Ketamine-induced Neurotoxicity Research Articles

Curcumin-ZnO conjugated nanoparticles confer neuroprotection against ketamine-induced neurotoxicity.

Nanotechnology and its application to manipulate herbal compounds to design new neuroprotective agents to manage neurotoxicity has recently increased. Cur-ZnO conjugated nanoparticles were synthesized and used in an experimental model of ketamine-induced neurotoxicity. Cur-ZnO conjugated nanoparticles were chemically characterized, and the average crystalline size was determined. Forty-nine adult mice were divided into seven groups of seven animals each. Normal saline was given to control mice (group 1). Ketamine (25 mg/kg) was given to a second group. A third group of mice was given ketamine (25 mg/kg) in combination with curcumin (40 mg/kg), while mice in groups 4, 5, and 6 received ketamine (25 mg/kg) plus Cur-ZnO nanoparticles (10, 20, and 40 mg/kg). Group 7 received only ZnO (5 mg/kg). All doses were ip for 14 days. Hippocampal mitochondrial quadruple complex enzymes, oxidative stress, inflammation, and apoptotic characteristics were assessed. Cur-ZnO nanoparticles and curcumin decreased lipid peroxidation, GSSG content, IL-1β, TNF-α, and Bax levels while increasing GSH and antioxidant enzymes like GPx, GR, and SOD while increasing Bcl-2 level and mitochondrial quadruple complex enzymes in ketamine treatment groups. The neuroprotective properties of Cur-ZnO nanoparticles were efficient in preventing ketamine-induced neurotoxicity in the mouse brain. The nanoparticle form of curcumin (Cur-ZnO) required lower doses to produce neuroprotective effects against ketamine-induced toxicity than conventional curcumin.

Lipoxin A4 methyl ester attenuated ketamine-induced neurotoxicity in SH-SY5Y cells via regulating leptin pathway

Ketamine, the widely used intravenous anesthetic, has been reported to cause neurotoxicity and disturbs normal neurogenesis. However, the efficacy of current treatment strategies targeting ketamine's neurotoxicity remains limited. Lipoxin A4 methyl ester (LXA4 ME) is relatively stable lipoxin analog, which serves an important role in protecting against early brain injury. The purpose of this study was to investigate the protective effect of LXA4 ME on ketamine-caused cytotoxicity in SH-SY5Y cells, as well as the underlying mechanisms. Cell viability, apoptosis and endoplasmic reticulum stress (ER stress) were detected by adopting experimental techniques including CCK-8 assay, flow cytometry, western blotting and transmission electron microscope. Furthermore, examining the expression of leptin and its receptor (LepRb), we also measured the levels of activation of the leptin signaling pathway. Our results showed that LXA4 ME intervention promoted the cell viability, inhibited cell apoptosis, and reduced the expression of ER stress related protein and morphological changes induced by ketamine. In addition, inhibition of leptin signaling pathway caused by ketamine could be reversed by LXA4 ME. However, as the specific inhibitor of leptin pathway, leptin antagonist triple mutant human recombinant (leptin tA) attenuated the cytoprotective effect of LXA4 ME against ketamine-induced neurotoxicity. In conclusion, our findings demonstrated LXA4 ME could exert a neuroprotective effect on ketamine-induced neuronal injury via activation of the leptin signaling pathway.

LncRNA TUG1 Promoted Stabilization of BAG5 by Binding DDX3X to Exacerbate Ketamine-Induced Neurotoxicity.

