Serotonin protects C6 glioma cells from glutamate toxicity

doi:10.1016/0306-4522(94)90304-2

Neuroscience

Volume 59, Issue 4, April 1994, Pages 1043-1050

https://doi.org/10.1016/0306-4522(94)90304-2 Get rights and content

Abstract

It was recently shown that addition ofl-glutamate in millimolar amounts to a culture of C6 glioma cells induced cell death within 24 h. The mechanism for glutamate toxicity in the C6 glioma cells is linked to the inhibition of cystine uptake, leading to glutathione depletion through the cystine/glutamate antiporter (Xc¯) system. In the present study, neurotransmitters, whose receptors were localized on the glioma (glial) cells, were evaluated for their ability to protect C6 cells from glutamate toxicity through this amino acid antiporter. Among them, only 100 μM serotonin suppressed cell death by glutamate in a constant co-existence culture. The suppressive dose of serotonin was relatively low and the half-effective dose was about 35 μM. 8-Hydroxy-2-(dl-n-propylamino)tetralin, a specific serotonin_1A agonist, showed a comparable suppression to glutamate damage, while 1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane, a specific serotonin₂ agonist, and quipazine, a non-selective serotonin_1B agonist, did not suppress it. Furthermore, propranolol and pindolol significantly blocked the serotonin effect, but spiperone, mianserin and ketanserin did not block it. These results strongly indicate that this protective action of serotonin to glutamate toxicity was receptor (serotonin_1A) mediated.

Serotonin did not protect the C6 cells from glutathione depletion by glutamate. The cellular level of glutathione was depleted even under the co-existence of serotonin and glutamate. Serotonin induced a significant inhibition of lipid peroxide accumulation in the C6 glioma cells to glutamate exposure and the low rate of lipid peroxide accumulation was controlled. These results suggest that the serotonin action is not due to restoration of cellular glutathione, but due to suppression of some generating processes of lipid peroxide accumulation. This protective function of serotonin against glutamate toxicity could not be seen in the primary cortical and cerebellar neuronal culture to glutamate toxicity, which is thought to be triggered through the N-methyl-d-aspartate receptor. This novel function of serotonin on the glioma (glial) cell death by glutamate was characterized pharmacologically and its cellular mechanism for protection was discussed in comparison with that of a powerful antioxidant, vitamin E.

References (38)

AzmitiaE.C. et al.
S-100B but not NGF, EGF, insulin or calmodulin, is a CNS serotonergic growth factor
Brain Res.
(1990)
BabsonA.L. et al.
A rapid colorimetric assay for serum lactic dehydrogenase
Clin. chim. Acta
(1965)
ChoiD.W.
Glutamate neurotoxicity and diseases of the nervous system
Neuron
(1988)
FullerR.W.
The pharmacology and therapeutic potential of serotonin receptor agonists and antagonists
Adv. Drug Res.
(1988)
HamonM. et al.
Are there selective ligands for 5HT_1A and 5HT_1B receptor binding sites in brain?
Trends pharmac. Sci.
(1986)
KatoS. et al.
Sensitive and insensitive states of cultured glioma cells to glutamate damage
Brain Res.
(1984)
KatoS. et al.
Cystine/glutamate antiporter expression in retinal Müller glial cells: implications ford,l-alpha-aminoadipate toxicity
Neuroscience
(1993)
KatoS. et al.
A mechanism for glutamate toxicity in the C6 glioma cells involving inhibition of cystine uptake leading to glutathione depletion
Neuroscience
(1992)
KatoS. et al.
Gliotoxic effects of α-aminoadipic acid isomers on the carp retina: a long-term observation
Neuroscience
(1990)
LowryO.H. et al.
Protein measurement with the Folin phenol reagent
J. biol. Chem.
(1951)

MiddlemissD.N. et al.

8-Hydroxy-2-(di-n-propylamino)-tetralin discriminates between subtypes of the 5-HT recognition site

Eur. J. Pharmac.

(1983)

SchousboeA.

Transport and metabolism of glutamate and GABA in neurons and glial cells

Int. Rev. Neurobiol

(1981)

ToyookaT. et al.

