and ex vivo diffusion tensor imaging (DTI). The NCSE was induced by electrical stimulation with intrahippocampal electrodes and terminated with pentobarbital anesthesia. Video-electroencephalographic recordings were performed throughout the experiment. RESULTS: DTI of mice 7 weeks post-NCSE showed no robust long-lasting changes in fractional anisotropy within the hippocampal epileptic focus. Instead, we found pathophysiological changes developing over time when measuring protein levels and cell counts in extracted brain tissue. At 6 and 24 hours post-NCSE in rats, few changes were observed within the hippocampus and cortical or subcortical structures in Western blot analyses of key components of the cellular immune response and synaptic protein expression, while neurodegeneration had started. However, 1 week post-NCSE, both excitatory and inhibitory synaptic protein levels were decreased in hippocampus, concomitant with an excessive microglial and astrocytic activation. At 4 weeks, a continuous immune response in the hippocampus was accompanied with neuronal loss. Levels of the excitatory synaptic adhesion molecule N-cadherin were decreased specifically in rats that developed unprovoked spontaneous seizures (epileptogenesis) within 1 month following NCSE, compared to rats only exhibiting acute symptomatic seizures within 1 week post-NCSE. SIGNIFICANCE: These findings provide evidence for a significant brain pathology following NCSE in an experimental rodent model.

Immune response in the eye following epileptic seizures.

BACKGROUND: Epileptic seizures are associated with an immune response in the brain. However, it is not known whether it can extend to remote areas of the brain, such as the eyes. Hence, we investigated whether epileptic seizures induce inflammation in the retina. METHODS: Adult rats underwent electrically induced temporal status epilepticus, and the eyes were studied 6 h, 1, and 7 weeks later with biochemical and immunohistochemical analyses. An additional group of animals received CX3CR1 antibody intracerebroventricularly for 6 weeks after status epilepticus. RESULTS: Biochemical analyses and immunohistochemistry revealed no increased cell death and unaltered expression of several immune-related cytokines and chemokines as well as no microglial activation, 6 h post-status epilepticus compared to non-stimulated controls. At 1 week, again, retinal cytoarchitecture appeared normal and there was no cell death or micro- or macroglial reaction, apart from a small decrease in interleukin-10. However, at 7 weeks, even if the cytoarchitecture remained normal and no ongoing cell death was detected, the numbers of microglia were increased ipsi- and contralateral to the epileptic focus. The microglia remained within the synaptic layers but often in clusters and with more processes extending into the outer nuclear layer. Morphological analyses revealed a decrease in surveying and an increase in activated microglia. In addition, increased levels of the chemokine KC/GRO and cytokine interleukin-1ß were found. Furthermore, macroglial activation was noted in the inner retina. No alterations in numbers of phagocytic cells, infiltrating macrophages, or vascular pericytes were observed. Post-synaptic density-95 cluster intensity was reduced in the outer nuclear layer, reflecting seizure-induced synaptic changes without disrupted cytoarchitecture in areas with increased microglial activation. The retinal gliosis was decreased by a CX3CR1 immune modulation known to reduce gliosis within epileptic foci, suggesting a common immunological reaction. CONCLUSIONS: Our results are the first evidence that epileptic seizures induce an immune response in the retina. It has a potential to become a novel non-invasive tool for detecting brain inflammation through the eyes.

Role of fractalkine-CX3CR1 pathway in seizure-induced microglial activation, neurodegeneration, and neuroblast production in the adult rat brain.

Temporal lobe seizures lead to an acute inflammatory response in the brain primarily characterized by activation of parenchymal microglial cells. Simultaneously, degeneration of pyramidal cells and interneurons is evident together with a seizure-induced increase in the production of new neurons within the dentate gyrus of the hippocampus. We have previously shown a negative correlation between the acute seizure-induced inflammation and the survival of newborn hippocampal neurons. Here, we aimed to evaluate the role of the fractalkine-CX3CR1 pathway for these acute events. Fractalkine is a chemokine expressed by both neurons and glia, while its receptor, CX3CR1 is primarily expressed on microglia. Electrically-induced partial status epilepticus (SE) was induced in adult rats through stereotaxically implanted electrodes in the hippocampus. Recombinant rat fractalkine or CX3CR1 antibody was infused intraventricularly during one week post-SE. A significant increase in the expression of CX3CR1, but not fractalkine, was observed in the dentate gyrus at one week. CX3CR1 antibody treatment resulted in a reduction in microglial activation, neurodegeneration, as well as neuroblast production. In contrast, fractalkine treatment had only minor effects. This study provides evidence for a role of the fractalkine-CX3CR1 signaling pathway in seizure-induced microglial activation and suggests that neuroblast production following seizures may partly occur as a result of microglial activation.

