Dopamine and noradrenaline control distinct functions in rodent microglial cells

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Abstract

Microglial cells are the immune-competent elements of the brain. They not only express receptors for chemokines and cytokines but also for neurotransmitters such as GABA [Charles et al., Mol. Cell Neurosci. 24 (2003)214], glutamate [(Noda et al., J. Neurosci. 20 (2000) 251], and adrenaline [Mori et al., Neuropharmacology 43 (2002) 1026]. Here we report the functional expression of dopamine receptors in mouse and rat microglia, in culture and brain slices. Using the patch clamp technique as the functional assay we identified D1- and D2-like dopamine receptors using subtype-specific ligands. They triggered the inhibition of the constitutive potassium inward rectifier and activated potassium outward currents in a subpopulation of microglia. Chronic dopamine receptor stimulation enhanced migratory activity and attenuated the lipopolysaccharide (LPS)-induced nitric oxide (NO) release similar as by stimulation of adrenergic receptors. While, however, noradrenaline attenuated the LPS-induced release of TNF-alpha and IL-6, dopamine was ineffective in modulating this response. We conclude that microglia express dopamine receptors which are distinct in function from adrenergic receptors.

Introduction

Microglial cells are the immune competent elements of the brain and express a large variety of receptors for cytokines, chemokines, and growth factors (Wesemann and Benveniste, 2004). Moreover, they can release a variety of immunomodulators upon their activation by any type of injury or pathologic process and thereby communicate with intrinsic brain cells and invading immune cells. There is some evidence that this communication is mediated via soluble factors such as chemokines, cytokines, and neurotransmitters. It was proven that microglial cells express receptors for the classical neurotransmitters such as glutamate (Noda et al., 2000) and GABA (Kuhn et al., 2004). The presence of adrenergic receptors on microglia was established by Chang and Liu (2000), Colton and Chernyshev (1996), Fujita et al. (1998) and Thery et al. (1994).

Neurotransmitters seem to control glial immune function because activation of GABA (B) receptors affected the release of interleukin-6 (IL-6) and interleukin-12+p40 (IL-12+p40) after activation, induced experimentally by the bacterial endotoxin lipopolysaccharide (LPS) (Kuhn et al., 2004). A similar down regulation of LPS-induced cytokine release was observed by chronic metabotropic purinergic receptor activation (Boucsein et al., 2003). In contrast, activation of glutamate receptors of the AMPA subtype enhanced the production of the cytokine TNF-alpha (Noda et al., 2000). Noradrenaline and a β2 adrenergic agonist elevated intracellular cAMP levels (Mori et al., 2002, Prinz et al., 2001). Salbutamol, an adrenergic agonist dose-dependently inhibited LPS-induced IL-12+p40 release and the involvement of β2 adrenoceptors, was inferred by the observation that a specific antagonist abolished this inhibitory effect (Prinz et al., 2001). β2 adrenoceptors furthermore modulated microglial proliferation (Fujita et al., 1998). In contrast to noradrenaline less is known about the action of dopamine on microglial immune function. Only in peritoneal macrophages it was examined that dopamine decreased the release of IL-12+p40 production but the inhibitory effect was mediated via β adrenoceptors (Hasko et al., 2002). If microglia also express dopamine receptors is unknown, but several peripheral cell types such as lymphocytes and macrophages express dopamine receptors (Ricci et al., 1994, Santambrogio et al., 1993). So it was obvious to look for dopamine receptors on microglial cells.

The present work presents evidence that microglial cells express dopamine receptors and that activation of these receptors modulate microglial properties. Moreover, we compared the dopamine responses with those triggered by noradrenaline and found different features as well as many common effects.

Section snippets

Dopamine and noradrenaline modulate distinct K+ conductances in cultured microglial cells

We used the patch-clamp technique to test for the presence of functional dopaminergic and adrenergic receptors on microglia in culture. To study changes in conductance over time we repetitively clamped the membrane from a holding potential of −70 mV to a series of de- and hyperpolarizing voltage steps and repeated this clamp protocol every 5 s (Fig. 1). This procedure allows us to follow the reversal potential of the response as well as the resting membrane potential.

Dopamine (1 mM) triggered

Evidence for the presence of dopamine and noradrenaline receptors on microglial cells

In the present study we used the patch-clamp technique to pharmacologically identify dopamine and adrenergic receptors in microglial cells. While the functional expression of adrenergic receptors in microglial cells has been reported before (Chang and Liu, 2000, Colton and Chernyshev, 1996, Fujita et al., 1998, Prinz et al., 2001, Thery et al., 1994), we provide the first evidence for the microglial expression of dopamine receptors. While dopamine is known to trigger signaling also via

Microglial cell culture

Microglial cultures were prepared from cerebral cortex of newborn NMRI mice and Wistar rats as described previously (Prinz et al., 1999). In brief, brain tissue was carefully freed of blood vessels and meninges. Cortical tissue was trypsinized for 2 min, dissociated with a fire-polished pipette, and washed twice. Mixed glial cells were cultured for 9–12 days in Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% fetal calf serum (FCS), 2 mM l-glutamine, and antibiotics (100 units/ml

Acknowledgments

We thank Michael Berlin, Irene Haupt, Madlen Driesdner and Ina Lauterbach for excellent technical assistance. This work was supported by grants from the German Research Foundation (DFG Schwerpunkt and SFB507).

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