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Updated: Aug 4, 2026
Primary Microglia Isolation from Mixed Glial Cell Cultures of Neonatal Rat Brain Tissue
Published on: August 15, 2012
P2 Purinoceptor expression and functional changes of hypoxia-activated cultured rat retinal microglia
K Morigiwa1, M Quan, M Murakami
1Department of Physiology, Osaka University Graduate School of Medicine, 2-2 Yamadaoka, Suita, Osaka, Japan. km@phys2.med.osaka-u.ac.jp
Abstract:
P(2) purinoceptors appear to modulate microglia function, but their role in hypoxic microglia has not been investigated. We examined in postnatal rat retinal microglia cultured under hypoxic (1% oxygen) condition, their P2 expression, proliferation and cytokine release in the presence or absence of the P2 receptor agonists and antagonists. Fura-2 fluorescence measurements of intracellular Ca(2+) rises to P2 receptor agonists and antagonists indicated that both P(2U) and P(2Z) were expressed in hypoxic microglia. Hypoxia induced BrdU incorporation and release of interleukin-1beta (IL-1beta) and tumor necrosis factor-alpha (TNF-alpha) as well. The P(2U) agonist, UTP, maintained the BrdU incorporation, whereas the P(2Z) agonist, BzATP, suppressed it, but significantly enhanced IL-1beta and TNF-alpha release, suggesting that the P(2U) response may underlie the mitotic activity, and that of P(2Z), the IL-1beta and TNF-alpha release of hypoxia-activated microglia.
Insights
Hypoxic microglia express P2 purinoceptors. P2U receptors promote proliferation, while P2Z receptors enhance inflammatory cytokine release, revealing distinct roles in microglial responses to low oxygen.
Area of Science:
- Neuroscience
- Immunology
- Cell Biology
Background:
- Microglia play crucial roles in central nervous system immunity and homeostasis.
- P2 purinoceptors are implicated in modulating microglial function.
- The specific role of P2 purinoceptors in microglia under hypoxic conditions remains largely uncharacterized.
Purpose of the Study:
- To investigate the expression and function of P2 purinoceptors in rat retinal microglia under hypoxia.
- To determine the effects of P2 purinoceptor activation on microglial proliferation and cytokine release during hypoxia.
Main Methods:
- Primary cultures of postnatal rat retinal microglia were established and subjected to hypoxic conditions (1% oxygen).
- P2 purinoceptor expression was assessed.
- Intracellular calcium (Ca2+) levels were measured using Fura-2 fluorescence in response to P2 receptor agonists and antagonists.
- Bromodeoxyuridine (BrdU) incorporation was used to evaluate cell proliferation.
- The release of interleukin-1beta (IL-1beta) and tumor necrosis factor-alpha (TNF-alpha) was quantified.
Main Results:
- Hypoxic microglia expressed both P2U and P2Z purinoceptors.
- Hypoxia induced microglial proliferation (BrdU incorporation) and the release of IL-1beta and TNF-alpha.
- The P2U agonist (UTP) maintained hypoxia-induced BrdU incorporation.
- The P2Z agonist (BzATP) suppressed BrdU incorporation but significantly enhanced the release of IL-1beta and TNF-alpha.
Conclusions:
- P2U purinoceptors are involved in mediating the proliferative response of microglia under hypoxic conditions.
- P2Z purinoceptors contribute to the inflammatory response of microglia by promoting the release of IL-1beta and TNF-alpha during hypoxia.
- These findings elucidate distinct functional roles for P2U and P2Z purinoceptors in hypoxia-activated microglia.
