Neurobiology of the Rapid-Acting Antidepressant Effects of Ketamine: Impact and Opportunities.

The discovery of the rapid-acting antidepressant effects of ketamine has 1) led to a paradigm shift in our perception of what is possible in treating severe depression; 2) spurred a wave of basic, translation, and clinical research; and 3) provided an unprecedented investigational tool to conduct longitudinal mechanistic studies that may capture behavioral changes as complex as clinical remission and relapse within hours and days of treatment. Unfortunately, these advances did not yet translate into clinical biomarkers or novel treatments, beyond ketamine. In contrast to slow-acting antidepressants, in which targeting monoaminergic receptors identified several efficacious drugs with comparable mechanisms, the focus on the receptor targets of ketamine has failed in several clinical trials over the past decade. Thus, it is becoming increasingly crucial that we concentrate our effort on the downstream molecular mechanisms of ketamine and their effects on the brain circuitry and networks. Honoring the legacy of our mentor, friend, and colleague Ron Duman, we provide a historical note on the discovery of ketamine and its putative mechanisms. We then detail the molecular and circuits effect of ketamine based on preclinical findings, followed by a summary of the impact of this work on our understanding of chronic stress pathology across psychiatric disorders, with particular emphasis on the role of synaptic connectivity and its brain network effects in the pathology and treatment of clinical depression.

GLYX-13 Produces Rapid Antidepressant Responses with Key Synaptic and Behavioral Effects Distinct from Ketamine.

GLYX-13 is a putative NMDA receptor modulator with glycine-site partial agonist properties that produces rapid antidepressant effects, but without the psychotomimetic side effects of ketamine. Studies were conducted to examine the molecular, cellular, and behavioral actions of GLYX-13 to further characterize the mechanisms underlying the antidepressant actions of this agent. The results demonstrate that a single dose of GLYX-13 rapidly activates the mTORC1 pathway in the prefrontal cortex (PFC), and that infusion of the selective mTORC1 inhibitor rapamycin into the medial PFC (mPFC) blocks the antidepressant behavioral actions of GLYX-13, indicating a requirement for mTORC1 similar to ketamine. The results also demonstrate that GLYX-13 rapidly increases the number and function of spine synapses in the apical dendritic tuft of layer V pyramidal neurons in the mPFC. Notably, GLYX-13 significantly increased the synaptic responses to hypocretin, a measure of thalamocortical synapses, compared with its effects on 5-HT responses, a measure of cortical-cortical responses mediated by the 5-HT2A receptor. Behavioral studies further demonstrate that GLYX-13 does not influence 5-HT2 receptor induced head twitch response or impulsivity in a serial reaction time task (SRTT), whereas ketamine increased responses in both tests. In contrast, both GLYX-13 and ketamine increased attention in the SRTT task, which is linked to hypocretin-thalamocortical responses. The differences in the 5-HT2 receptor synaptic and behavioral responses may be related to the lack of psychotomimetic side effects of GLYX-13 compared with ketamine, whereas regulation of the hypocretin responses may contribute to the therapeutic benefits of both rapid acting antidepressants.

Synaptic plasticity and depression: new insights from stress and rapid-acting antidepressants.

Depression is a common, devastating illness. Current pharmacotherapies help many patients, but high rates of a partial response or no response, and the delayed onset of the effects of antidepressant therapies, leave many patients inadequately treated. However, new insights into the neurobiology of stress and human mood disorders have shed light on mechanisms underlying the vulnerability of individuals to depression and have pointed to novel antidepressants. Environmental events and other risk factors contribute to depression through converging molecular and cellular mechanisms that disrupt neuronal function and morphology, resulting in dysfunction of the circuitry that is essential for mood regulation and cognitive function. Although current antidepressants, such as serotonin-reuptake inhibitors, produce subtle changes that take effect in weeks or months, it has recently been shown that treatment with new agents results in an improvement in mood ratings within hours of dosing patients who are resistant to typical antidepressants. Within a similar time scale, these new agents have also been shown to reverse the synaptic deficits caused by stress.

Optogenetic stimulation of infralimbic PFC reproduces ketamine's rapid and sustained antidepressant actions.