Inflammation without neuronal death triggers striatal neurogenesis comparable to stroke.

Ischemic stroke triggers neurogenesis from neural stem/progenitor cells (NSPCs) in the subventricular zone (SVZ) and migration of newly formed neuroblasts toward the damaged striatum where they differentiate to mature neurons. Whether it is the injury per se or the associated inflammation that gives rise to this endogenous neurogenic response is unknown. Here we showed that inflammation without corresponding neuronal loss caused by intrastriatal lipopolysaccharide (LPS) injection leads to striatal neurogenesis in rats comparable to that after a 30 min middle cerebral artery occlusion, as characterized by striatal DCX+ neuroblast recruitment and mature NeuN+/BrdU+ neuron formation. Using global gene expression analysis, changes in several factors that could potentially regulate striatal neurogenesis were identified in microglia sorted from SVZ and striatum of LPS-injected and stroke-subjected rats. Among the upregulated factors, one chemokine, CXCL13, was found to promote neuroblast migration from neonatal mouse SVZ explants in vitro. However, neuroblast migration to the striatum was not affected in constitutive CXCL13 receptor CXCR5(-/-) mice subjected to stroke. Infarct volume and pro-inflammatory M1 microglia/macrophage density were increased in CXCR5(-/-) mice, suggesting that microglia-derived CXCL13, acting through CXCR5, might be involved in neuroprotection following stroke. Our findings raise the possibility that the inflammation accompanying an ischemic insult is the major inducer of striatal neurogenesis after stroke.

Reduced epilepsy development in synapsin 2 knockout mice with autistic behavior following early systemic treatment with interleukin-6 receptor antibody.

BACKGROUND: Individuals with autism spectrum disorder (ASD) have an increased risk of developing epilepsy. Both ASD and epilepsy have been associated with increased levels of immune factors in the blood, including the proinflammatory cytokine interleukin 6 (IL-6). Mice lacking the synapsin 2 gene (Syn2 KO) exhibit ASD-like behavior and develop epileptic seizures. Their brains display neuroinflammatory changes including elevated IL-6 levels. We aimed to investigate the effect of systemic IL-6 receptor antibody (IL-6R ab) treatment on seizure development and frequency in Syn2 KO mice. MATERIAL AND METHODS: Weekly systemic (i.p.) injections of IL-6R ab or saline were given to Syn2 KO mice starting either early in life at 1 month of age, before seizure debut or at 3 months of age, directly after seizure debut and continued for 4 or 2 months, respectively. Seizures were provoked by handling the mice three times per week. The neuroinflammatory response and synaptic protein levels in the brain were determined by ELISA, immunohistochemistry and western blots. In an additional group of Syn2 KO mice, with IL-6R ab treatment early in life, ASD-related behavioral tests including social interaction and repetitive self-grooming, as well as cognitive memory and depressive-/anxiety-like tests, and actigraphy measurements of circadian sleep-awake rhythm were analyzed. RESULTS: The IL-6R ab treatment reduced seizure development and frequency in Syn2 KO mice when initiated before, but not after, seizure debut. However, early treatment did not reverse the neuroinflammatory response or the imbalance in synaptic protein levels in the brain previously reported in Syn2 KO mice. The treatment did not affect social interaction, performance in memory, depressive-/anxiety-like tests or the sleep-awake rhythm of Syn2 KO mice. CONCLUSION: These findings suggest the involvement of IL-6 receptor signaling during epilepsy development in Syn2 KO mice, without significant alterations of the immune reaction in the brain, and independently of cognitive performance, mood and circadian sleep-awake rhythm.

Immune response in blood before and after epileptic and psychogenic non-epileptic seizures.

