Data and statistical analysis
Data collection and statistical analysis comply with the recommendations of the British Journal of Pharmacology on experimental design and analysis in pharmacology (Curtis et al., 2018). The number of experiments in each experimental group was selected on the basis of observing a statistically significant effect while using the minimum number of animals (3R principles) and on the experience from the previous studies. Animals were randomly assigned in the different experimental groups with each group having the same number of animals by design. Blinding of the operator was not feasible, but data analysis was performed semi-blinded by an independent analyst.
Statistical analysis was performed using Statistica 6.1 Base (Tulsa, OK, USA). Shapiro–Wilk normality test was used to analyze the data distribution. Null-hypothesis testing was performed by ANOVA. One-way ANOVA followed by Dunnett’s or Tukey’s test for multiple comparison post hoc was used. For related groups one-way repeated ANOVA test was performed. Data are presented as mean ± SEM. Values of P < 0.05 were considered significant.