As a positive control for COX-2, LPS (2.5 μg) was stereotaxically injected into the mouse striatum and RNA was isolated 6 h later. To compare the expression of inflammatory mediators in the different experimental groups the amount of mRNA was estimated as the ratio of GAPDH. Blood samples Alpelisib cell line (∼500 μl) were taken by cardiac puncture in terminally anaesthetized mice and collected in microfuge tubes. Samples were spun down and serum kept at −20 °C until further use. IL-1β, IL-6 and TNF-α serum levels were assessed with a sandwich-type ELISAs using a matched antibody pair

(duoset ELISA development assay, R&D) according to the manufacturer’s instructions with minor modification. Serum levels of prostaglandin E metabolites were measured according manufacture’s instruction (Cayman, USA). Brain levels of prostaglandin E2 (PGE2) were measured according manufactures instruction (Assay designs, USA), with minor modification. Briefly, serum samples (50 μl) were diluted 1:10 in assay buffer

provided by the manufacturer. Samples and standard were derivatized by adding 150 μl of carbonate buffer followed by overnight incubation at 37 °C. Samples and standards were then analysed according to manufactures’ instructions. Brain tissue was homogenized in 100 μl PBS and mixed with 1 ml 100% ethanol. After centrifugation at 3000 rpm for 10 min at 4 °C, supernatant was transferred to an empty tube and ethanol evaporated under a stream of nitrogen. Samples were resuspended in 500 μl of assay buffer and PGE2 levels measured according to manufacturer’s Selleckchem Trametinib instructions. Burrowing and open-field activity were analysed by one-way analysis of variance (ANOVA) followed, if significant, by Dunnett’s post-test versus controls. Data were analysed for normality using the Kolmogorov–Smirnov test and for equal variances using the Bartlett’s test. Changes in body temperature were assessed by paired Student’s t-test. The intervention studies were analysed by one-way analysis of variance (ANOVA) or two-way

ANOVA, followed, if significant, by Bonferroni’s post-test using Graphpad Prism software. Values were expressed as mean ± SEM. A p-value <0.05 was considered to indicate statistical significant difference. Clomifene We previously showed that pre-treatment of mice with indomethacin is sufficient to inhibit LPS-induced changes in burrowing activity (Teeling et al., 2007). In the present study, we aimed to further investigate these observations. We tested various well known anti-inflammatory drugs, including: indomethacin, ibuprofen, acetaminophen (paracetamol) and dexamethasone (Table 1), and measured their effect on LPS-induced changes in body temperature, burrowing and open-field activity, and production of inflammatory mediators. Mice were habituated to burrowing prior to the experiment.