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Novel modes of communication between neuronal activity and microglial process dynamics

University of British Columbia

Microglia are morphologically dynamic cells that survey neuronal dendrites and rapidly respond to ATP. However, the role of ATP in mediating neuron-microglia communication remains to be determined. We therefore investigated the question whether high neuronal activity would evoke ATP release and thereby trigger a change in microglial process dynamics. To address this we used acute hippocampal brain slices and two-photon laser scanning microscopy and we developed a novel method for fixation and immunolabeling of microglia processes. We discovered that multiple brief applications of NMDA triggered a transient outgrowth of microglia processes similar to application of ATP. The outgrowth was reversible and repeatable, indicating that it was not due to excitotoxic damage. ATP release, secondary to NMDAR activation, was the key mediator as blocking purinergic receptors abolished outgrowth. Hemichannel opening is a well-defined mechanism for ATP release, but outgrowth still occurred in the absence of the hemichannel protein pannexin 1 and in the presence of the hemichannel blocker carbenoxolone. Utilizing whole cell patch clamping we demonstrated that activation of dendritic NMDAR on single neurons was sufficient to trigger microglia process outgrowth. These results suggest that dendritic neuronal NMDAR activation triggers ATP release via a hemichannel-independent mechanism. It is well established that high neuronal activity leads to a reduction in extracellular Ca²+ which causes opening of astrocytic Cx43 hemichannels and subsequent ATP release. We therefore investigated whether hemichannel opening could trigger a change in microglial process dynamics. Indeed, removal of extracellular Ca²+ triggered a microglial response, which we refer to as microglial process focalization because it was distinctively different from the NMDA-evoked process outgrowth. This focalization was also mediated by ATP as it was blocked by selective blockade of microglial purinergic receptors and we observed a strong inverse relationship between the concentration of extracellular Ca²+ and microglial responses. Carbenoxolone, which did not block NMDA-evoked process outgrowth, resulted in a dose-dependent block of microglial process focalization which is consistent with the mechanism of ATP release being opening of Cx43 hemichannels. Taken together, our data provide novel insight into how high neuronal activity triggers release of ATP as a mechanism for enhancing neuron-microglia communication.

Text

2015-04-28

Attribution-NonCommercial-NoDerivs 2.5 Canada

http://hdl.handle.net/2429/52982

Neuroscience

University of British Columbia

UBCV

http://creativecommons.org/licenses/by-nc-nd/2.5/ca/