Patricia Parnet

Cytokine-induced sickness behaviour: mechanisms and implications

Sickness behaviour represents the expression of the adaptive reorganization of the priorities of the host during an infectious episode. This process is triggered by pro-inflammatory cytokines produced by peripheral phagocytic cells in contact with invading micro-organisms. The peripheral immune message is relayed to the brain via a fast neural pathway and a slower humoral pathway, resulting in the expression of pro-inflammatory cytokines in macrophage-like cells and microglia in the brain. The cellular and molecular components of this previously unsuspected system are being progressively identified. These advances are opening new avenues for understanding brain disorders, including depression.

Peripheral administration of lipopolysaccharide induces the expression of cytokine transcripts in the brain and pituitary of mice

The reverse transcription polymerase chain reaction (RT-PCR) was used to assess the induction of mRNA of the proinflammatory cytokines IL-1 beta, IL-6 and TNF alpha in the spleen, pituitary, hypothalamus and hippocampus of mice after an intraperitoneal injection of lipopolysaccharide (LPS, 10 micrograms/mouse). The kinetics of cytokine gene expression induced by peripheral LPS in the pituitary and brain structures were different from that observed in the spleen. For IL-1 beta the dose-response curve was also measured and also found to be different. These results support the idea that one pathway by which peripheral immune stimuli affect brain functions includes local synthesis of proinflammatory cytokines in certain brain structures.

Synergy between tumor necrosis factor α and interleukin-1 in the induction of sickness behavior in mice

Like interleukin-1, recombinant human tumor necrosis factor alpha (TNF alpha) has been found to decrease social exploration and induce weight loss in mice in a dose and time-dependent manner. The present study was carried out to study the interaction between these two cytokines. Mice were injected IP with subthreshold doses of TNF alpha (2.5 micrograms/mouse) and IL-1 beta (50 ng/mouse). Social exploration was decreased 2 and 4 h after injection of TNF and IL-1, but body weight was not affected. Subthreshold doses of TNF alpha (90 ng/mouse) and IL-1 beta (100 pg/mouse) were also injected intracerebroventricularly (ICV). Social exploration was decreased 1.5 and 3 h after injections of the two cytokines and body weight was decreased for 6 h. To test the possibility of central induction of IL-1 by TNF alpha, mice pretreated with IL-1 receptor antagonist (IL-1ra, 1.8 micrograms/mouse, ICV) were injected with 90 ng TNF alpha. Pretreatment with IL-1ra antagonized the depressive effect of TNF alpha on behavior, but had no effect on weight loss induced by this cytokine. These results suggest that TNF alpha-induced behavioral alterations are mediated by endogenously released IL-1, whereas metabolic changes are dependent on the release of other cytokines.

Expression of type I and type II interleukin-1 receptors in mouse brain

Although binding sites for IL-1 have been identified in the mouse brain, it is still unknown whether these binding sites correspond to the type I or type II IL-1 receptor. Quantitative autoradiography was used to confirm the presence of specific binding sites for radiolabelled recombinant human IL-1 alpha (125I-HuIL-1 alpha) in the brain of DBA/2 mice. IL-1 binding was highest in the dentate gyrus, consisting of a single class of high affinity binding sites with a Kd of 0.1 nM and a Bmax of 57 fmol/mg protein. A similar Kd of 0.2 nM was obtained using isolated membranes from the whole hippocampus, although the number of binding sites was lower (2 fmol/mg protein). Affinity cross-linking of 125I-Hu-IL-1 alpha to hippocampal membranes revealed the existence of two types of IL-1 receptor proteins, consistent with the sizes of the type I (85 kD) and type II (60 kD) IL-1 receptor. Oligonucleotide probes were then synthesized and used in RT-PCR followed by Southern blotting to show that the whole brain expresses transcripts for both the type I and type II IL-1 receptors. The murine neuroblastoma cell line, C1300, expresses type I rather than type II IL-1 receptor mRNA. The type I receptor protein can be identified by flow cytometry on the membrane of the C1300 neuronal cell line using indirect immunofluorescence with a rat anti-mouse type I IL-1 receptor MoAb. These data show that mouse brain expresses both type I and type II IL-1 receptor mRNA and proteins and offer further support to the idea that type I IL-1 receptors are synthesized and expressed by neurons.