As a clinically widely used anesthetic, ketamine (KET) has been reported to cause neurotoxicity in patients. Our work aimed to probe the function of long-chain non-coding RNA taurine-upregulated gene 1 (lncRNA TUG1) in KET-induced neurotoxicity. HT22 cells were subjected to KET to build the cell model. 3-(4, 5-Dimethylthiazolyl2)-2, 5-diphenyltetrazolium bromide (MTT) assay was employed to determine cell viability. Additionally, cell apoptosis was evaluated by flow cytometry. The binding relationships among TUG1, DEAD-box RNA helicase 3X (DDX3X), and Bcl-2-associated athanogene 5 (BAG5) were verified by RIP and RNA pull-down assays. Cell viability was impaired and cell apoptosis was increased in KET-treated HT22 cells accompanied by increased TUG1, DDX3X, and BAG5 expressions. TUG1 knockdown dramatically enhanced cell viability and repressed the of KET-induced apoptosis in HT22 cells, while TUG1 overexpression presented the opposite effects. In addition, we found that TUG1 promoted DDX3X expression via directly binding with DDX3X. As expected, DDX3X overexpression abolished the palliative effect of TUG1 knockdown on KET-induced neurotoxicity. Further research proved that TUG1 increased the stability of BAG5 through interacting with DDX3X. Finally, as expected, the moderating effect of TUG1 knockdown on KET-induced neuron injury was abolished by BAG5 overexpression. Taken together, TUG1 promoted BAG5 expression by binding DDX3X to exacerbate KET-induced neurotoxicity.

Ketamine-induced neurotoxicity is mediated through endoplasmic reticulum stress in vitro in STHdhQ7/Q7 cells

Ketamine has traditionally been used as a dissociative anesthetic agent and more recently as a treatment for treatment-resistant depression. However, there is growing concern over the increased use of ketamine in recreational and therapeutic settings due to the potential neurotoxic effects. Recent studies have demonstrated that ketamine is cytotoxic in several cell types, such as fibroblasts, hepatocytes, uroepithelial cells, and adult induced pluripotent stem cells (iPSCs). Ketamine has been shown to dysregulate calcium signalling, increase reactive oxygen species (ROS) production, and impair mitochondrial function, ultimately leading to apoptosis. However, it is unclear whether endoplasmic reticulum (ER) stress plays a role in ketamine associated neurotoxicity in striatal neurons. Disruption to ER homeostasis can initiate ER-mediated cell death, which has been implicated in several neurodegenerative diseases. Thus, the purpose of this study was to determine whether ketamine's neurotoxic effects involve an ER stress-dependent pathway and to elucidate the underlying mechanisms involved in its neurotoxic effects. Mouse striatal cells were treated with various concentrations of ketamine (10 μM, 100 μM, 1 mM) or DMEM for 9–72 hrs. Cell viability was assessed using the MTT assay, and changes in gene expression of ER stress markers were evaluated using RT-qPCR. MTT results revealed that 1 mM ketamine decreased cell viability in striatal cells after 24 h of treatment. Gene expression studies complemented these findings such that ketamine upregulated pro-apoptotic ER stress markers, including X-box binding protein 1 (XBP1), activating transcription factor 4 (ATF4), and C/EBP homologous protein (CHOP) and downregulated pro-survival ER stress proteins such as GRP78, MANF and CDNF. Ketamine activated all three stress sensing pathways including PERK, IRE1, and ATF6. Taken together, our results show that ketamine-induced neurotoxicity is mediated through an ER stress-dependent apoptotic pathway.

MiRNA-384-5p Targets GABRB1 to Regulate Ketamine-Induced Neurotoxicity in Neurons.

To determine the function of miR-384-5p in ketamine-induced neurotoxicity. Neonatal hippocampal neurons were isolated from rats and treated with varying doses of ketamine. RT-qPCR was utilized to measure the miR-384-5p level in ketamine-treated neurons. Neuronal viability was evaluated by MTT assay. TUNEL staining and flow cytometry were applied to measure neuronal apoptosis. H2-DCFDA staining was utilized to detect the intracellular ROS level. Protein levels were measured using Western blotting. A luciferase reporter experiment was used in HEK293T cells to verify the interaction of miR-384-5p with GABRB1. Ketamine induced neurotoxicity and miR-384-5p upregulation in hippocampal neurons. miR-384-5p downregulation mitigated ketamine-induced neurotoxicity by restraining apoptosis and ROS activity in neurons. GABRB1 was demonstrated to be targeted by miR-384-5p. GABRB1 depletion worsened ketamine-induced neurotoxicity. Moreover, GABRB1 depletion lessened the protective effect of miR-384-5p inhibition against ketamine-mediated neurotoxicity. miR-384-5p regulates ketamine-induced neurotoxicity in hippocampal neurons by targeting GABRB1.