Simultaneous determination of thiols and disulfides by high-performance liquid chromatography with fluorescence detection

Analyt. chim. Acta

(1988)

YagiK.

A simple fluorometric assay for lipoperoxide in blood plasma

Biochem. Med.

(1976)

CurtisD.R. et al.

Amino acid transmitters in the mammalian central nervous system

Ergebn. Physiol. Biol. Chem. exp. Pharmac.

(1974)

De DuveC. et al.

Tocopherol, Oxygen and Biomembranes

(1978)

FrazerA. et al.

Subtypes of receptors for serotonin

A. Rev. Pharmac. Toxic.

(1990)

KimelbergH.K.

Glial Cell Receptors

(1988)

Levi-MontalciniR.

The nerve growth factor 35 years later

Science

(1987)

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Citation Excerpt :
Significant alterations in the monoamines and oxidative stress were induced only when the two food additives MSG and ASM were administered in a combination dose where monoamines like DA, DOPAC and 5-HT and oxidative stress parameters like GSH were decreased, while TBARS were increased. Such neurotransmitters and oxidative stress indices have also been reported to be affected by MSG and ASM exposures in earlier studies (Dawson, 1983; Johnston et al., 1984; Dawson et al., 1989; Shinagawa, 1994; Bamforth et al., 1993) and these studies support the present findings keeping in mind the fact that the synergistic effects of MSG and ASM when administered in combination may be due to the interaction and cumulative effects of their individually caused effects that statistically remained insignificant in the present study. MSG and ASM alone showed no significant changes in biochemical parameters, most likely due to variations in the time period and age of exposures and the doses used herein.
The present study was designed to investigate the in vivo effects of monosodium glutamate (MSG) and aspartame (ASM) individually and in combination on the cognitive behavior and biochemical parameters like neurotransmitters and oxidative stress indices in the brain tissue of mice. Forty male Swiss albino mice were randomly divided into four groups of ten each and were exposed to MSG and ASM through drinking water for one month. Group I was the control and was given normal tap water. Groups II and III received MSG (8 mg/kg) and ASM (32 mg/kg) respectively dissolved in tap water. Group IV received MSG and ASM together in the same doses. After the exposure period, the animals were subjected to cognitive behavioral tests in a shuttle box and a water maze. Thereafter, the animals were sacrificed and the neurotransmitters and oxidative stress indices were estimated in their forebrain tissue. Both MSG and ASM individually as well as in combination had significant disruptive effects on the cognitive responses, memory retention and learning capabilities of the mice in the order (MSG + ASM) > ASM > MSG. Furthermore, while MSG and ASM individually were unable to alter the brain neurotransmitters and the oxidative stress indices, their combination dose (MSG + ASM) decreased significantly the levels of neurotransmitters (dopamine and serotonin) and it also caused oxidative stress by increasing the lipid peroxides measured in the form of thiobarbituric acid-reactive substances (TBARS) and decreasing the level of total glutathione (GSH). Further studies are required to evaluate the synergistic effects of MSG and ASM on the neurotransmitters and oxidative stress indices and their involvement in cognitive dysfunctions.
Reactive oxygen species involved in the glutamate toxicity of C6 glioma cells via x̄ antiporter system
1996, Neuroscience
We recently demonstrated that continuous t-glutamate exposure led to cell death in C6 glioma cells over a period of 24–36 h, due to inhibition of cystine uptake through the cystine/glutamate (xc¯) antiporter. The antioxidant vitamin E provided protection against this effect, supporting the hypothesis that depletion of glutathione might be responsible, resulting from insufficient cystine uptake. To clarify the content of oxidative stress after glutathione depletion, the present study was done to investigate accumulation and target molecules of reactive oxygen species induced by glutamate treatment. The accumulation of reactive oxygen species was increased three-fold as compared to a control culture. Membrane oxidation, as judged by lipid peroxidation, was increased two-fold after glutamate treatment. Cellular ATP content