1. Crosstalk of cAMP to the ERK1/2 cascade emerges during postnatal maturation.
Crosstalk of cAMP to the ERK1/2 cascade emerges during postnatal maturation. (A,B) Basal levels of pRII and pERK1/2 in sensory neurons from newborn rats (P0), and rats that were 10 days (P10), 6 weeks (W6), 12 months (M12) and 24 months (M24) old. Mean values were normalized to the adult group (W6) and scaled to P0. Basal pRII levels slightly increased after birth and then remained constant in all analyzed age classes. In contrast, basal pERK1/2 levels doubled from P10 to W6. Values are means±s.e.m.; n=8–16; >3000 neurons/condition; one-way ANOVA with Tukey's test. (C) The distribution of baseline pRII intensities remained similar in all analyzed ages. (D) Baseline pERK1/2 intensity distributions were significantly broader in sensory neurons from adult versus newborn and neonatal rats. (E) Relative fold changes of pRII intensity after stimulation with PGI2 (2 µM, 5 min), 5-HT (2 µM, 5 min), Fsk (10 µM, 30 min) and NGF (1 nM, 30 min). Responses to GPCR agonists and Fsk were similar in sensory neurons of all analyzed ages. As expected, NGF did not induce activation of PKA-II. Values are means±s.e.m.; n=4–8; >1500 neurons/condition; two-way ANOVA with Dunnett's test. (F) Induction of ERK1/2 phosphorylation in the same cells as shown in E. Stimulation with GPCR agonists or Fsk resulted in age-dependent cross-activation of the ERK1/2 cascade. (G) Kinetics of pRII intensity changes upon stimulation with PGI2 (2 µM), 5-HT (2 µM), Fsk (10 µM) and NGF (1 nM). PGI2 and 5-HT induced a rapid and transient pRII response, whereas the effect of Fsk was long lasting. The magnitude and kinetics of the responses were similar for all age classes. Values are means±s.e.m.; n=4; total of >1500 neurons/condition; two-way ANOVA with Bonferroni's test. (H) Relative changes in pERK1/2 levels in the same cells as shown in G. The amplitude, but not the kinetics, of ERK1/2 activation by PGI2, 5-HT, and Fsk increased until 6 weeks of age. AU, arbitrary units; PDE, probability density estimate. *P<0.05; **P<0.01; ***P<0.001.
Joerg Isensee et al. J Cell Sci 2017;130:
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