Lipoxin A4 methyl ester protects PC12 cells from ketamine-induced neurotoxicity via the miR-22/BAG5 pathway.

Ketamine is an anesthetic that induces neurotoxicity when administered at high doses. In this work, we explored the protective effects of lipoxin A4 methyl ester (LXA4 ME) against ketamine-induced neurotoxicity and the underlying protective mechanism in pheochromocytoma (PC12) cells. PC12 cells were treated with 50μM of ketamine and different LXA4 ME concentrations of LXA4 ME (5-50nM) for 24h, and their viability, apoptosis, and oxidative status were assessed. Quantitative real-time polymerase chain reaction experiments showed that ketamine downregulated miR-22 expression and upregulated Bcl-2-associated athanogene 5 (BAG5) in PC12 cells in a concentration-dependent manner. LXA4 ME induced the opposite effects, thus attenuating ketamine-induced neurotoxicity. Further in vitro assays showed that miR-22 directly targeted BAG5, thus promoting cell viability by suppressing cell apoptosis and oxidative stress. Under expression miR-22 or upregulation of BAG5 antagonized the effects of LXA4 ME. LXA4 ME can protect PC12 cells from ketamine-induced neurotoxicity by activating the miR-22/BAG5 signaling pathway. Thus, LXA4 ME can be used as a protective drug against ketamine-induced neural damage.

Blockade of the NLRP3/caspase-1 axis attenuates ketamine-induced hippocampus pyroptosis and cognitive impairment in neonatal rats

BackgroundMultiple studies have revealed that repeated or long-term exposure to ketamine causes neurodegeneration and cognitive dysfunction. Pyroptosis is an inflammatory form of programmed cell death that has been linked to various neurological diseases. However, the role of NLRP3/caspase-1 axis-related pyroptosis in ketamine-induced neurotoxicity and cognitive dysfunction remains uncertain.MethodsTo evaluate whether ketamine caused NLRP3/caspase1-dependent pyroptosis, flow cytometry analysis, western blotting, ELISA test, histopathological analysis, Morris water maze (MWM) test, cell viability assay, and lactate dehydrogenase release (LDH) assay were carried out on PC12 cells, HAPI cells, and 7-day-old rats. In addition, the NLRP3 inhibitor MCC950 or the caspase-1 inhibitor VX-765 was used to investigate the role of the NLRP3/caspase-1 axis in ketamine-induced neurotoxicity and cognitive dysfunction.ResultsOur findings demonstrated that ketamine exposure caused cell damage and increased the levels of pyroptosis in PC12 cells, HAPI cells, and the hippocampus of neonatal rats. After continuous exposure to ketamine, targeting NLRP3 and caspase-1 with MCC950 or VX765 improved pyroptosis, reduced neuropathological damages, and alleviated cognitive dysfunction.ConclusionNLRP3/Caspase-1 axis-dependent pyroptosis is involved in ketamine-induced neuroinflammation and cognitive dysfunction, and it provides a promising strategy to treat ketamine-related neurotoxicity.

ADNOC and Petronas ink deal to explore collaboration opportunities

Currently, an increasing proportion of adolescent ketamine users simultaneously consume alcohol. However, the potential behavioural and neurological alterations induced by such a drug combination and the underlying mechanisms have not been systematically examined. Therefore, in the present study, the behavioural and morphological changes and the underlying mechanisms were studied in adolescent rats after repeated alcohol and/or ketamine treatment. This study provided the first evidence that co-administration of alcohol (2 and 4 g/kg, i.g.) in adolescent rats significantly potentiated the neurotoxic properties of repeated ketamine (30 mg/kg, i.p.) treatments over 14 days, manifesting as increased locomotor activity, stereotypic behaviour, ataxia and morphological changes. This potentiation was associated with the enhancement by alcohol of ketamine-induced glutamate (Glu) and dopamine (DA) release in the cortex and hippocampus. Further mechanistic study demonstrated that alcohol potentiated ketamine-induced neurotoxicity through down-regulation of Akt (a serine/threonine kinase or protein kinase, PKB), protein kinase A (PKA), calmodulin-dependent kinase IV (CaMK-IV)-mediated cyclic AMP-responsive element binding protein (CREB) pathways and induction of neuronal apoptosis in the cortex and hippocampus of the adolescent rats. As this study provides strong evidence that repeated alcohol and ketamine co-exposure may cause serious neurotoxicity, attention needs to be drawn to the potential risk of this consumption behaviour, especially for adolescents.

