Magi‐1 scaffolds Na _v 1‐8 and Slack K _Na channels in dorsal root ganglion neurons regulating excitability and pain

Abstract

Voltage‐dependent sodium (Nav) 1.8 channels regulate action potential generation in nociceptive neurons, identifying them as putative analgesic targets. Here, we show that Nav1.8 channel plasma membrane localization, retention, and stability occur through a direct interaction with the postsynaptic density‐95/discs large/zonula occludens‐l‐and WW domain‐containing scaffold protein called membrane‐associated guanylate kinase with inverted orientation (Magi)‐l. The neurophysiological roles of Magi‐1 are largely unknown, but we found that dorsal root ganglion (DRG)‐specific knockdown of Magi‐1 attenuated thermal nociception and acute inflammatory pain and produced deficits in Nav1.8 protein expression. A competing cell‐penetrating peptide mimetic derived from the Nav1.8 WW binding motif decreased sodium currents, reduced Nav1.8 protein expression, and produced hypoexcitability. Remarkably, a phosphorylated variant of the very same peptide caused an opposing increase in Nav1.8 surface expression and repetitive firing. Likewise, in vivo, the peptides produced diverging effects on nocifensive behavior. Additionally, we found that Magi‐1 bound to sequence like a calcium‐activated potassium channel sodium‐activated (Slack) potassium channels, demonstrating macrocomplexing with Nav1.8 channels. Taken together, these findings emphasize Magi‐1 as an essential scaffold for ion transport in DRG neurons and a central player in pain.—Pryce, K. D., Powell, R., Agwa, D., Evely, K. M., Sheehan, G. D., Nip, A., Tomasello, D. L., Gururaj, S., Bhattacharjee, A. Magi‐1 scaffolds Nav1.8 and Slack KNa channels in dorsal root ganglion neurons regulating excitability and pain. FASEB J. 33, 7315–7330 (2019). http://www.fasebj.org

Slack K + channels attenuate NMDA‐induced excitotoxic brain damage and neuronal cell death

“…As expected, cultured Slack KO CGCs remained unstained (Figure 2D). CGC Slack channels were functionally involved in the generation of outward currents that require activation of Na V channels in WT CGCs, which is in line with previous studies implying a functional coupling between Slack and Na V in dorsal root ganglia 81,82 . In line with others, our data suggest Slack channel activation by TTX‐sensitive inward Na + currents 81 and that the TTX‐sensitive, and thus Na V ‐dependent, K + outward current fraction in central neurons is largely lost in the absence of endogenous Slack channels (Figure 2F).…”

“…CGC Slack channels were functionally involved in the generation of outward currents that require activation of Na V channels in WT CGCs, which is in line with previous studies implying a functional coupling between Slack and Na V in dorsal root ganglia. 81,82 In line with others, our data suggest Slack channel activation by TTX-sensitive inward Na + currents 81 and that the TTX-sensitive, and thus Na Vdependent, K + outward current fraction in central neurons is largely lost in the absence of endogenous Slack channels (Figure 2F). Moreover, given that glutamate excitotoxicity in CGCs is mediated through NMDAR overstimulation, [83][84][85] we are the first to show that the amount of excitotoxicity closely depends on the expression of Slack (Figure 3).…”

Functional Coupling of Slack Channels and P2X3 Receptors Contributes to Neuropathic Pain Processing

“…In an earlier study we demonstrated that Slack is highly expressed in DRG neurons and that the vast majority (86.5%) of Slack-positive cells bind IB4, whereas 12.6% are positive for calcitonin gene-related peptide (CGRP), two markers of non-peptidergic and peptidergic C fiber sensory neurons, respectively [ 15 ]. Moreover, Slack has been shown to generate a sodium-activated outward potassium current (I KNa ) in whole-cell voltage-clamp recordings on IB4-positive sensory neurons of naive mice [ 15 , 18 , 22 ]. To examine whether Slack-mediated I KNa is altered after peripheral nerve injury, we performed whole-cell patch-clamp recordings on IB4-positive sensory neurons of WT and Slack -/- mice 14–19 days after SNI.…”

“…Our study uncovered P2X3 as a previously unrecognized signaling pathway that regulates Slack activity. Previously identified Na + -permeable ion channels that may activate Slack include AMPA receptors [ 41 ] and Na v 1.8 channels [ 22 ]. It has been shown that Na + entering a neuron via a small but persistent sodium current may be effective in activating I KNa , even in the absence of bulk internal Na + [ 10 ].…”

Growth Differentiation Factor-15 Produces Analgesia by Inhibiting Tetrodotoxin-Resistant Nav1.8 Sodium Channel Activity in Rat Primary Sensory Neurons

“…Among them, the tetrodotoxinresistant (TTX-R) Na ? channel Nav1.8, mainly expressed by small-and medium-diameter DRG neurons [27,28], substantially contributes to the upstroke of the action potential (AP) [29,30]. Nav1.8-null mice show an increased threshold to noxious mechanical and thermal stimuli as well as delayed development of inflammatory pain [31].…”