Purinergic receptors on microglial cells: functional expression in acute brain slices and modulation of microglial activation in vitro

Abstract

Microglial cells are the pathologic sensors in the brain. ATP released from damaged cells is a candidate for signalling neural injury to microglia. Moreover, ATP is an extracellular messenger for propagating astrocyte activity in the form of Ca2+ waves. To test for the functional expression of purinoreceptors in microglial cells we employed the patch-clamp technique in acute slices of adult mouse brain. ATP triggered a nonselective cationic and a K+ current. Pharmacological screening with purinergic ligands indicated the presence of P2Y1 and P2Y2/4 receptors linked to the activation of a K+ current and P2X receptors, including P2X7, linked to the activation of a nonselective cationic current. These findings suggest that microglial cells in situ express different purinergic receptors with distinct sensitivity and functional coupling. To test for the involvement of purinoreceptors in microglial activation, we stimulated cultured microglial cells with lipopolysaccharide and measured the release of tumour necrosis factor alpha, interleukin-6, interleukin-12 and macrophage inflammatory protein 1alpha, induction of K+ outward currents and nitric oxide release. All these parameters were reduced in the presence of purinergic ligands, indicating that purinergic receptor activation attenuated indicators of microglial activation.

The inflammatory effects of UDP-glucose in N9 microglia are not mediated by P2Y14 receptor activation

“…Previously, we and others have reported that adenine di-and triphosphate nucleotides, the endogenous ligands for P2 receptors, exert anti-inflammatory effects in microglia [3,4]; however, the specific P2 receptors involved in these actions are not yet clear. The P2Y14 receptor, previously called GPR105 (an orphan G-protein coupled receptor), has only recently been identified as a member of the purinergic receptor family [1]; hence, comparatively little is known about its function in any cell type.…”

Hypoxia induced amoeboid microglial cell activation in postnatal rat brain is mediated by ATP receptor P2X4

“…It has been reported that ramified microglia recruitment to the site of ischemia and hypoxia impairment is associated with cellular loss and P2X4-R are involved in cellular activation [19-21]. In this regard, the neuroprotective actions exerted by TNP-ATP and PPADS and the possible use of purinergic antagonist in the pharmacological treatment of oxygen/glucose deprivation have been considered [19-21,31]. Although P2X4 expression and inflammation after hypoxia-ischemia in the rat brain have been investigated after intraperitoneally injection of minocycline [22], the use of selected P2X4 antagonists to suppress AMC activation and associated inflammation in the postnatal brain after hypoxia has not been explored.…”

“…There is ample evidence to suggest that ATP is involved in microglial activation specifically in the ramified phenotype in the adult brain [24,25,31]. On the other hand, the role of ATP in AMC activation especially the mechanisms in hypoxia have remained elusive.…”

P2X7 receptors in cerebral ischemia

“…There are several possible modes of ATP release: (1) damaged neurons are depolarized by the elevated extracellular K + due to impairment of the Na + -K + pump. ATP passively flows out along the electrochemical gradient through membrane channels of glial cells [23] or from the presynaptic vesicles exocytotically [35] ; (2) organelles release ATP from astroglia in a Ca 2+ -independent manner via P2X 7 R channels [25,36] ; and (3) P2X 7 Rs on activated amoeboid microglia and reactive microglia amplify the ATP signal and trigger ATP release through an autocrine process [37,38] . In short, cerebral ischemia results in a sustained progressive elevation of ATP levels in the penumbra [22,23] .…”