The incision was started 2 mm from the proximal edge of the heel and extended toward the toes. The underlying muscle was elevated with a curved forceps, leaving the muscle origin and insertion intact. The skin was apposed with a single mattress suture (6.0 nylon). The preemptive effects of intrathecal administered drugs were evaluated in mice that received Phα1β (100 and 200 pmol/site), ω-conotoxin MVIIA (1 and 10 pmol/site), morphine (1000 pmol/site) or PBS (5 μl/site) 2 h before the incision. In another group of animals, intrathecal Phα1β (100
and 200 pmol/site), PFT�� manufacturer ω-Conotoxin MVIIA (1 and 10 pmol/site), morphine (1000 pmol/site) or PBS (5 μl/site) were administered 1 h after the incision. Mechanical allodynia was evaluated as previously described (Bortalanza et al., 2002). Briefly, it was measured by a marked increase in withdrawal response frequency to 10 applications of Von Frey filament as compared to baseline values. Preliminary studies carried out in our laboratory showed that 0.09 g of Von Frey filament produce a mean withdrawal response frequency of about 10%, which is considered an innocuous stimulus. Therefore, only 0.09 g of von Frey filament was used in our experiments. Mice were
placed individually learn more in clear Pexiglass boxes (9 × 7 × 11 cm) on elevated wire meshed platforms to allow access to the ventral surface of the hind paws. The frequency of mechanical withdrawal was determined before (baseline) and after incision procedure. The experiments to evaluate the cardiovascular effects of toxins and morphine were done in rats since the cardiovascular monitoring used was better suited for rats than for mice. They were anesthetized with ketamine 10% and xilazyne 2% (0.1 ml/100 g i.p.).
A small incision was made in the inguinal region, and the femoral artery was exposed. A polyethylene catheter (0.011 ID, Clay Adams, Parsippany, NJ, USA) filled with heparinized saline and sealed with a stylet was inserted in the abdominal aorta through the femoral artery (4 cm) for recording mean arterial pressure (MAP) and heart rate (HR). The catheter was routed s.c. to the nape of the neck where it was exteriorized and secured. Rats were then allowed to Tolmetin recover in their home cages for at least 24 h before experiments began. All animals for which data were reported remained in good health conditions throughout the course of surgical procedures and experimental protocol as assessed by appearance, behavior, and maintenance of body weight. The animals received intrathecal administration of the test agents using a 10 μl microsyringe containing vehicle (PBS), Phα1β (200 pmol/site), ω-conotoxin MVIIA (100 pmol/site) or morphine (433 pmol/site). This concentration of morphine was used based in studies showing that an intrathecal morphine concentration of 300–500 pmol produced a significant analgesic response in rats (Kream et al., 1993).
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