Psilocybin has no immediate or persistent analgesic effect in acute and chronic mouse pain models

Abstract

The psychedelic psilocybin may have lasting therapeutic effects for patients with chronic pain syndromes. Some clinical and preclinical data suggest these putative benefits derive from direct analgesic effects. However, this possibility has not been comprehensively tested in preclinical models. Here, we show that psilocybin is not analgesic over a range of doses across multiple pain assays and models of acute and chronic inflammatory, neuropathic, or musculoskeletal pain in mice.

Figure 1.. Survey of sensory and analgesic properties of psilocybin.

(A) Psilocybin at 0.3, 2, and 10 mg/kg is physiologically active as shown by quantification of Head Twitch Response (males n=3–7 and females n=3 per group). One-way ANOVA of total head twitches F (3, 28) = 42.98 P<0.0001. Post-hoc Tukey Test, * P < 0.05 versus saline. Mean ± SEM. (B) Psilocybin reduces rearing bouts in a dose-dependent manner (males n=3–7 and females n=3 per group). One-way ANOVA of total rearing bouts F (3, 28) = 15.96, P<0.0001. * Post-hoc Tukey Test, P <0.05 versus saline. Mean ± SEM. Psilocybin did not improve mechanical (C) or thermal hypersensitivity (D), whereas morphine produced significant antinociception in both (n=5 males/group). Two-way repeated measures ANOVA (Mechanical: F(2.904, 23.2) = 103.0, P<0.0001; Thermal: F(2.207, 39.72) = 89.85, P<0.0001). Post-hoc Bonferroni test, * P<0.05 across pre-morphine vs 30m post-morphine for mechanical and thermal tests. All data are represented as mean ± SEM. (E) Psilocybin reduces marble burying performance at 2 mg/kg (males n=7–10 and females 7–11 per group). One-way ANOVA of buried marbles, F(5, 90) = 83.07, P<0.0001 P<0.0001. Post-hoc Tukey Test, * P < 0.05 versus control; # P <0.05 versus CFA and control. (F) Psilocybin 2 mg/kg has no effect on mouse grimace scale after laparotomy (males n = 6–7, females = 4–6 per group). One-way ANOVA of mouse grimace scale difference score (post-laparotomy minus baseline mouse grimace scale values), F (2, 33) = 4.535, P=0.0182. Post-hoc Tukey test, * P < 0.05 versus saline and psilocybin.

Figure 2.. Psilocybin does not improve mechanical or thermal hypersensitivity in male or female mice in three pain models.

(A) Legend and timeline for pain models and assessments. Relative to baseline testing: SNI surgery was performed 7 days prior; AIMP involves two acidic normal saline injections, 6 days and 1 day prior; CFA injection to the paw occurs 1 day prior. Psilocybin injections were administered after baseline testing. Hargreaves test (thermal) is quantified by paw withdrawal latency. Von Frey test (mechanical) is quantified by paw withdrawal threshold (PWT). Muscle sensitivity to tweezer force is quantified by muscle withdrawal threshold (MWT). (B,C) SNI produces neuropathic pain and long-lasting thermal and mechanical hyperalgesia. 7 days after surgery, in the acute phase, mice (males and females n=6 per group) were injected with either psilocybin or vehicle control and tested out to 24 hours. Two weeks later, in the chronic pain phase, mice received a second injection and again were tested out to 24 hours. Psilocybin has no effect on either thermal or mechanical hyperalgesia at the doses tested. Two-way repeated measures ANOVA (Thermal: F (32, 440) = 4.861 P<0.0001; Mechanical: F (32, 440) = 7.979 P<0.0001). Post-hoc Tukey Test, * P < 0.0001 versus control. (D,E) AIMP produces long-lasting cutaneous thermal and muscle mechanical hyperalgesia. 24 h after initiation of pain, in the acute phase, mice (males and females n=5–7 per group) were injected with either psilocybin or vehicle control and tested out to 24 hours. Two weeks later, in the chronic pain phase, mice received a second injection and again were tested out to 24 hours. Psilocybin has no effect on either cutaneous thermal or muscle mechanical hyperalgesia at the doses tested. Two-way repeated measures ANOVA (Thermal: F (32, 440) = 2.809 P<0.0001; Mechanical: F (32, 440) = 11.99, P<0.0001) Post-hoc Tukey Test, * P < 0.0001 versus control. (F,G) CFA produces long-lasting thermal and mechanical hyperalgesia due to inflammation of the paw. 24 h after CFA injection, in the acute pain phase, mice (males and females n = 5–7 per group) were injected with either psilocybin or vehicle control and tested out to 24 hours. Two weeks later, in the chronic pain phase, the mice received a second injection and were again tested out to 24 hours. Psilocybin has no effect on either thermal or mechanical hyperalgesia at the doses tested. Two-way repeated measures ANOVA (Thermal: F (32, 280) = 3.664 P<0.0001; Mechanical: (32, 280) = 5.242). Post-hoc Tukey Test, * P < 0.0001 versus control.

