Unveiling the molecular crosstalk between the purinergic P2X7 receptor and cannabinoid subtype 2 receptor in human microglial HMC3 cells: New insights for multitarget therapies in neuroinflammation

Background Microglia-mediated neuroinflammation plays a pivotal role in the progression and possible onset of numerous neurodegenerative diseases. A promising therapeutic approach involves shifting microglial polarization from a pro-inflammatory to an anti-inflammatory phenotype. Two receptors central to this process are the purinergic P2X7 receptor, an ATP-gated ion channel that promotes pro-inflammatory signaling, and the cannabinoid receptor subtype 2, a G i/o protein-coupled receptor associated with anti-inflammatory effects. Methods Here, we investigated the therapeutic potential of pharmacological modulation of these receptors in human HMC3 microglial cells stimulated with LPS, IFNγ, and BzATP. Specifically, we evaluated three selective P2X7R antagonists (A740003, JNJ-54173717, and GI-39) in combination with the CB2R agonist GW405833. Functional outcomes were assessed by measuring intracellular calcium accumulation, plasma membrane permeabilization, and cytokine release. Results Co-treatment with P2X7R antagonists and GW405833 markedly attenuated intracellular calcium accumulation, membrane permeabilization, and IL-6 secretion, while enhancing IL-4 release. When CP55,940, a non-selective cannabinoid receptor agonist, was used in combination with P2X7R antagonists, the pharmacological activity of the selective CB2R agonist was substantially reduced. This suggests that the observed effects result from a combined interaction between CB2R activation and P2X7R inhibition rather than from alternative mechanisms. Docking analyses were used as a qualitative tool to explore possible ligand accommodation within P2X7R and CB2R binding pockets. Conclusions Concurrent modulation of P2X7R and CB2R produces enhanced anti-inflammatory effects in activated HMC3 cells. This multitarget strategy offers a promising therapeutic avenue for controlling neuroinflammation and for the development of improved interventions for neurodegenerative disorders.