Overstimulation of NMDA type of glutamate receptors is accompanied by calcium influx that initiates a cascade of events involving mitochondrial dysfunction, activation of enzymes associated with the release and oxidation of arachidonic acid (ARA) , and generation of reactive oxygen species (Farooqui and Horrocks 1994; Farooqui and Horrocks 2006). These enzymes include isoforms of PLA₂ , cyclooxygenase-2 (COX-2) , lipoxygenases (LOX) , and epoxygenases (EPOX) (Phillis et al. 2006) (Fig. 4.3). An uncontrolled and sustained increase in cytosolic calcium levels also contributes to the stimulation. calpains, nitric oxide synthase, protein phosphatases, and various protein kinases (Arundine and Tymianski 2004; Farooqui et al. 2008). Accumulation of oxygenated ARA metabolites and elevation in platelet activating factor (PAF) along with abnormal ion homeostasis, changes in redox status, and lack of energy generation is associated with neural cell injury and cell death in acute neural trauma (ischemia, epilepsy, head injury, and spinal cord trauma) and neurodegenerative diseases such as Alzheimer disease (AD), and Parkinson disease (PD) (Farooqui et al. 2007; Farooqui et al. 2008) .

Stimulation of NMDA receptors increases the generation of prostaglandins, leukotrienes, and thromboxanes from ARA. These metabolites are collectively called as eicosanoids (Phillis et al. 2006). Neurons and glia generate prostaglandins, whereas cerebral microvessels and the choroid plexus mainly produce thromboxanes. Eicosanoids act through their receptors and modulate signal transduction pathways and gene transcription. Thus, PGD₂ activates the Prostaglandin D₂ receptor (DP receptors), PGE₂ activates the PGE2-activated receptors (EP receptors), and PGF_2α, PGI₂, and TXA₂, respectively, stimulate the Prostaglandin F2α receptor, PGI2 Receptor and Thromboxane receptor (FP, IP, and TP receptors) (Coleman et al. 1994; Phillis et al. 2006). These receptors are linked to the generation of cyclic AMP, diacylglycerol, and phosphatidylinositol 1,4,5-trisphosphate and modulation of Ca²⁺ influx. Availability of glutamate in the synaptic cleft increases neuronal excitability and synaptic transmission at the presynaptic level whereas the uptake of glutamate by astrocytes prevents glutamate accumulation in the synaptic cleft and retards glutamate-mediated neurotoxicity. Thus, interplay among glutamate, prostaglandin, leukotriene, and thromboxane receptors in the brain may modulate neuronal cell survival. Under normal conditions, this cross talk refines the communication among receptors, but under pathological situations, the interplay among prostaglandin, leukotriene, and thromboxane receptors may promote neuronal injury, which depends upon the levels of glutamate, magnitude of PLA₂ expression, levels of eicosanoids and the intensity of neuroinflammation and oxidative stress (Farooqui and Horrocks 2006; Farooqui et al. 2008). In addition, glutathione, a tripeptide involved in maintaining redox status binds to its own receptors and modulates glutamatergic excitatory neurotransmission by displacing glutamate from NMDA receptors (Janaky et al. 1999) resulting in reduction of reduced glutathione. Collective evidence suggests that in neural cells, glutamate-mediated modulation of eicosanoid generation, activities of glutamate transports, depletion of glutathione, and decreased ATP levels are closely associated with neuroinflammation and oxidative stress-mediated neuronal cell death (Gilroy et al. 2004; Manev et al. 2000) .

