Phosphoinositide 3-kinase {gamma} is not a predominant regulator of ATP-dependent directed microglial process motility or experience-dependent ocular dominance plasticity

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Microglia are dynamic cells whose extensive interactions with neurons and glia during development allow them to regulate neuronal development and function. The microglial P2Y12 receptor is crucial for microglial responsiveness to extracellular ATP and mediates numerous microglial functions, including ATP-dependent directional motility, microglia-neuron interactions, and experience-dependent synaptic plasticity. However, little is known about the downstream signaling effectors that mediate these diverse actions of P2Y12. Phosphoinositide-3-kinase gamma (PI3K), a lipid kinase activated downstream of Gi protein-coupled receptors such as P2Y12, could translate localized extracellular ATP signals into directed microglial action and serve as a broad effector of P2Y12-dependent signaling. Here, we used pharmacological and genetic methods to manipulate P2Y12 and PI3K signaling, to determine whether inhibiting PI3K phenocopied the loss of P2Y12 signaling in mouse microglia. While pan-inhibition of all PI3K activity substantially affected P2Y12-dependent microglial responses, our results suggest that PI3K specifically is only a minor part of the P2Y12 signaling pathway. PI3K was not required to maintain homeostatic microglial morphology or their dynamic surveillance in vivo. Further, PI3K was not strictly required for P2Y12-dependent microglial responses ex vivo or in vivo, although we did observe subtle deficits in the recruitment of microglial process towards sources of ATP. Finally, PI3K was not required for ocular dominance plasticity, a P2Y12-dependent form of experience-dependent synaptic plasticity that occurs in the developing visual cortex. Overall, our results demonstrate that PI3K is not the major mediator of P2Y12 function in microglia, but may have a role in amplifying or fine-tuning the chemotactic response.

Significance Statement Extracellular ATP acts as a chemoattractant for microglial processes via the microglial P2Y12 receptor, facilitating interactions between neurons and microglia. However, the intracellular pathways underlying this directional motility remain unknown. While phosphoinositide-3-kinase (PI3K) activity is required for ATP-mediated microglial motility, PI3K specifically is largely dispensable but plays a subtle modulatory role. Because PI3K is the only isoform directly activated by G-protein coupled receptors, P2Y12 likely activates PI3K indirectly to promote directed microglial motility.

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