NMDA Receptor Activation-Dependent Antidepressant-Relevant Behavioral and Synaptic Actions of Ketamine

Abstract

Ketamine is a well-characterized NMDA receptor (NMDAR) antagonist, although the relevance of this pharmacology to its rapid (within hours of administration) antidepressant actions, which depend on mechanisms convergent with strengthening of excitatory synapses, is unclear. Activation of synaptic NMDARs is necessary for the induction of canonical long-term potentiation (LTP) leading to a sustained expression of increased synaptic strength. We tested the hypothesis that induction of rapid antidepressant effects requires NMDAR activation, by using behavioral pharmacology, western blot quantification of hippocampal synaptoneurosomal protein levels, and ex vivo hippocampal slice electrophysiology in male mice. We found that ketamine exerts an inverted U-shaped dose-response in antidepressant-sensitive behavioral tests, suggesting that an excessive NMDAR inhibition can prevent ketamine's antidepressant effects. Ketamine's actions to induce antidepressant-like behavioral effects, up-regulation of hippocampal AMPAR subunits GluA1 and GluA2, as well as metaplasticity measured ex vivo using electrically-stimulated LTP, were abolished by pretreatment with other non-antidepressant NMDAR antagonists, including MK-801 and CPP. Similarly, the antidepressant-like actions of other putative rapid-acting antidepressant drugs (2R,6R)-hydroxynorketamine (ketamine metabolite), MRK-016 (GABA_Aα5 negative allosteric modulator), and LY341495 (mGlu_2/3 receptor antagonist) were blocked by NMDAR inhibition. Ketamine acted synergistically with an NMDAR positive allosteric modulator to exert antidepressant-like behavioral effects and activation of the NMDAR subunit GluN2A was necessary and sufficient for such relevant effects. We conclude rapid-acting antidepressant compounds share a common downstream NMDAR-activation dependent effector mechanism, despite variation in initial pharmacological targets. Promoting NMDAR signaling or other approaches that enhance NMDAR-dependent LTP-like synaptic potentiation may be an effective antidepressant strategy.SIGNIFICANCE STATEMENT The anesthetic and antidepressant drug ketamine is well-characterized as an NMDA receptor (NMDAR) antagonist; though, the relevance and full impact of this pharmacology to its antidepressant actions is unclear. We found that NMDAR activation, which occurs downstream of their initial actions, is necessary for the beneficial effects of ketamine and several other putative antidepressant compounds. As such, promoting NMDAR signaling, or other approaches that enhance NMDAR-dependent long-term potentiation (LTP)-like synaptic potentiation in vivo may be an effective antidepressant strategy directly, or acting synergistically with other drug or interventional treatments.

Keywords: AMPA receptor; LTP; NMDA receptor; antidepressant; hydroxynorketamine; ketamine.

Figure 1.

High doses of ketamine do not elicit antidepressant-relevant behavioral actions in mice. Mice received an injection of vehicle or different doses of racemic ketamine and were assessed for antidepressant-like responses 24 h later. A, Ketamine at the dose of 10 mg/kg significantly decreased escape failures in helpless mice, whereas the doses of 1, 3, 30, and 100 mg/kg did not exert significant antidepressant-relevant reductions in escape failures in the learned helplessness paradigm. B, Similarly, ketamine at the dose of 10 mg/kg reversed the decrease in sucrose preference of mice that underwent chronic social defeat stress whereas the high dose of 100 mg/kg did not elicit such an antidepressant-related response. Mice were tested for sucrose versus water preference during the 24-h period following drug administration. Data are the mean ± SEM; *p < 0.05, ***p < 0.001 as indicated by Holm–Šídák post hoc comparisons. See Table 1 for complete details on the statistical analyses and precise group sizes. CSDS, chronic social defeat stress; KET, racemic ketamine; SAL, saline; Treat, treatment.

Figure 2.

Blocking NMDAR activity prevents the antidepressant-relevant behavioral effects of ketamine. A, Mice received an injection of vehicle or the NMDAR channel blocker MK-801 (0.1 mg/kg) and 10 min later they received an injection of vehicle or racemic ketamine (10 mg/kg). MK-801 pretreatment prevented the antidepressant-relevant behavioral actions of ketamine in the learned helplessness paradigm. B, Similarly, pretreatment with the competitive NMDAR blocker (±)-CPP (10 mg/kg) blocked ketamine's antidepressant-relevant actions in the learned helplessness paradigm. C, MK-801 (0.1 mg/kg) pretreatment also prevented the anti-anhedonic actions of ketamine in mice that underwent chronic social defeat stress, as measured by the sucrose preference test. D, MK-801 (0.03 mg/kg) pretreatment prevented ketamine's actions on immobility time in the forced-swim test. E, Co-administration of subeffective doses of ketamine with the NMDAR positive modulator rapastinel induced a synergistic reduction of escape failures in helpless mice in the learned helplessness paradigm. In all paradigms, mice were tested 24 h following drug administration. Data are the mean ± SEM; *p < 0.05, **p < 0.01, ***p < 0.001 as indicated by Holm–Šídák post hoc comparisons. See Table 1 for complete details on the statistical analyses and precise group sizes. CSDS, chronic social defeat stress; KET, racemic ketamine; SAL, saline; Treat, treatment.

