Luminal communication between intracellular calcium stores modulated by GTP and the cytoskeleton

György Hajnóczky, Chi Lin, Andrew P. Thomas

Research output: Contribution to journalArticle

72 Citations (Scopus)

Abstract

The activation properties of inositol 1,4,5-trisphosphate (InsP3)- sensitive Ca2+ stores in permeabilized and intact hepatocytes were investigated by monitoring Mn2+ quench of fura-2 compartmentalized within these stores, as Mn2+ passed through InsP3-activated channels in a retrograde manner. In cells permeabilized in suspension the InsP3-sensitive pool size was dependent on InsP3 dose, and there was a large unresponsive compartment. By contrast, essentially all of the compartmentalized dye was accessible following activation of a small fraction of the InsP3 receptors in carefully permeabilized attached cells. After loading the cytosol of intact hepatocytes with Mn2+, both submaximal and maximal vasopressin doses caused complete quench of the entire intracellular pool of compartmentalized fura-2. Vasopressin-induced Mn2+ quench occurred in a stepwise manner at doses that gave cytosolic Ca2+ oscillations, reflecting periodic opening of intracellular Ca2+ channels. Pretreatment with thapsigargin to eliminate feedback effects of Ca2+ fluxes converted the steps to a continuous quench. The data suggest that Ca2+ stores in attached permeabilized and intact hepatocytes are luminally connected, making the entire store accessible to InsP3. In cells permeabilized in suspension, GTP increased InsP3-sensitive pool size, and this effect was inhibited by cytochalasin B. GTP did not change the initial rate of Mn2+ quench but increased the proportion of slowly accessible stores in the InsP3-sensitive compartment, apparently by recruitment of InsP3-insensitive stores. Preincubation on ice or with cytoskeletal inhibitors dissociated slowly accessible compartments from the InsP3-sensitive stores in both intact and subsequently permeabilized attached hepatocytes. Addition of GTP to permeabilized cells reversed this disruption of store continuity. It is suggested that GTP- and cytoskeleton- dependent luminal communication between Ca2+-stores is an important determinant of function, which could modulate the availability of Ca2+ for release.

Original languageEnglish (US)
Pages (from-to)10280-10287
Number of pages8
JournalJournal of Biological Chemistry
Volume269
Issue number14
StatePublished - Apr 8 1994

Fingerprint

Guanosine Triphosphate
Cytoskeleton
Hepatocytes
Calcium
Fura-2
Communication
Vasopressins
Suspensions
Chemical activation
Cytochalasin B
Inositol 1,4,5-Trisphosphate
Thapsigargin
Ice
Cytosol
Coloring Agents
Availability
Fluxes
Feedback
Monitoring

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Luminal communication between intracellular calcium stores modulated by GTP and the cytoskeleton. / Hajnóczky, György; Lin, Chi; Thomas, Andrew P.

In: Journal of Biological Chemistry, Vol. 269, No. 14, 08.04.1994, p. 10280-10287.

Research output: Contribution to journalArticle

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