Mircea Oprica

Inflammatory mechanisms associated with brain damage induced by kainic acid with special reference to the interleukin-1 system.

The evidence of inflammatory processes in the clinical manifestations and neuropathological sequelae of epilepsy have accumulated in the last decade. Administration of kainic acid, an analogue of the excitatory amino acid glutamate, induces a characteristic behavioural syndrome and a reproducible pattern of neurodegeneration in several brain areas, closely resembling human temporal lobe epilepsy. Results from studies using the kainic acid model indicate that manipulation of pro‐ and anti‐inflammatory cytokines can modify the outcome with regard to the behavioural syndrome as well as the neuropathological consequences. Interleukin‐1 is one of the most important cytokines and has several actions in the brain that are critical for the host defense against injury and infection, and it is involved in the initiation of early stages of inflammation. It is believed that interleukin‐1 plays a pivotal role in the neuroinflammation associated with certain forms of neurodegeneration, including cerebral ischemia, trauma and excitotoxic brain injury. In this review, we have summarized the experimental data available with regard to the involvement of the interleukin‐1 system in kainic acid‐induced changes in the brain and emphasized the modulatory role of interleukin‐1β in this model of epilepsy.

Impaired long term memory consolidation in transgenic mice overexpressing the human soluble form of IL-1ra in the brain

Interleukin-1 (IL-1) is expressed following LTP induction and is required for long-term memory consolidation. We demonstrate that the long-term, but not short-term memory is impaired in a transgenic mouse strain overexpressing the human soluble interleukin-1 receptor antagonist (hsIL-1ra) in the brain. Overexpression of IL-1ra was found to reduce the basal as well as the novelty-induced upregulation of activity-regulated cytoskeleton-associated protein (Arc) in the dentate gyrus and in the retrosplenial cortex. Together with the finding that blocking IL-1 receptors interferes with the BDNF-ERK1/2 pathway, our data suggest an essential role played by physiological levels of IL-1 in long-term memory consolidation.

Blunted neurogenesis and gliosis due to transgenic overexpression of human soluble IL‐1ra in the mouse

Interleukin‐1 (IL‐1) is one of the most important cytokines in neuroinflammation, in acute conditions as well as during natural ageing and neurodegenerative disorders. Using a transgenic mouse strain with brain‐directed overexpression of IL‐1 receptor antagonist (Tg hsIL‐1ra), we show that blocking IL‐1 receptor‐mediated activity resulted in abolishing the alterations in neurogenesis in response to acute and chronic neuroinflammation. In addition, using a novel approach to quantifying glial activation, we show that expression of the astrocyte cytoskeletal marker glial fibrillary acidic protein (GFAP) following kainic acid (KA)‐induced seizures or during ageing did not change in Tg hsIL‐1ra animals. Nevertheless, the astrocyte morphology showed major alterations, consisting of fragmentation of the processes in Tg hsIL‐1ra mice. Similarly, although there was a higher degree of basal microglial activation in the transgenic mice than wild‐type animals, there was no change following KA‐induced seizures or with ageing. Taken together, our results indicate that IL‐1 is crucial for the adaptability of the brain to acute and chronic neuroinflammation.

Inflammation in the nervous system--physiological and pathophysiological aspects.

There is ample evidence for the occurrence of inflammatory processes in most major neurodegenerative disorders, both in acute conditions such as traumatic brain injury and stroke, and in chronic disorders such as Alzheimers disease, epilepsy, amyotrophic lateral sclerosis and Parkinsons disease. Studies on inflammatory factors such as pro- and antiinflammatory cytokines in experimental models of neurodegenerative disorders suggest that they are not merely bystanders, but may be involved in the neurodegenerative process. In addition, there are findings indicating that inflammatory factors may have beneficial effects on the nervous system, particularly during development of the nervous system. The challenge is to understand when, where and during which circumstances inflammation and inflammatory factors are positive or negative for neuronal survival and functioning. Some of our studies on cytokines, particularly the interleukin-1 system, are summarised and discussed in relation to neurodegeneration, cognition, and temperature changes.

