A Study Of The Mechanisms By Which Neuritin Regulates The Expression Of Neuronal Potassium Channel Kv4.2 α-subunite And Neuronal Excitability

Neuritin is a new neurotrophic factor discovered in a screen to identify genes involved in activity-dependent synaptic plasticity. Neuritin also plays multiple roles in the process of neural development and synaptic plasticity. The receptors for binding neuritin and its downstream signaling effectors, however, remain unclear. Here, we report that neuritin specifically increases the densities of transient outward K+ currents (IA) in both rat and mice cerebellar granule neurons (CGNs) in a time-and concentration-dependent manner. Neuri in-induced amplification of IAis mediated by increased mRNA and protein expression of Kv4.2, the main a-subunit of IA. Exposure of CGNs to neuritin markedly induces phosphorylation of ERK (pERK), Akt (pAkt), and mammalian target of rapamycin (pmTOR). Neuritin-induced IA and increased expression of Kv4.2 are attenuated by ERK, Akt, or mTOR inhibitors. Unexpectedly, pharmacological blockade of insulin receptor, but not the insulin-like growth factor 1 receptor, abrogates the effect of neuritin on IAamplification and Kv4.2 induction. Indeed, neuritin activates downstream signaling effectors of the insulin receptor in CGNs and HeLa.After proving that Neuritin increases Kv4.2-mediated IAdensities in rat cerebellar granule neurons (CGNs) by activating insulin receptor pathway and up-regulated the transcription of Kv4.2. Here, we further shown the mechanism and function of upregulation of Kv4.2 induced by Neuritin. We found that treatments with specific chemical inhibitors against of calcineurin (CaN), Cyclosporin A (CsA) or FK520, significantly offset the increase of Kv4.2 expression and IA density induced by Neuritin. Immunocytochemistry and luciferase reporter studies with Hela cell shown that Neuritin does activate NFATc4 and increase the activation of Kv4.2 promoter. In NFATc4-/- mice, Neuritin could not increase the expression of Kv4.2 and IAdensity in CGNs and cortical neuron with the absence of this transcriptional factor. By using AAV-mediated over expression of Neuritin, we found that Neuritin decreases the frequency of action potential recorded from the mouse prefrontal cortical pyramid neurons and this function could not be repeated on NFATc4-/- mice.Our study, for the first time revealed that Neuritin could activate IR and followed by the activation of MEK-ERK, Akt-mTOR, CaN-NFATc4 these three signaling pathways to up-regulate the expression of Kv4.2 at both transcription and translation levels. Finally, Neuritin could increase the densities of IAcurrents. Also, Neuritin could modulate neuronal excitability with the help of NFATc4. These discoveries not only provide us a basis for understanding the role of Neuritin, but also provide a reference for studying other neurotrophic factors.