Long Noncoding RNA SPRY4-IT1 Modulates Ketamine-Induced Neurotoxicity in Human Embryonic Stem Cell-Derived Neurons through EZH2

Objective: This study aimed to investigate whether long noncoding RNA sprouty receptor tyrosine kinase signaling antagonist 4-intronic transcript 1 (SPRY4-IT1) is involved in the regulation of ketamine-induced neurotoxicity. Methods: Human embryonic stem cells (hESCs) were induced into neurons in vitro and treated with ketamine. Apoptosis and neurite degeneration assays were used to determine ketamine-induced neurotoxicity and qRT-PCR to determine SPRY4-IT1 expression. SPRY4-IT1 was downregulated in hESC-induced neurons to examine its regulation on ketamine-induced neurotoxicity. The correlation between enhancer of zeste homolog 2 (EZH2) and SPRY4-IT1 was also examined. EZH2 was upregulated in SPRY4-IT1-downregualted hESC-induced neurons to further examine its participation in SPRY4-IT1-mediated ketamine neurotoxicity. Results: Ketamine-induced dose-dependent apoptosis, neurite degeneration, and SPRY4-IT1 upregulation in hESC-induced neurons. Lentivirus-mediated SPRY4-IT1 downregulation protected ketamine neurotoxicity. EZH2 expression was positively correlated with SPRY4-IT1 in hESC-induced neurons. EZH2 overexpression markedly reversed the protective effects of SPRY4-IT1 knockdown on ketamine neurotoxicity. Conclusions: SPRY4-IT1 is involved in anesthesia-induced neurotoxicity, possibly through the regulation on EZH2 gene.

Antidepressant Actions of Ketamine: Potential Role of L-Type Calcium Channels.

Recently, the United States Food and Drug Administration approved esketamine, the S-enantiomer of ketamine, as a fast-acting therapeutic drug for treatment-resistant depression. Although ketamine is known as an N-methyl-d-aspartate (NMDA) receptor antagonist, the underlying mechanisms of how it elicits an antidepressant effect, specifically at subanesthetic doses, are not clear and remain an advancing field of research interest. On the other hand, high-dose (more than the anesthetic dose) ketamine-induced neurotoxicity in animal models has been reported. There has been progress in understanding the potential pathways involved in ketamine-induced antidepressant effects, some of which include NMDA-receptor antagonism, modulation of voltage-gated calcium channels, and brain-derived neurotrophic factor (BDNF) signaling. Often these pathways have been shown to be linked. Voltage-gated L-type calcium channels have been shown to mediate the rapid-acting antidepressant effects of ketamine, especially involving induction of BDNF synthesis downstream, while BDNF deficiency decreases the expression of L-type calcium channels. This review focuses on the reported studies linking ketamine's rapid-acting antidepressant actions to L-type calcium channels with an objective to present a perspective on the importance of the modulation of intracellular calcium in mediating the effects of subanesthetic (antidepressant) versus high-dose ketamine (anesthetic and potential neurotoxicant), the latter having the ability to reduce intracellular calcium by blocking the calcium-permeable NMDA receptors, which is implicated in potential neurotoxicity.

Ketamine-induced neurotoxicity in neurodevelopment: A synopsis of main pathways based on recent in vivo experimental findings.

Ketamine, a phencyclidine derivative and N-methyl-D-aspartate (NMDA) receptor antagonist, is widely used as an anesthetic, analgesic, and sedative agent in daily pediatric practice. Experimental studies have suggested that early prenatal or postnatal exposure to ketamine can induce neuroapoptosis, and establish neurobehavioral deficits that are evident in adulthood. However, most of the currently available clinical evidence is derived from retrospective and observational clinical studies. We, herein, attempt a brief review of the cellular and molecular mechanisms suggested to mediate ketamine-induced developmental neurotoxicity, utilizing a selected number of recent in vivo experimental evidence.