Inflammatory processes may provoke epileptic seizures and seizures may promote an immune reaction. Hence, the systemic immune reaction is a tempting diagnostic and prognostic marker in epilepsy. We explored the immune response before and after epileptic and psychogenic non-epileptic seizures (PNES). Serum samples collected from patients with videoEEG-verified temporal or frontal lobe epilepsy (TLE or FLE) or TLE + PNES showed increased interleukin-6 (IL-6) levels in between seizures (interictally), compared to controls. Patients with PNES had no increase in IL-6. The IL-6 levels increased transiently even further within hours after a seizure (postictally) in TLE but not in FLE patients. The postictal to interictal ratio of additionally five immune factors were also increased in TLE patients only. We conclude that immune factors have the potential to be future biomarkers for epileptic seizures and that the heterogeneity among different epileptic and non-epileptic seizures may be disclosed in peripheral blood sampling independent of co-morbidities.

Inhibition of microglial activation protects hippocampal neurogenesis and improves cognitive deficits in a transgenic mouse model for Alzheimer's disease.

BACKGROUND: Activated microglia with macrophage-like functions invade and surround ß-amyloid (Aß) plaques in Alzheimer's disease (AD), possibly contributing to the turnover of Aß, but they can also secrete proinflammatory factors that may be involved in the pathogenesis of AD. Microglia are known to modulate adult hippocampal neurogenesis. OBJECTIVES/METHODS: To determine the role of microglia on neurogenesis in brains with Aß pathology, we inhibited microglial activation with the tetracycline derivative minocycline in doubly transgenic mice expressing mutant human amyloid precursor protein (APP) and mutant human presenilin-1 (PS1). RESULTS: Minocycline increased the survival of new dentate granule cells in APP/PS1 mice indicated by more BrdU+/NeuN+ cells as compared to vehicle-treated transgenic littermates, accompanied by improved behavioral performance in a hippocampus-dependent learning task. Both brain levels of Aß and Aß-related morphological deficits in the new neurons labeled with GFP-expressing retrovirus were unaffected in minocycline-treated mice. CONCLUSIONS: These results suggest a role for microglia in Aß-related functional deficits and in suppressing the survival of new neurons, and show that modulation of microglial function with minocycline can protect hippocampal neurogenesis in the presence of Aß pathology.

Levetiracetam and N-Cadherin Antibody Alleviate Brain Pathology Without Reducing Early Epilepsy Development After Focal Non-convulsive Status Epilepticus in Rats.

Focal non-convulsive status epilepticus (fNCSE) is a neurological condition characterized by a prolonged seizure that may lead to the development of epilepsy. Emerging experimental evidence implicates neuronal death, microglial activation and alterations in the excitatory and inhibitory synaptic balance as key features in the pathophysiology following fNCSE. We have previously reported alterations in the excitatory adhesion molecule N-cadherin in rats with fNCSE originating from the hippocampus that subsequently also develop spontaneous seizures. In this study, fNCSE rats were treated intraperitoneally with the conventional anti-epileptic drug levetiracetam in combination with intraparenchymal infusion of N-cadherin antibodies (Ab) for 4 weeks post-fNCSE. The N-cadherin Ab was infused into the fornix and immunohistochemically N-cadherin Ab-stained neurons were detected within the dorsal hippocampal structures as well as in superjacent somatosensory cortex. Continuous levetiracetam treatment for 4 weeks post-fNCSE reduced microglia activation, including cell numbers and morphological changes, partly decreased neuronal cell loss, and excitatory post-synaptic scaffold protein PSD-95 expression in selective hippocampal structures. The additional treatment with N-cadherin Ab did not reverse neuronal loss, but moderately reduced microglial activation, and further reduced PSD-95 levels in the dentate hilus of the hippocampus. Despite the effects on brain pathology within the epileptic focus, neither monotherapy with systemic levetiracetam nor levetiracetam in combination with local N-cadherin Ab administration, reduced the amount of focal or focal evolving into bilateral convulsive seizures, seizure duration, or interictal epileptiform activity during 1 month of continuous electroenephalogram recordings within the hippocampus after fNCSE. Behavioral tests for spatial memory, anxiety, social interaction and anhedonia did not detect gross behavioral differences between fNCSE rats with or without treatment. The results reveal the refractory features of the present rodent model of temporal lobe epilepsy following fNCSE, which supports its clinical value for further therapeutic studies. We identify the persistent development of epilepsy following fNCSE, in spite of partly reduced brain pathology within the epileptic focus.

