Influence of mTOR signaling pathway on ketamine-induced injuries in the hippocampal neurons of rats

ABSTRACT

Objective: To explore the influences of mammalian target of rapamycin (mTOR) signaling pathway on ketamine-induced apoptosis, oxidative stress and Ca²⁺ concentration in the hippocampal neurons of rats.

Methods: The primary hippocampal neurons isolated from fetal Sprague Dawley rats were treated with ketamine (0, 50, 100 and 500 μM) for 4 days to observe its effect on mTOR signaling pathway and apoptosis of rat hippocampal neurons. Then, the hippocampal neurons were divided into C (Control), R (Rapamycin, an inhibitor of mTOR signaling pathway), K (Ketamine) and R + K (Rapamycin + Ketamine) groups to detect the apoptosis, reactive oxygen species (ROS) production, and Ca²⁺ concentration via the terminal transferase uridyl nick end labelling (TUNEL) assay, dichloro-dihydro-fluorescein diacetate (DCFH-DA) method and Fluo-3 acetoxymethyl ester (Fluo-3AM) staining, respectively. The expressions of mTOR signaling pathway and apoptosis-related proteins in hippocampal neurons were examined by qRT-PCR and Western blot.

Results: Ketamine could dose-dependently promote the apoptosis of rat hippocampal neurons with upregulation of p-mTOR and its downstream regulators (p-4E-BP-1 and p-p70S6K). However, ketamine-induced apoptosis in hippocampal neurons was reversed significantly by the administration of rapamycin, as evident by the decrease in expressions of pro-apoptotic proteins (Bax and cleaved Caspase-3) and the increase in anti-apoptotic protein (Bcl-2). Meanwhile, the ROS generation and Ca²⁺ concentration was inhibited accompanied with reduced malonildialdehyde levels but elevated superoxide and glutathione peroxidase activities.

Conclusion: Inhibition of mTOR signaling pathway protected rat hippocampal neurons from ketamine-induced injuries via reducing apoptosis, oxidative stress, as well as Ca²⁺ concentration.

Abbreviations: mTOR: mammalian target of rapamycin; SD: Sprague-Dawley; SPF: Specific-pathogen free; ROS: reactive oxygen species; TUNEL: terminal transferase uridyl nick end labelling; DCFH-DA: Dichloro-dihydro-fluorescein diacetate; Fluo-3A: Fluo-3 acetoxymethyl ester; NMDAR: non-competitive N-methyl-D-aspartame glutamate receptor; 4E-BP1: 4E binding protein 1; p70S6K: p70 S6 Kinase; PCR: Polymerase chain reaction; MDA: malonildialdehyde; GSH-PX: glutathione peroxidase; ANOVA: One-way Analysis of Variance.

KEYWORDS:

• mTOR signaling pathway
• ketamine
• hippocampal neurons
• cell apoptosis

Acknowledgments

We would like to give our sincere appreciation to the reviewers for their suggestions on this work.

Disclosure statement

No potential conflict of interest was reported by the authors.

Additional information

Notes on contributors

Fei-fei Liu

Fei-fei Liu He worked in the Third Hospital of Hebei Medical University with the main research areas of memory dysfunction and anaesthesia in the elderly.

Shuang Zhao

Shuang Zhao He graduated from Hebei Medical University, and is currently the deputy chief physician, specializing in anesthesia for clinical senior patients.

Peng Liu

Peng Liu He graduated from Hebei Medical University, and works in the Third Hospital of Hebei Medical University with neuropathic pain as his research field.

Shu-ping Huo

Shuping Huo He graduated from Hebei Medical University,specializing in anesthesia and nerve block anesthesia in clinical senior patients.