The PKC/NOX/ROS and PYK2/MEK/ERK/PARP signalling pathways drive TRPM2 channel activation induced by non-cytolytic oxidative stress in microglial cells.

OBJECTIVES

The study aimed to investigate the signalling mechanism for TRPM2 channel activation by non-cytolytic oxidative stress in microglia.

METHODS

Microglia from wild-type (WT) and TRPM2-knockout (KO) mice were exposed to 10-30 mM HO for up to 24 hours. Morphological changes characteristic of microglial activation, [Ca], ROS generation and the effects of inhibiting particular signalling pathways were examined.

RESULTS

Exposure of WT microglia to HO for 24 hours caused no cell death but induced salient morphological changes, which was prevented by TRPM2-KO. Exposure of WT microglia to HO to 2 hours failed, and extension to 8 hours was required, to induce an increase in [Ca], which was abolished by TRPM2-KO. Exposure of microglia to HO for 8 hours induced ROS generation, which was suppressed by inhibition of PKC and NADPH oxidases (NOX). HO-induced PARP activation in TRPM2-KO cells was lower than that in WT cells. Furthermore, HO-induced activation of PARP and TRPM2 and morphological changes were attenuated by inhibition of PCK and NOX as well as PYK2 and MEK/ERK.

CONCLUSION

Our results support that PKC/NOX-mediated ROS generation and TRPM2-mediated Ca-induced activation of the PYK2/MEK/ERK pathway form a positive feedback mechanism to drive TRPM2 channel activation by non-cytolytic oxidative stress.