Ketamine produces rapid and sustained antidepressant actions in depressed patients, but the precise cellular mechanisms underlying these effects have not been identified. Here we determined if modulation of neuronal activity in the infralimbic prefrontal cortex (IL-PFC) underlies the antidepressant and anxiolytic actions of ketamine. We found that neuronal inactivation of the IL-PFC completely blocked the antidepressant and anxiolytic effects of systemic ketamine in rodent models and that ketamine microinfusion into IL-PFC reproduced these behavioral actions of systemic ketamine. We also found that optogenetic stimulation of the IL-PFC produced rapid and long-lasting antidepressant and anxiolytic effects and that these effects are associated with increased number and function of spine synapses of layer V pyramidal neurons. The results demonstrate that ketamine infusions or optogenetic stimulation of IL-PFC are sufficient to produce long-lasting antidepressant behavioral and synaptic responses similar to the effects of systemic ketamine administration.

Ketamine Strengthens CRF-Activated Amygdala Inputs to Basal Dendrites in mPFC Layer V Pyramidal Cells in the Prelimbic but not Infralimbic Subregion, A Key Suppressor of Stress Responses.

A single sub-anesthetic dose of ketamine, a short-acting NMDA receptor blocker, induces a rapid and prolonged antidepressant effect in treatment-resistant major depression. In animal models, ketamine (24 h) reverses depression-like behaviors and associated deficits in excitatory postsynaptic currents (EPSCs) generated in apical dendritic spines of layer V pyramidal cells of medial prefrontal cortex (mPFC). However, little is known about the effects of ketamine on basal dendrites. The basal dendrites of layer V cells receive an excitatory input from pyramidal cells of the basolateral amygdala (BLA), neurons that are activated by the stress hormone CRF. Here we found that CRF induces EPSCs in PFC layer V cells and that ketamine enhanced this effect through the mammalian target of rapamycin complex 1 synaptogenic pathway; the CRF-induced EPSCs required an intact BLA input and were generated primarily in basal dendrites. In contrast to its detrimental effects on apical dendritic structure and function, chronic stress did not induce a loss of CRF-induced EPSCs in basal dendrites, thereby creating a relative imbalance in favor of amygdala inputs. The effects of ketamine were complex: ketamine enhanced apical EPSC responses in all mPFC subregions, anterior cingulate (AC), prelimbic (PL), and infralimbic (IL) but enhanced CRF-induced EPSCs only in AC and PL-responses were unchanged in IL, a critical area for suppression of stress responses. We propose that by restoring the strength of apical inputs relative to basal amygdala inputs, especially in IL, ketamine would ameliorate the hypothesized disproportional negative influence of the amygdala in chronic stress and major depression.

REDD1 is essential for stress-induced synaptic loss and depressive behavior.

Major depressive disorder (MDD) affects up to 17% of the population, causing profound personal suffering and economic loss. Clinical and preclinical studies have revealed that prolonged stress and MDD are associated with neuronal atrophy of cortical and limbic brain regions, but the molecular mechanisms underlying these morphological alterations have not yet been identified. Here, we show that stress increases levels of REDD1 (regulated in development and DNA damage responses-1), an inhibitor of mTORC1 (mammalian target of rapamycin complex-1; ref. 10), in rat prefrontal cortex (PFC). This is concurrent with a decrease in phosphorylation of signaling targets of mTORC1, which is implicated in protein synthesis-dependent synaptic plasticity. We also found that REDD1 levels are increased in the postmortem PFC of human subjects with MDD relative to matched controls. Mutant mice with a deletion of the gene encoding REDD1 are resilient to the behavioral, synaptic and mTORC1 signaling deficits caused by chronic unpredictable stress, whereas viral-mediated overexpression of REDD1 in rat PFC is sufficient to cause anxiety- and depressive-like behaviors and neuronal atrophy. Taken together, these postmortem and preclinical findings identify REDD1 as a critical mediator of the atrophy of neurons and depressive behavior caused by chronic stress exposure.

Medial prefrontal D1 dopamine neurons control food intake.