Brain pathology in focal status epilepticus: evidence from experimental models.

Status Epilepticus (SE) is often a neurological emergency characterized by abnormally sustained, longer than habitual seizures. The new ILAE classification reports that SE "…can have long-term consequences including neuronal death, neuronal injury…depending on the type and duration of seizures". While it is accepted that generalized convulsive SE exerts detrimental effects on the brain, it is not clear if other forms of SE, such as focal non-convulsive SE, leads to brain pathology and contributes to long-term deficits in patients. With the available clinical and experimental data, it is hard to discriminate the specific action of the underlying SE etiologies from that exerted by epileptiform activity. This information is highly relevant in the clinic for better treatment stratification, which may include both medical and surgical intervention for seizure control. Here we review experimental studies of focal SE, with an emphasis on focal non-convulsive SE. We present a repertoire of brain pathologies observed in the most commonly used animal models and attempt to establish a link between experimental findings and human condition(s). The extensive literature on focal SE animal models suggest that the current approaches have significant limitations in terms of translatability of the findings to the clinic. We highlight the need for a more stringent description of SE features and brain pathology in experimental studies in animal models, to improve the accuracy in predicting clinical translation.

Inflammatory reaction in the retina after focal non-convulsive status epilepticus in mice investigated with high resolution magnetic resonance and diffusion tensor imaging.

Pathophysiological consequences of focal non-convulsive status epilepticus (fNCSE) have been difficult to demonstrate in humans. In rats fNCSE pathology has been identified in the eyes. Here we evaluated the use of high-resolution 7 T structural T1-weighted magnetic resonance imaging (MRI) and 9.4 T diffusion tensor imaging (DTI) for detecting hippocampal fNCSE-induced retinal pathology ex vivo in mice. Seven weeks post-fNCSE, increased number of Iba1+ microglia were evident in the retina ipsilateral to the hemisphere with fNCSE, and morphologically more activated microglia were found in both ipsi- and contralateral retina compared to non-stimulated control mice. T1-weighted intensity measurements of the contralateral retina showed a minor increase within the outer nuclear and plexiform layers of the lateral retina. T1-weighted measurements were not performed in the ipsilateral retina due to technical difficulties. DTI fractional anisotropy(FA) values were discretely altered in the lateral part of the ipsilateral retina and unaltered in the contralateral retina. No changes were observed in the distal part of the optic nerve. The sensitivity of both imaging techniques for identifying larger retinal alteration was confirmed ex vivo in retinitis pigmentosa mice where a substantial neurodegeneration of the outer retinal layers is evident. With MR imaging a 50 % decrease in DTI FA values and significantly thinner retina in T1-weighted images were detected. We conclude that retinal pathology after fNCSE in mice is subtle and present bilaterally. High-resolution T1-weighted MRI and DTI independently did not detect the entire pathological retinal changes after fNCSE, but the combination of the two techniques indicated minor patchy structural changes.

Environment matters: synaptic properties of neurons born in the epileptic adult brain develop to reduce excitability.

Neural progenitors in the adult dentate gyrus continuously produce new functional granule cells. Here we used whole-cell patch-clamp recordings to explore whether a pathological environment influences synaptic properties of new granule cells labeled with a GFP-retroviral vector. Rats were exposed to a physiological stimulus, i.e., running, or a brain insult, i.e., status epilepticus, which gave rise to neuronal death, inflammation, and chronic seizures. Granule cells formed after these stimuli exhibited similar intrinsic membrane properties. However, the new neurons born into the pathological environment differed with respect to synaptic drive and short-term plasticity of both excitatory and inhibitory afferents. The new granule cells formed in the epileptic brain exhibited functional connectivity consistent with reduced excitability. We demonstrate a high degree of plasticity in synaptic inputs to adult-born new neurons, which could act to mitigate pathological brain function.

Abeta immunotherapy protects morphology and survival of adult-born neurons in doubly transgenic APP/PS1 mice.

