Tetrodotoxin-sensitive sodium channels in rat muscle cells developing in vitro

S. J. Sherman, J. C. Lawrence, D. J. Messner, K. Jacoby, W. A. Catterall

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42 Scopus citations

Abstract

Primary cultures of skeletal muscle cells from fetal rat have high affinity receptors for tetrodotoxin (TTX) and saxitoxin (STX). Specific binding of [ 3H]STX indicates the presence of a single class of high affinity sites with a K(D) = 9.6 nM at 36 °C and a B(max) = 69 fmol/mg of protein after 12 days in culture. The K(D) for TTX determined by competitive displacement of [ 3H]STX is 39 nM at 36 °C. These high affinity binding sites are associated with a functionally active sodium channel since spontaneous contractile activity is blocked by TTX with a K(I) of 26 nM. Sodium ion influx mediated by this TTX-sensitive channel was quantitated by using neurotoxins which cause persistent activation. Inhibition by TTX of 22Na + influx through channels activated by 200 μM veratridine plus 30 nM scorpion toxin could be resolved into two components. A TTX-sensitive component (K(I) = 21 nM) accounted for 40% of the Na + influx while the remaining influx was carried by the previously characterized TTX-insensitive sodium channel (K(I) = 1.7 μM). The TTX-sensitive component could be activated by veratridine alone (K 0.5 = 51 μM). Scorpion toxin enhanced the activation of TTX-sensitive Na + influx by 50 μM veratridine (K 0.5 = 6 nM) but not the activation produced by a saturating concentration (200 μM) of veratridine. Sea anemone toxin II did not have a detectable effect in enhancing veratridine action on the TTX-sensitive sodium channels. The time course of sodium channel development in vitro was investigated. TTX-insensitive 22Na + influx increased from 0 at day 1 to 250 nmol/min/mg at day 10 and remained near this maximum level for at least 10 more days. Specific [ 3H]STX binding increased from 0 to 59 fmol/mg of protein on day 10 but then declined to 21 fmol/mg of protein over the next 10 days. It is concluded that functionally active TTX-sensitive Na + channels coexist with TTX-insensitive channels in primary cultures of fetal rat muscle cells. These two channels have different toxin sensitivities and separate development regulation.

Original languageEnglish (US)
Pages (from-to)2488-2495
Number of pages8
JournalJournal of Biological Chemistry
Volume258
Issue number4
StatePublished - Aug 25 1983
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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    Sherman, S. J., Lawrence, J. C., Messner, D. J., Jacoby, K., & Catterall, W. A. (1983). Tetrodotoxin-sensitive sodium channels in rat muscle cells developing in vitro. Journal of Biological Chemistry, 258(4), 2488-2495.