Figure 3.. Effects of psilocybin on temperature preference and cold hypersensitivity

(A-C) Thermal Place Preference. Mice are given free access to 30 C and either 40 C or 20 C surfaces. (A) Saline-injected mice have no preference, whereas CFA-injected mice avoid the 40 C plate. This is effect is abolished by 10 mg/kg morphine. Psilocybin produces a dose-dependent preference for the 40 C plate (males n=4–7 and females n=4–6 each per group). One-way ANOVA. F (5, 49) = 9.147 P<0.0001. Post-hoc Tukey Test, * P < 0.05 versus control. (B) The dose-dependent preference is present in mice that are not injected with CFA (males and females, n=3–4 each per group). One-way ANOVA F (3, 26) = 10.34 P=0.0001. Post-hoc Tukey Test, * P < 0.001 versus control. (C) Psilocybin induces a dose-dependent avoidance of 20 C (males and females, n=4–6 each per group). One-way ANOVA F (3, 34) = 7.211, P=0.0007. Post-hoc Tukey Test, * P < 0.001 versus control. (D) Mice (male and female, n=4–6 each per group) were injected with formalin or sham into the paw and given i.p. injection of saline or psilocybin (0.3, 2, 10 mg/kg). 30 minutes later they were assessed for cold sensitivity using the cold plate test. There was a significant decrease in paw lifts compared to SNI controls at 10 mg/kg, but not at 0.3 or 2 mg/kg (F (5, 53) = 8.311, P < 0.0001; Post-hoc Tukey Test * P < 0.05, different from SNI). (E-J) Mice (male and female, n = 6 each per group) underwent SNI and 7 days later were injected with psilocybin or saline control and underwent behavioral testing (acute phase; [E-G]). Two weeks later they received a second injection of the same and underwent repeat behavioral testing (chronic phase; [H-J]). Cold sensitivity was assessed using the cold plate test. In the acute phase (E-G), SNI treated mice had significantly more paw lifts than controls. There was no significant decrease in paw lifts at any dose of psilocybin at the (E) 1 hour, (F) 4 hour, or 24 hour (G) time points (One-way ANOVA, 1 hour: F (4, 55) = 4.840 P=0.0020, 4 hours: F(4, 55) = 5.571, P=0.0008, 24 hours F (4, 55) = 3.520 P=0.0125; Post-hoc Tukey Test * P < 0.05, versus SNI). Two weeks later, in the chronic phase (H-J), psilocybin similarly had no effect at (H) 1 hour, (I) 4 hours, or 24 hours(J) after injection (One-way ANOVA, 1 hour: F (4, 55) = 5.370 P=0.0010, 4 hours: F (4, 55) = 5.571 P=0.0008, 24 hours: F(4, 55) = 6.190, P=0.0003; Post-hoc Tukey Test *, P < 0.05 versus SNI).

Figure 4.. Psilocybin produces dose-dependent hypothermia.

(A) Psilocybin produces a dose-dependent decrease in body temperature that emerges at 17 minutes, reaches a nadir around 30 minutes, and normalizes by 70 minutes (male and female mice, n=2 each) Two-way repeated measures ANOVA F (330, 1320) = 2.692 P<0.0001. Post-Hoc Tukey Test (not shown). (B,C) Thermal camera image of female mouse immediately after (B), and 30 minutes after (C) 10 mg/kg psilocybin injection. (D) 30 minutes after injection, body temperature is significantly lower in 10 mg/kg psilocybin as compared to control (males and females, n=2 each per group). One-way ANOVA F (3, 12) = 6.651 P=0.0068. Post-hoc Tukey Test, * P < 0.05, different from control.