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.