Although the prefrontal cortex influences motivated behavior, its role in food intake remains unclear. Here, we demonstrate a role for D1-type dopamine receptor-expressing neurons in the medial prefrontal cortex (mPFC) in the regulation of feeding. Food intake increases activity in D1 neurons of the mPFC in mice, and optogenetic photostimulation of D1 neurons increases feeding. Conversely, inhibition of D1 neurons decreases intake. Stimulation-based mapping of prefrontal D1 neuron projections implicates the medial basolateral amygdala (mBLA) as a downstream target of these afferents. mBLA neurons activated by prefrontal D1 stimulation are CaMKII positive and closely juxtaposed to prefrontal D1 axon terminals. Finally, photostimulating these axons in the mBLA is sufficient to increase feeding, recapitulating the effects of mPFC D1 stimulation. These data describe a new circuit for top-down control of food intake.

Scopolamine rapidly increases mammalian target of rapamycin complex 1 signaling, synaptogenesis, and antidepressant behavioral responses.

BACKGROUND: Clinical studies report that scopolamine, an acetylcholine muscarinic receptor antagonist, produces rapid antidepressant effects in depressed patients, but the mechanisms underlying the therapeutic response have not been determined. The present study examines the role of the mammalian target of rapamycin complex 1 (mTORC1) and synaptogenesis, which have been implicated in the rapid actions of N-methyl-D-aspartate receptor antagonists. METHODS: The influence of scopolamine on mTORC1 signaling was determined by analysis of the phosphorylated and activated forms of mTORC1 signaling proteins in the prefrontal cortex (PFC). The numbers and function of spine synapses were analyzed by whole cell patch clamp recording and two-photon image analysis of PFC neurons. The actions of scopolamine were examined in the forced swim test in the absence or presence of selective mTORC1 and glutamate receptor inhibitors. RESULTS: The results demonstrate that a single, low dose of scopolamine rapidly increases mTORC1 signaling and the number and function of spine synapses in layer V pyramidal neurons in the PFC. Scopolamine administration also produces an antidepressant response in the forced swim test that is blocked by pretreatment with the mTORC1 inhibitor or by a glutamate alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptor antagonist. CONCLUSIONS: Taken together, the results demonstrate that the antidepressant actions of scopolamine require mTORC1 signaling and are associated with increased glutamate transmission, and synaptogenesis, similar to N-methyl-D-aspartate receptor antagonists. These findings provide novel targets for safer and more efficacious rapid-acting antidepressant agents.

GSK-3 inhibition potentiates the synaptogenic and antidepressant-like effects of subthreshold doses of ketamine.

A single dose of the short-acting NMDA antagonist ketamine produces rapid and prolonged antidepressant effects in treatment-resistant patients with major depressive disorder (MDD), which are thought to occur via restoration of synaptic connectivity. However, acute dissociative side effects and eventual fading of antidepressant effects limit widespread clinical use of ketamine. Recent studies in medial prefrontal cortex (mPFC) show that the synaptogenic and antidepressant-like effects of a single standard dose of ketamine in rodents are dependent upon activation of the mammalian target of rapamycin (mTOR) complex 1 (mTORC1) signaling pathway together with inhibitory phosphorylation of glycogen synthase kinase-3 (GSK-3), which relieves its inhibitory in influence on mTOR. Here, we found that the synaptogenic and antidepressant-like effects of a single otherwise subthreshold dose of ketamine were potentiated when given together with a single dose of lithium chloride (a nonselective GSK-3 inhibitor) or a preferential GSK-3ß inhibitor; these effects included rapid activation of the mTORC1 signaling pathway, increased inhibitory phosphorylation of GSK-3ß, increased synaptic spine density/diameter, increased excitatory postsynaptic currents in mPFC layer V pyramidal neurons, and antidepressant responses that persist for up to 1 week in the forced-swim test model of depression. The results demonstrate that low, subthreshold doses of ketamine combined with lithium or a selective GSK-3 inhibitor are equivalent to higher doses of ketamine, indicating the pivotal role of the GSK-3 pathway in modulating the synaptogenic and antidepressant responses to ketamine. The possible mitigation by GSK-3 inhibitors of the eventual fading of ketamine's antidepressant effects remains to be explored.

