Separation of cardiac plasmalemma into cell surface and T-tubular components. Distribution of saxitoxin- and nitrendipine-binding sites

D. D. Doyle, T. J. Kamp, H. C. Palfrey, R. J. Miller, E. Page

Research output: Contribution to journalArticlepeer-review

26 Scopus citations

Abstract

To compare surface sarcolemmal with T-tubular distributions of [3H]saxitoxin (STX)- and [3H]nitrendipine (NTD)-binding sites, we centrifuged membrane vesicles from sheep and bovine ventricles on a 10-40% linear sucrose gradient from which fractions were assayed for STX and NTD binding; for markers of surface sarcolemma (ouabain-sensitive Na,K-ATPase activity, [3H]quinuclidinyl benzilate binding); and for markers of junctional sarcoplasmic reticulum known to be preferentially associated with T-tubules (ryanodine-sensitive Ca2+ uptake, calsequestrin, an M(r) 300,000 putative phosphorylatable 'foot' protein, and electron microscopically visible junctional sarcoplasmic reticulum-plasmalemma complexes). We identified three distinct peaks in the sucrose gradient, each characterized by significant high and low affinity STX- and high affinity NTD-binding: Peak I (~19% sucrose), highly enriched in surface sarcolemma; Peak III (~36% sucrose), enriched in junctional sarcoplasmic reticulum markers and hence in junctional sarcoplasmic reticulum complexes with T-tubule; and Peak II (~27% sucrose), showing greatest specific STX binding and only moderate NTD binding, enriched in T-tubular membrane, unassociated with junctional sarcoplasmic reticulum. For ventricular myocytes, the ratio NTD sites/STX sites was 2.5 for surface sarcolemma, but only ~1.0 for T-tubules. Unlike data published for mammalian skeletal muscle, sheep and beef cardiac NTD receptors were not significantly more concentrated in T-tubular than in surface plasmalemma.

Original languageEnglish (US)
Pages (from-to)6556-6563
Number of pages8
JournalJournal of Biological Chemistry
Volume261
Issue number14
StatePublished - 1986

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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