The RSK2-RPS6 axis promotes axonal regeneration in the peripheral and central nervous systems
Fig 3
RSK controls the preconditioning effect in mature DRG neurons.
(A) Representative microphotographs of preconditioned cultures of mature DRG neurons treated with DMSO (control), translation inhibitor (Cycloheximide, 2 nM), mTOR inhibitors (Torin1, 5 nM or Rapamycin, 0.1 nM), S6K1 inhibitor (PF-4708671-8uM). Scale bar: 250 μm. (B) Quantification of longest neurite length per neuron 16 h after plating (mean ± SEM, two-way ANOVA, 3–4 independent DRG cultures, approximately 50–100 cells counted per condition per culture (except for Cycloheximide)). (C) Percentage of neurons growing a neurite 16 h after plating (mean ± SEM, two-way ANOVA, 10 random microscopy fields quantified per condition). (D) Representative microphotographs of naive and preconditioned cultures of mature DRG neurons treated with DMSO (control) or RSK inhibitor (BRD-7389 (3 μm)). Scale bar: 250 μm. (E) Quantification of longest neurite length per neuron 16 h after plating in naive condition (mean ± SEM, two-way ANOVA, 3 independent DRG cultures, approximately 50–100 cells counted per condition per culture for DMSO condition; all neurons found with a neurite were quantified in BRD7389 condition). (F) Percentage of neurons growing a neurite 16 h after plating in naive DRG treated with DMSO or BRD7389 (mean ± SEM, paired t test, 10 random microscopy fields quantified per condition). (G) Quantification of the longest neurite length 16 h after plating in PC DRG treated with DMSO or BRD7389 (mean ± SEM, two-way ANOVA, approximately 50–100 cells per condition per culture for DMSO condition; all neurons growing a neurite were quantified in BRD7389 condition). (H). Percentage of neurons growing a neurite 16 h after plating in naive DRG treated with DMSO or BRD7389 (mean ± SEM, paired t test, 5 independent DRG cultures, 10 random microscopy fields were quantified per condition). ⁎⁎⁎p < 0.001, ⁎⁎p < 0.01, ⁎p < 0.05. Raw data can be found in Supporting information (S1 Data). DRG, dorsal root ganglion; mTOR, mammalian target of rapamycin; PC, precondtionned.