Figure 3.

Blocking NMDAR activity prevents the antidepressant-relevant behavioral effects of distinct rapid-acting antidepressant compounds. A, B, Mice received an injection of vehicle or the NMDAR channel blocker MK-801 and 10 min later were given an additional injection of vehicle or ketamine's metabolite (2R,6R)-hydroxynorketamine (HNK; 10 mg/kg) and tested in the forced swim test 24 h later. While (A) the dose of 0.1 mg/kg MK-801 completely prevented the antidepressant-like behavioral actions of (2R,6R)-HNK, (B) 0.03 mg/kg MK-801 did not prevent (2R,6R)-HNK's actions to decrease immobility time in the forced-swim test. MK-801 pretreatment (0.1 mg/kg) prevented the antidepressant-relevant behavioral actions of (C) the negative allosteric modulator of GABA_A receptors containing α5 subunits (GABA-NAM) MRK-016 in the forced-swim test. MK-801 pretreatment (0.1 mg/kg) prevented the antidepressant-like effects of (D) (2R,6R)-HNK, (E) MRK-016, and (F) the mGlu2/3 receptor antagonist LY341495 in the learned helplessness paradigm 24 h following drug administration. Data are the mean ± SEM; *p < 0.05, **p < 0.01, ***p < 0.001 as indicated by Holm–Šídák post hoc comparisons. See Table 1 for complete details on the statistical analyses and precise group sizes. KET, racemic ketamine; SAL, saline.

Figure 4.

Blocking NMDAR activity prevents the procognitive and synaptic actions of ketamine. A, Mice received an injection of vehicle or different doses of racemic ketamine and were tested for short-term novel object recognition memory 24 h after drug administration. Ketamine (10 mg/kg) enhanced the discrimination index, indicative of a procognitive effect, whereas a higher dose of ketamine (100 mg/kg) impaired object recognition memory. B, Administration of the NMDAR channel blocker MK-801 (0.1 mg/kg) 10 min before ketamine (10 mg/kg) prevented the procognitive effect of ketamine in the novel object recognition test. C, Representative western blot images for GluA1 and GluA2 AMPAR subunits from hippocampal synaptoneurosomes. D, E, Pretreatment with MK-801 prevented the ketamine-induced enhancement of synaptoneurosomal levels of GluA1 and GluA2 AMPAR subunits. F, Traces composed of representative sweeps from 5 min pre-tetanus (gray) and 56–60 min post-tetanus (black) from SAL-SAL, SAL-KET, MK-801-SAL, and MK-801-KET treatment groups. G, H, Pretreatment with MK-801 prevented the metaplastic effect of ketamine on long-term potentiation magnitude at the SC-CA1 synapse. Data are the mean ± SEM; *p < 0.05, **p < 0.01, ***p < 0.001 as indicated by Holm–Šídák post hoc comparisons. See Table 1 for complete details on the statistical analyses and precise group sizes. HFS, high-frequency stimulation; KET, racemic ketamine; LTP, long-term potentiation; MK, MK-801; SAL, saline.

Figure 5.

The antidepressant-like actions of ketamine are mediated through GluN2A activity. A, Mice received an injection of vehicle or the GluN2A-selective NMDAR negative allosteric modulator PEAQX (5 or 30 mg/kg) followed by an injection of vehicle or ketamine (10 mg/kg) 10 min later, and then were tested for reversal of helpless behavior 24 h later. PEAQX pretreatment, at both doses administered, prevented the antidepressant-relevant actions of ketamine to decrease escape failures of helpless mice. B, The GluN2A-selective NMDAR positive allosteric modulator GNE-5729 induced a decrease in locomotor activity of mice in the open-field test only at the highest dose administered (3 mg/kg). C, In the forced-swim test, GNE-5729 at the dose of 3 mg/kg significantly reduced immobility time of mice, indicative of an antidepressant response. D, Similarly, the dose of 1 mg/kg of GNE-5729 significantly reduced escape failures of helpless mice. Data are the mean ± SEM; *p < 0.05, ***p < 0.001 as indicated by Holm–Šídák post hoc comparisons. See Table 1 for complete details on the statistical analyses and precise group sizes. KET, ketamine; SAL, saline; VEH, vehicle.