Involvement of P2X and P2Y receptors in microglial activation in vivo
H Franke1, C Schepper, P Illes
1Rudolf Boehm Institute of Pharmacology and Toxicology, University of Leipzig, Härtelstr. 16-18, D-04107, Leipzig, Germany, frah@medizin.uni-leipzig.de.
Abstract:
Microglial cells are the primary immune effector cells in the brain. Extracellular ATP, e.g., released after brain injury, may initiate microglial activation via stimulation of purinergic receptors. In the rat nucleus accumbens (NAc), the involvement of P2X and P2Y receptors in the generation of microglial reaction in vivo was investigated. A stab wound in the NAc increased immunoreactivity (IR) for P2X(1,2,4,7) and P2Y(1,2,4,6,12) receptors on microglial cells when visualized with confocal laser scanning microscopy. A prominent immunolabeling of P2X(7) receptors with antibodies directed against the ecto- or endodomain was found on Griffonia simplicifolia isolectin-B4-positive cells. Additionally, the P2X(7) receptor was colocalized with active caspase 3 but not with the anti-apoptotic marker pAkt. Four days after local application of the agonists alpha,betameATP, ADPbetaS, 2MeSATP, and BzATP, an increase in OX 42- and G. simplicifolia isolectin-IR was observed around the stab wound, quantified both densitometrically and by counting the number of ramified and activated microglial cells, whereas UTPgammaS appeared to be ineffective. The P2 receptor antagonists PPADS and BBG decreased the injury-induced increase of these IRs when given alone and in addition inhibited the agonist effects. Further, the intra-accumbally applied P2X(7) receptor agonist BzATP induced an increase in the number of caspase-3-positive cells. These results indicate that ATP, acting via different P2X and P2Y receptors, is a signaling molecule in microglial cell activation after injury in vivo. The up-regulation of P2X(7)-IR after injury suggests that this receptor is involved in apoptotic rather than proliferative effects.
Insights
Extracellular ATP activates brain microglial cells via P2X and P2Y receptors following injury. P2X7 receptor signaling is linked to microglial apoptosis after brain damage.
Area of Science:
- Neuroscience
- Immunology
- Cell Biology
Background:
- Microglial cells are the brain's primary immune cells.
- Extracellular ATP, released during brain injury, can activate microglia through purinergic receptors.
Purpose of the Study:
- To investigate the role of P2X and P2Y receptors in microglial activation in the rat nucleus accumbens (NAc) following stab wound injury.
- To determine the specific P2 receptor subtypes involved and their downstream effects, including apoptosis.
Main Methods:
- Induction of stab wound injury in the rat NAc.
- Immunohistochemical analysis using confocal laser scanning microscopy to visualize P2 receptor subtypes (P2X1,2,4,7 and P2Y1,2,4,6,12) on microglial cells.
- Application of P2 receptor agonists and antagonists.
- Quantification of microglial activation markers (OX42, Griffonia simplicifolia isolectin-B4) and apoptotic markers (caspase-3).
Main Results:
- Stab wound injury increased immunoreactivity for multiple P2X and P2Y receptors on microglia.
- The P2X7 receptor was prominently expressed on microglial cells and colocalized with active caspase-3, indicating involvement in apoptosis.
- P2 receptor agonists increased microglial activation, while antagonists reduced injury-induced activation.
- The P2X7 receptor agonist BzATP increased the number of caspase-3 positive cells.
Conclusions:
- ATP acts as a signaling molecule mediating microglial activation in the brain after injury via P2X and P2Y receptors.
- The upregulation of P2X7 receptor expression post-injury suggests its role in microglial apoptotic processes rather than proliferation.