The hippocampus is heavily affected by progressive neurodegeneration and beta-amyloid pathology in Alzheimer's disease (AD). The hippocampus is also one of the few brain regions that generate new neurons throughout adulthood. Because hippocampal neurogenesis is regulated by both endogenous and environmental factors, we determined whether it benefits from therapeutic reduction of beta-amyloid peptide (Abeta)-related toxicity induced by passive Abeta immunotherapy. Abeta immunotherapy of 8-9-month-old mice expressing familial AD-causing mutations in the amyloid precursor protein and presenilin-1 genes with an antibody against Abeta decreased compact beta-amyloid plaque burden and promoted survival of newly born neurons in the hippocampal dentate gyrus. As these neurons matured, they exhibited longer dendrites with more complex arborization compared with newly born neurons in control-treated transgenic littermates. The newly born neurons showed signs of functional integration indicated by expression of the immediate-early gene Zif268 in response to exposure to a novel object. Abeta immunotherapy was associated with higher numbers of synaptophysin-positive synaptic boutons. Labeling dividing progenitor cells with a retroviral vector encoding green fluorescent protein (GFP) showed that Abeta immunotherapy restored the impaired dendritic branching, as well as the density of dendritic spines in new mature neurons. The presence of cellular prion protein (PrP(c)) on the dendrites of the GFP(+) newly born neurons is compatible with a putative role of PrP(c) in mediating Abeta-related toxicity in these cells. In addition, passive Abeta immunotherapy was accompanied by increased angiogenesis. Our data establish that passive Abeta immunotherapy can restore the morphological maturation of the newly formed neurons in the adult hippocampus and promote angiogenesis. These findings provide evidence for a role of Abeta immunotherapy in stimulating neurogenesis and angiogenesis in transgenic mouse models of AD, and they suggest the possibility that Abeta immunotherapy can recover neuronal and vascular functions in brains with beta-amyloidosis.

Inflammation regulates functional integration of neurons born in adult brain.

Inflammation influences several steps of adult neurogenesis, but whether it regulates the functional integration of the new neurons is unknown. Here, we explored, using confocal microscopy and whole-cell patch-clamp recordings, whether a chronic inflammatory environment affects the morphological and electrophysiological properties of new dentate gyrus granule cells, labeled with a retroviral vector encoding green fluorescent protein. Rats were exposed to intrahippocampal injection of lipopolysaccharide, which gave rise to long-lasting microglia activation. Inflammation caused no changes in intrinsic membrane properties, location, dendritic arborization, or spine density and morphology of the new cells. Excitatory synaptic drive increased to the same extent in new and mature cells in the inflammatory environment, suggesting increased network activity in hippocampal neural circuitries of lipopolysaccharide-treated animals. In contrast, inhibitory synaptic drive was more enhanced by inflammation in the new cells. Also, larger clusters of the postsynaptic GABA(A) receptor scaffolding protein gephyrin were found on dendrites of new cells born in the inflammatory environment. We demonstrate for the first time that inflammation influences the functional integration of adult-born hippocampal neurons. Our data indicate a high degree of synaptic plasticity of the new neurons in the inflammatory environment, which enables them to respond to the increase in excitatory input with a compensatory upregulation of activity and efficacy at their afferent inhibitory synapses.

Long-term accumulation of microglia with proneurogenic phenotype concomitant with persistent neurogenesis in adult subventricular zone after stroke.

Neural stem cells (NSCs) in the adult rat subventricular zone (SVZ) generate new striatal neurons during several months after ischemic stroke. Whether the microglial response associated with ischemic injury extends into SVZ and influences neuroblast production is unknown. Here, we demonstrate increased numbers of activated microglia in ipsilateral SVZ concomitant with neuroblast migration into the striatum at 2, 6, and 16 weeks, with maximum at 6 weeks, following 2 h middle cerebral artery occlusion in rats. In the peri-infarct striatum, numbers of activated microglia peaked already at 2 weeks and declined thereafter. Microglia in SVZ were resident or originated from bone marrow, with maximum proliferation during the first 2 weeks postinsult. In SVZ, microglia exhibited ramified or intermediate morphology, signifying a downregulated inflammatory profile, whereas amoeboid or round phagocytic microglia were frequent in the peri-infarct striatum. Numbers of microglia expressing markers of antigen-presenting cells (MHC-II, CD86) increased in SVZ but very few lymphocytes were detected. Using quantitative PCR, strong short- and long-term increase (at 1 and 6 weeks postinfarct) of insulin-like growth factor-1 (IGF-1) gene expression was detected in SVZ tissue. Elevated numbers of IGF-1-expressing microglia were found in SVZ at 2, 6, and 16 weeks after stroke. At 16 weeks, 5% of microglia but no other cells in SVZ expressed the IGF-1 protein, which mitigates apoptosis and promotes proliferation and differentiation of NSCs. The long-term accumulation of microglia with proneurogenic phenotype in the SVZ implies a supportive role of these cells for the continuous neurogenesis after stroke.