Synaptic dysfunction in depression: potential therapeutic targets.

Basic and clinical studies demonstrate that depression is associated with reduced size of brain regions that regulate mood and cognition, including the prefrontal cortex and the hippocampus, and decreased neuronal synapses in these areas. Antidepressants can block or reverse these neuronal deficits, although typical antidepressants have limited efficacy and delayed response times of weeks to months. A notable recent discovery shows that ketamine, a N-methyl-D-aspartate receptor antagonist, produces rapid (within hours) antidepressant responses in patients who are resistant to typical antidepressants. Basic studies show that ketamine rapidly induces synaptogenesis and reverses the synaptic deficits caused by chronic stress. These findings highlight the central importance of homeostatic control of mood circuit connections and form the basis of a synaptogenic hypothesis of depression and treatment response.

Signaling pathways underlying the rapid antidepressant actions of ketamine.

Currently available medications have significant limitations, most notably low response rate and time lag for treatment response. Recent clinical studies have demonstrated that ketamine, an NMDA receptor antagonist produces a rapid antidepressant response (within hours) and is effective in treatment resistant depressed patients. Molecular and cellular studies in rodent models demonstrate that ketamine rapidly increases synaptogenesis, including increased density and function of spine synapses, in the prefrontal cortex (PFC). Ketamine also produces rapid antidepressant actions in behavioral models of depression, and reverses the deficits in synapse number and behavior resulting from chronic stress exposure. These effects of ketamine are accompanied by stimulation of the mammalian target of rapamycin (mTOR), and increased levels of synaptic proteins. Together these studies indicate that ketamine rapidly reverses the atrophy of spines in the PFC and thereby causes a functional reconnection of neurons that underlies the rapid behavioral responses. These findings identify new targets for rapid acting antidepressants that are safer than ketamine. This article is part of a Special Issue entitled 'Anxiety and Depression'.

Brain-derived neurotrophic factor Val66Met allele impairs basal and ketamine-stimulated synaptogenesis in prefrontal cortex.

BACKGROUND: Knock-in mice with the common human brain-derived neurotrophic factor (BDNF) Val66Met polymorphism have impaired trafficking of BDNF messenger RNA to dendrites. It was hypothesized, given evidence that local synapse formation is dependent on dendritic translation of BDNF messenger RNA, that loss-of-function Met allele mice would show synaptic deficits both at baseline and in response to ketamine, an N-methyl-D-aspartate antagonist that stimulates synaptogenesis in prefrontal cortex (PFC). METHODS: Whole-cell recordings from layer V medial PFC pyramidal cells in brain slices were combined with two-photon laser scanning for analysis of wildtype, Val/Met, and Met/Met mice both at baseline and in response to a low dose of ketamine. RESULTS: Val/Met and Met/Met mice were found to have constitutive atrophy of distal apical dendrites and decrements in apically targeted excitatory postsynaptic currents in layer V pyramidal cells of PFC. In addition, spine density and diameter were decreased, indicative of impaired synaptic formation/maturation (synaptogenesis). In Met/Met mice the synaptogenic effect of ketamine was markedly impaired, consistent with the idea that synaptogenesis is dependent on dendritic translation/release of BDNF. In parallel behavioral studies, we found that the antidepressant response to ketamine in the forced swim test was blocked in Met/Met mice. CONCLUSIONS: The results demonstrate that expression of the BDNF Met allele in mice results in basal synaptic deficits and blocks synaptogenic and antidepressant actions of ketamine in PFC, suggesting that the therapeutic response to this drug might be attenuated or blocked in depressed patients who carry the loss of function Met allele.

Glutamate N-methyl-D-aspartate receptor antagonists rapidly reverse behavioral and synaptic deficits caused by chronic stress exposure.

