Rates of insulin secretion in INS-1 cells are enhanced by coupling to anaplerosis and Kreb's cycle flux independent of ATP synthesis

Gary W. Cline, Rebecca L. Pongratz, Xiaojian Zhao, Klearchos K Papas

Research output: Contribution to journalArticle

12 Citations (Scopus)

Abstract

Mechanistic models of glucose stimulated insulin secretion (GSIS) established in minimal media in vitro, may not accurately describe the complexity of coupling metabolism with insulin secretion that occurs in vivo. As a first approximation, we have evaluated metabolic pathways in a typical growth media, DMEM as a surrogate in vivo medium, for comparison to metabolic fluxes observed under the typical experimental conditions using the simple salt-buffer of KRB. Changes in metabolism in response to glucose and amino acids and coupling to insulin secretion were measured in INS-1 832/13 cells. Media effects on mitochondrial function and the coupling efficiency of oxidative phosphorylation were determined by fluorometrically measured oxygen consumption rates (OCRs) combined with 31P NMR measured rates of ATP synthesis. Substrate preferences and pathways into the TCA cycle, and the synthesis of mitochondrial 2nd messengers by anaplerosis were determined by 13C NMR isotopomer analysis of the fate of [U- 13C] glucose metabolism.Despite similar incremental increases in insulin secretion, the changes of OCR in response to increasing glucose from 2.5 to 15. mM were blunted in DMEM relative to KRB. Basal and stimulated rates of insulin secretion rates were consistently higher in DMEM, while ATP synthesis rates were identical in both DMEM and KRB, suggesting greater mitochondrial uncoupling in DMEM. The relative rates of anaplerosis, and hence synthesis and export of 2nd messengers from the mitochondria were found to be similar in DMEM to those in KRB. And, the correlation of total PC flux with insulin secretion rates in DMEM was found to be congruous with the correlation in KRB. Together, these results suggest that signaling mechanisms associated with both TCA cycle flux and with anaplerotic flux, but not ATP production, may be responsible for the enhanced rates of insulin secretion in more complex, and physiologically-relevant media.

Original languageEnglish (US)
Pages (from-to)30-35
Number of pages6
JournalBiochemical and Biophysical Research Communications
Volume415
Issue number1
DOIs
StatePublished - Nov 11 2011
Externally publishedYes

Fingerprint

Adenosine Triphosphate
Insulin
Fluxes
Metabolism
Glucose
Oxygen Consumption
Nuclear magnetic resonance
Oxygen
Mitochondria
Oxidative Phosphorylation
Metabolic Networks and Pathways
Buffers
Salts
Amino Acids
Substrates
Growth

Keywords

  • Anaplerosis
  • ATP synthesis
  • Beta-cells
  • Glucose-stimulated insulin secretion
  • INS-1
  • Mitochondrial metabolism
  • Second messengers
  • Substrate cycling

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Cell Biology
  • Molecular Biology

Cite this

Rates of insulin secretion in INS-1 cells are enhanced by coupling to anaplerosis and Kreb's cycle flux independent of ATP synthesis. / Cline, Gary W.; Pongratz, Rebecca L.; Zhao, Xiaojian; Papas, Klearchos K.

In: Biochemical and Biophysical Research Communications, Vol. 415, No. 1, 11.11.2011, p. 30-35.

Research output: Contribution to journalArticle

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