Tetramethylenedisulfotetramine alters Ca²⁺ dynamics in cultured hippocampal neurons: mitigation by NMDA receptor blockade and GABA(A) receptor-positive modulation.
Tetramethylenedisulfotetramine (TETS) is a potent convulsant that is considered a chemical threat agent. We characterized TETS as an activator of spontaneous Ca²⁺ oscillations and electrical burst discharges in mouse hippocampal neuronal cultures at 13-17 days in vitro using FLIPR Fluo-4 fluorescence measurements and extracellular microelectrode array recording. Acute exposure to TETS (≥ 2 µM) reversibly altered the pattern of spontaneous neuronal discharges, producing clustered burst firing and an overall increase in discharge frequency. TETS also dramatically affected Ca²⁺ dynamics causing an immediate but transient elevation of neuronal intracellular Ca²⁺ followed by decreased frequency of Ca²⁺ oscillations but greater peak amplitude. The effect on Ca²⁺ dynamics was similar to that elicited by picrotoxin and bicuculline, supporting the view that TETS acts by inhibiting type A gamma-aminobutyric acid (GABA(A)) receptor function. The effect of TETS on Ca²⁺ dynamics requires activation of N-methyl-D-aspartic acid (NMDA) receptors, because the changes induced by TETS were prevented by MK-801 block of NMDA receptors, but not nifedipine block of L-type Ca²⁺ channels. Pretreatment with the GABA(A) receptor-positive modulators diazepam and allopregnanolone partially mitigated TETS-induced changes in Ca²⁺ dynamics. Moreover, low, minimally effective concentrations of diazepam (0.1 µM) and allopregnanolone (0.1 µM), when administered together, were highly effective in suppressing TETS-induced alterations in Ca²⁺ dynamics, suggesting that the combination of positive modulators of synaptic and extrasynaptic GABA(A) receptors may have therapeutic potential. These rapid throughput in vitro assays may assist in the identification of single agents or combinations that have utility in the treatment of TETS intoxication.
Department of Molecular Biosciences, School of Veterinary Medicine, University of California, Davis, California 95616, USA.
Zhengyu
Z
Cao
Bruce D
BD
Hammock
Mark
M
McCoy
Michael A
MA
Rogawski
Pamela J
PJ
Lein
Isaac N
IN
Pessah
D000200:Action Potentials; D000818:Animals; D000831:Animals, Newborn; D000927:Anticonvulsants / Q000494:pharmacology*; D001640:Bicuculline / Q000633:toxicity; D001952:Bridged-Ring Compounds / Q000633:toxicity*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D020013:Calcium Signaling / Q000187:drug effects*; D002478:Cells, Cultured; D003292:Convulsants / Q000633:toxicity*; D003975:Diazepam / Q000494:pharmacology; D016291:Dizocilpine Maleate / Q000633:toxicity; D004305:Dose-Response Relationship, Drug; D018691:Excitatory Amino Acid Antagonists / Q000633:toxicity*; D058787:GABA-A Receptor Antagonists / Q000633:toxicity; D057166:High-Throughput Screening Assays / Q000295:instrumentation / Q000379:methods; D006624:Hippocampus / Q000187:drug effects* / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D008839:Microelectrodes; D009474:Neurons / Q000187:drug effects* / Q000378:metabolism; D010852:Picrotoxin / Q000633:toxicity; D011280:Pregnanolone / Q000494:pharmacology; D011963:Receptors, GABA-A / Q000187:drug effects* / Q000378:metabolism; D016194:Receptors, N-Methyl-D-Aspartate / Q000037:antagonists & inhibitors* / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D013997:Time Factors
D000927:Anticonvulsants; D001640:Bicuculline; D001952:Bridged-Ring Compounds; D002121:Calcium Channel Blockers; D003292:Convulsants; D003975:Diazepam; D016291:Dizocilpine Maleate; D018691:Excitatory Amino Acid Antagonists; D058787:GABA-A Receptor Antagonists; D010852:Picrotoxin; D011280:Pregnanolone; D011963:Receptors, GABA-A; D016194:Receptors, N-Methyl-D-Aspartate; C010349:tetramethylenedisulfotetramine