Korean J Anesthesiol.  2000 Feb;38(2):350-364. 10.4097/kjae.2000.38.2.350.

Effects of Morphine on Free Radical-induced Apoptosis of Human Neuroblastoma SH-SY5Y Cells

• Ha JR
• Cheong YP
• Kim J
• Choi DH
• Lee CS
• Son Y
• Kim TY
• Song YK
• Yoon JS

• KMID: 1936362
• DOI: http://doi.org/10.4097/kjae.2000.38.2.350

Abstract

BACKGROUND: The effect of opioids on nitric oxide (NO)- and peroxynitrite-induced neuronal cell death is largely unknown. In the present study, we examined the effect of morphine on NO- and peroxynitrite-induced cell death using a human neuroblastoma SH-SY5Y cell line, which abundantly expresses micro, delta, kappa-opioid receptors.
METHODS
The cultured cells were pretreated with morphine and exposed to 3-morpholinosydnonimine (SIN-1) that simultaneously generates NO and superoxide, thus possibly forming peroxynitrite. The cell damage was assessed by using MTT assay and crystal violet staining. Morphological nuclear changes and enzymatic evidences of apoptosis of the cells after exposure to SIN-1 for 24 hours were evaluated by using 4', 6-diamidino-2-phenylindole (DAPI) staining and the measurement of pro-apoptotic protease (caspase-3) activity, respectively. Levels of reduced glutathion (GSH) were measured by monochloronimane (MCB) assay.
RESULTS
Pretreatment of SH-SY5Y with morphine significantly inhibited the apoptotic cell death. Morphine also inhibited SIN-1-induced caspase-3 (pro-apoptotic protease) activity in a dose-dependent manner. However, naloxone (20 microM) could not antagonize completely the effect of morphine in SIN- 1-induced cell death. Pre-administered GSH and N-acetylcysteine (NAC) have been found to protect SIN-induced apoptosis, and the neuroblastoma cells treated with morphine had significantly elevated the levels of GSH.
CONCLUSIONS
The present study shows that morphine protects the human neuroblastoma cell line SH- SY5Y from peroxynitrite-induced apoptotic cell death through elevated GSH levels. The protective actionof morphine seems to be associated with inhibition of the apoptotic pathway. However, it is suggested that morphine protects the cells possibly via other unknown mechanisms in addition to the activation of opioid receptors.

Keyword

Analgesics: morphine; opioid; Cells: neuroblastoma cell; SH-SY5Y; Toxicity: apoptosis; nitric oxide; peroxynitrite; superoxide

MeSH Terms

Acetylcysteine
Analgesics, Opioid
Apoptosis*
Caspase 3
Cell Death
Cell Line
Cells, Cultured
Gentian Violet
Humans*
Morphine*
Naloxone
Neuroblastoma*
Neurons
Nitric Oxide
Peroxynitrous Acid
Receptors, Opioid
Superoxides
Acetylcysteine
Analgesics, Opioid
Caspase 3
Gentian Violet
Morphine
Naloxone
Nitric Oxide
Peroxynitrous Acid
Receptors, Opioid
Superoxides