Toll-like receptors (TLRs) are a family of pattern recognition receptors that are characterized by an extracellular leucine-rich repeat domain and an intracellular Toll/IL-1 receptor (TIR) domain. TLRs recognize a variety of highly conserved structural motifs expressed by microbial pathogens, called pathogen-associated molecular patterns (PAMPs) (Kaisho and Akira 2004). Mammals contain at least ten TLRs. Despite a high degree of structural similarity, each receptor has a distinct function in innate immune recognition. Each TLR has some degree of ligand specificity, which specificity is extended through dimerization of TLRs. Some TLRs employ additional co-receptors that assist in pathogen recognition, such as CD14 for TLR4 (Kielian 2006). In the brain tissue, Toll-like receptors (TLRs) are mainly expressed in microglial cells (Kettenmann et al. 2011) . TLRs are essential for mounting an immune response against infection. Thus, TLRs serve as pathogen-associated molecular pattern recognition receptors that bind microbial molecular motif with high affinity, and play a central role in the initiation of cellular innate immune responses (Aravalli et al. 2007; Kawai and Akira 2010). The stimulation of TLRs in microglial cells triggers the release of several cytokines and chemokines (Aravalli et al. 2007; Okun et al. 2009). Thus, detailed investigations have indicated that the stimulation of TLR3 causes the release of IL-6, IL-12, chemokine ligand 10, TNF-α , and IFNβ (Alexopoulou et al. 2001; Jack et al. 2005); TLR2 modulates the secretion of IL-6 and IL-10 (Jack et al. 2005); TLR9 regulates the production of NO and TNF-α (Iliev et al. 2004), and stimulation of TLR4 triggers the release of IL-6 and TNF-α (Okun et al. 2009). Most of above mentioned cytokines and their receptors (see below) are associated with the release of ARA and ceramide from neural membrane phospholipids and sphingolipid through the activation of PLA₂s and SMases leading to the generation of proinflammatory lipid mediators (eicosanoids, platelet activating factor, and ceramide) (Farooqui 2011). As stated above, the cytoplasmic domains of the TLRs share a high degree of homology with the intracellular portion of the IL-1R (Takeda et al. 2003). TLR activation results in the recruitment of the adaptor protein MyD88, which is associated with the serine/threonine kinase IL-1R-associated kinase (IRAK). IRAK then interacts with the tumor necrosis factor (TNF) receptor-associated factor (TRAF) adaptor protein TRAF6, which provides a bridge to the protein kinase NF-κB-inducing kinase (NIK) (Fig. 4.4). NIK then phosphorylates IKK (IκB kinase), resulting in the phosphorylation of IκB. As stated above, IκB phosphorylation targets the protein for ubiquitination and proteasome-mediated degradation, resulting in the release and nuclear translocation of NF-κB, where it can influence the expression of numerous immune response genes (Kielian 2006) .

Eicosanoids are released outside of the cells immediately after their synthesis. They act through eicosanoids receptors that are located on plasma and nuclear membranes. Eicosanoid receptors include EP₁, EP₂, EP₃, and EP₄. These receptors are encoded by different genes and differ in their responses to various agonists and antagonists. Among eicosanoid receptors, EP₂ activation is involved in microglial-mediated paracrine neurotoxicity as well as suppression of microglia internalization of aggregated neurotoxic peptides along with neuroinflammation (Coleman et al. 1994; Cimino et al. 2008). In addition, brain also contains PGF receptor (FP), PGI receptor (IP), and TXA receptor (TP), which may contribute to neurotransmitter release, sleep, and vasodialation and vasoconstriction of cerebral vessels in the brain .

RAGE is a type I transmembrane protein composed of three extracellular immunoglobulin-like domains (V, C1, and C2), a single transmembrane domain and a short cytoplasmic tail thought to be important in signal transduction (Hudson et al. 2008). RAGEs are expressed in the brain in neurons, microglia, and astrocytes (Lue et al. 2001; Sasaki et al. 2001; Yan et al. 1996). Aβ is a specific ligand for RAGE, which interacts with the N-terminal domain of RAGE (Chaney et al. 2005). RAGE expression is elevated in AD pathology-enriched brain regions, including hippocampus and inferior frontal cortex, when compared to cerebellum where AD pathology is limited. The interactions of RAGE with its diverse ligands (AGEs, Aβ, amphoterin and S100B protein) mediate multiple physiological and pathological functions, including inflammation, oxidative stress, neurodegeneration, maintenance of homeostasis, tumorigenesis, promotion of neurite outgrowth, cell survival, cell migration, and neuronal differentiation (Ramasamy et al. 2005; Meneghini et al. 2010). The binding of AGEs with RAGE not only results in the activation of the mitogen-activated protein kinases (MAPKs) and the phosphatidylinositol-3 kinase (PtdIns 3K) pathways, but also in activation of NADPH oxidases, generation of superoxide, and activation of NF-κB. This transcription factor migrates to the nucleus and induces the induction of inflammatory cytokines and adhesion molecule 1 (Ramasamy et al. 2005; Meneghini et al. 2010) .

