Antidepressant-relevant concentrations of the ketamine metabolite (2 <i>R</i> ,6 <i>R</i> )-hydroxynorketamine do not block NMDA receptor function

Abstract

Preclinical studies indicate that (2<i>R</i>,6<i>R</i>)-hydroxynorketamine (HNK) is a putative fast-acting antidepressant candidate. Although inhibition of NMDA-type glutamate receptors (NMDARs) is one mechanism proposed to underlie ketamine's antidepressant and adverse effects, the potency of (2<i>R</i>,6<i>R</i>)-HNK to inhibit NMDARs has not been established. We used a multidisciplinary approach to determine the effects of (2<i>R</i>,6<i>R</i>)-HNK on NMDAR function. Antidepressant-relevant behavioral responses and (2<i>R</i>,6<i>R</i>)-HNK levels in the extracellular compartment of the hippocampus were measured following systemic (2<i>R</i>,6<i>R</i>)-HNK administration in mice. The effects of ketamine, (2<i>R</i>,6<i>R</i>)-HNK, and, in some cases, the (2<i>S</i>,6<i>S</i>)-HNK stereoisomer were evaluated on the following: (<i>i</i>) NMDA-induced lethality in mice, (<i>ii</i>) NMDAR-mediated field excitatory postsynaptic potentials (fEPSPs) in the CA1 field of mouse hippocampal slices, (<i>iii</i>) NMDAR-mediated miniature excitatory postsynaptic currents (mEPSCs) and NMDA-evoked currents in CA1 pyramidal neurons of rat hippocampal slices, and (<i>iv</i>) recombinant NMDARs expressed in <i>Xenopus</i> oocytes. While a single i.p. injection of 10 mg/kg (2<i>R</i>,6<i>R</i>)-HNK exerted antidepressant-related behavioral and cellular responses in mice, the ED₅₀ of (2<i>R</i>,6<i>R</i>)-HNK to prevent NMDA-induced lethality was found to be 228 mg/kg, compared with 6.4 mg/kg for ketamine. The 10 mg/kg (2<i>R</i>,6<i>R</i>)-HNK dose generated maximal hippocampal extracellular concentrations of ∼8 µM, which were well below concentrations required to inhibit synaptic and extrasynaptic NMDARs in vitro. (2<i>S</i>,6<i>S</i>)-HNK was more potent than (2<i>R</i>,6<i>R</i>)-HNK, but less potent than ketamine at inhibiting NMDARs. These data demonstrate the stereoselectivity of NMDAR inhibition by (2<i>R</i>,6<i>R</i>;2<i>S</i>,6<i>S</i>)-HNK and support the conclusion that direct NMDAR inhibition does not contribute to antidepressant-relevant effects of (2<i>R</i>,6<i>R</i>)-HNK.