Metabolic Pools of ATP in Cultured Bovine Adrenal Medullary Chromaffin Cells
- 1 September 1986
- journal article
- research article
- Published by Wiley in Journal of Neurochemistry
- Vol. 47 (3), 945-952
- https://doi.org/10.1111/j.1471-4159.1986.tb00702.x
Abstract
Cultured bovine adrenal chromaffin cells contain a pool of ATP sequestered within the chromaffin vesicles and an extravesicular pool of ATP. In a previous study it was shown that the turnover of ATP in the extravesicular pool was biphasic. One phase occurred with a t1/2 of 3.5-4.5 h whereas the second phase occurred with a t1/2 of several days. The studies described here were undertaken to characterize further vesicular and extravesicular pools of ATP by examining the effects of metabolic inhibitors, adenosine, and digitonin on ATP utilization and subcellular localization immediately after and 48 h after labeling with [3H]adenosine and 32Pi. Immediately after labeling a combination of cyanide, 2-deoxy-D-glucose, the .beta.-glucono-1,5-lactone resulted in a 90-95% depletion of the labeled ATP but only a 25% depletion of the endogenous ATP within 30 min. Forty-eight hours after labeling, addition of the inhibitors resulted in a 70% depletion of the [3H]ATP but only a 25% depletion of the [32P]ATP and endogenous ATP. Addition of 10 .mu.M adenosine to the media resulted in a similar loss of [3H]ATP in cells examined immediately after or 48 h after labeling. Adenosine increased the amounts of [32P]ATP when added immediately after labeling but had no effect on the [32P]ATP content when added 48 h after labeling. Permeabilization of the plasma membrane by treatment with digitonin resulted in a loss of 90-95% of the labeled ATP but only 20% of the endogenous ATP in cells examined immediately after labeling and in a 79% loss of [3H]ATP, a 32% loss of [32P]ATP, and a 23% loss of endogenous ATP in cells examined 48 h after labeling. These studies show that ATP contained within chromaffin vesicles is metabolically inert and does not exchange readily with cytosolic ATP, but they do not differentiate extravesicular pools of labeled ATP. Calculation of ATP disappearance after abrupt cessation of ATP production indicates that cultured adrenal chromaffin cells utilize ATP at a rate of 0.24 nmol/min/106 cells, corresponding to a turnover time of 12.5 min for the extravesicular pool.Keywords
This publication has 14 references indexed in Scilit:
- Turnover and Storage of Newly Synthesized Adenine Nucleotides in Bovine Adrenal Medullary Cell CulturesJournal of Neurochemistry, 1986
- Purine Nucleotide Synthesis in Adrenal Chromaffin CellsJournal of Neurochemistry, 1985
- Arachidonic Acid Release and Catecholamine Secretion from Digitonin‐Treated Chromaffin Cells: Effects of Micromolar Calcium, Phorbol Ester, and Protein Alkylating AgentsJournal of Neurochemistry, 1985
- Metabolic energy is required in human platelets at any stage during optical aggregation and secretionBiochimica et Biophysica Acta (BBA) - General Subjects, 1984
- Phosphorus-31 nuclear magnetic resonance studies of pig adrenal glandsNeuroscience, 1984
- Improved method for high energy nucleotide analysis of canine cardiac muscle using reversed-phase high-performance liquid chromatographyJournal of Chromatography B: Biomedical Sciences and Applications, 1983
- Dynamic aspects of chromaffin granule structureNeuroscience, 1982
- Quantification of Pathways of Glucose Utilization and Balance of Energy Metabolism of Rabbit ReticulocytesEuropean Journal of Biochemistry, 1982
- [40] Thin-layer separation methods for nucleic acid derivativesPublished by Elsevier ,1967
- A semiautomatic method for determination of catecholaminesAnalytical Biochemistry, 1963