MiRNA-429 alleviates ketamine-induced neurotoxicity through targeting BAG5.

Ketamine is a kind of anesthetic broadly applied in clinic. However, growing evidence has indicated that ketamine may induce neurotoxicity. Previous studies showed that mircoRNAs (miRNAs) participate in various aspects of biological regulations. In our work, we aimed to reveal the role of miR-429 in ketamine-induced neurotoxicity. The qRT-PCR was used to measure the miR-429 levels in ketamine-treated PC12 cells. TUNEL staining and caspase 3 activity detection assays were performed to assess cell apoptosis. A Cellular Reactive Oxygen Species Detection Assay Kit was utilized to detect ROS activity. A luciferase reporter assay was conducted in HEK-293T cells to test the binding between miR-429 and BAG5. Herein, we found that ketamine could induce the apoptosis and ROS activity in PC12 cells. The qRT-PCR results showed that miR-429 expression was downregulated by treatment of ketamine in a dose-dependent manner. Overexpression of miR-429 alleviated ketamine-induced neurotoxicity in PC12 cells. Mechanically, BAG5 was identified to be a target of miR-429 and negatively regulated by miR-429. Moreover, BAG5 expression was upregulated after ketamine treatment. Rescue assays revealed that overexpression of BAG5 reversed the suppressive effects of miR-429 upregulation on ketamine-induced neurotoxicity in PC12 cells. In summary, miR-429 attenuates ketamine-induced neurotoxicity in PC12 cells by the downregulation of BAG5.

Yes‑associated protein protects and rescues SH‑SY5Y cells from ketamine‑induced apoptosis.

Ketamine is a widely used intravenous anesthetic; however, basic and clinical studies have demonstrated that prolonged exposure can cause irreversible injury to the immature human brain. Yes-associated protein (YAP) is the main effector of the Hippo signaling pathway, which serves an important role in regulating tissue homeostasis and organ size during development. However, whether YAP mediates ketamine-induced apoptosis is not completely understood. Based on the functions of YAP during apoptosis resistance and cell self-renewal regulation, the present study hypothesized that YAP serves a role during ketamine-induced apoptosis. An in vitro model was utilized to investigate the effects of ketamine on neurotoxicity and to further investigate the role of YAP in ketamine-induced apoptosis using techniques including CCK-8 assay, flow cytometry and western blotting. The present study assessed the effects of YAP overexpression and knockdown on the expression of typical apoptotic markers in SH-SY5Y cells. Ketamine induced apoptosis in a dose-dependent manner, which was regulated by YAP. Following YAP overexpression, ketamine-treated SH-SY5Y cells displayed increased activity and viability, whereas expression levels of the apoptotic markers were decreased compared with the negative control group. By contrast, ketamine-induced apoptosis was enhanced following YAP knockdown. Collectively, the results of the present study indicated that YAP may serve an important role during ketamine-induced neurotoxicity, and alterations to YAP signaling may counteract ketamine-induced apoptosis. The neuroprotective effect of YAP activation may serve as a novel pharmacological target for the treatment of ketamine-induced neurotoxicity via neurogenesis normalization.

Overexpression of long non-coding RNA SNHG16 against cerebral ischemia-reperfusion injury through miR-106b-5p/LIMK1 axis