Immune Profile in Blood Following Non-convulsive Epileptic Seizures in Rats.

Non-convulsive status epilepticus (NCSE) is a prolonged epileptic seizure with subtle symptoms that may delay clinical diagnosis. Emerging experimental evidence shows brain pathology and epilepsy development following NCSE. New diagnostic/prognostic tools are therefore needed for earlier and better stratification of treatment. Here we examined whether NCSE initiates a peripheral immune response in blood serum from rats that experienced electrically-induced NCSE. ELISA analysis showed an acute transient increase in serum protein levels including interleukin-6 6 h post-NCSE, similar to the immune reaction in the brain. At 4 weeks post-NCSE, when 75% of rats subjected to NCSE had also developed spontaneous seizures, several immune proteins were altered. In particular, markers associated with microglia, macrophages and antigen presenting cells, such as CD68, MHCII, and galectin-3, were increased and the T-cell marker CD4 was decreased in serum compared to both non-stimulated controls and NCSE rats without spontaneous seizures, without correlation to interictal epileptiform activity. Analyses of serum following intracerebral injection of lipopolysaccharide (LPS) showed an acute increase in interleukin-6, but at 4 weeks unaltered levels of MHCII and galectin-3, an increase in CD8 and CD11b and a decrease in CD68. None of the increased serum protein levels after NCSE or LPS could be confirmed in spleen tissue. Our data identifies the possibility to detect peripheral changes in serum protein levels following NCSE, which may be related to the development of subsequent spontaneous seizures.

Physical Activity Reduces Epilepsy Incidence: a Retrospective Cohort Study in Swedish Cross-Country Skiers and an Experimental Study in Seizure-Prone Synapsin II Knockout Mice.

BACKGROUND: Epilepsy patients commonly exercise less than the general population. Animal studies indicate beneficial effects of physical activity in established epilepsy, while its effect on the development is currently less known. METHODS: Here, we investigated the incidence of epilepsy during 20 years in a cohort of participants from the long-distance Swedish cross-country ski race Vasaloppet (n = 197,685) and compared it to the incidence of non-participating-matched controls included in the Swedish population register (n = 197,684). Individuals diagnosed with diseases such as stroke and epilepsy before entering the race were excluded from both groups. Experimentally, we also determined how physical activity could affect the development of epilepsy in epilepsy-prone synapsin II knockout mice (SynIIKO), with and without free access to a running wheel. RESULTS: We identified up to 40-50% lower incidence of epilepsy in the Vasaloppet participants of all ages before retirement. A lower incidence of epilepsy in Vasaloppet participants was seen regardless of gender, education and occupation level compared to controls. The participants included both elite and recreational skiers, and in a previous survey, they have reported a higher exercise rate than the general Swedish population. Sub-analyses revealed a significantly lower incidence of epilepsy in participants with a faster compared to slower finishing time. Dividing participants according to specified epilepsy diagnoses revealed 40-50% decrease in focal and unspecified epilepsy, respectively, but no differences in generalized epilepsy. Voluntary exercise in seizure-prone SynIIKO mice for 1 month before predicted epilepsy development decreased seizure manifestation from > 70 to 40%. Brain tissue analyses following 1 month of exercise showed increased hippocampal neurogenesis (DCX-positive cells), while microglial (Iba1) and astrocytic activation (GFAP), neuronal Map2, brain-derived neurotrophic factor and its receptor tyrosine receptor kinase B intensity were unaltered. Continued exercise for additionally 2 months after predicted seizure onset in SynIIKO mice resulted in a 5-fold reduction in seizure manifestation (from 90 to 20%), while 2 months of exercise initiated at the time of predicted seizure development gave no seizure relief, suggesting exercise-induced anti-epileptogenic rather than anti-convulsive effect. CONCLUSION: The clinical study and the experimental findings in mice indicate that physical activity may prevent or delay the development of epilepsy.

Tumor necrosis factor receptor 1 is a negative regulator of progenitor proliferation in adult hippocampal neurogenesis.