was significantly reduced by glutamate exposure. For the two cytosolic enzymes examined, activity of glyceraldehyde 3-phosphate dehydrogenase was slightly enhanced by glutamate treatment, while activity of glutamine synthetase was not changed. Impairment of nuclear DNA after glutamate exposure was also revealed by nuclear chromatin condensation with DNA fragmentation. Thus, the multiple targets (membrane, cytoplasm and nuclei) of oxygen radicals in glutamate toxicity through the xc¯antiporter system were evaluated for the first time. Furthermore, prevention from cell death and from cellular toxicity induced by oxygen radicals could be seen using three specific oxygen radical scavengers, catalase, 3,3,5,5-tetramethyl-pyrroline N-oxide and α-phenyl-N-t-butylnitrone, without restoring the glutathione deficit. This indicates that radical scavengers did not interact with the xc¯antiporter system, but directly scavenged the oxygen radicals.
Taken together, the data strongly suggest that O₂⁻, H₂0₂ and ^.OH accumulate in response to oxidative stress after glutathione depletion, resulting in glutamate cell death of C6 glioma cells.
Activation of serotonergic 5-HT<inf>1A</inf> receptor reduces Ca<sup>2+</sup>- and glutamatergic receptor-evoked arachidonic acid and NO/cGMP release in adult hippocampus
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Stimulation of glutamatergic NMDA receptor in adult rat hippocampal synaptoneurosomes induces statistically significant Ca²⁺-dependent liberation of arachidonic acid (AA) and nitric oxide (NO)-activated cGMP synthesis. NMDA acting for 5 min at 100 μM markedly increases, by approx. 25%, Ca²⁺-mediated AA release from phospholipids of hippocampal synaptoneurosomes. Prolonged stimulation of NMDA receptor up to 10 min has smaller stimulatory effect and enhances AA release by about 6%. Moreover, NMDA activates NO-dependent cGMP production by approx. 5 times more than the Ca²⁺ itself. Release of both these second messengers is completely blocked by the competitive NMDA antagonist, APV (100 μM). The NMDA-mediated cGMP elevation completely depends on NO action, and is abolished by the specific inhibitor of NO synthase, N^G-nitro-l-arginine. Moreover, serotonin at 10 μM in the presence of 10 μM pargyline, potently decreases both Ca²⁺- and NMDA receptor-mediated AA and cGMP release in hippocampal synaptoneurosomes. The agonist of 5-HT_1A receptor, buspirone, in a way similar to serotonin itself, counteracts the Ca²⁺- and also NMDA receptor-evoked AA release and cGMP accumulation. An antagonist of 5-HT_1A receptor, NAN-190, eliminates the effect of serotonin and buspirone on AA and NO/cGMP liberation. An antagonist of serotonergic 5-HT₂ receptor, ketanserin, has no effect on the Ca²⁺ and serotonin action. These results indicate that serotonin, through 5-HT_1A receptor, potently antagonizes the action of excitatory amino acid for AA release and NO/cGMP synthesis in the adult rat hippocampus. In conclusion, the interaction of serotonin with the glutamatergic system in the hippocampus may play an important role in the modulation of a signal transduction pathway, and by this molecular mechanism serotonin may exert a neuroprotective effect on hippocampal neurons. Copyright © 1996 Elsevier Science Ltd.
DL-α-aminoadipate is a toxin to Muller cells
1996, Progress in Retinal and Eye Research
Antidepressants increase glial cell line-derived neurotrophic factor production through monoamine-independent activation of protein tyrosine kinase and extracellular signal-regulated kinase in glial cells
2007, Journal of Pharmacology and Experimental Therapeutics
Mechanisms of antidepressants and serotonin (5-HT)-induced glial cell line-derived neurotrophic factor (GDNF) releases in rat C6 gliobrastoma cells
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Serotonin protects C6 glioma cells from glutamate toxicity

Abstract

Brain Res.

Clin. chim. Acta

Neuron

Adv. Drug Res.

Trends pharmac. Sci.

Brain Res.

Neuroscience

Neuroscience

Neuroscience

J. biol. Chem.

Eur. J. Pharmac.

Int. Rev. Neurobiol

Analyt. chim. Acta

Biochem. Med.

Amino acid transmitters in the mammalian central nervous system

Ergebn. Physiol. Biol. Chem. exp. Pharmac.

Tocopherol, Oxygen and Biomembranes

Subtypes of receptors for serotonin

A. Rev. Pharmac. Toxic.

Glial Cell Receptors

The nerve growth factor 35 years later

Science