Ca2+-associated triphasic pH changes in mitochondria during brown adipocyte activation

Yanyan Hou, Tetsuya Kitaguchi, Rókus Kriszt, Yu Hua Tseng, Michael Raghunath, Madoka Suzuki

Research output: Contribution to journalArticle

2 Citations (Scopus)

Abstract

Objective Brown adipocytes (BAs) are endowed with a high metabolic capacity for energy expenditure due to their high mitochondria content. While mitochondrial pH is dynamically regulated in response to stimulation and, in return, affects various metabolic processes, how mitochondrial pH is regulated during adrenergic stimulation-induced thermogenesis is unknown. We aimed to reveal the spatial and temporal dynamics of mitochondrial pH in stimulated BAs and the mechanisms behind the dynamic pH changes. Methods A mitochondrial targeted pH-sensitive protein, mito-pHluorin, was constructed and transfected to BAs. Transfected BAs were stimulated by an adrenergic agonist, isoproterenol. The pH changes in mitochondria were characterized by dual-color imaging with indicators that monitor mitochondrial membrane potential and heat production. The mechanisms of pH changes were studied by examining the involvement of electron transport chain (ETC) activity and Ca2+ profiles in mitochondria and the intracellular Ca2+ store, the endoplasmic reticulum (ER). Results A triphasic mitochondrial pH change in BAs upon adrenergic stimulation was revealed. In comparison to a thermosensitive dye, we reveal that phases 1 and 2 of the pH increase precede thermogenesis, while phase 3, characterized by a pH decrease, occurs during thermogenesis. The mechanism of pH increase is partially related to ETC. In addition, the pH increase occurs concurrently with an increase in mitochondrial Ca2+. This Ca2+ increase is contributed to by an influx from the ER, and it is further involved in mitochondrial pH regulation. Conclusions We demonstrate that an increase in mitochondrial pH is implicated as an early event in adrenergically stimulated BAs. We further suggest that this pH increase may play a role in the potentiation of thermogenesis.

Original languageEnglish
Pages (from-to)797-808
Number of pages12
JournalMolecular Metabolism
Volume6
Issue number8
DOIs
Publication statusPublished - 2017 Aug 1

Fingerprint

Brown Adipocytes
Mitochondria
Thermogenesis
Electron Transport
Endoplasmic Reticulum
Adrenergic Agents
Mitochondrial Dynamics
Adrenergic Agonists
Mitochondrial Membrane Potential

Keywords

  • Brown adipocytes
  • Ca
  • Confocal microscopy
  • Endoplasmic reticulum
  • Fluorescence imaging
  • Mitochondria-associated ER membrane

ASJC Scopus subject areas

  • Molecular Biology
  • Cell Biology

Cite this

Hou, Y., Kitaguchi, T., Kriszt, R., Tseng, Y. H., Raghunath, M., & Suzuki, M. (2017). Ca2+-associated triphasic pH changes in mitochondria during brown adipocyte activation. Molecular Metabolism, 6(8), 797-808. https://doi.org/10.1016/j.molmet.2017.05.013

Ca2+-associated triphasic pH changes in mitochondria during brown adipocyte activation. / Hou, Yanyan; Kitaguchi, Tetsuya; Kriszt, Rókus; Tseng, Yu Hua; Raghunath, Michael; Suzuki, Madoka.

In: Molecular Metabolism, Vol. 6, No. 8, 01.08.2017, p. 797-808.

Research output: Contribution to journalArticle

Hou, Y, Kitaguchi, T, Kriszt, R, Tseng, YH, Raghunath, M & Suzuki, M 2017, 'Ca2+-associated triphasic pH changes in mitochondria during brown adipocyte activation', Molecular Metabolism, vol. 6, no. 8, pp. 797-808. https://doi.org/10.1016/j.molmet.2017.05.013
Hou, Yanyan ; Kitaguchi, Tetsuya ; Kriszt, Rókus ; Tseng, Yu Hua ; Raghunath, Michael ; Suzuki, Madoka. / Ca2+-associated triphasic pH changes in mitochondria during brown adipocyte activation. In: Molecular Metabolism. 2017 ; Vol. 6, No. 8. pp. 797-808.
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N2 - Objective Brown adipocytes (BAs) are endowed with a high metabolic capacity for energy expenditure due to their high mitochondria content. While mitochondrial pH is dynamically regulated in response to stimulation and, in return, affects various metabolic processes, how mitochondrial pH is regulated during adrenergic stimulation-induced thermogenesis is unknown. We aimed to reveal the spatial and temporal dynamics of mitochondrial pH in stimulated BAs and the mechanisms behind the dynamic pH changes. Methods A mitochondrial targeted pH-sensitive protein, mito-pHluorin, was constructed and transfected to BAs. Transfected BAs were stimulated by an adrenergic agonist, isoproterenol. The pH changes in mitochondria were characterized by dual-color imaging with indicators that monitor mitochondrial membrane potential and heat production. The mechanisms of pH changes were studied by examining the involvement of electron transport chain (ETC) activity and Ca2+ profiles in mitochondria and the intracellular Ca2+ store, the endoplasmic reticulum (ER). Results A triphasic mitochondrial pH change in BAs upon adrenergic stimulation was revealed. In comparison to a thermosensitive dye, we reveal that phases 1 and 2 of the pH increase precede thermogenesis, while phase 3, characterized by a pH decrease, occurs during thermogenesis. The mechanism of pH increase is partially related to ETC. In addition, the pH increase occurs concurrently with an increase in mitochondrial Ca2+. This Ca2+ increase is contributed to by an influx from the ER, and it is further involved in mitochondrial pH regulation. Conclusions We demonstrate that an increase in mitochondrial pH is implicated as an early event in adrenergically stimulated BAs. We further suggest that this pH increase may play a role in the potentiation of thermogenesis.

AB - Objective Brown adipocytes (BAs) are endowed with a high metabolic capacity for energy expenditure due to their high mitochondria content. While mitochondrial pH is dynamically regulated in response to stimulation and, in return, affects various metabolic processes, how mitochondrial pH is regulated during adrenergic stimulation-induced thermogenesis is unknown. We aimed to reveal the spatial and temporal dynamics of mitochondrial pH in stimulated BAs and the mechanisms behind the dynamic pH changes. Methods A mitochondrial targeted pH-sensitive protein, mito-pHluorin, was constructed and transfected to BAs. Transfected BAs were stimulated by an adrenergic agonist, isoproterenol. The pH changes in mitochondria were characterized by dual-color imaging with indicators that monitor mitochondrial membrane potential and heat production. The mechanisms of pH changes were studied by examining the involvement of electron transport chain (ETC) activity and Ca2+ profiles in mitochondria and the intracellular Ca2+ store, the endoplasmic reticulum (ER). Results A triphasic mitochondrial pH change in BAs upon adrenergic stimulation was revealed. In comparison to a thermosensitive dye, we reveal that phases 1 and 2 of the pH increase precede thermogenesis, while phase 3, characterized by a pH decrease, occurs during thermogenesis. The mechanism of pH increase is partially related to ETC. In addition, the pH increase occurs concurrently with an increase in mitochondrial Ca2+. This Ca2+ increase is contributed to by an influx from the ER, and it is further involved in mitochondrial pH regulation. Conclusions We demonstrate that an increase in mitochondrial pH is implicated as an early event in adrenergically stimulated BAs. We further suggest that this pH increase may play a role in the potentiation of thermogenesis.

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