The mechanistic basis for the rapid antidepressant-like effects of ketamine: From neural circuits to molecular pathways

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The identification of the specific molecular targets related to the activity of rapid antidepressants provides an opportunity to better understand the etiological basis for their efficacy.In the clinic, the application of ketamine is hampered by the risk of misuse and the potential development of adverse effects, including neurotoxicity and dissociative reactions (Vieira et al., 2021;Yang et al., 2022;Gastaldon et al., 2021;Short et al., 2018;Kasper et al., 2021).In an effort to better manage or abrogate these side effects, researchers have sought to clarify the key proteins and brain circuits through which ketamine and related drugs influence the pathogenesis of MDD, providing opportunities for the development of new and more precise therapies for affected patients.The neural circuitry consists of a complex web of densely interconnected neurons within the brain that simultaneously coordinate information processing through activity that undergoes constant reorganization and refinement due to synaptic changes driven by experience.
An interest in neurostimulatory and neuromodulatory therapies aimed at resolving these aberrant connections has increased recently due to evidence suggesting that abnormal neuroconnectivity within brain networks responsible for the regulation of mood, behavior, and cognition contributes to the pathogenesis of MDD.Both translational neuroscience and psychiatric research have long focused on identifying more effective treatment options for MDD patients.There has been a shift away from models of depression focused solely on neurochemical deficits and toward models in which broader changes in the underlying neural circuitry enable key limbic nodes to induce durable changes in patient affect (Spellman and Liston, 2020).The construction of these models has been enabled by complementary human neuroimaging results and neurophysiological studies of animal models using chemogenetic and optogenetic approaches (Spellman and Liston, 2020).
Stable functional connectivity patterns have been detected through resting-state functional magnetic resonance imaging studies, even though the precise mechanisms through which rapid antidepressants affect functional brain networks are not well understood.For example, in a randomized, placebo-controlled functional connectome study of ketamine-treated anesthetized monkeys, the drug was found to primarily influence targets within cortical-limbic-striatal circuits while also reversing the effects of depression.The areas of the brain with the greatest decreases in functional connection were the nucleus accumbens (Nac), the posterior and subgenual cingulate cortices, and the orbital prefrontal cortex.Overall, these analyses suggested that NMDAR blockade was sufficient to trigger both local synaptic plasticity and the persistent reconfiguration of cortico-limbic-striatal circuit networks, highlighting opportunities for the development of therapeutic agents targeting particular circuits or loci to achieve long-term antidepressant efficacy (Lv et al., 2016).
Evidence presented by Anita E Autry et al. indicated that ketamine can rapidly block spontaneous NMDAR-mEPSCs, resulting in antidepressant-like effects on behavior.Evoked neurotransmission or behavioral circuitry disinhibition cannot elicit this fast-acting antidepressant activity, which instead relies on the enhancement of neurotransmission following increases in plasticity induced by NMDAR antagonist treatment (Autry et al., 2011).Much like ketamine, memantine functions as a noncompetitive antagonist of NMDAR activity but has not been found to exert any rapid antidepressant-like effects when used to treat individuals affected by MDD, suggesting that ketamine must function at least in part through mechanisms independent of NMDAR inhibition (Zarate Jr. et al., 2006).Consistently, several studies have found that the neurobiological basis for the activity of rapid antidepressants is closely tied to synaptic structural remodeling and plasticity (Krishnan and Nestler, 2008;Martinowich et al., 2013).
In this review, we discuss how ketamine and similar drugs rapidly alleviate depression by acting on specific neuronal circuits, brain regions, and molecular targets.These research results, primarily derived from preclinical studies, highlight the many different pathways that likely function synergistically in the treatment of MDD.Through this article, we hope to provide readers with new insights into these neurophysiological mechanisms to provide a foundation for efforts to overcome the barriers currently limiting the clinical deployment of rapid antidepressants.

Ketamine's rapid antidepressant-like effects are mainly mediated via prefrontal cortex circuits
The current generation of antidepressants used in clinical practice is hypothesized to work by normalizing activity within key brain circuits that have been dysregulated in depressed individuals.The low effectiveness of these medications, if they are not administered frequently for sufficient time, may be explained by the recurrent activation of downstream biochemical and signaling pathways that eventually permit the adaptive effects of these treatments (Martinowich et al., 2013).The roles of specific brain regions associated with the effects of rapid antidepressants in MDD are not independent of one another, as they are members of neural circuits linked together by nerve fibers.Depression is associated with structural and functional abnormalities impacting these circuits, and the restoration of normal neural pathway connectivity may provide an effective means of efficiently alleviating depressive symptoms.As such, drugs with rapid antidepressant activity are believed to function by rapidly reshaping and remediating the neural circuits in appropriate areas of the brain (Peng et al., 2020).
The prefrontal cortex (PFC) is crucial to the rapid antidepressant-like effects of ketamine.This has also been identified in animal research.For 70 years, the prefrontal cortex has in essence been defined as the part of the cerebral cortex that receives projections from the mediodorsal nucleus of the thalamus.Mice and rats possess fewer areas in the frontal lobe than primates, and all areas in the prefrontal cortex of mice and rats are agranular.At times the agranular cytoarchitecture is used as a definition of the rodent prefrontal cortex (Carlen, 2017).The medial PFC (mPFC) functions as an integrator of neural inputs from many different areas in the brain and it sends neuronal projections that extend to multiple limbic and cortical structures.These functional links provide the mPFC with the ability to shape a wide range of behavioral and cognitive processes, such as habit formation, attention, long-term memory, working memory, and emotional and inhibitory control (Hiser and Koenigs, 2018;Riga et al., 2014).Rapid behavioral effects of ketamine administration have been shown by investigations in animal research.The mPFC functions as a relay station, relaying information from the limbic system, midbrain and hindbrain involved in controlling emotional, reward, and stress responses.These regions include hippocampus (HP) (Georgiou et al., 2022), amygdala (Hare et al., 2019), ventral tegmental area (VTA) (Wu et al., 2021a), and the dorsal raphe nucleus (DRN) (Pham et al., 2020).In addition, the neural projections from entorhinal cortex (EC) to CA1 also involve in the rapid antidepressant effects of ketamine (Georgiou et al., 2022) (Fig. 1).
In a rapid-antidepressant neural-circuit study of ketamine, the PFC is among the most important brain circuits associated with this activity, serving as the center of a variety of neural loops that link various brain regions following the administration of ketamine.The primary regions sending input into the PFC is the limbicstructure, which include the HP.The primary output regions from the mPFC are the limbic, the midbrain and hindbrain, which include the BLA, VTA and DRN.The circuit which independent on PFC is the EC to CA1 projection.