Long non-coding RNA (LncRNA) involved in types of physiological insults and diseases via regulating the responses of complex molecular, including cerebral ischemia-reperfusion (I/R) injury. LncRNA SNHG16 played a potential role in ketamine-induced neurotoxicity. In this study, we utilized an in vitro cell model of I/R to examine the specific function and mechanism of LncRNA SNHG16 in oxygen-glucose deprivation and reperfusion (OGD/R) induced SH-SY5Y cells. After in vitro treatment of OGD/R, the lower the SH-SY5Y cell survival, the higher cell the apoptosis and increased caspase-3 activity was observed. Also, OGD/R induced endoplasmic reticulum stress (ERS) through increasing GRP78 and CHOP expressions and down-regulated LncRNA SNHG16 in SH-SY5Y cells. Conversely, LncRNA SNHG16 overexpression promoted OGD/R induced SH-SY5Y cell survival, suppressed its apoptosis, and caspase-3 activity. GRP78 and CHOP expressions were significantly suppressed in LncRNA SNHG16 overexpressing cells. MiR-106b-5p expression was increased and LIMK1 expression was down-regulated in OGD/R induced SH-SY5Y cells, and these effects were reversed by LncRNA SNHG16 overexpression, respectively. Moreover, LIMK1 is a direct target of MiR-106b-5p, and knockdown of LIMK1 reversed the effects of LncRNA SNHG16 on OGD/R-induced SH-SY5Y cells biology. Altogether, these results confirmed an important neuroprotection role of LncRNA SNHG16 in OGD/R induced SH-SY5Y cells injury, and miR-106b-5p/LIMK1 signal axis was involved in the action of LncRNA SNHG16.

Exogenous GM1 Ganglioside Attenuates Ketamine-Induced Neurocognitive Impairment in the Developing Rat Brain

A prolonged exposure to ketamine triggers significant neurodegeneration and long-term neurocognitive deficits in the developing brain. Monosialotetrahexosylganglioside (GM1) can limit the neuronal damage from necrosis and apoptosis in neurodegenerative conditions. We aimed to assess whether GM1 can prevent ketamine-induced developmental neurotoxicity. Postnatal day 7 (P7) rat pups received 5 doses of intraperitoneal ketamine (20 mg/kg per dose) at 90-minute intervals for 6 hours. Cognitive functions, determined by using Morris water maze (MWM) including escape latency (at P32-36) and platform crossing (at P37), were compared among the ketamine-exposed pups treated with or without exogenous GM1 (30 mg/kg; n = 12/group). The effect of GM1 on apoptosis in hippocampus was determined by terminal deoxynucleotidyl transferase-mediated 2'-deoxyuridine 5'-triphosphate nick end labeling (TUNEL) staining and activated caspase 3 measurement. The hippocampal expression of brain-derived neurotrophic factor (BDNF), along with the phosphorylation of protein kinase B (AKT) and extracellular signal-related kinases 1 and 2 (ERK1/2), was detected by western blotting (n = 6/group). Anti-BDNF antibody (2 μg per rat) administered before GM1 treatment was applied to determine the neuroprotective mechanisms of GM1. The rats receiving ketamine exposure experinced cognitive impairment in MWM test compared to the control rats, indicated by prolonged escape latency at P34 (P = .006), P35 (P = .002), and P36 (P = .005). However, in GM1-pretreated rats, ketamine exposure did not induce prolonged escape latency. The exogenous GM1 increased the platform-crossing times at P37 (3.00 ± 2.22 times vs 5.40 ± 1.53 times, mean ± standard deviation; P = .041) and reduced the hippocampal TUNEL-positive cells and cleaved-caspase 3 expression in ketamine-exposed young rats. Ketamine decreased BDNF expression and phosphorylation of AKT and ERK in the hippocampus, whereas exogenous GM1 blocked these ketamine-caused effects. However, for the ketamine-exposed rat pups receiving exogenous GM1, compared to immunoglobulin Y (IgY) isotype control, the BDNF-neutralizing antibody treatment counteracted the exogenous GM1-induced improvement of the escape latency at P36 (41.32 ± 12.37 seconds vs 25.14 ± 8.97 seconds, mean ± standard deviation; P = .036), platform-crossing times at P37 (2.16 ± 1.12 times vs 3.92 ± 1.97 times, mean ± standard deviation; P < .036), apoptotic activity, as well as AKT and ERK1/2 phosphorylation in the hippocampus of ketamine-challenged young rats. Our data suggest that the exogenous GM1 acts on BDNF signaling pathway to ameliorate the cognitive impairment and hippocampal apoptosis induced by ketamine in young rats. Our study may indicate a potential use of GM1 in preventing the cognitive deficits induced by ketamine in the young per se.

