Figure 1.
Fatty acid amide concentrations in brain following treatment with FAAH inhibitors, WIN 55,212-2 and vehicle in SCI and uninjured rats.
Rats were treated with either URB597 (URB; 3 mg/kg, i.p.), WIN 55,212-2 (WIN; 3 mg/kg, s.c.) or vehicle (Veh; 1.5 ml/kg, i.p.) for seven days and euthanized four hours following the last treatment. One group of URB597 rats were euthanized two hours (2 h) following a single treatment. Rats that received PF-3845 (PF3, PF10; 3, 10 mg/kg, p.o) or vehicle (Veh, p.o.) were treated once and euthanized four hours following dosing. Levels of AEA, OEA and PEA from each rat are shown, the thick horizontal line is the mean and the thin horizontal lines are the S.E.M. n = 4–10/group. * p<0.05, **p<0.01, ***p<0.001 vs. vehicle (Student's t-test).
Figure 2.
Fatty acid amide concentrations in thoracic spinal cord segments rostral to the spinal injury from SCI rats or comparable thoracic spinal cord segments from uninjured rats following treatment with FAAH inhibitors, WIN 55,212-2 and vehicle.
Both SCI and uninjured rats were treated with either URB597 (URB; 3 mg/kg, i.p.), WIN 55,212-2 (WIN; 3 mg/kg, s.c.) or vehicle (Veh; 1.5 ml/kg, i.p.) for seven days and euthanized four hours following the last treatment. Rats that received PF-3845 (PF3, PF10) were treated once (3, 10 mg/kg, p.o.) and euthanized four hours following dosing. One group of uninjured rats was treated once with URB597 and euthanized two hours (2 h) following treatment. The FAA levels of thoracic spinal cord of uninjured rats from this figure are repeated for Figures 3 and 4. Levels of AEA, OEA and PEA from each rat are shown, the thick horizontal line is the mean and the thin horizontal lines are the S.E.M. n = 4–10/group. * p<0.05, **p<0.01, ***p<0.001 vs. vehicle (Student's t-test).
Figure 3.
Fatty acid amide concentrations in thoracic spinal cord segments from the epicenter of injury from SCI rats or comparable spinal cord segments from uninjured rats following treatment with FAAH inhibitors, WIN 55,212-2 and vehicle.
Both SCI and uninjured rats were treated with either URB597 (URB; 3 mg/kg, i.p.), WIN 55,212-2 (WIN; 3 mg/kg, s.c.) or vehicle (Veh; 1 ml/kg, i.p.) for seven days and euthanized four hours following the last treatment. One group of uninjured rats was treated once with URB597 and euthanized two hours (2 hr) following treatment. Rats that received PF-3845 (PF3, PF10) were treated once (3, 10 mg/kg, p.o.) and euthanized four hours following dosing. Levels of AEA, OEA and PEA from each rat are shown, the thick horizontal line is the mean and the thin horizontal lines are the S.E.M. n = 4–10/group. * p<0.05, **p<0.01, ***p<0.001 vs. vehicle (Student's t-test).
Figure 4.
Fatty acid amide concentrations in thoracic spinal cord segments caudal to the spinal injury from SCI rats or comparable spinal cord segments from uninjured rats following treatment with FAAH inhibitors, WIN 55,212-2 and vehicle.
Rats were treated with either URB597 (URB; 3 mg/kg, i.p.), WIN 55,212-2 (WIN; 3 mg/kg, s.c.) or vehicle (Veh; 1 ml/kg, i.p.) for seven days and euthanized four hours following the last treatment. One group of uninjured rats was treated once with URB597 and euthanized two hours (2 h) following treatment. Rats that received PF-3845 (PF3, PF10) were treated once (3, 10 mg/kg, p.o.) and euthanized four hours following dosing. Levels of AEA, OEA and PEA from each rat are shown, the dark horizontal line is the mean and the light horizontal lines are the S.E.M. n = 4–10/group. * p<0.05, **p<0.01, ***p<0.001 vs. vehicle (Student's t-test).
Figure 5.
Fatty acid amide concentrations in lumbar spinal cord from SCI rats or uninjured rats following treatment with FAAH inhibitors, WIN 55,212-2 and vehicle.
Rats were treated with either URB597 (URB; 3 mg/kg, i.p.), WIN 55,212-2 (WIN; 3 mg/kg, s.c.) or vehicle (Veh; 1.5 ml/kg, i.p.) for seven days and euthanized four hours following the last treatment. One group of uninjured rats was treated once with URB597 and euthanized two hours (2 h) following treatment. Rats that received PF-3845 (PF3, PF10) were treated once (3, 10 mg/kg, p.o.) and euthanized four hours following dosing. Levels of AEA, OEA and PEA from each rat are shown, the thick horizontal line is the mean and the thin horizontal lines are the S.E.M. n = 4–10/group. * p<0.05, **p<0.01, ***p<0.001 vs. vehicle (Student's t-test).
Table 1.
Summary of Changes in Fatty Acid Amides Four Weeks Following a Spinal Cord Injury in Rats.
Table 2.
Fold-change of tissue fatty acid amide levels following treatment with URB597 or WIN 55,212-2 relative to vehicle treatment.
Table 3.
Fold-change of tissue fatty acid amide levels following a single oral treatment with PF-3845 compared to oral vehicle treatment.
Figure 6.
Effects of URB597 and WIN 55,212-2 treatment over seven days on below-level cutaneous hypersensitivity in rats with neuropathic SCI pain.
Baseline hind paw withdrawal thresholds were measured prior to treatment with either URB597 (3 mg/kg, i.p.), WIN 55,212-2 (3 mg/kg, s.c.) or vehicle (Veh, 1.5 ml/kg, i.p.). Rats were treated twice daily and tested following the first daily injection. On the first day of testing, a robust antinociception was observed beginning 30 min post-injection of WIN 55,212-2, which was observed also observed on days 3 and 7. By contrast, no antinociceptive effects were observed following treatment with either URB597 or vehicle. Data presented as mean ± S.E.M. n = 8–10/group. * p<0.05 vs. vehicle (Two-way repeated measures ANOVA, Student-Newman-Keuls test).
Figure 7.
Effect of PF-3845 treatment on below-level cutaneous hypersensitivity in rats with neuropathic SCI pain.
Following baseline hind paw withdrawal threshold measurement, SCI rats were treated with either PF-3845 (3, 10 mg/kg, p.o.) or vehicle (Veh; 5 ml/kg, p.o.). An antinociceptive effect was observed 1 and 3 hours following administration of 10 mg/kg, but not 3 mg/kg. Data presented as mean ± S.E.M. n = 8–9/group. * p<0.05 vs. vehicle (Two-way repeated measures ANOVA, Student-Newman-Keuls test).
Figure 8.
Effects of URB597 and WIN 55,212-2 on BBB Locomotor Scores in SCI rats.
Prior to spinal compression surgery (“Baseline”), BBB Locomotor Scores were 21, indicating normal hind limb function. (A Score of “0” indicates no hind limb function.) The mean BBB Locomotor Score obtained the day before initiation of the seven-day treatment procedure (“4 wks” post-SCI) was 9.8±0.4. Spinal cord-injured rats were evaluated again on the sixth day of treatment (“5 wks” post-SCI) and there were no significant changes in BBB Scores. Data presented as mean ± S.E.M. n = 8–10/group. *p<0.05 vs. pre-SCI (Two-way repeated measures ANOVA, Student-Newman-Keuls test).