Calcium transport mechanisms in membrane vesicles from guinea pig brain synaptosomes

Donald Gill, E. F. Grollman, L. D. Kohn

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Abstract

Ca2+ transport mechanisms were investigated using membrane vesicles prepared from guinea pig brain synaptosomes by hypotonic lysis. Two major mechanisms of Ca2+ transport exist, Na+-Ca2+ exchange and ATP-dependent Ca2+ uptake. A third although minor component of Ca2+ uptake occurs under hyperpolarizing conditions (determined by increased uptake of [3H]tetraphenylphosphonium+). Na+-Ca2+ exchange results in a rapid increase of [Ca2+](i) (up to 100-fold above [Ca2+]0), has a K(m) for Ca2+ of 40 μM, is fully reversed by added external Na+, is inhibited by agents dissipating Na+ gradients (monensin or veratridine), and is uninfluenced by mitochondrial inhibitors. ATP-dependent Ca2+ uptake has a higher affinity for Ca2+ (K(m) = 12 μM), is dependent on Mg2+ or Mn2+, and is inhibited by β, γ-imidoadenosine 5'-triphosphate and VO43-, although only slightly (20%) inhibited by high concentrations of mitochondrial inhibitors. Both mechanisms are temperature-dependent, fully reversed by A23187, and higher in the presence of external K+. Ca2+ loaded in vesicles via ATP-dependent Ca2+ uptake is rapidly effluxed upon addition of external Na+ (as for Na+-Ca2+ exchange). Therefore a single population of vesicles exists containing both Ca2+ transport mechanisms. The two mechanisms are independent since they accumulate Ca2+ additively, are selectively inhibited by monensin and VO43-, and show distinct specificity toward other divalent cations and La3+. Although independent, Na+ (100 mM) inhibits ATP-dependent Ca2+ uptake (K(m) for ATP increased from 40 to 300 μM) in the absence of any net Na+ movement. Since Na+-Ca2+ exchange functions in the synaptosomal plasma membrane, the results suggest that both Ca2+ transport mechanisms originate from this membrane and function in the present experiments in inverted plasma membrane vesicles.

Original languageEnglish (US)
Pages (from-to)184-192
Number of pages9
JournalJournal of Biological Chemistry
Volume256
Issue number1
StatePublished - Dec 1 1981

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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