Traditional Chinese medicine, Kami-Shoyo-San protects ketamine-induced neurotoxicity in human embryonic stem cell-differentiated neurons through activation of brain-derived neurotrophic factor.

Anesthesia-induced neurotoxicity may cause permanent dysfunctions in human brains. In this work, we used a cell-based in-vitro model to demonstrate that traditional Chinese medicine, Kami-Shoyo-San may protect ketamine-induced neuronal apoptosis in human embryonic stem cell-differentiated neurons. Human embryonic stem cell-differentiated neurons were cultured in vitro and treated with high-concentration ketamine to induce neuronal apoptosis. Pre-incubation of Kami-Shoyo-San was conducted to evaluate its neuroprotection on ketamine-injured neurons. Quantitative real-time PCR and western blot assays were used to assess brain-derived neurotrophic factor and its receptor, tropomyosin receptor kinase B, in response to Kami-Shoyo-San and ketamine treatment. Brain-derived neurotrophic factor/tropomyosin receptor kinase B signaling pathway was then deactivated, by siRNA application, to further explore its functional role in Kami-Shoyo-San-mediated protection on ketamine-induced apoptosis among human embryonic stem cell-differentiated neurons. High concentration of ketamine-induced significant apoptosis, whereas pre-incubation of Kami-Shoyo-San markedly rescued ketamine-induced apoptosis, in human embryonic stem cell-differentiated neurons. Kami-Shoyo-San activated brain-derived neurotrophic factor/tropomyosin receptor kinase B signaling pathway by upregulating brain-derived neurotrophic factor and inducing tropomyosin receptor kinase B phosphorylation. Conversely, siRNA-mediated brain-derived neurotrophic factor/tropomyosin receptor kinase B signaling pathway deactivation reversed the neuroprotection of Kami-Shoyo-San in ketamine-injured human embryonic stem cell-differentiated neurons. Kami-Shoyo-San could protect ketamine-induced neurotoxicity, and the underlying mechanism may involve brain-derived neurotrophic factor/tropomyosin receptor kinase B signaling pathway.

Effects of Perinatal Exposure to Ketamine on the Developing Brain.

Initially used as an analgesic and anesthetic, ketamine has unfortunately been abused as a popular recreational party drug due to its psychotropic effects. Over the last decade, ketamine has also emerged as an effective rapid-onset anti-depressant. The increasingly widespread use and misuse of the drug in infants and pregnant women has posed a concern about the neurotoxicity of ketamine to the immature brains of developing fetuses and children. In this review, we summarize recent research findings on major possible mechanisms of perinatal ketamine-induced neurotoxicity. We also briefly summarize the neuroprotective effects of ketamine in the presence of noxious stimuli. Future actions include implementation of more drug abuse education and prevention campaigns to raise the public’s awareness of the harmful effects of ketamine abuse; further investigations to justify the clinical use of ketamine as analgesic, anesthetic and anti-depressant; and further studies to develop alternatives to ketamine or treatments that can alleviate the detrimental effects of ketamine use, especially in infants and pregnant women.

Inhibition of microRNA-375 ameliorated ketamine-induced neurotoxicity in human embryonic stem cell derived neurons

Excessive exposure to commonly used anesthetic agents, such as ketamine, may induce permanent damage to immature human brains. In this work, we used a human embryonic stem cell (hESC)-derived neuron model to assess the expression and function of human microRNA 735 (hsa-miR-375) in regulating ketamine-induced neural cell death and neural toxicity in vitro. In the in vitro culture, hESC-derived neurons were incubated with ketamine for 72 h. After that, cell viability, reactive oxygen species activity, neural apoptosis, neurite degeneration, and hsa-miR-375 gene expression were assessed, respectively. We found ketamine induced neural death, reactive oxygen species augmentation, neural apoptosis, neurite degeneration and hsa-miR-375 upregulation, in hESC-derived neurons. In addition, we discovered that, lentivirus-mediated mR-375 downregulation protected ketamine-induced neural cell death and neural toxicity. Also, human brain derived neurotrophic factor (BDNF) was found to be directly and reversely regulated by hsa-miR-375. Moreover, BDNF downregulation was shown to functionally reverse the protective effect of miR-375 downregulation on ketamine-induced neural cell death and neural toxicity. Overall, this work provided strong evidence showing hsa-miR-375 is an active regulator in anesthesia-induced neural cell death and neural toxicity, possibly via inverse regulation on BDNF gene.