Input regions of PFC
The HP forms part of the limbic system and plays a vital role in cognition, memory consolidation, emotional regulation, spatial navigation, and associative learning (Zhong et al., 2020;Biane et al., 2023;Qasim et al., 2023;Garvert et al., 2023;Terada et al., 2022).The HP is closely associated with both the development and treatment of MDD, as MDD patients exhibit pronounced reductions in hippocampal volume, and functional connectivity.The effects of antidepressants on HP and their ability to reverse stress-and MDD-related changes in this region may underlie the clinical benefits of ketamine (Lin et al., 2021).The HP-PFC circuit serves as a central mediator of memory and cognitive function, and MDD is associated with the dysregulation of this connectivity.As reported by Natalia Gass et al., an increase in HP-mPFC functional connectivity was observed to be associated with ketamine exposure and dosage during acute treatment in rats (Gass et al., 2014).Moreover, low-dose ketamine treatment can reportedly reduce inhibitory inputs into hippocampal pyramidal cells, thereby contributing to their disinhibition (Fan et al., 2018;Gerhard et al., 2020;Widman and McMahon, 2018), lowering the barrier to action potential generation in response to excitatory synaptic inputs (Widman and McMahon, 2020).Further investigation into the role of ventral hippocampus (vHP)-mPFC activity in the rapid effect of ketamine as an antidepressant was conducted by F R Carreno et al (Carreno et al., 2016) When lidocaine was used to inactivate the vHP, the authors found that this had no impact on the antidepressant-like effects of ketamine in model animals in a forced swimming test (FST), although this inhibition abrogated the sustained benefits of such treatment.Optogenetic DREADD-mediated vHP-mPFC pathway activation or specific pharmacogenetic activation of this circuit was sufficient to yield antidepressant-like effects mimicking those induced by ketamine, whereas the optogenetic inactivation of this vHP-mPFC axis fully reversed the antidepressant benefits of ketamine when performed at the time of FST analyses.These results demonstrate the importance of the vHP-mPFC circuit as a mediator of the effects of ketamine as a rapid antidepressant (Carreno et al., 2016).

Output regions of the PFC
The amygdala coordinates the activity of cortical networks to determine the physiological significance of emotional stimuli.Reversing depressive-like behaviors and boosting amygdala inputs to layer V cells in the medial prefrontal cortex have both been linked to the action of ketamine (Liu et al., 2015).Brendan D Hare et al. demonstrated that mPFC-BLA circuit invovled in the rapid onset effects of ketamine.They used an optogenetics study to show that activating the projection from the mPFC to the basolateral amygdala (BLA) resulted in rapid antidepressant-like effects.By increasing neuronal activity only in the BLA, light stimulation of D1 dopamine receptor (Drd1) neurons in the mPFC region of the brain suggested that the Drd1 neurons facilitated BLA area involvement in the fast anti-depressant-like effects (Hare et al., 2019).
Several stress-related MDD behaviors, including their onset and resolution, were found to be directly associated with dopaminergic neurons (DA neuron) in the midbrain, such as anhedonia and a drop in motivation (Wu et al., 2021a;Chaudhury et al., 2013;Tye et al., 2013).VTA DA neurons in vivo display both tonic firing at low frequency and phasic firing at high frequency.Repeated social-defeat stress, which is a widely used mouse model of MDD, induces an increase in phasic firing, which encodes reward-related signals (Chaudhury et al., 2013).Anatomical tracing and electron microscopy studies suggest that mPFC pyramidal neurons provide inputs to the VTA DA neurons projecting to the mPFC, thereby establishing excitatory connections (Beier et al., 2015;Carr and Sesack, 2000).trans-Synaptic tracing and optogenetic analyses have further confirmed thisprojections (Beier et al., 2015;Lodge, 2011;Xiao et al., 2018).The release of glutamate from mPFC terminals can elicit excitatory responses in VTA DA neurons, provoking activity that is synchronized across the VTA, mPFC, and other limbic structures (Gariano and Groves, 1988;Kumar et al., 2013;Tong et al., 1996;You et al., 2007).The VTA and mPFC are thought to play a pivotal role in modulating the effect of treatment on behavior.Restoring VTA DA activity and correcting behavioral impairments, as shown by Mingzheng Wu et al., requires a local infusion of ketamine in the mPFC.In vivo administration of ketamine led to a fast rise in mPFC Drd1-positive cell activity (Wu et al., 2021a).These authors also demonstrated that Drd1 activation-mediated DA signaling is essential for the ability of ketamine to restore glutamate-evoked dendritic spinogenesis in the mPFC, prolonging the effects of ketamine on behavior following corticosterone treatment.Chemogenetic Gαs-coupled signaling cascade activation downstream of the activation of Drd1 within the mPFC can also eliminate escape behaviors following learned helplessness (Wu et al., 2021b).Mingzheng Wu et al. posited that recurrent mPFC-VTA circuit activation may amplify initial ketamine-induced increases in mPFC activity, contributing to the prolonged behavioral benefits of ketamine administration even when it is no longer detectable in vivo, although further work will be necessary to test this model directly (Wu et al., 2021a).
The DRN is a brain area that modulates stress-related physiology and behavior, aggression, and affective disorders (Abrams et al., 2004;Pucilowski and Kostowski, 1983;Lowry et al., 2008).Researchers Thu Ha Pham et al. employed a preclinical method to investigate the function of the mPFC-DRN circuit, as well as the AMPAR/DRN and GABAAR/ mPFC, in the maintenance of the antidepressant-like effects of ketamine.They found that these effects were promoted by AMPAR activation in the DRN and 5-HT-glutamate release in the mPFC, whereas GABAAR was restricted by GABAAR activation GABA release in the mPFC (Pham et al., 2020).Neural projections from different brain areas, including the PFC, govern the serotoninergic neuron (5-HT neuron) in the DRN (Pollak Dorocic et al., 2014).It was found that 5-HT neurons in the DRN are involved in the antidepressant-like effects of ketamine, with regulation coming from the mPFC-DRN projections.The researchers discovered that the antidepressant-like effects of a mGlu2/3 receptor antagonist and ketamine may be due to the activation of 5-HT neurons in the DRN, which is controlled by stimulation of AMPA receptors in the mPFC (Fukumoto et al., 2016).

Other circuit
Polymnia Georgiou et al. showed that exposure of mice to the scent of male experimenter before ketamine administration activates corticotropin-releasing factor (CRF)-expressing cells in the EC that project to CA1, priming this pathway to ketamine's antidepressantrelevant effects both in vivo and in vitro.They also found that ketamine administered by a female experimenter will have antidepressantlike effects provided the injection is preceded by a stressor or other manipulation that activates the CRF EC-CA1 projection.In this study, Polymnia Georgiou et al. suggested that acute ketamine and (2R,6R)-HNK combined with CRF receptor agonists may be a novel treatment approach for mood disorders.They also showed that activation of the CRF EC-CA1 pathway is an important determinant of the antidepressant effects of ketamine (Georgiou et al., 2022).

