In Vitro Dose-Dependent Inhibition of the Intracellular Spontaneous Calcium Oscillations in Developing Hippocampal Neurons by Ketamine

Abstract

Spatial and temporal abnormalities in the frequency and amplitude of the cytosolic calcium oscillations can impact the normal physiological functions of neuronal cells. Recent studies have shown that ketamine can affect the growth and development and even induce the apoptotic death of neurons. This study used isolated developing hippocampal neurons as its study subjects to observe the effect of ketamine on the intracellular calcium oscillations in developing hippocampal neurons and to further explore its underlying mechanism using Fluo-4-loaded laser scanning confocal microscopy. Using a semi-quantitative method to analyze the spontaneous calcium oscillatory activities, a typical type of calcium oscillation was observed in developing hippocampal neurons. In addition, the administration of NMDA (N-Methyl-D-aspartate) at a concentration of 100 µM increased the calcium oscillation amplitude. The administration of MK801 at a concentration of 40 µM inhibited the amplitude and frequency of the calcium oscillations. Our results demonstrated that an increase in the ketamine concentration, starting from 30 µM, gradually decreased the neuronal calcium oscillation amplitude. The inhibition of the calcium oscillation frequency by 300 µM ketamine was statistically significant, and the neuronal calcium oscillations were completely eliminated with the administration of 3,000 µM Ketamine. The administration of 100, 300, and 1,000 µM NMDA to the 1 mM ketamine-pretreated hippocampal neurons restored the frequency and amplitude of the calcium oscillations in a dose-dependent manner. In fact, a concentration of 1,000 µM NMDA completely reversed the decrease in the calcium oscillation frequency and amplitude that was induced by 1 mM ketamine. This study revealed that ketamine can inhibit the frequency and amplitude of the calcium oscillations in developing hippocampal neurons though the NMDAR (NMDA receptor) in a dose-dependent manner, which might highlight a possible underlying mechanism of ketamine toxicity on the rat hippocampal neurons during development.