Acetyl l-carnitine protects motor neurons and Rohon-Beard sensory neurons against ketamine-induced neurotoxicity in zebrafish embryos

Abstract

Ketamine, a non-competitive antagonist of N-methyl-D-aspartate (NMDA) type glutamate receptors is commonly used as a pediatric anesthetic. Multiple studies have shown ketamine to be neurotoxic, particularly when administered during the brain growth spurt. Previously, we have shown that ketamine is detrimental to motor neuron development in the zebrafish embryos. Here, using both wild type (WT) and transgenic (hb9:GFP) zebrafish embryos, we demonstrate that ketamine is neurotoxic to both motor and sensory neurons. Drug absorption studies showed that in the WT embryos, ketamine accumulation was approximately 0.4% of the original dose added to the exposure medium. The transgenic embryos express green fluorescent protein (GFP) localized in the motor neurons making them ideal for evaluating motor neuron development and toxicities in vivo. The hb9:GFP zebrafish embryos (28 h post fertilization) treated with 2 mM ketamine for 20 h demonstrated significant reductions in spinal motor neuron numbers, while co-treatment with acetyl L-carnitine proved to be neuroprotective. In whole mount immunohistochemical studies using WT embryos, a similar effect was observed for the primary sensory neurons. In the ketamine-treated WT embryos, the number of primary sensory Rohon-Beard (RB) neurons was significantly reduced compared to that in controls. However, acetyl L-carnitine co-treatment prevented ketamine-induced adverse effects on the RB neurons. These results suggest that acetyl L-carnitine protects both motor and sensory neurons from ketamine-induced neurotoxicity.

Zebrafish as a New Platform Used in Exploration of Ketamine-Induced Neurodevelopmental Toxicity

“…On the one hand lower dosage ketamine only slow down the development or differentiation of neural cells and on the other hand neurons may have degenerated by a process other than apoptosis, likely, necrosis. Thus we cannot see any differences from apoptosis staining [31].…”

“…And the axon length of motor neuron has an approximated 20% reduction compared to the control group. In embryos treated with ketamine plus acetyl L-carnitine, the number of spinal motor neurons was restored, almost to control levels, which suggests that ketamine's adverse effect on the motor neurons is prevented by acetyl L-carnitine [31].…”

“…Some researchers found out that ketamine exposure resulted in a reduction in the GFP intensity in the motor neurons resided in the cranial and spinal area of zebrafish embryos. To be more specific, there was a significant reduction (30%) in the number of spinal motor neurons in ketamine-treated embryos [29,31]. And the axon length of motor neuron has an approximated 20% reduction compared to the control group.…”

“…But after acridine orange (AO) staining, experimental groups, which were exposed to lower concentration of ketamine, showed no more cellular deaths than the control group [29][30][31]. However, exposure to higher concentration of ketamine will cause apoptosis in the region of thalamus and the connection part between midbrain and after brain.…”

“…Hb9:green-fluorescentprotein(hb9:GFP) transgenic zebrafish, in which the promoter of the transcription factor hb9 that is found in developing motor neurons of zebrafish, drives GFP expression specific ally in motor neurons [17,18,20,29,31]. Some researchers found out that ketamine exposure resulted in a reduction in the GFP intensity in the motor neurons resided in the cranial and spinal area of zebrafish embryos.…”