Antinociceptive and Analgesic Effects of (2 R,6 R)-Hydroxynorketamine

Abstract

Commonly used pain therapeutics, such as opioid medications, exert dangerous side effects and lack effectiveness in treating some types of pain. Ketamine is also used to treat pain, but side effects limit its widespread use. (2R,6R)-hydroxynorketamine (HNK) is a ketamine metabolite that potentially shares some beneficial behavioral effects of its parent drug without causing significant side effects. This study compared the profile and potential mechanisms mediating the antinociception activity of ketamine and (2R,6R)-HNK in C57BL/6J mice. Additionally, this study compared the reversal of mechanical allodynia by (2R,6R)-HNK with gabapentin in a model of neuropathic pain. Unlike the near-immediate and short-lived antinociception caused by ketamine, (2R,6R)-HNK produced late-developing antinociception 24 hours following administration. Pharmacological blockade of α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) receptors with 2,3-dioxo-6-nitro-7-sulfamoyl-benzo[f]quinoxaline (NBQX) prevented the initiation and expressionof (2R,6R)-HNK antinociception, suggesting the involvement of α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptor-dependent glutamatergic mechanisms in the pain reduction-like responses. Blockade of opioid receptors with naltrexone partially prevented the antinociceptive effect of ketamine but was ineffective against (2R,6R)-HNK. Furthermore, (2R,6R)-HNK did not produce dystaxia, even when tested at doses five times greater than those needed to produce antinociception, indicating a superior safety profile for (2R,6R)-HNK over ketamine. Additionally, (2R,6R)-HNK reversed mechanical allodynia in a spared nerve injury model of neuropathic pain with similar short-term efficacy to gabapentin (within 4 hours) while outperforming gabapentin 24 hours after administration. These findings support the further study of (2R,6R)-HNK as a potentially valuable agent for treating different types of pain and establish certain advantages of (2R,6R)-HNK treatment over ketamine and gabapentin in corresponding assays for pain. SIGNIFICANCE STATEMENT: The ketamine metabolite (2R,6R)-HNK produced antinociception in male and female mice 24 hours after administration via activation of AMPA receptors. The effects of (2R,6R)-HNK differed in time course and mechanism and presented a better safety profile than ketamine. (2R,6R)-HNK also reversed allodynia in SNI-operated animals within 4 hours of treatment onset, with a duration of effect lasting longer than gabapentin. Taken together, (2R,6R)-HNK demonstrates the potential for development as a non-opioid analgesic drug.

Fig. 1.

(2R,6R)-HNK produced delayed antinociception (Panel B), while ketamine produced rapid and short-lived antinociception (Panel A). (2S,6S)-HNK did not produce antinociception at any time point (Panel B). The data were analyzed together using a repeated measures two-way ANOVA but are shown in two separate graphs for clarity. The error bars represent group means ± S.D. Comparisons are made between drug/dose and saline at the same time point: **P < 0.01, ***P < 0.001. N = 20 for saline control, N = 21 for ketamine 10 mg/kg, N = 10 for ketamine 30 mg/kg, and N = 22 for both HNK groups.

Fig. 2.

Male (Panel A) and female (Panel B) time course for latency to respond to hot plate stimulus following (2R,6R)-HNK 10 mg/kg treatment. Comparisons are made between the treatment and saline groups at the same time point: *P < 0.05. N = 30 for males and N = 36 for females. Panel C (males) and Panel D (females) represent the quarter log dose-response for (2R,6R)-HNK mediated antinociception measured 24 hours after treatment. (2R,6R)-HNK produced antinociception at doses of 10 and 18 mg/kg in both sexes, while the 32 mg/kg dose produced antinociception in female mice only. Asterisks indicate significant differences compared with the saline control. *P < 0.05, **P < 0.01, ***P < 0.001. N = 10–12 for males, and N = 17 for females. The error bars represent group means ± S.D. For the dose-response figures, the mean of the saline group is represented by the dashed line, while the ± S.D. are represented by dotted lines.

Fig. 3.

The effects of AMPA receptor antagonism with NBQX (10 mg/kg) or opioid receptor antagonism with naltrexone (1 mg/kg) on the initiation or expression of antinociception by (2R,6R)-HNK. The effects of (2R,6R)-HNK (10 mg/kg) were measured 24 hours after injection. Panel A: The NBQX given 30 minutes before (2R,6R)-HNK treatment blocked the initiation of (2R,6R)-HNK antinociception. Panel B: Naltrexone given 30 minutes before (2R,6R)-HNK treatment did not block the initiation of (2R,6R)-HNK antinociception. Panel C: NBQX given 24 hours following (2R,6R)-HNK treatment blocked the expression of (2R,6R)-HNK mediated antinociception. Panel D: Naltrexone given 24 hours following (2R,6R)-HNK treatment did not alter (2R,6R)-HNK antinociception. Panel E: Ketamine (10 mg/kg) mediated antinociception measured 10 minutes following treatment was unaffected by pretreatment with NBQX (10 mg/kg given 30 minutes prior), while naltrexone (1 mg/kg given 30 minutes prior) blocked ketamine antinociception. The error bars represent group means ± S.D. Group comparisons are indicated with horizontal lines. Asterisks represent treatment/saline to saline/saline comparisons: **P < 0.01, ***P < 0.001, and ****P < 0.0001. The # symbol represents treatment/NBQX to treatment/saline comparisons: # P < 0.05, ## P < 0.01, and ### P < 0.001. The @ symbol represents NBQX/saline compared with NBQX/ketamine: @@@@, P < 0.0001. N = 11–12 for the (2R,6R)-HNK experiments, N = 10 for the ketamine experiment.

Fig. 4.

The SNI condition produced mechanical allodynia 11 days after surgery. Time 0 represents mechanosensitivity results on day 11 before treatment. Results of a baseline measurement before surgery demonstrated no significant difference between groups (data not shown, average baseline before surgery for all groups was 4.3 ± 1.10 g f). Panel A: The experimental paradigm comprised once-daily treatment of three days and mechanosensitivity testing before daily treatment. Panel B: (2R,6R)-HNK reversed mechanical allodynia associated with the neuropathic pain condition within 4 hours of treatment, and the effect persisted for at least 24 hours. Repeat dosing did not appear to lessen the pain reduction-like effect. Gabapentin reversed mechanical allodynia 4 hours following treatment but did not produce a significant effect 24 hours following the first or third treatment. Vertical bars represent the group means ± S.D. Drug doses are compared with saline at the same time point, **P < 0.01, ***P < 0.001, ****P < 0.0001, N = 9-10 (N = 9 for saline and gabapentin 30 mg/kg groups, N = 10 for all other groups).