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Electrical and neurotransmitter activity of mature neurons derived from mouse embryonic stem cells by Sox‐1 lineage selection and directed differentiation
Author(s) -
Lang R. J.,
Haynes J. M.,
Kelly J.,
Johnson J.,
Greenhalgh J.,
O'Brien C.,
Mulholland E. M.,
Baker L.,
Munsie M.,
Pouton C. W.
Publication year - 2004
Publication title -
european journal of neuroscience
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.346
H-Index - 206
eISSN - 1460-9568
pISSN - 0953-816X
DOI - 10.1111/j.1460-9568.2004.03782.x
Subject(s) - glutamate receptor , microbiology and biotechnology , excitatory postsynaptic potential , inhibitory postsynaptic potential , population , biology , nmda receptor , neuroscience , cnqx , chemistry , ampa receptor , receptor , biochemistry , demography , sociology
Sx1TV2/16C is a mouse embryonic stem (ES) cell line in which one copy of the Sox1 gene, an early neuroectodermal marker, has been targeted with a neomycin (G418) selection cassette. A combination of directed differentiation with retinoic acid and G418 selection results in an enriched neural stem cell population that can be further differentiated into neurons. After 6–7 days post‐plating (D6–7PP) most neurons readily fired tetrodotoxin (TTX)‐sensitive action potentials due to the expression of TTX‐sensitive Na + and tetraethylammonium (TEA)‐sensitive K + channels. Neurons reached their maximal cell capacitance after D6–7PP; however, ion channel expression continued until at least D21PP. The percentage of cells receiving spontaneous synaptic currents (s.s.c.) increased with days in culture until 100% of cells received a synaptic input by D20PP. Spontaneous synaptic currents were reduced in amplitude and frequency by TTX, or upon exposure to a Ca 2+ ‐free, 2.5 m m Mg 2+ saline. S.s.c. of rapid decay time constants were preferentially blocked by the nonNMDA glutamatergic receptor antagonists CNQX or NBQX. Ca 2+ levels within ES cell‐derived neurons increased in response to glutamate receptor agonists l ‐glutamate, AMPA, N ‐methyl‐ d ‐aspartate (NMDA) and kainic acid and to acetylcholine, ATP and dopamine. ES cell‐derived neurons also generated cationic and Cl – ‐selective currents in response to NMDA and glycine or GABA, respectively. It was concluded that ES‐derived neurons fire action potentials, receive excitatory and inhibitory synaptic input and respond to various neurotransmitters in a manner akin to primary central neurons.