Electron microscopy techniques employed to explore mitochondrial defects in the developing rat brain following ketamine treatment

Ketamine, an FDA-approved N-methyl-D-aspartate (NMDA) receptor antagonist, is commonly used for general pediatric anesthesia. Accumulating evidence has indicated that prolonged exposure to ketamine induces widespread apoptotic cell death in the developing brains of experimental animals. Although mitochondria are known to play a pivotal role in cell death, little is known about the alterations in mitochondrial ultrastructure that occur during ketamine-induced neurotoxicity. The objective of this pilot study was to utilize classic and contemporary methods in electron microscopy to study the impact of ketamine on the structure of mitochondria in the developing rat brain. While transmission electron microscopy (TEM) was employed to comprehensively study mitochondrial inner membrane topology, serial block-face scanning electron microscopy (SBF-SEM) was used as a complementary technique to compare the overall mitochondrial morphology from a representative treated and untreated neuron. In this study, postnatal day 7 (PND-7) Sprague-Dawley rats were treated with ketamine or saline (6 subcutaneous injections × 20 mg/kg or 10 ml/kg, respectively, at 2-h intervals with a 6-h withdrawal period after the last injection, n=6 each group). Samples from the frontal cortex were harvested and analyzed using TEM or SBF-SEM. While classic TEM revealed that repeated ketamine exposure induces significant mitochondrial swelling in neurons, the newer technique of SBF-SEM confirmed the mitochondrial swelling in three dimensions (3D) and showed that ketamine exposure may also induce mitochondrial fission, which was not observable in the two dimensions (2D) of TEM. Furthermore, 3D statistical analysis of these reconstructed mitochondria appeared to show that ketamine-treated mitochondria had significantly larger volumes per unit surface area than mitochondria from the untreated neuron. The ultrastructural mitochondrial alterations demonstrated here by TEM and SBF-SEM support ketamine’s proposed mechanism of neurotoxicity in the developing rat brain.

Long noncoding RNA SNHG16 reduced ketamine-induced neurotoxicity in human embryonic stem cell-derived neurons

BackgroundClinical evidence demonstrates that prolonged exposure to ketamine may cause irreversible injury to immature human brains. In this study, we utilized an in vitro model to examine the function of long noncoding RNA (lncRNA) SNHG16 in ketamine-induced neurotoxicity in human embryonic stem cell (hESC)-derived neurons. MethodsHESCs were induced toward neuronsin vitro, and treated with ketamine, at various concentrations, for 48 h. Viability, apoptosis, caspase-3 activity and ROS activity were then examined among hESC-derived neurons. Ketamine-induced gene expression change of SNHG16 was assessed by qRT-PCR. SNHG16 was overexpressed in hESC-derived neurons, which were then treated with ketamine, followed by biochemical assays to assess the effects of SNHG16 upregulation on ketamine-induced neurotoxicity. Correlation between SNHG16 and NeuroD1 gene was assess by qRT-PCR. In SNHG16-upregulated hESC-derived neurons, they were double transfected with siRNA to knock down NeuroD1. The functions of NeuroD1 inhibition on SNHG16-associated neural protection on ketamine-induced neurotoxicity were further assessed. Results48-h in vitro treatment of ketamine induced significant neurotoxicity, and downregulated SNHG16 among hESC-derived neurons. Conversely, SNHG16 upregulation reduced ketamine-induced neurotoxicity. NeuroD1 expression was downregulated by ketamine in hESC-derived neurons, and concomitantly upregulated by SNHG16 overexpression. SiRNA-mediated NeuroD1 inhibition reversed the protection of SNHG16 upregulation on ketamine-induced neurotoxicity. ConclusionsSNHG16 is an important epigenetic factor which may functionally modulate ketamine-induced neurotoxicity through NeuroD1.