NF-κB is a family of transcription factors that regulates multiple cellular functions associated with immune responses, neuroinflammation , apoptosis, cell growth and survival (Celec 2004). The mammalian NF-κB family members include NF-κB1 (p105 and p50), NF-κB2 (p100 and p52), RelA (p65), RelB and c-Rel, which is activated by upstream signals from diverse immune receptors, such as ligand-triggered Toll-like receptors (TLRs) , interleukin-1 receptor (IL-1R), tumor necrosis factor receptor (TNF-R) (see below) and antigen receptors (Hayden and Ghosh 2004; Rahman et al. 2009). The activation of NF-κB is tightly regulated by inhibitors of NF-κB (IκBs). These proteins are also involved in the regulation of these diverse cellular processes, such as neuroinflammation, apoptosis, cell growth and survival (Baeuerle and Baltimore 1988). In unstimulated cells, NF-κB proteins are localized in an inactive in the cytoplasm, usually as homodimers or as heterodimers with RelA, RelB or c-Rel and complexed with the inhibitory IκBs (Figs. 4.5 and 4.6). The IκBs are in turn regulated by another group of regulatory proteins called IκB kinases (IKKs). The IKK complex is composed of two catalysis subunits (IKKα and IKKβ) and a regulatory subunit IKKγ, or NEMO (NF-κB essential modulator) (Perkins 2007). Upon ligand engagement, the receptors initiate signal transduction events that lead to the activation of IKK complex by activated cellular protein kinases like NF-κB-inducing kinase (NIK), mitogen-activated protein/extracellular signal-regulated kinase1 (MEKK1), transforming growth factor-β (TGFβ)- activated kinase 1 (TAK1), MEKK2 or MEKK3. Activated IKK phosphorylates IκBα using specific serine residue within the IκBα proteins, triggering their ubiquitination via ubiquitin ligase (Traencker et al. 1995) . The IκBα protein is then degraded by the 26S proteasome, allowing the release and translocation of the active NF-κB dimer into the nucleus, where it binds to target sequences in the genome and facilitates the expression of a number of proteins including many enzymes (sPLA₂, COX-2, NADPH oxidase and inducible nitric oxide synthase) and cytokines (TNF-α, IL-1β, and IL-6) (Figs. 4.1and 4.2) (O’Neill 2003; Karin and Ben-Neriah 2000). Collective evidence suggests that NF-κB can be activated by distinct mechanisms. These include microbial invasion recognized by TLR, generation of ROS, cellular generation of inflammatory eicosanoids , and interaction with inflammatory cytokines via defined cell surface receptors (see below). It is also known that several of these initiating events are modulated by dietary factors. This also means that appropriate use of the diet can either turn on or turn off the activation of NF-κB. This new knowledge is the foundation of anti-inflammatory nutrition (Sears 2008; Sears and Riccordi 2011) . Antioxidant, M40403, not only blocks excitotoxicity, but also prevents NF-κB translocation to the nucleus (McInnis et al. 2002). NMDA receptor-mediated activation of NF-κB involves ΙκB-α degradation by a caspase-3-like cysteine protease. In addition to NF-κB, other transcription factors such as AP-1, STATs, HIF-1, Egr-1, and IRF1also contribute to neuroinflammation (Kwon et. al. 2004; Kim et al. 2002; Wang et al. 2008; Zhang et al. 2006; Friedle et al. 2011; Iadecola et al. 1999). In contrast, transcription factors such as PPARs or Nrf2 play an anti-inflammatory role in glial activation (Drew et al. 2006; Lee et al. 2005; Shih et al. 2003). In addition, CCAAT/enhancer binding protein β (C/EBPβ), a group of the C/EBP subfamily of bZIP transcription factors also contributes to proinflammatory gene expression during glial cell activation. At least three N-terminally truncated isoforms of C/EBPβ are known to occur in various tissues (Ossipow et al. 1993; Welm et al. 1999). C/EBPβ not only plays an important role in inflammation, but also functions in modulation of cell energy metabolism, cell proliferation and differentiation (Ramji and Foka 2002). It is becoming increasingly evident that promoters of many pro-inflammatory genes contain putative C/EBPβ consensus sequences (Reddy et al. 2000) and C/EBPβ levels are upregulated in response to pro-inflammatory stimuli in macrophages (Bradley et al. 2003) and glial cells (Cardinaux et al. 2000). It is interesting to note that C/EBPβ deficiency provides neuroprotection following ischemic (Kapadia et al. 2006) and excitotoxic injuries (Cortes-Canteli et al. 2008) .