BACKGROUND: Despite widely reported clinical and preclinical studies of rapid antidepressant actions of glutamate N-methyl-D-aspartate (NMDA) receptor antagonists, there has been very little work examining the effects of these drugs in stress models of depression that require chronic administration of antidepressants or the molecular mechanisms that could account for the rapid responses. METHODS: We used a rat 21-day chronic unpredictable stress (CUS) model to test the rapid actions of NMDA receptor antagonists on depressant-like behavior, neurochemistry, and spine density and synaptic function of prefrontal cortex neurons. RESULTS: The results demonstrate that acute treatment with the noncompetitive NMDA channel blocker ketamine or the selective NMDA receptor 2B antagonist Ro 25-6981 rapidly ameliorates CUS-induced anhedonic and anxiogenic behaviors. We also found that CUS exposure decreases the expression levels of synaptic proteins and spine number and the frequency/amplitude of synaptic currents (excitatory postsynaptic currents) in layer V pyramidal neurons in the prefrontal cortex and that these deficits are rapidly reversed by ketamine. Blockade of the mammalian target of rapamycin protein synthesis cascade abolishes both the behavioral and biochemical effects of ketamine. CONCLUSIONS: The results indicate that the structural and functional deficits resulting from long-term stress exposure, which could contribute to the pathophysiology of depression, are rapidly reversed by NMDA receptor antagonists in a mammalian target of rapamycin dependent manner.

mTOR-dependent synapse formation underlies the rapid antidepressant effects of NMDA antagonists.

The rapid antidepressant response after ketamine administration in treatment-resistant depressed patients suggests a possible new approach for treating mood disorders compared to the weeks or months required for standard medications. However, the mechanisms underlying this action of ketamine [a glutamate N-methyl-D-aspartic acid (NMDA) receptor antagonist] have not been identified. We observed that ketamine rapidly activated the mammalian target of rapamycin (mTOR) pathway, leading to increased synaptic signaling proteins and increased number and function of new spine synapses in the prefrontal cortex of rats. Moreover, blockade of mTOR signaling completely blocked ketamine induction of synaptogenesis and behavioral responses in models of depression. Our results demonstrate that these effects of ketamine are opposite to the synaptic deficits that result from exposure to stress and could contribute to the fast antidepressant actions of ketamine.

Regulation of nucleus accumbens activity by the hypothalamic neuropeptide melanin-concentrating hormone.

The lateral hypothalamus and the nucleus accumbens shell (AcbSh) are brain regions important for food intake. The AcbSh contains high levels of receptor for melanin-concentrating hormone (MCH), a lateral hypothalamic peptide critical for feeding and metabolism. MCH receptor (MCHR1) activation in the AcbSh increases food intake, while AcbSh MCHR1 blockade reduces feeding. Here biochemical and cellular mechanisms of MCH action in the rodent AcbSh are described. A reduction of phosphorylation of GluR1 at serine 845 (pSer(845)) is shown to occur after both pharmacological and genetic manipulations of MCHR1 activity. These changes depend upon signaling through G(i/o), and result in decreased surface expression of GluR1-containing AMPA receptors (AMPARs). Electrophysiological analysis of medium spiny neurons (MSNs) in the AcbSh revealed decreased amplitude of AMPAR-mediated synaptic events (mEPSCs) with MCH treatment. In addition, MCH suppressed action potential firing MSNs through K(+) channel activation. Finally, in vivo recordings confirmed that MCH reduces neuronal cell firing in the AcbSh in freely moving animals. The ability of MCH to reduce cell firing in the AcbSh is consistent with a general model from other pharmacological and electrophysiological studies whereby reduced AcbSh neuronal firing leads to food intake. The current work integrates the hypothalamus into this model, providing biochemical and cellular mechanisms whereby metabolic and limbic signals converge to regulate food intake.

Modeling "psychosis" in vitro by inducing disordered neuronal network activity in cortical brain slices.