Molecular targets associated with rapid antidepressant activity
Many different targets associated with a diverse range of signaling cascades in cells have been found to play an important role in mediating the rapid antidepressant of ketamine and related drugs, with many such targets playing roles associated with synaptic plasticity and structural remodeling (Martinowich et al., 2013) (Table 1).One of the most essential features of neurons is their ability to generate new synapses, a process known as synaptogenesis.By altering its subcellular structure in response to synaptic activity, synaptogenesis facilitates the integration of novel information to produce a suitable adaptive response in the future.Long-term potentiation (LTP) is one cellular model of memory and learning that has been used to investigate synaptogenesis and its underlying processes.Glutamate receptor insertion and synapse development in the spine occur in response to elevated neuronal activity.The rapid effect of ketamine as an antidepressant is widely believed to be mediated through glutamatergic transmission, which controls synaptic plasticity (Lener et al., 2017;Kavalali and Monteggia, 2012;Suzuki et al., 2021).Ketamine's pharmacokinetics and in vivo pharmacodynamics are complex; therefore, it is debatable whether or not it suppresses or stimulates glutamate.A competitive NMDA antagonist 2amino-7-phosphonoheptanoic acid a non-competitive NMDA antagonist Dizolcipine and a partial agonist at strychnine-insensitive glycine receptors 1-aminocylopropanecarboxylic acid showed antidepressantlike effects in rats in the early 1990s (Trullas and Skolnick, 1990).Dendritic atrophy caused by prolonged stress was prevented by inhibiting NMDARs or glutamate release (McEwen, 1999).These first results lent credence to the hypothesis that ketamine's mood-lifting effects arise from the NMDAR antagonistic effects on glutamatergic neurons, which in turn induce homeostatic synaptic plasticity and increase synaptic drive.The disinhibition theory contends that subanesthetic dosages of ketamine temporarily promote rather than block glutamate neurotransmission, in contrast to the glutamate inhibition (Moghaddam et al., 1997).It is interesting to note that the elevation of several neurotrophic/ growth factors was first correlated with acute stimulation of glutamate neurotransmission rather than inhibition (Patterson et al., 1992;Zafra et al., 1991).According to recent research, this process may also include the 5-hydroxytryptamine (5-HT) and γ-Aminobutyric acid (GABA) systems (Gigliucci et al., 2013;Zanos and Gould, 2018b).Through its disinhibitory function, the GABA system controls the glutamate system.According to the current preclinical research, the glutamate and 5-HT systems may work together to reduce the symptoms of MDD (Fig. 2).
The rapid antidepressant-like effects of ketamine is mediated through glutamatergic transmission, which induce homeostatic synaptic plasticity.The antidepressant-like effects of ketamine may emerge directly as a downstream consequence of NMDA glutamate receptor antagonism.These NMDA receptors activate eEF2 and decrease BDNF expression.Blockade of these receptors raises BDNF expression and shuttles AMPA receptors, enhancing synaptic efficacy (Krystal et al., 2019).Ketamine also antagonizes N-methyl-D-aspartate (NMDA) receptors on GABAergic interneurons and on post-synaptic neurons; the former disinhibits cortical glutamatergic neurons and the latter increases synthesis of BDNF (Lener et al., 2017).Ketamine increases extracellular glutamate.This leads to activity-dependent release of BDNF and stimulation of signaling cascades, including the mTOR translational system, the Erk pathway, and other pathways in dendrites of neurons.Induction of translation results in increased levels of PSD95, GluA1 and other synaptic proteins, providing the machinery required for increased synaptogensis and spine formation (Li et al., 2010).Apart from targeting neuronal NMDARs essential for synaptic transmission, ketamine also interacts with astrocytes (Stenovec et al., 2021).The level of Kir4.1 on astrocytes tightly regulates the degree of membrane hyperpolarization and the amount of bursting activity of neurons.KCNQ2 is essential for the sustained antidepressant-like effects of ketamine in glutamatergic neurons (Ma and Hashimoto, 2022).Ketamine was able to upregulate KCNQ2 mRNA expression in GLUT neurons, which could potentially occur via the CaM/CaN/AKAP5/NFAT transcriptional axis in GLUT neurons of the vHP, thereby improved the depressive symptoms.Neuronal hyperpolarization may de-inactivate T-type voltage-sensitive calcium channels (T-VSCCs), which in turn initiate NMDAR-dependent bursts and thereby increase suppression of downstream monoaminergic centres (Cui et al., 2018).Ketamine can also indirectly modulate serotonergic signaling through the inhibition of glutamatergic activity (Fukumoto et al., 2014).

NMDARs
Ketamine can function through the direct inhibition of extra-synaptic NMDAR or the suppression of spontaneous neurotransmission mediated by NMDAR signaling.Shuangshuang Ma et al. demonstrated that after a single systemic injection, ketamine continues to suppress burst firing and blocked NMDARs in the lateral habenula (LHb) for up to 24 h.This long inhibition of NMDARs is not due to endocytosis but depends on the use-dependent trapping of ketamine in NMDARs.The rate of untrapping is regulated by neural activity.Harnessing the dynamic equilibrium of ketamine-NMDAR interactions by activating the LHb and opening local NMDARs at different plasma ketamine concentrations, they were able to either shorten or prolong the antidepressant-like effects of ketamine in vivo (Ma et al., 2023).Panos Zanos et al. reported that ketamine worked in concert with an NMDAR-positive allosteric modulator to induce antidepressant-like behavioral effects that relied on the activation of the GluN2A subunit (Zanos et al., 2023).Furthermore, it is widely believed that GluN2B is responsible for the antidepressant-like effects of ketamine.Santosh Pothula et al. showed that knockdown of GluN2B in neurons expressing Gad1 but not Camk2a prevented the effects of ketamine.Ketamine have antidepressant-like effects, and this action is mediated via pyramidal neurons in the mPFC that express the Drd1 gene.These results highlight the distinctive physiological triggers and convergence on Drd1-pyramidal cell signaling, that underlie the antidepressant-like effects associated with the blockade or manipulation of NMDAR signaling (Pothula et al., 2021).
The effect of ketamine on the expression of GluA2 in the brain has been addressed by a few research studies.Ex vivo electrically-stimulated LTP analyses revealed that GluA2 upregulation and hippocampal metaplasticity were associated with ketamine's antidepressant-like behavioral effects.In the mesolimbic circuit, Olga Skiteva et al. found that modest doses of ketamine caused long-lasting alterations in GluA2 contributions to synaptic AMPARs and calcium permeability, whereas the opposite was true for VTA DA neurons and NAc SPNs.Thus, the removal of GluA2-deficient CP-AMPARs is linked to ketamine-associated glutamatergic synapse metaplasticity onto VTA DA neurons, and ketamine may enhance the insertion of these GluA2-deficient CP-AMPARs into NAc SPNs (Skiteva et al., 2021).

Metabotropic glutamate receptor (mGluR)
Ketamine research has demonstrated the significance of mGluRs in the MDD development and treatment (Lener et al., 2017).There is strong evidence that agents that target mGlu2/3 and mGlu5 exhibit rapid antidepressant activity (Chaki and Fukumoto, 2018).Panos Zanos et al. found that the ketamine metabolite (2R,6R)-HNK exerts mGlu2 receptor-dependent antidepressant actions.The (2R,6R)-HNK mechanism of action as an antidepressant is that it synergistically exerts antidepressant-relevant activities and increases gamma qEEG oscillations when taken with subeffective dosages of a mGlu2/3 receptor antagonist (Zanos et al., 2019).One clinical study found that ketamine infusion was related to decreased mGluR5 availability, which persisted for 24 h after administration in both the control and depression-model groups.These rapid reductions in mGluR5 availability are likely a result of rapid surges in extracellular glutamate that drive the downregulation or internalization of mGluR5.Consistently, a single ketamine dose can promote a rapid but transient increase in glutamate efflux and cycling.It is believed that the rapid effects of ketamine as an antidepressant depend on the glutamate surge caused by the medication, as well as the internalization of mGluR5 (Esterlis et al., 2018).