Tumor necrosis factor-alpha (TNF-alpha) is a proinflammatory cytokine, acting through the TNF-R1 and TNF-R2 receptors. The two receptors have been proposed to mediate distinct TNF-alpha effects in the CNS, TNF-R1 contributing to neuronal damage and TNF-R2 being neuroprotective. Whether TNF-alpha and its receptors play any role for neurogenesis in the adult brain is unclear. Here we used mouse models with loss of TNF-R1 and TNF-R2 function to establish whether signaling through these receptors could influence hippocampal neurogenesis in vivo under basal conditions, as well as after status epilepticus (SE), which is associated with inflammation and elevated TNF-alpha levels. Notably, in the intact brain, the number of new, mature hippocampal neurons was elevated in TNF-R1(-/-) and TNF-R1/R2(-/-) mice, whereas no significant changes were detected in TNF-R2(-/-) mice. Also after SE, the TNF-R1(-/-) and TNF-R1/R2(-/-) mice produced more new neurons. In contrast, the TNF-R2(-/-) mice showed reduced SE-induced neurogenesis. Cell proliferation in the dentate subgranular zone was elevated in TNF-R1(-/-) and TNF-R1/R2(-/-) mice both under basal conditions and after SE. The TNF-R2(-/-) mice either showed no change or minor decrease of cell proliferation. TNF-R1 and TNF-R2 receptors were expressed by hippocampal progenitors, as assessed with reverse transcription-PCR on sorted or cultured cells and immunocytochemistry on cultures. Our data reveal differential actions of TNF-R1 and TNF-R2 signaling in adult hippocampal neurogenesis and identify for the first time TNF-R1 as a negative regulator of neural progenitor proliferation in both the intact and pathological brain.

Interactions Between Microglia and Newly Formed Hippocampal Neurons in Physiological and Seizure-Induced Inflammatory Environment.

Adult hippocampal neurogenesis is modulated by physiological and pathological stimuli, including seizures and inflammation. Here, we describe stable interactions between microglia and newborn neurons using two-photon and confocal microscopy. On 3 weeks-old neurons, these interactions exhibit preferences for distal dendrites under physiological conditions. Conversely, after status epilepticus, ramified microglia, in particular, interact more with the proximal dendrites of new neurons. No such differences were found on 6 weeks-old neurons. Our study demonstrates regional and temporal specificity of the interactions between newborn neurons and microglia during a critical period for homeostasis and synaptic integration.

Alterations in Brain Inflammation, Synaptic Proteins, and Adult Hippocampal Neurogenesis during Epileptogenesis in Mice Lacking Synapsin2.

Synapsins are pre-synaptic vesicle-associated proteins linked to the pathogenesis of epilepsy through genetic association studies in humans. Deletion of synapsins causes an excitatory/inhibitory imbalance, exemplified by the epileptic phenotype of synapsin knockout mice. These mice develop handling-induced tonic-clonic seizures starting at the age of about 3 months. Hence, they provide an opportunity to study epileptogenic alterations in a temporally controlled manner. Here, we evaluated brain inflammation, synaptic protein expression, and adult hippocampal neurogenesis in the epileptogenic (1 and 2 months of age) and tonic-clonic (3.5-4 months) phase of synapsin 2 knockout mice using immunohistochemical and biochemical assays. In the epileptogenic phase, region-specific microglial activation was evident, accompanied by an increase in the chemokine receptor CX3CR1, interleukin-6, and tumor necrosis factor-α, and a decrease in chemokine keratinocyte chemoattractant/ growth-related oncogene. Both post-synaptic density-95 and gephyrin, scaffolding proteins at excitatory and inhibitory synapses, respectively, showed a significant up-regulation primarily in the cortex. Furthermore, we observed an increase in the inhibitory adhesion molecules neuroligin-2 and neurofascin and potassium chloride co-transporter KCC2. Decreased expression of γ-aminobutyric acid receptor-δ subunit and cholecystokinin was also evident. Surprisingly, hippocampal neurogenesis was reduced in the epileptogenic phase. Taken together, we report molecular alterations in brain inflammation and excitatory/inhibitory balance that could serve as potential targets for therapeutics and diagnostic biomarkers. In addition, the regional differences in brain inflammation and synaptic protein expression indicate an epileptogenic zone from where the generalized seizures in synapsin 2 knockout mice may be initiated or spread.