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Updated: Aug 24, 2026
Organotypic Slice Cultures for Studies of Postnatal Neurogenesis
Published on: March 4, 2015
Synaptic P2X7 and oxygen/glucose deprivation in organotypic hippocampal cultures
Fabio Cavaliere1, Susanna Amadio, Giuseppe Sancesario
1Fondazione Santa Lucia, Rome, Italy.
Abstract:
The P2X7 receptor for extracellular ATP is the main candidate, among P2 receptors, inducing cell death in the immune system. Here, we demonstrate the direct participation of this receptor to cell damage induced by oxygen/glucose deprivation, in the ex vivo model of organotypic hippocampal cultures. By pharmacological and immunological approaches, we show that P2X7 is rapidly and transiently up regulated in hippocampal areas eliciting metabolism impairment. Moreover, the P2 antagonists 2',3',-dialdehyde ATP and reactive blue 2 prevent both up regulation of this receptor and hypoxic/hypoglycemic damage. By confocal laser microscopy, we show that P2X7 is present at the synaptic level of fibers extending from the CA1-2 pyramidal cell layer throughout the strata oriens and radiatum, but absent on oligodendrocytes, astrocytes or neuronal cell bodies. Colocalization of P2X7 is obtained with neurofilament-L protein and with synaptophysin, not with myelin basic protein, glial fibrillary acidic protein or a marker for neuronal nuclei. P2X7 up regulation and diffuse cellular damage are also induced by 3'-O-(4-benzoyl) benzoyl-ATP, an agonist selective but not exclusive for P2X7. In summary, our study demonstrates that P2X7 not only directly participates to the hypoxic/hypoglycemic process, but also owns specific phenotypic localization. We do not exclude that it might serve as a sensor of dysregulated neuronal activity and ATP release, both occurring during oxygen/glucose deprivation.
Insights
The P2X7 receptor, crucial for immune cell death, directly contributes to brain cell damage from oxygen/glucose deprivation. This study reveals its specific synaptic localization and role in hypoxic/hypoglycemic injury.
Area of Science:
- Neuroscience
- Cell Biology
- Immunology
Background:
- Extracellular ATP acts via P2 receptors, with P2X7 a key mediator of immune cell death.
- Oxygen/glucose deprivation (OGD) causes significant neuronal damage, particularly in the hippocampus.
- The precise role of P2X7 in OGD-induced neuronal injury remains to be fully elucidated.
Purpose of the Study:
- To investigate the direct involvement of the P2X7 receptor in cell damage induced by oxygen/glucose deprivation in hippocampal cultures.
- To determine the localization and regulation of P2X7 during hypoxic/hypoglycemic conditions.
Main Methods:
- Organotypic hippocampal slice cultures were subjected to oxygen/glucose deprivation.
- Pharmacological inhibition using P2 antagonists (2',3'-dialdehyde ATP, reactive blue 2) and a P2X7 agonist (3'-O-(4-benzoyl) benzoyl-ATP).
- Immunological approaches and confocal laser microscopy to assess P2X7 expression, localization, and colocalization with neuronal and glial markers (neurofilament-L, synaptophysin, myelin basic protein, GFAP, neuronal nuclei).
Main Results:
- P2X7 receptor is rapidly and transiently upregulated in hippocampal areas experiencing metabolic impairment during OGD.
- P2X7 antagonists effectively prevent both receptor upregulation and hypoxic/hypoglycemic damage.
- Confocal microscopy revealed P2X7 localization at the synaptic level of CA1-2 pyramidal cell fibers, colocalizing with synaptic markers but absent from glial cells and neuronal bodies.
- P2X7 upregulation and cellular damage were also induced by a selective P2X7 agonist.
Conclusions:
- The P2X7 receptor directly participates in hypoxic/hypoglycemic injury in the hippocampus.
- P2X7 exhibits specific synaptic localization within hippocampal neuronal networks.
- P2X7 may function as a sensor for neuronal activity and ATP release during oxygen/glucose deprivation, contributing to cell damage.