Role of the GABA system in ketamine antidepressant treatment
In mouse model systems, ketamine administration has been demonstrated to increase signaling by GABA, the brain's principal inhibitory neurotransmitter (Duman et al., 2019).The disinhibition hypothesis posits that the initial inhibition of GABAergic interneurons by ketamine results in the subsequent disinhibition of key glutamatergic neuronal activity.As an NMDAR open-channel blocker, ketamine has the potential to increase the sensitivity of tonic-firing GABA interneurons, leading to a rapid disinhibition of pyramidal neurons and consequent transitory glutamate burst at low dosages.Homeostatic selftuning processes that maintain the balance of excitation and inhibition (E/I) and the average firing rates of neurons are crucial to the stable functioning of neuronal networks in the face of abrupt fluctuations in neural excitability (Turrigiano, 2012).Ketamine acts on GABA receptors as an indirect control of the glutamate system.XiaoHui Tang et al. suggested that ketamine might enhance the synthesis of GABA and modify the plasticity of astrocytes via GABAAR α1 downregulation.This raises levels of GABA and facilitates the GABA conversion into ATP, thereby leading to a rapid-acting antidepressant-like action (Tang et al., 2023).Zhen Ren et al. observed behavioral changes consistent with depression in mice with a heterozygous deletion of GABAAR γ2 (γ2 +/− ), accompanied by decreased expression and function of NMDAR and AMPAR in homeostatic hippocampal and PFC NMDAR and AMPAR (Ren et al., 2016).In these mice, ketamine was sufficient to reverse these behavioral and glutamate receptor deficits while enhancing GABA synaptic function.Incubating cultured GABAAR γ2 +/− neurons in the presence of ketamine was associated with an increase in levels of VGAT and gephyrin, which are markers of GABA synapses (Duman et al., 2019;Ren et al., 2016).

5-HT receptors enhance the rapid antidepressant effect
The 5-HT transporter (SERT) serves as the primary target for many effective antidepressants on the market, highlighting the link between 5-HT and MDD incidence.Through its ability to regulate the levels of free active neurotransmitters within the synaptic cleft, SERT mediates serotonergic neurotransmission and thus serves as a key target for conventional antidepressants such as SSRIs (Wan et al., 2021).Enhancing the strength and length of 5-HT-mediated signaling, SSRIs raise extracellular 5-HT levels, providing effective treatment in many patients, although the therapeutic effects generally only manifest after a several weeks of treatment (Blier et al., 1987;Yamanaka et al., 2014).
The current investigations characterised the rapid antidepressantlike effects of ketamine are related to 5-HT system (Gigliucci et al., 2013;du Jardin et al., 2016a;du Jardin et al., 2016b).Ketamine's antidepressant-like effects may be attributed to the activation of AMPARs in the medial prefrontal cortex and subsequent activation of certain 5-HT neurons in the DRN (Fukumoto et al., 2016).The rapid antidepressant-like effects of ketamine have been linked to the 5-HT1B receptor more often than any other 5-HT receptor.The participation of additional 5-HT receptors in this process is poorly understood (Fukumoto et al., 2014;Yamanaka et al., 2014;du Jardin et al., 2016b;du Jardin et al., 2017;Grieco et al., 2017;Kos et al., 2006).Increased AMPAR activity underlies the effects of ketamine on 5-HT release and 5-HT1B receptor binding in the neostriatum and ventral pallidum (VP) of rats and rhesus monkeys, respectively (Yamanaka et al., 2014).Using voxel-based analysis of standardized brain images, researchers found that ketamine treatment increased binding of 5-HT1B receptors in the Nac and VP and reduced binding of SERT in these same areas.These effects were blocked by the pretreatment of the AMPAR antagonist, and 2, 3-dihydroxy -6-nitro -7-sulfamoylbenzo(f)quinoxaline (NBQX), indicating that ketamine-induced modifications in 5-HT1B receptor binding are linked to AMPAR activation.NBQX can disrupt the antidepressant activity of ketamine in rodent models, supporting a model where altered serotonergic neurotransmission underlies such activity, particularly through postsynaptic 5-HT1B receptor upregulation in the Nac and VP (Yamanaka et al., 2014).
As the serotonergic and glutamatergic neurotransmission pathways interact strongly with one another, ketamine may indirectly modulate serotonergic signaling through the inhibition of NMDAR activity.Indeed, several preclinical studies have documented the importance of serotonergic neurotransmission in the effects of ketamine as an antidepressant (Gigliucci et al., 2013;Fukumoto et al., 2014;Celada et al., 2004).As such, ketamine can alleviate the symptoms of depression by modulating this serotonergic signaling and/or through the activation of 5-HT receptor signaling and the consequent engagement of appropriate downstream signaling pathways (du Jardin et al., 2016a).

