Lipopolysaccharide-induced microglial activation in culture: temporal profiles of morphological change and release of cytokines and nitric oxide

Nakamura, Yoichi; Si, Qiu-Shang; Kataoka, Kiyoshi

doi:10.1016/s0168-0102(99)00071-1

Cited by 190 publications

(149 citation statements)

References 14 publications

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“…First, pure microglia cultures were stimulated with a range of LPS concentrations (1-1000 ng/ml) for 3-24 h to determine an appropriate concentration and duration of treatment. Consistent with other studies (Nakamura et al, 1999;Dijkstra et al, 2001), 10 ng/ml LPS was adequate and produced a detectable increase in NO within 3 h. Because there was no additional increase after 12 h, this time point was used for subsequent experiments (Fig. 6 D).…”

Section: Kv13-mediated Neuron Killing Involves the Respiratory Burstsupporting

confidence: 78%

Microglia Kv1.3 Channels Contribute to Their Ability to Kill Neurons

Fordyce¹,

Jagasia²,

Zhu³

et al. 2005

J. Neurosci.

190

189

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Many CNS disorders involve an inflammatory response that is orchestrated by cells of the innate immune system: macrophages, neutrophils, and microglia (the endogenous CNS immune cell). Hence, there is considerable interest in anti-inflammatory strategies that target these cells. Microglia express Kv1.3 (KCNA3) channels, which we showed previously are important for their proliferation and the NADPH-mediated respiratory burst. Here, we demonstrate the potential for targeting Kv1.3 channels to control CNS inflammation. Rat microglia express Kv1.2, Kv1.3, and Kv1.5 transcripts and protein, but only a Kv1.3 current was detected. When microglia were activated with lipopolysaccharide or a phorbol ester, only the Kv1.3 transcript (but not protein) expression changed. Using a Transwell cell-culture system that allows separate drug treatment of microglia or neurons, we found that activated microglia killed postnatal hippocampal neurons through a process that requires Kv1.3 channel activity in microglia but not in neurons. A major neurotoxic molecule in this model was peroxynitrite, which is formed from superoxide and nitric oxide; thus, it is significant that Kv1.3 channel blockers reduced the respiratory burst, but not nitric oxide production, by the activated microglia. In addressing the biochemical pathway affected by Kv1.3 channel activity, we found that Kv1.3 acts via a different cellular mechanism from the broad-spectrum drug minocycline, which is often used in animal models of neuroinflammation. That is, the dose-dependent reduction in neuron killing by minocycline corresponded with a reduction in p38 mitogen-activated protein kinase activation in microglia; however, none of the Kv1.3 blockers affected p38 activation.

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Section: Kv13-mediated Neuron Killing Involves the Respiratory Burstsupporting

confidence: 78%

Microglia Kv1.3 Channels Contribute to Their Ability to Kill Neurons

Fordyce¹,

Jagasia²,

Zhu³

et al. 2005

J. Neurosci.

190

189

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“…First, tumor necrosis factor (TNF) a was released with 1-h lag time, second interleukin (IL)-1b within 3 h and, third IL-6. Finally, NO is released with about a 6-h lag time, and linear NO production continues for more than 48 h. 13) A marked morphologic change is also observed: upon the addition of LPS, a change from an amoeboid to a bipolar rod shape occurs for 3 to 6 h and then returns gradually to a small round shape. 13) Before the release of cytokines, various cellular changes occur upon LPS addition.…”

Section: Microglial Activation In Various Neuro-logical Diseasesmentioning

confidence: 97%

“…Finally, NO is released with about a 6-h lag time, and linear NO production continues for more than 48 h. 13) A marked morphologic change is also observed: upon the addition of LPS, a change from an amoeboid to a bipolar rod shape occurs for 3 to 6 h and then returns gradually to a small round shape. 13) Before the release of cytokines, various cellular changes occur upon LPS addition. The mRNA level of myristoylated alanine-rich C kinase substrate (MARCKS) and MARCKSrelated protein (MRP) increase with the increase in protein kinase C (PKC).…”

Section: Microglial Activation In Various Neuro-logical Diseasesmentioning

confidence: 97%

Regulating Factors for Microglial Activation.

Nakamura

2002

Biological & Pharmaceutical Bulletin

222

114

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“…Microglia generally exhibit two distinct types of morphology categorized as ramified and ameboid and which, in a simplified designation, may represent inactivated and activated states, respectively (Kreutzberg, 1996;Nakamura et al, 1999). During the course of the RT-PCR studies described above, it was apparent that pharmacological manipulation of purinergic receptors with chronic ATP treatment had effects to alter morphology of cultured human microglia.…”

Section: Effects Of Purinergic Agents On Morphology Of Microgliamentioning

confidence: 99%

Differential activation of subtype purinergic receptors modulates Ca²⁺ mobilization and COX‐2 in human microglia

Choi

Hong

Ryu

et al. 2003

Glia

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KEY WORDShuman microglia; adenosine triphosphate; P 2Y and P 2X receptors; COX-2, store-operated channel Ca 2ϩ entry ABSTRACTWe have studied modulation of purinergic receptors (P 2Y and P 2X subtypes) on changes in intracellular Ca 2ϩ [Ca 2ϩ ] i and expression and production of COX-2 in human microglia. Measurements using Ca 2ϩ -sensitive spectrofluorometry showed adenosine triphosphate (ATP) to cause rapid transient increases in [Ca 2ϩ ] i . Application of ATP plus the P 2X antagonist, pyridoxal-phosphate-6-azophenyl-2Ј,4Ј-disulfonic acid (PPADS), or treatment with adenosine diphosphate-␤-S (ADP-␤-S), a selective P 2Y agonist, led to a considerable prolongation in [Ca 2ϩ ] i responses compared with ATP. The prolonged time courses were consistent with sustained activation of store-operated channels (SOC) since SKF96365, an inhibitor of SOC, blocked this component of the response. RT-PCR data showed that microglia expressed no COX-2 either constitutively or following treatment of cells with ATP (100 M for 8 h). However, treatment using ATP plus PPADS or with ADP-␤-S led to marked expression of COX-2. The enhanced COX-2 with ATP plus PPADS treatment was absent in the presence of SKF96365 or using Ca 2ϩ -free solution. Immunocytochemistry, using a specific anti-COX-2 antibody, also revealed a pattern of purinergic modulation whereby lack of P 2X activation enhanced the production of COX-2 protein. These results suggest that modulation of subtypes of purinergic receptors regulates COX-2 in human microglia with a link involving SOCmediated influx of Ca 2ϩ .

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Lipopolysaccharide-induced microglial activation in culture: temporal profiles of morphological change and release of cytokines and nitric oxide

Cited by 190 publications

References 14 publications

Microglia Kv1.3 Channels Contribute to Their Ability to Kill Neurons

Microglia Kv1.3 Channels Contribute to Their Ability to Kill Neurons

Regulating Factors for Microglial Activation.

Differential activation of subtype purinergic receptors modulates Ca²⁺ mobilization and COX‐2 in human microglia

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Lipopolysaccharide-induced microglial activation in culture: temporal profiles of morphological change and release of cytokines and nitric oxide

Cited by 190 publications

References 14 publications

Microglia Kv1.3 Channels Contribute to Their Ability to Kill Neurons

Microglia Kv1.3 Channels Contribute to Their Ability to Kill Neurons

Regulating Factors for Microglial Activation.

Differential activation of subtype purinergic receptors modulates Ca2+ mobilization and COX‐2 in human microglia

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Product

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Differential activation of subtype purinergic receptors modulates Ca²⁺ mobilization and COX‐2 in human microglia