Nutritional programming affects hypothalamic organization and early response to leptin.

Nutritional programming, taking place in utero or early after birth, is closely linked with metabolic and appetite disorders in adulthood. Following the hypothesis that nutritional programming impacts hypothalamic neuronal organization, we report on discrepancies of multiple molecular and cellular early events that take place in the hypothalamus of rats submitted to intrauterine growth restriction (IUGR). Expression screening performed on hypothalami from IUGR rats at birth and at postnatal d 12 identified changes in gene expression of neurodevelopmental process (cell differentiation and cytoskeleton organization). Additionally, a slight reduction of agouti-related protein and a strong reduction of alpha-MSH-immunoreactive efferent fibers were demonstrated in the paraventricular nucleus of IUGR rats. Rapid catch-up growth of IUGR rats, 5 d after birth, had a positive effect on neurodevelopmental factors and on neuronal projections emanating from the arcuate nucleus. The molecular and cellular anomalies detected in IUGR rats can be related to the reduced and delayed plasma leptin surge from d 0-16 when compared with control and IUGR rats with catch-up growth. However, the ability of leptin to activate intracellular signaling in arcuate nucleus neurons was not reduced in IUGR rats. Other mechanism such as epigenetic regulation of the major appetite-regulating neuropeptides genes was analyzed in parallel with their mRNA expression during postnatal development. This study reveals the importance of an early catch-up growth that reduces abnormal organization of hypothalamic pathways involved in energy homeostasis, whereas protein restriction, maintained during postnatal development leads to an important immaturity of the hypothalamus.

Expression and regulation of interleukin-1 receptors in the brain. Role in cytokines-induced sickness behavior

Sickness behavior refers to the coordinated set of behavior changes that develop in sick individuals during the course of an infection. At the molecular level, these changes are due to the effects of proinflammatory cytokines as interleukin-1 on the brain. The purpose of this article is not to review the entire field of cytokines and behavior, but rather to address the role of interleukin-1 receptors (IL-1Rs) in sickness behavior. We briefly describe the notion of sickness behavior and present the distribution of IL-1Rs in the central nervous system of the human, mouse and rat. We then bring arguments in favor of the functionality of the various subtypes of receptors and evaluate the nature of the signaling pathways activated by brain IL-1Rs to initiate central modifications leading to symptoms of sickness. Finally, modulation of IL-1 action on its receptor by various opposing factors including glucocorticoids and anti-inflammatory cytokines is discussed.

Adrenalectomy enhances pro-inflammatory cytokines gene expression, in the spleen, pituitary and brain of mice in response to lipopolysaccharide.

To assess the possible influence of endogenous glucocorticoids on cytokine expression in the brain, adrenalectomized mice and sham operated mice were injected with saline or lipopolysaccharide (LPS, 10 micrograms/mouse, subcutaneously) and the levels of transcripts for IL-1 alpha, IL-1 beta, IL-1ra, IL-6 and tumor necrosis factor-alpha (TNF alpha) were determined 2 h after treatment in the spleen, pituitary, hypothalamus, hippocampus and striatum, using semi-quantitative reverse transcription polymerase chain reaction (RT-PCR). Levels of IL-1 beta were measured by ELISA in plasma and tissues of mice sacrificed after the administration of LPS or saline. LPS induced the expression of pro-inflammatory cytokines at the mRNA level in all tissues under investigation, except for TNF alpha in the hippocampus. This effect was potentiated by adrenalectomy in the spleen for IL-1 alpha and IL-1ra, the pituitary for cytokines other than IL-1ra, the hypothalamus for all cytokines, the hippocampus for cytokines other than TNF alpha, and the striatum for IL-1 alpha and IL-6. In saline-treated mice, adrenalectomy increased IL-1 alpha and IL-1 beta gene expression in the hypothalamus and IL-1 alpha gene expression in the hippocampus and striatum. LPS increased plasma and tissue levels of IL-1 beta, as determined by ELISA, and this effect was potentiated by adrenalectomy in plasma and tissues other than the spleen. These results can be interpreted to suggest that endogenous glucocorticoids regulate the neural components of the host response to infection and inflammation by inhibiting cytokine expression in peripheral organs and the brain.

