Bupivacaine hcl

Primary antibodies targeting the following proteins were used for western blot and tissue sections: FGF9 (Abcam, EPR19937), Olig1 (Millipore, MAB5540), NeuN (Millipore, ABN78), Iba-1 (Abcam, ab5076), anti-GFAP (Millipore, MAB360), anti-Calbindin (Sigma, C2724), Caspase-3 (Santa Cruz, sc-373730), GABA (Sigma, A2052), VGAT (SYSY, 131011), Calbindin (Sigma, C2724), GAPDH (Abcam,ab128915), Adcy5/6 (Santa Cruz, sc-514785), p-ERK (Cell Signaling, #4370), ERK (Cell Signaling, #4695), p-Akt (Immunoway, YP006), Akt (Immunoway, YT0176), and β-actin (Proteintech, 60008-1-Ig). Secondary antibodies for western blot were from Rockland Immunochemicals, USA, and fluorescein isothiocyanate secondary antibodies were from Jackson ImmunoResearch, West Grove, PA. Hoechst stain (1:100, Invitrogen), RNeasy Lipid Tissue Mini kit (QIAGEN, 74804), SQ22536, bupivacaine-HCl, pentylenetetrazol, GABA, and Glu were purchased from Sigma (Madrid, Spain).

Thirteen female BALB/c mice conformed to the mock treatment. The animals were injected with 0.1 mL of 0.25% bupivacaine-HCl (Sigma-Chemical Co., St Louis, MO, USA) in the left quadriceps at day 0 and on day 2 and 10 received an intramuscular injection of 100μg of empty vector pcDNA™3.1 D/V5-His-TOPO®. 13 female BALB/c mice received 0.1 mL of Phosphate Buffer Solution (PBS; Gibco® Invitrogen™) for the control group (Fig 1).

The experimental animals were divided into eight groups of 13 female BALB/c mice per treatment and received an injection of 0.1 mL of 0.25% bupivacaine-HCl (Sigma-Chemical Co., St Louis, MO, USA) in DPBS (Gibco® Invitrogen™) in the left quadriceps at day 0. On days 2 and 10, the mice received intramuscular injections of 100 μg of a cocktail mixture of pcDNA™3.1 D/V5-His-TOPO® ™ (Invitrogen™ Carlsbad, CA) encoding either: D1_83-119, D2, B´/B or B´/B_COOH co-vaccinated IFN-γ or IL-10 encoded into pcDNA™3.1D/V5-His-TOPO® (Fig 1). The sample size was calculated based on tolerizing DNA vaccines designed experiments [21 (link), 25 (link)], and the exact value of n in each experiment was described in Fig 1.