Cytokines and chemokines are proteins and peptides produced and released by different cells, for example: leukocytes, muscle cells, and neurons . These proteins act in a pleiotropic way or in synergy with other substances and can modulate the production of other cytokines. Structural studies have allowed the classification of these proteins into different structural classes such as the helical cytokines, the trimeric tumor necrosis factor (TNF) family, the cysteine knot growth factors and the β-trefoil growth factors (Farooqui and Horrocks 2007). Cytokines can also be classified according to the type of receptor they bind, which comprise six major families: class I cytokine receptors (the largest family), class II cytokine receptors, TNF receptors, tyrosine kinase receptors, and chemokine receptors (Tansey and Wyss-Coray 2008). In addition, cytokine families can also be named differently according to other aspects such as the sharing of a receptor subunit (i.e. the gp130 family) or its physiological roles (i.e. neuropoietic family, for its effects on hematopoietic and nervous system).

In brain tissue, cytokines (TNF-α, IL-1β, and IFN-γ) and chemokines (MCP1and CXCL3) mediate cellular intercommunication through autocrine (self), paracrine (local) or endocrine (distant) mechanisms (Tansey and Wyss-Coray 2008). Their actions involve a complex network linked to feedback loops and cascades. Cytokines and chemokines mediate their effects by binding to specific membrane-associated receptors that are composed of an extracellular ligand-binding region, a membrane-spanning region, and an intracellular region that is activated by binding of cytokines, and chemokines that facilitate the delivery of a signal to the nucleus (Rothwell and Relton 1993). Cytokine and chemokine-mediated signal transduction processes are extremely complex . These processes depend on several factors, such as the target cell type and state of activation, the local cytokine concentration, receptor type and interaction with other cytokine. Due to these factors, the precise effect of cytokines on neurons, astrocytes, oligodendrocytes, and microglial cells is difficult to predict. However, many studies have indicated that the magnitude and persistence elevations in cytokine and chemokine levels may be related to the intensity of neuroinflammation (Farooqui et al. 2007; Farooqui 2010a). Aging is accompanied by enhanced basal expression of proinflammatory cytokines and these same proinflammatory mediators often are associated with neural injury-related activation of microglia and astroglia . Taken together, the aging, inflammation and glial activation phenotypes serve as the basis for the “inflam-aging” hypothesis (Salvioli et al. 2006; Franceschi et al. 2007; Giunta 2008). According to this hypothesis, increased inflammation during the aging process results from dysregulation of the immune system and a progressive inability to properly handle pathological stimuli (Giunta 2008). Studies on aging confirm that proinflammatory cytokines in the IL-6 and TNF-α families are participants in the complex relationship between aging and chronic morbidity (Giunta 2008). Through JAK2/STAT3 signaling, these cytokines activate target genes involved in immune responses, differentiation, survival, apoptosis, and proliferation. Activation of the JAK2/STAT3 pathway is not only associated with trauma, but also with stroke (Yamashita et al. 2005; Xie et al. 2007; Shyu et al. 2008). Accumulating evidence suggests that aging may affect the proinflammatory and astrocytic response to stroke .