P2X₇Receptor Expression after Ischemia in the Cerebral Cortex of Rats

Abstract

Large amounts of adenosine 5'-triphosphate (ATP) released from cellular sources under pathological conditions such as ischemia may activate purinoceptors of the P2X and P2Y types. In the present study, the expression of the P2X7 receptor-subtype in the brain cortex of spontaneously hypertensive rats was investigated using a permanent focal cerebral ischemia model. Immunocytochemistry with antibodies raised against the intracellular C-terminus of the P2X7 receptor showed a time-dependent upregulation of labeled cells in the peri-infarct region after right middle cerebral artery occlusion (MCAO) in comparison to controls. Double immunofluorescence visualized with confooal laser scanning microscopy indicated the localization of the P2X7 receptor after ischemia on microglial cells (after 1 and 4 days), on tubulin betaIII-labeled neurons (after 4 and 7 days), and on glial fibrillary acidic protein (GFAP)-positive astrocytes (after 4 days). In the following experiments, changes occurring 4 days after MCAO were investigated in detail. Western blot analysis of the cortical tissue around the area of necrosis indicated an increase in the P2X7 receptor protein. Immunoelectron microscopy revealed the receptor localization on synapses (presynaptically), on dendrites, as well as on the nuclear membrane of neurons (postsynaptically) and glial cells. Terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick end labeling in combination with P2X7 receptor immunocytochemistry indicated a co-expression on the apoptotic cells. Active caspase 3 was especially observed on GFAP-positive astrocytes. In conclusion, the present data demonstrate a postischemic, time-dependent upregulation of the P2X7 receptor-subtype on neurons and glial cells and suggest a role for this receptor in the pathophysiology of cerebral ischemia in vivo.

Physiological and pathological functions of P2X7 receptor in the spinal cord

“…Consistent with this hypothesis an increase in P2X7R immunoreactivity has been observed in activated microglia of two transgenic models of Alzheimer's disease [71,72] in microglia/ macrophages of spinal cord undergoing multiple or amyotrophic lateral sclerosis [73], in ischemic cortical tissue [74], and in a model of kainite-induced seizures [62] suggesting that microglial P2X7R, together with P2X4R, might be a general mediator of stress during pathological states. Besides uptake and release of inflammatory molecules, other pathways might contribute to microglial effects on neuronal excitability and disease outcome.…”

Astroglial P2X7 receptor current density increased following long-term exposure to rotenone

“…P2X7 receptors were primarily localised along astrocyte processes in the hippocampus [14], while P2X7 receptors were observed on the bodies of GFAP-labelled reactive astrocytes in the nucleus accumbens after mechanical damage, but no labelling was detected on the processes [16]. In the cerebral cortex after ischaemia, P2X7 receptor immunoreactivity was equally present on cell bodies and processes of astrocytes [17]. In our study, astrocytes were harvested from the ventral portion of the midbrain.…”

“…Immunocytochemical studies have shown the localisation of P2X7 receptor subunits on astrocytes in various hippocampal subregions [14], in the cerebellum [15], in the nucleus accumbens after mechanical damage [16] and in the cerebral cortex after ischaemia [17]. In the latter two brain regions, P2X7 receptors were localised in GFAPlabelled reactive astrocytes [16,17].…”

P2X7 receptors in cerebral ischemia

“…Immediately after ischemia, the cerebral blood flow dramatically decreases in the ischemic core, leading to rapid energy failure, ATP depletion, ionic imbalance, and neuronal death [1] . The cells in the penumbra (tissues surrounding the ischemic core), however, can retain their activity for a prolonged period.…”

“…Seven distinct ionotropic P2X (P2X [1][2][3][4][5][6][7] and eight distinct metabotropic P2Y receptor subunits (P2Y 1,2,4,6,[11][12][13][14], classified by electrophysiological properties and their amino-acid sequence homology, have been cloned from mammals [4] .…”