Neuroprotective effects of tetrodotoxin as a Na+ channel modulator and glutamate release inhibitor in cultured rat cerebellar neurons and in gerbil global brain ischemia.
OBJECTIVE
Studies examining the role of tetrodotoxin-sensitive ion channels in hypoxic-ischemic neuronal damage have concluded that sodium influx is an important initiating event. We examined the neuroprotectant effect of tetrodotoxin on both cultured cerebellar neurons and on CA1 hippocampal neurons of gerbils exposed to brain ischemia.


METHODS
We studied neuroprotective mechanisms using cultured rat cerebellar granule cells exposed to veratridine, which induced cytotoxicity, neurotransmitter release, and calcium influx. Survival of gerbil CA1 neurons was examined by direct neuron counts 7 days after 6 minutes of global ischemia with reperfusion.


RESULTS
Tetrodotoxin protected cultured neurons in a dose-dependent manner from veratridine-induced toxicity (protective concentration [PC50] = 22 nmol/L). Veratridine induced [3H]aspartate efflux that was sodium dependent, only 25% calcium dependent, and was inhibited by tetrodotoxin (inhibitory concentration [IC50] = 60 nmol/L). Veratridine initiated increases in intracellular calcium that were also reversed by tetrodotoxin (IC50 = 63 nmol/L); reversal was dependent on the sodium-calcium exchanger and the sodium-potassium pump. Neuroprotection of 90% (n = 10; P = .001 versus vehicle) of gerbil CA1 hippocampal neurons was achieved by pretreatment with 2 ng of tetrodotoxin delivered three times intracerebroventricularly, without causing hypothermia.


CONCLUSIONS
Sodium channel blockers like tetrodotoxin may have utility in treatment of ischemic neuronal injury by preventing excessive neuronal depolarizations, limiting excitotoxic glutamate release through reversal of the sodium-dependent glutamate transporter, preventing intracellular calcium overload, preserving cellular energy stores, and allowing recovery of ionic homeostasis through operation of the sodium-calcium exchanger.
Department of Cardiovascular Pharmacology, SmithKline Beecham Pharmaceuticals, King of Prussia, Pa 19406-0939.
P G
PG
Lysko
C L
CL
Webb
T L
TL
Yue
J L
JL
Gu
G
G
Feuerstein
D000818:Animals; D001224:Aspartic Acid / Q000037:antagonists & inhibitors / Q000378:metabolism; D001921:Brain / Q000187:drug effects*; D002545:Brain Ischemia / Q000378:metabolism* / Q000503:physiopathology; D002118:Calcium / Q000378:metabolism; D000252:Calcium-Transporting ATPases / Q000187:drug effects; D002478:Cells, Cultured; D002531:Cerebellum / Q000166:cytology / Q000187:drug effects*; D004305:Dose-Response Relationship, Drug; D018691:Excitatory Amino Acid Antagonists / Q000008:administration & dosage / Q000494:pharmacology*; D005849:Gerbillinae; D018698:Glutamic Acid / Q000378:metabolism*; D006624:Hippocampus / Q000187:drug effects / Q000378:metabolism; D017136:Ion Transport / Q000187:drug effects; D009474:Neurons / Q000187:drug effects*; D018377:Neurotransmitter Agents / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D015424:Reperfusion; D026941:Sodium Channel Blockers; D015222:Sodium Channels / Q000187:drug effects*; D000254:Sodium-Potassium-Exchanging ATPase / Q000187:drug effects; D013779:Tetrodotoxin / Q000008:administration & dosage / Q000494:pharmacology*; D014701:Veratridine / Q000633:toxicity
D018691:Excitatory Amino Acid Antagonists; D018377:Neurotransmitter Agents; D026941:Sodium Channel Blockers; D015222:Sodium Channels; D001224:Aspartic Acid; D018698:Glutamic Acid; D013779:Tetrodotoxin; D014701:Veratridine; D000252:Calcium-Transporting ATPases; D000254:Sodium-Potassium-Exchanging ATPase; D002118:Calcium