Rapid antidepressant molecular mechanism centered on neurotrophic factors
The neuroprotective and mood-lifting properties of antidepressants are associated with brain-derived neurotrophic factor (BDNF) (Pattwell et al., 2012;Martinowich et al., 2007).According to the neurotrophic factor hypothesis, BDNF is closely involved in the rapid antidepressant action of ketamine.BDNF regulates synaptic plasticity, such as LTP, by interacting with its high-affinity receptor, tropomyosin receptor kinase B (TrkB), and triggering subsequent intracellular signaling (Minichiello, 2009).Lisa M Monteggia et al. confirmed that ketamine's rapid effects on behavior rely on BDNF-TrkB signaling, which requires quick upregulation of BDNF at the protein level, resulting in a rapid rise of dendritic BDNF levels post-treatment (Monteggia et al., 2013).In a similar vein, PeiYi Lin et al. investigated the effects of ketamine on depression by deleting TrkB or BDNF in postsynaptic CA1 or presynaptic CA3 areas of the Schaffer collateral pathway.While BDNF deletion in both the CA3 and CA1 regions was sufficient to ablate synaptic potentiation in response to ketamine, only the deletion of postsynaptic CA1 TrkB expression interfered with ketamine activity.These results thus confirmed that CA1 BDNF-TrkB signaling activity is essential for the rapid behavioral effects of ketamine, facilitating synaptic potentiation and underscoring the role of this synaptic locus in this therapeutic context (Lin et al., 2021).Evidence suggests that the BDNF-TrkB system is involved in the rapid-acting antidepressant-like actions of TGF-β1, thus showing comparative effectiveness.(R)-ketamine triggers the induction of TGF-β1 or TGF-β1 by binding via interacting with its receptor TGF-β receptor 1/2 in the microglia.Following this, the released BDNF binds to its receptor TrkB, initiating the MEK-ERK-CREB signaling pathway, which in turn promotes synaptogenesis and elicits antidepressant-like effects (Wei et al., 2022).
Another key molecule that acts in conjunction with BDNF is Eukaryotic elongation factor 2 (eEF2).eEF2 is one of several key proteins associated with protein translation, initiation, and elongation expressed in post-synaptic dendrites (Suzuki and Monteggia, 2020).Anita E Autry et al. found that eEF2 kinase activity can be disrupted at rest by NMDAR blockade through the administration of ketamine, thereby suppressing eEF2 phosphorylation and preventing the translational upregulation of BDNF.Rapid antidepressant-like behavioral effects in response to eEF2K inhibitor treatment (Autry et al., 2011).The spontaneous activation of NMDAR signaling can trigger eEF2Kmediated phosphorylation of eEF2.This causes this protein to dissociates from the translational machinery, suppressing translational activity.This effect is suppressed by inhibiting the NMDA pathway, resulting in eEF2 dephosphorylation and increase in target transcript translation (Autry et al., 2011;Monteggia et al., 2013).Chinnakkaruppan Adaikkan et al. further revealed a link between rapid ketamine-induced antidepressant activity and a decrease in eEF2K-mediated eEF2 phosphorylation through analyses of biochemical and behavioral changes in eEF2Kknockout mice.The deletion of this kinase was associated with the absence of the Thr56 phosphorylation of eEF2.This absence played a role in enhancing protein synthesis and increasing both biochemical and behavioral resistance to the antidepressant-like effects of ketamine in these animals (Adaikkan et al., 2018).Consistent with these findings, Lisa M Monteggia et al. found that the treatment of naïve mice with ketamine or other NMDAR antagonists rapidly induced antidepressantlike behavioral changes, mediated by BDNF-and TrkB-dependent mechanisms.Specifically, ketamine was associated with a reduction in eEF2 phosphorylation, thereby de-repressing BDNF mRNA translation.These authors further confirmed the rapid antidepressant-like effects of eEF2K inhibitors.Overall, their findings highlight a clear cause-andeffect relationship between NMDAR blockade during resting states and rapid elevation in dendritic BDNF protein levels, suggesting a clinically and behaviorally significant role for the eEF2K-medicated BDNF regulation as a target associated with antidepressant activity (Monteggia et al., 2013).
The rapid effects of ketamine as an antidepressant have been also linked to many other neurotrophic factors and growth factors, such as neurotrophic growth factor VGF (non-acronymic) and vascular endothelial growth factor (VEGF) (Salton et al., 2000;Deyama et al., 2019a).VGF is strongly regulated by both BDNF and neuronal activity in the CNS.C Jiang et al. demonstrated that VGF downregulation in the hippocampus is controlled by the BDNF/TrkB signaling pathway.This observation is particularly significant due to VGF's critical involvement as a regulator of MDD-like behavior in the hippocampus.Notably, ketamine treatment induces rapid upsurges in VGF translation, a response that is suppressed by decreases in VGF expression.These effects are dependent on the rapid translation and secretion of BDNF.The rapid L. Ren translation of BDNF and VGF forms a positive feedback loop that regulates and mediates the effects of ketamine treatment (Jiang et al., 2018).Vascular endothelial cells, glial cells, and neuronal cells all express VEGF protein in significant quantities (Deyama et al., 2019a).A lower VEGF expression has been documented in the frontal cortex and hippocampus in the MDD (Duman and Monteggia, 2006;Elfving et al., 2010;Heine et al., 2005).Satoshi Deyama et al. revealed that the simultaneous administration of a VEGF-neutralizing antibody was sufficient to ablate both the rapid-acting and sustained antidepressant-like effects of BDNF within the mPFC.Furthermore, the antidepressant-like effects of BDNF signaling may also be mitigated by targeting VEGF deletion in mPFC neuronal populations.Selective injection of a VEGF-Flk-1 antagonist was found sufficient to prevent BDNF-mediated increases in dendritic complexity in primary cortical neurons.These findings emphasize the vital interplay between different entities and imply that BDNF is pivotal for enabling VEGF's behavioral and neurotrophic effects.Notably, a reciprocal regulatory interaction exists between BDNF and VEGF activity, as VEGF signaling is essential for the fast antidepressant-like and neurotrophic effects of BDNF (Deyama et al., 2019a).

The mTOR signaling pathway
Activation of the mTOR signaling pathway has been linked to the production of synaptic proteins in certain synapses, contributing to the expression of proteins essential for spine synapse formation, maturation, and function (Li et al., 2010).Nanxin Li et al. observed that the administration of ketamine to model rats rapidly activated the AKT-mTOR pathway and that this activation was associated with elevated levels of synaptic signaling proteins and an increase in the number and activity of spine synapses in the PFC.Remarkably, the blockade of mTOR signaling eliminated the synaptogenic and behavioral responses elicited by ketamine in mouse models of MDD (Li et al., 2010).Interestingly, the antidepressant-like effects of (S)-norketamine, the major metabolite of (S)-ketamine, were blocked by the mTOR inhibitor rapamycin (Yang et al., 2018a).In conclusion, this implies that mTOR plays an important role in the antidepressant-like effects of (S)-ketamine, however, its role is comparatively less for (R)-ketamine (Yang et al., 2018b).These ketamine-associated changes are the opposite of stressrelated synaptic deficits, and may thus underlie the rapid-acting antidepressant-like effects of NMDAR inhibition (Li et al., 2010).M M Harraz et al. further explored the role of interactions between NO/ GAPDH/Siah1 pathway components and the small G protein Rheb in the regulation of mTOR activity.The results of these studies showed that nitrosylated GAPDH, Siah1, and Rheb formed a ternary complex.In this complex, Siah1 has ubiquitin E3 ligase activity, thereby promoting the proteasomal degradation of Rheb and disrupting its ability to promote mTOR activation.NMDA signaling via this NO/GAPDH/Siah1 axis thus promotes Rheb degradation and suppresses mTOR signaling activity, whereas the NMDAR inhibitor ketamine can reverse these changes, contributing to its antidepressant efficacy (Harraz et al., 2016).
Direct evidence for ketamine's capacity to induce the activation of mTORC1-4E-BP signaling within cortical pyramidal excitatory cells supports the involvement of 4E-BP1 and 4E-BP2 in the ketamine's antidepressant-like effects and the accompanying synaptic plasticity in the hippocampus.Ketamine targets and activates mTORC1, which functions primarily by promoting cap-dependent mRNA translation via phosphorylation and deactivation of 4E-BPs (Aguilar-Valles et al., 2021).Members of the eIF4E-binding protein (4E-BP) family are responsible for both specific and general inhibition of translational activity within cells, and serve as key regulators of synaptic plasticity and various other cellular processes (Banko et al., 2005;Bidinosti et al., 2010).
Though several preclinical studies have shown that ketamine administration increases mTORC1 signaling (Li et al., 2010;Harraz et al., 2016;Zhou et al., 2014a;Yang et al., 2013), a study showed that a single dose rapamycin pretreatment failed to block the antidepressant effects of ketamine.This contradictory result also suggests that rapamycin may also play a role in prolonging ketamine (Abdallah et al., 2020).