Dantrolene protects neurons against kainic acid induced apoptosis in vitro and in vivo

Apoptotic cell death induced by kainic acid (KA) in cultures of rat cerebellar granule cells (CGC) and in different brain regions of Wistar rat pups on postnatal day 21 (P21) was studied. In vitro, KA (100–500 μM) induced a concentration‐dependent loss of cell viability in MTT assay and cell death had apoptotic morphology as studied by chromatin staining with propidium iodide (PI). In vivo, twenty‐four hours after induction of status epilepticus (SE) by an intraperitoneal KA injection (5 mg/kg) we quantified apoptotic cells in hippocampus (CA1 and CA3), parietal cortex and cerebellum using PI staining and terminal deoxynucleotidyl transferase‐mediated dUTP nick end labeling (TUNEL) technique. We report that dantrolene, a specific ryanodine receptor antagonist, was able to significantly reduce the apoptotic cell death in CGC cultures and in hyppocampal CA1 and parietal cortex regions. Our finding can be valuable for neuroprotective therapy strategies in patients with repeated generalized seizures or status epilepticus.

Transgenic overexpression of interleukin-1 receptor antagonist in the CNS influences behaviour, serum corticosterone and brain monoamines.

The effects of brain-directed overexpression of human soluble interleukin-1 receptor antagonist (hsIL-1ra) on behaviour, serum corticosterone (CST) levels and concentrations of dopamine (DA), 5-hydroxytryptamine (5-HT) and their metabolites in different brain regions, were investigated in six months old homozygotic transgenic male mice (Tg hsIL-1ra(+/+)). The transgenic and age-matched wild type (WT) mice were subjected to a battery of behavioural tests for analysis of open field (OF) behaviours, anxiety in elevated plus maze (EPM), and motor performance in rotarod. One week after the last behavioural test, half of the mice from each genotype were subjected to a mild stress, while the remaining mice served as controls for the determination of serum CST levels and monoamine concentrations in different brain regions. Tg hsIL-1ra(+/+) mice had higher locomotor scores and showed less habituation in the OF test, spent more time in the open arms of the EPM and had similar motor performance as compared to WT mice. The serum CST levels were comparable, both in basal conditions and upon stress, in the two genotypes. Tg hsIL-1ra(+/+) mice had lower concentrations of DA, 5-HT and their metabolites in several brain regions, with different effects on monoamine turnover upon stress. In conclusion, brain-directed overexpression of hsIL-1ra resulted in increased locomotion and decreased habituation, an anxiolytic effect, but did not influence motor performance. Finally, the activation of hypothalamo-pituitary-adrenal (HPA)-axis was comparable in the two genotypes, however Tg hsIL-1ra(+/+) mice had a modified metabolism of brain monoamines as compared to WT mice.

Studies on brain volume, Alzheimer-related proteins and cytokines in mice with chronic overexpression of IL-1 receptor antagonist

Inflammation is associated with both acute and chronic neurological disorders, including stroke and Alzheimers disease (AD). Cytokines such as interleukin (IL)‐1 have several activities in the brain both under physiological and pathophysiological conditions. The objective of this study was to evaluate consequences of the central blockade of IL‐1 transmission in a previously developed transgenic mouse strain with brain‐directed overexpression of human soluble IL‐1 receptor antagonist (Tg hsIL‐1ra). Effects on brain morphology and brain levels of the AD‐related proteins β‐amyloid precursor protein (APP) and presenilin 1(PS1), as well as the levels of IL‐1β, IL‐6 and tumour necrosis factor‐α (TNF‐α) were analysed in homozygotic and heterozygotic mice and wild type (WT) controls, of both genders and of young (30–40 days) and adult (13–14 months) age. A marked reduction in brain volume was observed in transgenic mice as determined by volumetry. Western blot analysis showed higher levels of APP, but lower levels of PS1, in adult animals than in young ones. In the cerebellum, heterozygotic (Tg hsIL‐1ra+/−) mice had lower levels of APP and PS1 than WT mice. With one exception, there were no genotypic differences in the levels of IL‐1β, IL‐6 and TNF‐α. The cytokine levels were generally higher in adult than in young mice. In conclusion, the chronic blockade of IL‐1 signalling in the brain was associated with an atrophic phenotype of the brain, and with modified levels of APP and PS1. Brain‐directed overexpression of hsIL‐1ra was not followed by major compensatory changes in the levels of pro‐inflammatory cytokines.

