Involvement of benzodiazepine receptors in neuroinflammatory and neurodegenerative diseases: evidence from activated microglial cells in vitro

doi:10.1016/j.nbd.2003.07.002

Neurobiology of Disease

Volume 14, Issue 3, December 2003, Pages 417-424

https://doi.org/10.1016/j.nbd.2003.07.002 Get rights and content

Abstract

Increased binding of a ligand for the peripheral benzodiazepine binding receptor is currently used in PET studies as an in vivo measurement of inflammation in diseases like multiple sclerosis and Alzheimer's disease. Although peripheral-type benzodiazepin receptors (PBRs) are abundant in many cell types and expressed in the CNS physiologically only at low levels, previous reports suggest that after experimental lesions in animal models and in human neurodegenerative/-inflammatory diseases upregulated PBR expression with increased binding of its ligand PK11195 is confined mainly to activated microglia in vivo/in situ. Because the functional role of the PBR is unknown, we confirm by immunohistochemistry and PCR (I) that this receptor is expressed on microglia in vitro and (II) that benzodiazepines modulate proliferation of microglial cells and the release of the inflammatory molecules nitric oxide (NO) and tumor necrosis factor-alpha (TNF-α) in cell culture supernatants of primary rat microglia. Compared to lipopolysaccharide-activated controls the release of NO was markedly decreased in cultures treated with benzodiazepines (clonazepam, midazolam, diazepam) and the PBR ligand PK11195. Moreover, release of TNF-α and proliferation was significantly inhibited in the benzodiazepine-treated groups. These findings link the in vivo data of elevated PBR levels in neurodegenerative/-inflammatory diseases to a functional role and opens up possible therapeutic intervention targeting the PBR in microglia.

Section snippets

Cell culture

For all experiments Wistar rats that were bred and held under constant conditions (12h/12h light/dark cycle) in the animal house of the University of Kiel were used Free-floating microglial cells were collected from the medium of primary cell cultures from neonatal rat cerebral cortex after 10 days, as detailed elsewhere Sievers and Parwaresch, 1994, Wilms and Hartmann, 1997, Wilms and Wollmer, 1999: Briefly, free-floating microglia was collected from the medium of primary cell cultures from

Cell morphology and expression of PBR

Within 3 days the seeded microglial cells developed the characteristic ramified shape of resting microglia in vitro with branched cellular processes and a small cell soma (Fig. 1A). After treatment with LPS the microglial cells adopted an amoeboid phenotype characteristic of activated myelomonocytic cells (Fig. 1B). Both resting and activated cells could be immunostained with a specific antibody directed against the PBR (Fig. 1D and E). However, it should be noted that resting microglia (Fig.

Discussion

Peripheral-type benzodiazepine receptors are widely distributed receptors that recognize a subset of benzodiazepine receptors. Although they are expressed in the undamaged CNS at only a low level, it appears that the relative density of PBR is a reliable marker for neuroinflammation and gliosis with neuronal damage (Banati, 2002): Animal studies indicate that neuronal damage experimentally induced by excitoxicity (Altar and Baudry, 1990), ischemia (Stephenson et al., 1995), experimental facial

Acknowledgements

The authors thank Mrs. Rosemarie Sprang for superior technical assistance. This work was supported by a grant from the Hensel-Foundation, University of Kiel (to R.L.) and a generous donation from the “Herbert und Inge Lampe Stiftung” (to H.W. and R.L.).

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¹: H.W. and J.C. contributed equally to this study.

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Regular articleInvolvement of benzodiazepine receptors in neuroinflammatory and neurodegenerative diseases: evidence from activated microglial cells in vitro

Abstract

Section snippets

Cell culture

Cell morphology and expression of PBR

Discussion

Acknowledgements

J. Biol. Chem.

J. Biol. Chem.

Brain Res.

Brain Res.

Neurosci. Lett.

Lancet

Eur. J. Pharmacol.

Neurosci. Lett.

J. Biol. Chem.

Cancer Lett.

Trends Neurosci.

Neurosci. Lett.

Brain Res.

Int. J. Immunopharmacol.

Eur. J. Pharmacol.

J. Biol. Chem.

Neurobiol. Aging

Trends Neurosci.

Neuroscience

Kidney Int.

J. Neuroimmunol.

Diazepam-binding inhibitora neuropeptide located in selected neuronal populations of rat brain

Science

Systemic injection of kainic acidgliosis in olfactory and limbic brain regions quantified with [3H]PK 11195 binding autoradiography

Exp. Neurol.

In vivo regulation of peripheral-type benzodiazepine receptor and glucocorticoid synthesis by Ginkgo biloba extract EGb 761 and isolated ginkgolides

Endocrinology

Visualising microglial activation in vivo

Glia

Cytotoxicity of microglia

Glia

Regular article
Involvement of benzodiazepine receptors in neuroinflammatory and neurodegenerative diseases: evidence from activated microglial cells in vitro

Systemic injection of kainic acidgliosis in olfactory and limbic brain regions quantified with [³H]PK 11195 binding autoradiography