The Erk signaling pathway
A relevant implication of the ERK signaling pathway in MDD has been highlighted.Activation of ERK1/2 has been suggested to be important for synaptic remodeling and plasticity (Di Benedetto et al., 2013).ERK has also been shown involvement in the rapid antidepressant mechanism of ketamine (Kang et al., 2022).Research has shown that low concentrations of ketamine increased levels of phospho-ERK in a dose and time dependent manner (Lepack et al., 2016).The Erk pathway has different antidepressant mechanisms in (S)-ketamine and (R)-ketamine.The ERK pathway may only be specific to the mechanism of (S)-ketamine, as (R)-ketamine did not show changes in levels of phosphorylation in the mouse PFC.Ketamine was also associated with transient increases in the levels of phosphorylated ERK1/ERK2 and PKB/Akt, both of which are components of growth factor signaling pathways related to mTOR signaling activity.Administration of an ERK inhibitor prior to treatment blocked the antidepressant-like effects of (R)-ketamine, while this effect was not observed with (S)-ketamine.Further, (R)-ketamine attenuated the ERK reduced phosphorylation in the hippocampus and prefrontal cortex of susceptible mice in the same model, which was not the case for (S)-ketamine (Yang et al., 2018b).Zhenzhong Ma et al. showed TrkB-dependent ERK activation is required for ketamine-induced progenitor differentiation and antidepressant response (Ma et al., 2017).

The GSK3 signaling pathway
Both GSK3α and GSK3β are regulators of myriad neuronal functions including neurogenesis, neuronal structure, synaptic plasticity, and gene expression.Consequently, the kinase GSK3 emerges as a promising therapeutic target for addressing MDD.In the absence of stimulation, GSK3 exhibits partial activation, mainly dictated by inhibitory phosphorylation at the N-terminal Ser21 in GSK3α and Ser9 in GSK3β.E Beurel et al. demonstrated the rapid suppression of GSK3 by ketamine in the brains of treated mice underscoring its necessity for the rapid induction of antidepressant activity in response to ketamine administration in a learned helplessness model of MDD (Beurel et al., 2011a).In rodent models, the administration of subanesthetic ketamine doses, known for their rapid antidepressant-like effects, is also associated with increases in serine phosphorylation of both GSK3α and GSK3β, effectively inhibiting their activities.This study used a depression mouse model to establish the necessity of GSK3 inhibition for the antidepressant outcomes of ketamine.Wei Zhou et al. identified that Akt mediates the phosphorylation of GSK-3β in rat PFC during the process of ketamine exerting rapid antidepressant actions (Zhou et al., 2014b).However, the complete role of GSK-3 is still not clearly understood.For example, in contract to ketamine administration, the GSK-3 inhibitor SB216763 did not induce a rapid antidepressant action in mice, when exposed to both acute screening tests and a chronic mild stress paradigm (Ma et al., 2013).On the other hand, the combined treatment of ketamine and SB 216763 demonstrated effectiveness at 24 h (Liu et al., 2013).E Beurel's experiment utilized a GSK3α 21A/21A /β 9A/9A knock-in mouse model with serine-to-alanine mutations.The study revealed that ketamine promote serine phosphorylation of GSK3, contributing to its antidepressant efficacy in these mice (Beurel et al., 2011b).Consequently, this paradoxical result may arise from disparities between the impacts of endogenous and exogenous GSK3 inhibition.Further investigation in this area is warranted.

Ion channel proteins
Recent preclinical evidence has highlighted potassium channels as promising novel molecular targets for ketamine treatment.Among these, Kir4.1 and KCNQ2 are two emergent players.Kir4.1 has been identified in studies as the primary astrocytic inwardly-rectifying potassium channel, using both genetic and pharmacological manipulations.Animal models of MDD have demonstrated an elevated expression of Kir4.1 in the LHb, specifically in the astrocytic membrane processes surrounding the neuronal soma.Notably, manipulating astrocyte expression of Kir4.1 either upwards or downwards has elicited opposing effects on neuronal bursting and depressive symptoms, underscoring the strong influence of Kir4.1's astrocyte expression on the activity of LHb neuron bursting and membrane hyperpolarization levels (Cui et al., 2018).
The voltage-dependent potassium channel KCNQ2 has emerged as a new target for treating MDD.The influence of ketamine is partially mediated by the activation of the kcnq2 gene in glutamatergic neurons located in the ventral hippocampus, as initially demonstrated by Juan Pablo Lopez et al (Lopez et al., 2022) As the main member of this protein family expressed in the CNS, KCNQ2, together with other variant isoforms, generate a distinctive M-current (Barrese et al., 2018).This current is responsible for regulating resting membrane potential and the dampening repetitive neuronal firing, thereby influencing the overall excitability of neurons (Baculis et al., 2020).The rapid antidepressantlike effects of ketamine were observed by Juan Pablo Lopez et al. in the FST as early as 30 min after administration.However, these effects were interfered in vivo when Kcnq2 expression was specifically silenced in vHP, or when KCNQ channel activity was systemically modulated through mediation of drug.Although changes in the expression of Kcnq2 at the mRNA level were detectable following a two-day treatment interval, no corresponding transcriptional shifts were evident at 30 min post-injection.This implies that Kcnq2 transcriptional regulation may be primarily responsible for mediating the long-term effects of ketamine treatment, rather than its rapid antidepressant efficacy.In contrast, the direct CaM-mediated activation of KCNQ2 protein appears to shape this rapid effect.Notably, Kcnq2 upregulation following ketamine treatment was impeded at all analyzed time points when CaM and L-type calcium channels (L-type VDCC) were blocked, while inhibition of CaN only suppressed ketamine-related changes in Kcnq2 expression at 6 h posttreatment.Ketamine thus appears to initiate a scascade of transcriptional mechanisms that ultimately drive changes in Kcnq2 transcription within cells, rather than having any immediate or direct effect on this gene.Furthermore, the authors established that ketamine was able to upregulate Kcnq2 mRNA expression in GLUT neurons in vitro (1, 2, and 6 h post-treatment) and in vivo (2 days post-treatment), which could potentially occur via the CaM/CaN/AKAP5/NFAT transcriptional axis in GLUT neurons of the vHP, thereby supporting the sustained alleviation of depression-related symptoms (Lopez et al., 2022).
L-type VDCC are members of the voltage-gated antidepressant-like effectscalcium channel family and play a vital role in various neuronal functions.Evidence has suggested that these channels are also involved in the rapid of ketamine.AMPA receptors increase intracellular calcium levels in mouse cortical and hippocampal neurons via the L-type VDCC activation (Giesen et al., 2020).Ketamine's antidepressant-like effects rely on the stimulation of calcium-permeable AMPA receptors, resulting in increased BDNF levels via the mTOR complex 1 activation (Lavender et al., 2020).Duman's group showed that administering verapamil or nifedipine, both L-type calcium channel antagonists with distinct structures, prior to ketamine administration, effectively inhibits the behavioral effects of ketamine in FST.Treatment with ketamine induces the release of BDNF in primary cortical neurons, and this effect is blocked by inhibiting L-type VDCCs or AMPA receptors.This indicates that the ketamine antidepressant-like effects are brought about through the activation of L-type VDCCs and the subsequent BDNF release (Lepack et al., 2014).BDNF-TrkB complexes initiate dendritic spine remodeling via calcium signaling (Adasme et al., 2011).ANA-12, a small-molecule and selective noncompetitive antagonist of TrkB, has been shown to inhibit the antidepressant-like effects of both (S)-and (R)ketamine in mice susceptible to chronic social defeat stress (Yang et al., 2015).The potential role of L-type VDCC has also been shown when increased calcium levels in spines and dendrites induced by BDNF in hippocampal dentate granule cells were decreased through the use of Ltype calcium channel blockers (Robinson et al., 2021).The entry of calcium through L-type VDCCs triggers the expression of the BDNF receptor, TrkB (Kingsbury et al., 2003).Activation of L-type VDCCs and NMDA receptors leads to the influx of calcium into cells, thus triggering the BDNF-TrkB complex's internalization and activating BDNF signaling.At low subanesthetic doses, ketamine's ability to block most of the NMDA receptors is limited, indicating that at such doses, ketamine may not fully inhibit calcium-permeable NMDA receptors.In this context, the BDNF synthesis and release are triggered by L-type VDCCs activation by the ketamine the subanesthetic concentrations, and not by the blockade of NMDA receptors.Hence, the calcium-BDNF link through L-type VDCCs may function as a positive feedback loop.Conversely, an excessive calcium influx through L-type VDCCs is recognized to cause neurotoxicity.Thus, the L-type VDCCs activation by ketamine subanesthetic doses is probably tightly controlled to induce an advantageous impact on depression without triggering neurotoxicity through prolonged calcium influx into the cells (Robinson et al., 2021).

