Inflammation in the brain has been increasingly associated with the development of a number of neurological diseases. The hallmark of neuroinflammation is the activation of microglia, the resident brain immune cells. Injection of bacterial endotoxin lipopolysaccharide (LPS) into the hippocampus, cortex, or substantia nigra of adult rats produced neurodegeneration only in the substantia nigra. Although LPS appeared to impact upon mesencephalic neurons in general, an extensive loss of dopaminergic neurons was observed. Analysis of the abundance of microglia revealed that the substantia nigra had the highest density of microglia. When mixed neuron-glia cultures derived from the rat hippocampus, cortex, or mesencephalon were treated with LPS, mesencephalic cultures became sensitive to LPS at a concentration as low as 10 ng/ml and responded in a dose-dependent manner with the production of inflammatory factors and a loss of dopaminergic and other neurons. In contrast, hippocampal or cortical cultures remained insensitive to LPS treatment at concentrations as high as 10 g/ml. Consistent with in vivo observations, mesencephalic cultures had fourfold to eightfold more microglia than cultures from other regions. The positive correlation between abundance of microglia and sensitivity to LPS-induced neurotoxicity was further supported by the observation that supplementation with enriched microglia derived from mesencephalon or cortex rendered LPS-insensitive cortical neuron-glia cultures sensitive to LPS-induced neurotoxicity. These data indicate that the region-specific differential susceptibility of neurons to LPS is attributable to differences in the number of microglia present within the system and may reflect levels of inflammation-related factors produced by these cells.

Increasing evidence has suggested that inflammation in the brain is closely associated with the pathogenesis of several degenerative neurologic disorders, including Parkinson's disease, Alzheimer's diseases, multiple sclerosis, amyotrophic lateral sclerosis, and AIDS dementia. The hallmark of brain inflammation is the activation of glial cells, especially that of microglia that produce a variety of proinflammatory and neurotoxic factors, including cytokines, fatty acid metabolites, free radicals--such as nitric oxide (NO) and superoxide. Excessive production of NO, as a consequence of nitric oxide synthase induction in activated glia, has been attributed to participate in neurodegeneration. Using primary mixed neuron-glia cultures and glia-enriched cultures prepared from embryonic rodent brain tissues, we have systemically studied the relationship between the production of NO and neurodegeneration in response to stimulation by the inflammagen lipopolysaccharide. This review summarizes our recent findings on the kinetics of NO generation, the relative contribution of microglia and astrocytes to NO accumulation, the relationship between NO production and neurodegeneration, and points of intervention along the pathways associated with NO generation to achieve neuroprotection. We also describe our results relating to the effect of several opioid-related agents on microglial activation and neuroprotection. Among these agents, the opioid receptor antagonist naloxone, especially its non-opioid enantiomer (+)-naloxone, promises to be of potential therapeutic value for the treatment of inflammation-related diseases.

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Immune stimulants, such as the bacterial endotoxin, lipopolysaccharide (LPS), the human immunodeficiency virus-1 coat protein gp120, or beta-amyloid peptides, lead to glial activation and production of various immune mediators, such as nitric oxide (NO) and proinflammatory cytokines in the brain. These mediators appear to contribute to neuronal cell death in neurodegenerative diseases. However, the signaling pathways, which mediate the neurotoxic effect by the endotoxin, are not understood. The purpose of this study was to determine the role of mitogen-activated protein kinase (MAPK) in LPS-induced neurodegeneration using mesencephalic dopaminergic neuron/glia cultures. We have found that the p38 MAPK is important in LPS-induced death of mesencephalic neurons in rat neuron-glia mixed cultures. Upon treatment with 10 ng/ml LPS, the number of dopaminergic neurons decreased by 80% within 48 h, preceded by a significant production of NO by glia. Neuroprotection by selective inhibition of p38 MAPK activity paralleled a decrease in LPS-induced inducible nitric oxide synthase (iNOS) expression. These events were significantly reduced by the selective p38 MAPK inhibitor, SB202190, but not by the inactive analogue SB202474. Inhibition of iNOS activity and NO production by treatment with GW274150 was also neuroprotective. Although the p38 MAPK inhibitor afforded significant neuroprotection from LPS toxicity in the neuron-glia mixed culture, it failed to protect dopaminergic neurons from 6-hydroxy-dopamine-induced toxicity, which acts directly on dopaminergic neurons by inducing hydroxyl radical formation from the mitochondria. The results suggest that p38 MAPK in glia plays a significant role in the LPS-induced death of mesencephalic neurons through induction of nitric oxide synthase and resulting NO production.

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A new phenylbutenoid dimer, (+/-)- trans-3-(4-hydroxy-3-methoxyphenyl)-4-[(E)-3,4-dimethoxystyryl]cyclohex-1-ene, was isolated from the rhizomes of Zingiber cassumunar along with the three known compounds, (+/-)- trans-3-(3,4-dimethoxyphenyl)-4-[(E)-3,4-dimethoxystyryl]cyclohex-1-ene, 4-(3,4-dimethoxyphenyl)but-1,3-diene, and 4-(2,4,5-trimethoxyphenyl)but-1,3-diene by bioassay-directed fractionation using the A549 human cancer cell line cytotoxicity assay. Structure of the new compound was elucidated by spectral analysis, including 1D and 2D NMR experiments.

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