INTRODUCTION: Dysregulation of neuronal networks has been suggested to underlie the cognitive and perceptual abnormalities observed schizophrenia. DISCUSSIONS: An in vitro model of psychosis is proposed based on the two different approaches to cause aberrant network activity in layer V pyramidal cells of prefrontal brain slices: (1) psychedelic hallucinogens such as lysergic acid diethylamide and (2) minimal GABA(A) receptor antagonism, modeling the GABA interneuron deficit in schizophrenia. A test of this model would be to determine if drugs that normalize aberrant networks in brain slices have efficacy in the treatment of schizophrenia. Selective agonists of glutamate mGlu2/3 metabotropic receptors, which are highly effective in suppressing aberrant network activity in slices, are the most advanced toward reaching that clinical endpoint. In accord with the model, a recent phase II clinical trial shows that an mGlu2/3 receptor agonist is equivalent in efficacy to a standard antipsychotic drug for both negative and positive symptoms in schizophrenic patients, but without the usual side effects. D1/5 dopamine receptor agonists are also effective in normalizing aberrant network activity induced by both hallucinogens and minimal GABA(A) antagonism; clinical efficacy remains to be determined. A general model of network regulation is presented, involving astrocytes, GABA interneurons, and glutamatergic pyramidal cells, revealing a wide range of potential sites hitherto not considered as therapeutic targets.

Distinct roles of adenylyl cyclases 1 and 8 in opiate dependence: behavioral, electrophysiological, and molecular studies.

BACKGROUND: Opiate dependence is a result of adaptive changes in signal transduction networks in several brain regions. Noradrenergic neurons of the locus coeruleus (LC) have provided a useful model system in which to understand the molecular basis of these adaptive changes. One of most robust signaling adaptations to repeated morphine exposure in this brain region is upregulation of adenylyl cyclase (AC) activity. Earlier work revealed the selective induction of two calmodulin-dependent AC isoforms, AC1 and AC8, after chronic morphine, but their role in opiate dependence has remained unknown. METHODS: Whole cell recordings from LC slices, behavioral paradigms for dependence, and gene array technology have been used to dissect the role of AC1 and AC8 in chronic morphine responses. RESULTS: Both AC1 and AC8 knockout mice exhibit reduced opiate dependence on the basis of attenuated withdrawal; however, partially distinct withdrawal symptoms were affected in the two lines. Loss of AC1 or AC8 also attenuated the electrophysiological effects of morphine on LC neurons: knockout of either cyclase attenuated the chronic morphine-induced enhancement of baseline firing rates as well as of regulation of neuronal firing by forskolin (an activator of ACs). The DNA microarray analysis revealed that both AC1 and AC8 affect gene regulation in the LC by chronic morphine and, in addition to common genes, each cyclase influences the expression of a distinct subset of genes. CONCLUSIONS: Together, these findings provide fundamentally new insight into the molecular and cellular basis of opiate dependence.

Stress blunts serotonin- and hypocretin-evoked EPSCs in prefrontal cortex: role of corticosterone-mediated apical dendritic atrophy.

Morphological studies show that repeated restraint stress leads to selective atrophy in the apical dendritic field of pyramidal cells in the medial prefrontal cortex (mPFC). However, the functional consequence of this selectivity remains unclear. The apical dendrite of layer V pyramidal neurons in the mPFC is a selective locus for the generation of increased excitatory postsynaptic currents (EPSCs) by serotonin (5-HT) and hypocretin (orexin). On that basis, we hypothesized that apical dendritic atrophy might result in a blunting of 5-HT- and hypocretin-induced excitatory responses. Using a combination of whole-cell recording and two-photon imaging in rat mPFC slices, we were able to correlate electrophysiological and morphological changes in the same layer V pyramidal neurons. Repeated mild restraint stress produced a decrement in both 5-HT- and hypocretin-induced EPSCs, an effect that was correlated with a decrease in apical tuft dendritic branch length and spine density in the distal tuft branches. Chronic treatment with the stress hormone corticosterone, while reducing 5-HT responses and generally mimicking the morphological effects of stress, failed to produce a significant decrease in hypocretin-induced EPSCs. Accentuating this difference, pretreatment of stressed animals with the glucocorticoid receptor antagonist RU486 blocked reductions in 5-HT-induced EPSCs but not hypocretin-induced EPSCs. We conclude: (i) stress-induced apical dendritic atrophy results in diminished responses to apically targeted excitatory inputs and (ii) corticosterone plays a greater role in stress-induced reductions in EPSCs evoked by 5-HT as compared with hypocretin, possibly reflecting the different pathways activated by the two transmitters.