Expression and localization of p80 and p68 interleukin-1 receptor proteins in the brain of adult mice

The biological effects of interleukin-1 (IL-1) are mediated by two distinct receptors, the p80 type I IL-1 and p68 type II IL-1 receptor proteins (IL-1RI and IL-1RII, respectively), both of which have been recently co-localized to the growth hormone synthesizing cells of the adenohypophysis. Previous studies have shown that IL-1 can bind to specific structures in the central nervous system, but the distribution of IL-1RI and IL-1RII proteins in the adult mouse brain has not been reported. Here we have used immunohistochemistry to study the expression, distribution and cellular localization of both isoforms of the IL-1 receptor proteins in the adult mouse brain. Using a combination of processing techniques (AMeX fixation and cryosectioning), we have immunolabeled brain sections for each isoform of the IL-1R. Both isoforms are expressed in the CNS, particularly in neuronal soma of the granular layer of the dentate gyrus and pyramidal cells of fields CA1-CA4 of Ammons horn of the hippocampus, in epithelial cells of the choroid plexus and ependymal layer, and in neuronal soma of Purkinje cells of the cerebellum. The IL-1RII isoform, but not IL-1RI, is expressed in specific neuronal soma and proximal cell processes of neurons of the paraventricular gray matter of the hypothalamus. These immunohistochemical data directly demonstrate the neuronal expression of both IL-1R proteins in situ. The distribution and cellular localization of IL-1R proteins in the CNS provide a molecular basis for understanding reciprocal interactions between the immune system and the brain.

Mechanisms of the Behavioural Effects of Cytokines

Sickness behavior refers to the coordinated set of behavioral changes that develop in sick individuals during the course of an infection. A sick individual typically displays depressed locomotor activity and little or no interest in his physical and social environment. Body care activities are usually absent and ingestive behavior is profoundly depressed despite the increased metabolism that is necessary for the fever response. Since the initial demonstration in the late eighties that similar symptoms are induced in healthy subjects by peripheral and central injection of the proinflammatory cytokines that are released by activated monocytes and macrophages during the host response to infection, the mechanisms of cytokine-induced sickness behaviour have been the subject of intense research, carried out at several levels of investigation. The purpose of the present chapter is to review the results that have been obtained in this field during the last decade. At the behavioural level, there is now clear evidence showing that sickness behaviour is not the result of weakness and physical debilitation affecting the sick individual, but the expression of a central motivational state that reorganizes the organism’s priorities to cope with pathogenic microorganisms. At the organ level, this motivational aspect of sickness behavior is important since it implies that the endogenous signals of sickness are likely to act on the brain to activate a set of neural structures that are at the origin of the subjective, behavioural and physiological components of sickness. The presence of cytokine receptors in the brain is in accordance with this view.

Nuclear factor κB nuclear translocation as a crucial marker of brain response to interleukin-1. A study in rat and interleukin-1 type I deficient mouse

The signalling pathways that mediate early central effects of interleukin‐1 (IL‐1) during the acute phase reaction have been poorly elucidated. Interaction of IL‐1β to its specific receptor interleukin‐1 receptor type I (IL‐1RI) leads to nuclear factor kappa B (ΝFκB) nuclear translocation and a robust transcriptional activation of inhibitor of kappa B alpha (IκBα) within the rat brain. Indeed, we demonstrated that IL‐1RI expressed in blood brain barrier (BBB) cells and in circumventricular organs (CVOs) is crucial for p65‐NFκB translocation induced by peripheral injection of IL‐1β. Moreover, it has been previously shown that monitoring IκBα mRNA synthesis is an effective tool to investigate the activity of the transcription factor NFκB into the CNS. However in the present study we observed time‐related and cell‐type differences between IκBα mRNA synthesis and p65‐NFκB translocation. This indicates that the expression of IκBα mRNA does not strictly parallel p65‐NFκB nuclear translocation, suggesting that these markers are not interchangeable to investigate NFκB activity but must be studied together. Thus, we hypothesize that IL‐1β reached the brain across the CVOs that lack a BBB and endothelial cells all over the brain and interacted with its receptors to induce NFκB translocation. The study of the consequences of the impairment of NFκB pathway activation in in vivo experimentation should bring important clues about the precise role of this transcription factor.