CaMKII-related molecules and pathways
The multifunctional enzyme CaMKII exhibits substantial presynaptic enrichment and controls the release of neurotransmitters through various mechanisms, including the regulation of synapsin-dependent translocation of synaptic vesicle from cellular that reserves into the vesicle pool readily for rapid release (Wang, 2008;Greengard et al., 1993).A pronounced drop in the autophosphorylation of CaMKIIa at Thr286 was observed in hippocampal synaptosomes following the administration of one dose of ketamine at subanesthetic levels, reducing CaMKIIa binding to syntaxin 1 A and thereby disrupting the assembly of the SNARE complex (Muller et al., 2013;Lazarevic et al., 2021).Chinnakkaruppan Adaikkan et al. further expended these investigations by treating WT C57BL/6 mice using the tatCN21 peptide, which specifically inhibits CaMKII.Delivery of tatCN21 throughout the body was linked to higher levels of behavioral resistance to ketamine therapy and an upswing in overall protein synthesis.Additionally, ketamine treatment was linked to changes in the regulation of CaMKII function, as shown by the autoinhibition (pT305 phosphorylation) and autoactivation (pT286) of hippocampal and cortical CaMKII.Additionally, the activation of CaMKII was found to control delays in GluA1 induction, as observed 24 h after ketamine therapy (Adaikkan et al., 2018).
The presynaptic activity of ketamine and its active metabolite (2 R, 6 R)-HNK was demonstrated by Vesna Lazarevic et al., employing a variety of pharmacological analyses.Both compounds triggered a retrograde action on inhibitory A1 receptors (A1Rs) in glutamatergic nerve terminals, achieving this by increasing adenosine release from presynaptic AMPARs.Activiation of these receptors effectively hampers downstream signaling mediated by synapsin and CaMKII, impeding the redistribution of synaptic vesicle from intracellular reserves to the rapidly releasable pool and subsequently diminishing glutamate release (Lazarevic et al., 2021).
Compelling evidence establishes a robust connection between the operation of synaptic proteins and the underlying neurobiological mechanisms of ketamine's action.Ketamine and a range of other antidepressants promote the upregulation of Homer1a, suggests that the induction of this gene potentially facilitates the antidepressant activities of these drugs.Homer proteins are closely associated with the postsynaptic density, forming a polymeric network and thereby regulating glutamate receptor functionality.Amrei Holz et al. demonstrated in a range of experiments that the intravenous administration of a cellmembrane-permeable form of TAT-Homer1a, simulating the upregulation of Homer1a, was sufficient to induce rapid antidepressant activity.Similar to ketamine, TAT-Homer1a upregulated mGlu5 signaling and, thereby, influencing protein phosphorylation in the mTOR pathway, as well as promoting AMPA receptor production and activation in vitro and in vivo.The ability of Homer1a to induce antidepressant activity was dependent on mGlu5 activity in excitatory CaMK2a neurons, while also requiring elevated levels of AMPA receptor translation, trafficking, and activity (Holz et al., 2019).
Yan Yang et al. discovered that the antidepressant-like effects of ketamine were associated with NMDAR-dependent bursting activity in the LHb.Animals with a depression-like phenotype exhibited significantly enhanced burst activity and theta-band synchronization in LHb.However, ketamine administration abrogated these changes.Consistently, burst-evoking LHb photostimulation was sufficient to induce anhedonia and despair-related behavior.Such LHb bursting was dependent on NMDARs and low-voltage-sensitive T-type calcium channels.The local blockade of either of these receptor types within the LHb resulted in the rapid induction of antidepressant-like effects (Yang et al., 2018c).Interestingly, Zheng Tian et al. reported that unlike (R)ketamine, the low-voltage-sensitive T-type calcium channel blocker ethosuximide did not have rapid or sustained antidepressant-like effects in a chronic social-defeat stress model (Tian et al., 2018).A preclinical study has also found that ethosuximide did not produce robust ketamine-like antidepressant actions in adult patients with major depressive disorder (Zhang et al., 2021b).These findings indicate that the relying solely on T-type calcium channels for rapid antidepressant therapy may not be sufficient.Indeed, Yan Yang et al. confirmed that the role of LHb bursting in ketamine rapid-antidepressant therapy relies on low-voltage-sensitive T-type calcium channels, however, the role of NMDARs cannot be ignored (Yang et al., 2018c).