Allergy influences the inflammatory status of the brain and enhances tau‐phosphorylation

Despite the existing knowledge regarding the neuropathology of Alzheimers disease (AD), the cause of sporadic forms of the disease is unknown. It has been suggested that systemic inflammation may have a role, but the exact mechanisms through which inflammatory processes influence the pathogenesis and progress of AD are not obvious. Allergy is a chronic inflammatory disease affecting more than 20% of the Western population, but the effects of allergic conditions on brain functions are largely unknown. The aim of this study was to investigate whether or not chronic peripheral inflammation associated with allergy affects the expression of AD‐related proteins and inflammatory markers in the brain. On the basis of previously described models for allergy in mice we developed a model of chronic airway allergy in mouse, with ovalbumin as allergen. The validity of the chronic allergy model was confirmed by a consistent and reproducible eosinophilia in the bronchoalveolar lavage (BAL) fluid of allergic animals. Allergic mice were shown to have increased brain levels of both immunoglobulin (Ig) G and IgE with a widespread distribution. Allergy was also found to increase phosphorylation of tau protein in the brain. The present data support the notion that allergy‐dependent chronic peripheral inflammation modifies the brain inflammatory status, and influences phosphorylation of an AD‐related protein, indicating that allergy may be yet another factor to be considered for the development and/or progression of neurodegenerative diseases such as AD.

Effects of chronic overexpression of interleukin-1 receptor antagonist in a model of permanent focal cerebral ischemia in mouse

Interleukin-1 receptor antagonist (IL-1ra) has been shown previously to have neuroprotective effects in animal models of stroke. The effects of chronic overexpression of human soluble IL-1ra (hsIL-1ra) were studied in a mouse model of permanent focal cerebral ischemia. A transgenic mouse strain (Tg hsIL-1ra+/–) has been developed using the promoter for glial fibrillary acidic protein (GFAP) to limit the overexpression to the CNS. Analysis of the neurological scores, infarct volume and edema formation revealed no differences between Tg hsIL-1ra+/– and wild-type (WT) mice. The cerebral ischemia resulted in pronounced astrocyte proliferation and microglial activation, as well as induction of inflammatory markers in both Tg hsIL-1ra+/– and WT mice, with no major differences between the two genotypes. Interestingly, hsIL-1ra expression in astrocytes was reduced in infarcted areas as compared to non-ischemic regions and sham-operated controls. In conclusion, transgenic overexpression of hsIL1-ra was not neuroprotective in this cerebral ischemia model, possibly due to insufficient levels for protection against the extensive lesion, or an up-regulation of compensatory inflammatory signals due to the lifetime blockade of IL-1 receptors.

Effects of α-MSH on kainic acid induced changes in core temperature in rats

Abstract The effects of intraperitoneal (i.p.) administration of kainic acid (KA) and α-melanocyte-stimulating hormone (α-MSH) alone or in combination, on core temperature of freely moving rats were examined. KA or saline was administered once (10 mg/kg) and α-MSH or saline was given repeatedly i.e. 10 min before and 10, 30 and 60 min after the administration of saline or KA. Two doses of α-MSH were used: 0.5 and 2.5 mg/kg. KA alone produced a biphasic effect on core temperature, i.e. an initial short-lasting hypothermia followed by hyperthermia that lasted about 6 h. The higher dose of α-MSH had a potentiating effect on KA-induced hypothermia, while the lower dose of α-MSH increased the hyperthermia produced by KA. α-MSH administered alone produced a late (3 h), dose-dependent increase in core temperature. It is conceivable that repeated administration of α-MSH in the doses used in our study may cause a cumulative effect in raising body temperature for a limited period of time. The previously described interactions between KA and α-MSH, respectively, with dopaminergic and serotoninergic systems may account for the effects on core temperature in rats observed in our study.