Heated metal plate

Thermal hypersensitivity after DEAB administration was evaluated with a hot plate test [34 (link)]. The animals (n = 8) were treated with DEAB (30 mg/kg, oral), vehicle (10 mL/kg, oral), or morphine as a positive control (5 mg/kg, subcutaneous). One hour after the oral and 30 min after the subcutaneous treatments, the mice were placed on a heated metal plate (Ugo Basile, Varese, Italy) with the temperature set at 56 ± 1 °C. The time (s) until the mouse manifested a nociceptive behavior (lifting or licking its hind-paw) was considered the latency response to the thermal stimuli. A cut-off time of 30 s was chosen to avoid tissue injury. This latency response was recorded 60, 90, and 120 min following oral treatment.

Thermal hypersensitivity after HEMh administration was evaluated using a hot plate test, as described by Eddy and Leimbach (1953) with modifications [15 (link),16 (link)]. Male Swiss mice (n = 8–10 animals per group) were treated with HEMh (62.5 mg/kg, by oral route), vehicle (saline, 10 mL/kg, by oral route), or morphine (5 mg/kg, subcutaneous) used as a positive control. One hour after oral and 30 min after the subcutaneous treatments the mice were placed on a heated metal plate (Ugo Basile, Gemonio VA, Italy) with a temperature set at 50 ± 1 °C (sensitizes more C and type II A δ nociceptors) or 56 ± 1 °C (sensitizes type I Aδ nociceptor). The time (in seconds) until the mouse manifested a nociceptive behavior (lifting or licking of the hind-paw) was considered as the latency response to the thermal stimuli. A cut-off time of 20 s was chosen to avoid tissue injury. The animals were pre-selected (24 h before starting the experiment), excluding those with a response time of less than 4 s or greater than 11 s. This latency response was recorded at 60, 90, 120, and 150 min following oral treatment and the prolongation of the latency time (s) compared with the values of the control group (vehicle) was used for statistical comparison.