Conclusion
Ketamine is a phencyclidine structural analog and a non-competitive antagonist of the NMDA receptor (Li and Vlisides, 2016).One of the first laboratories to investigate the synaptic plasticity of ketamine was the Duman laboratory, and the Duman team has provided a clearer interpretation of the temporal activation of the prefrontal mTOR signaling pathway induced by ketamine and the changes in downstream synaptic proteins that it triggers (Li et al., 2010).In particular, 24 h was proposed as a critical time point to detect the formation of synaptic plasticity in response to rapid antidepressants.The results of these experiments laid the framework for creating new faster antidepressant medications.The persistence of rapid antidepressant-like effects at time points beyond the drug's half-life, both in screened drug models and in animal diseasemodel assays, has important implications for research into rapid antidepressants, namely, the persistence of rapid antidepressant behavioral effects induced by the induction of synaptic plasticity.The persistence of the rapid antidepressant effect cannot be separated from its actual rapid antidepressant effect.An important point to consider is that the onset time of antidepressants in animal acute screening models can only serve as a reference.For example, the SSRI drug fluoxetine can also show antidepressant-like effects shortly after administration in an acute screening model (Roni and Rahman, 2015).This contrasts with the typical onset time of several weeks or more in clinical practice (Davey et al., 2019;Amsterdam et al., 1997).In preclinical research focusing on rapid-antidepressant drugs, more attention should be paid to experiments involving animal models referencing ketamine, the prototypic drug for rapid antidepressants.The characteristics reflected by ketamine in animal experiments form the basis for preclinical research aimed at developing new rapid-antidepressant drugs.More animal behavioral experimental evidence similar to ketamine may be more conducive to robust drug development.A great deal of research has followed around the synaptic plasticity of ketamine, most notably in the lab of Lisa M Monteggia who hypothesized that BDNF, eEF2, and TrkB were mostly responsible for ketamine's fast antidepressant-like effects (Autry et al., 2011;Lin et al., 2021).These rapid antidepressant-like effects of ketamine and similar drugs have been frequently verified in subsequent trials.
In terms of mechanistic studies, research on the glutamatergic system has been a hot topic since the beginning of ketamine rapidantidepressant-related studies.Numerous investigations have demonstrated that the classical route of the glutamatergic system is involved in the rapid antidepressant-like effects of ketamine.Research on ketamine receptors has expanded from the classical glutamatergic ion channel receptors, NMDA receptors, and AMPA receptors, to the metabolic receptors mGluR.Studies have shown that either inhibition or activation of the glutamatergic system can influence the rapid antidepressant effect of ketamine.At present, based on the evidence, blocking glutamate neurotransmission seems to be the most promising approach.The role of the 5-HT system as a synergistic mediator of the fast antidepressant effect is also attracting more attention from researchers.Although it may take a few weeks to see the full therapeutic effects of SSRIs, which work by increasing extracellular 5-HT concentrations and prolonging the duration and extent of 5-HT-mediated signals, they are routinely used as classsical antidepressants.There is also a lack of clarity about the time lag between SSRI-mediated increases in synaptic 5-HT levels and subsequent clinical improvements (Quentin et al., 2018).The amount of 5-HT in the brain tissue as well as the amount of extracellular 5-HT are both increased by ketamine administration in rats.The effects of ketamine, like SSRIs, on the serotonergic system have yet to be investigated and defined in primates.SERT and presynaptic autoreceptors make up this system (Pham and Gardier, 2019).Ketamine has been demonstrated to have antidepressant-like effects, and this has been attributed to the release of 5-HT (Gigliucci et al., 2013).It seems that its rapid antidepressant-like action in vivo may depend on boosting the firing of 5-HT neurons (Pham and Gardier, 2019).Additional research is needed to determine whether ketamine's rapid antidepressant effect requires the 5-HT rise that is caused by the drug, and if there is synergy between the increase in 5-HT and the glutamatergic system, a critical mechanism for rapid antidepressive effects.The focus of studies on ketamine's rapid antidepressant-like effects and metabolites has recently switched to the drug's mirror isomers.(R)-and (S)-ketamine, the mirror isomers of ketamine, have been studied more systematically in Hashimoto's laboratory for their rapid antidepressant-like effects.The clinical trials proved that (S)-ketamine has a more potent and effective rapid antidepressant-like effect than (R)-ketamine (Daly et al., 2018;Wajs et al., 2020;Leal et al., 2021;Leal et al., 2023).The (S)-ketamine-based drug, esketamine, is emergence as the first innovative drug for the rapid treatment of refractory depression.While esketamine shows promise for treating depression, its adverse effect such as dissociative, abuse, and stringent dosing regimen have prevented its widespread clinical use (Vieira et al., 2021;Yang et al., 2022;Gastaldon et al., 2021).The presence of these problems suggests that a large number of unanswered questions remain in the study of the rapid-antidepressant mechanisms of ketamine.Zarate Jr. and Todd D. Gould have done more detailed research on the metabolites of ketamine.They showed that the metabolism of (R,S)-ketamine to (2S,6S;2R,6R)-HNK is essential for its antidepressant-like effects and that the (2R,6R)-HNK enantiomer exerts antidepressant-related actions in mice.These antidepressant actions are independent of NMDAR inhibition but involve early and prolonged activation of α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptors (AMPARs) (Zanos et al., 2016).Although Hashimoto's research did not support the rapid antidepressant effect of ketamine metabolites (2R, 6R) -HNK (Shirayama and Hashimoto, 2018;Yang et al., 2017;Yamaguchi et al., 2018), a large number of studies confirm that the metabolites of ketamine have a rapid antidepressant-like effects similar to ketamine (Zanos et al., 2016;Zanos et al., 2019).The reasons for the different results of the experiment may be related to the specific animal model used, the strain and age of the experimental animals, and the source of the drug.The transition of ketamine research toward related enantiomers and metabolites reflects a shift in the focus of the field research from rapid onset of action to safety and efficacy.Because of the diversity of ketamine targets, it is often difficult to achieve both simultaneously.Therefore, the search for more specific targets is of great importance.The future direction of ketamine's safety mechanism may related to the excitatory toxicity of neurons, which is closely associated with NMDA receptors and calcium channels, especially L-type calcium channels (Robinson et al., 2021).
The search for new, more specific molecular targets of ketamine has remained the main direction within the realm of rapid antidepressant research.This pursuit holds the potential to enhance the effectiveness of new ketamine-derivative drugs while minimizing their toxicities.The metabotropic glutamate receptor mGluR, the synergistic effects of the 5-HT system, and related topics have received significant interest in recent years, casting light on ion channel proteins in the context of ketaminerelated molecular investigations.Such trends underscore the efforts of researchers to mitigate the side effects and ketamine's adverse effects.The identification of new molecular targets for ketamine's action may not only arise from research on ketamine itself; additional insights could emerge from novel targets in depression research, depression-related disease research, and in the development of new rapid antidepressantrelated drugs.In addition to exploring more specific drug targets and molecular pathways, future research should focus on elucidating the interplay between the neural circuits associated with these targets and the underlying signaling pathways.This approach can further clarify the interactions between the systems.In addition, searching for more effective rapid-antidepressant drugs with fewer side effects is another direction in research into rapid antidepressants.Advances in research on the mechanisms underlying rapid antidepressants will identify additional antidepressants with rapid onset of action, good efficacy, and fewer side effects that can be used to benefit patients in the future.

Declaration of Competing Interest
The author has no conflict of interest to declare.

Fig. 1 .
Fig. 1.Neural circuits involved in the rapid antidepressant action of ketamine.

Fig. 2 .
Fig. 2. The molecular mechanism underlying the rapid antidepressant-like effects of ketamine.

Table 1
Molecules targets for rapid antidepressant-like effects of ketamine.
(Zanos et al., 2016)Autry et al., 2011;Lin et al., 2021;Lepack et al., 2014;Garcia et al., 2008;Chen et al., 2006)(continued on next page) L. Ren the ketamine metabolite (2R,6R)-HNK can also induce acute enhancement of glutamatergic signaling, with the subsequent long-term adaptation that entails synaptic AMPAR upregulation(Zanos et al., 2016).Using rodent hippocampal slices, Ke Zhang et al. determined that ketamine was able to potentiate the excitatory synaptic transmission of Schaffer collateral-CA1 cells while also promoting the Ser845 phosphorylation of the AMPAR GluA1 subunit in the hippocampal CA1 region.While it alleviated behavioral symptoms of despair in WT mice,