Fig 2: αCaMKII phosphorylation levels in the nucleus accumbens (NAc), ventral tegmental area (VTA), and prefrontal cortex (PFC) after cocaine and social interaction CPP. (A) Representative western blot images of phosphorylated α-CaMKII (p-αCaMKII) (Thr286), αCaMKII, and β-III-Tubulin levels (used as protein loading control) in the NAc, VTA, and PFC of rats from the saline, cocaine, and social interaction CPP groups. (B) Relative intensity of NAc αCaMKII phosphorylation at Thr286 in rats that expressed saline, cocaine, and social interaction CPP. * p < 0.05, one-way ANOVA followed by Newman–Keuls multiple comparisons test; n = 5–6. (C) Relative intensity of the bands that express p-αCaMKII levels in the VTA of rats from the cocaine and social interaction CPP groups, as well as the control group—saline CPP; p = non-significant (ns), n = 4–5. (D) Relative intensity of the bands that express p-αCaMKII levels in the PFC of rats that expressed cocaine and social interaction CPP, and the control group—saline CPP; p = ns; n = 6–7. Results are presented as mean ± SEM.

Fig 3: Blockade of LTCC suppressed DOX-induced phosphorylation of CaMKII and NF-κB and increases in cleaved caspase 3 in cardiomyocytes. (a) Representative immunoblots of CaMKII, phosphorylated CaMKII, NF-κB, phosphorylated NF-κB, cleaved caspase 3, and GAPDH in NRVMs treated with or without nifedipine (Nif, 1 μM) in the presence or absence of DOX (10 μM) for 24 hour (n = 5). The experiment was conducted 3 times. (b–d) Quantitative analysis of phosphorylated CaMKII, phosphorylated NF-κB, and cleaved caspase 3 in each group (n = 5). (e) Representative immunoblots of CaMKII, phosphorylated CaMKII, NF-κB, phosphorylated NF-κB, cleaved caspase 3, and GAPDH in NRVMs treated with indicated small interfering RNA (siRNA) in the presence or absence of DOX (10 μM) for 24 hour (n = 5). (f–h) Quantitative analysis of phosphorylated CaMKII, phosphorylated NF-κB, and cleaved caspase 3 in each group (n = 5). The experiment was conducted 3 times. *P < 0.05, **P < 0.01: post-hoc Tukey’s comparison test.

Fig 4: Chem-iLTP induces CaMKII–dependent PX-RICS phosphorylation to promote the assembly of PX-RICS and 14-3-3 and cell surface transport of GABAARs.(A) WT and KO hippocampal neurons were moderately stimulated with NMDA in the presence or absence of KN93. The neuronal lysates were immunoprecipitated with an anti–PX-RICS antibody, and half of the immunoprecipitate was analyzed by Phos–tag (upper) or standard (lower) SDS-PAGE and immunoblotting. The intensities of unphosphorylated and phosphorylated PX-RICS bands (open and filled arrowheads, respectively) were corrected using those of total PX-RICS and the data were expressed as relative values with respect to the mock control (right panel).(B) WT hippocampal neurons were moderately stimulated with NMDA, and the lysates were analyzed by immunoprecipitation with an anti–PX-RICS antibody followed by immunoblotting with anti–14-3-3ζ and anti–14-3-3θ antibodies (upper panel). Total 14–3-3ζ/θ protein in the neuronal lysates was assessed by immunoblotting with anti–14-3-3ζ and anti–14-3-3θ antibodies (lower panel). Quantitative analysis of 14-3-3ζ/θ bound to PX-RICS (right panel). The ratio of the band intensity of 14-3-3ζ/θ bound to PX-RICS to that of total 14-3-3ζ/θ was assessed, and the data were expressed as relative values with respect to mock control.(C) Lysates of WT and KO hippocampal neurons modestly stimulated with NMDA in the presence or absence of KN93 were analyzed by immunoblotting with anti–β3 and anti–phospho-Ser383 β3 antibodies. α-tubulin was used as an internal control for the quantitative analysis.(D) WT hippocampal neurons were modestly stimulated with NMDA and subjected to live cell labeling of the surface γ2, followed by co-staining with anti-VGAT antibody to identify synaptically localized GABAARs (arrowheads). Arrows indicate large or elongated puncta of nonsynaptic γ2 often observed in NMDA-treated neurons. Scale bars, 10 μm. n = 5 (mock) and 5 (NMDA) across two independent experiments.Data are shown as the mean ± SEM from three (A, B, C) or two (D) independent experiments. NS, not significant; *P < 0.05; **P < 0.01 (Student's t-test).

Fig 5: Summary of key findings. Effects of moderate‐intensity interval running without (CON) or with blood flow restriction (BFR), or in normobaric, systemic hypoxia (HYP) on the mRNA content of Na+,K+‐ATPase (NKAα1‐3, NKAβ1‐3, FXYD1) and PGC‐1α (total, 1α1, 1α4) isoforms, indicators of responses to oxidative stress (HSP27 protein content in type I and II muscle fibres, catalase and heat‐shock protein 70,HSP70, mRNA content), muscle hypoxia (ie deoxygenated haemoglobin as measured by near‐infrared spectroscopy), lactate concentration, and AMPK and CaMKII signalling in the skeletal muscle of men. “p‐” denotes phosphorylation; ACC, Acetyl‐CoA carboxylase; AMPK, 5′ AMP‐activated protein kinase; CaMKII, Ca2+/calmodulin‐dependent protein kinase II; PLB, phospholamban; LT, lactate threshold