Time-lapse imaging of Ca2+-induced swelling and permeability transition: Single mitochondrion study

Shibata, Takahiro; Yoneda, Mayu; Morikawa, Daisuke; Ohta, Yoshihiro

doi:10.1016/j.abb.2019.01.016

Cited by 14 publications

(15 citation statements)

References 34 publications

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“…To discriminate between the enlargement of the same pore(s) and the opening of the new bigger one in the same organelle(s) (mechanisms 2 and 3) is methodically possible neither in mitochondrial suspension nor in single mitochondria. 57,58 Nevertheless, our results correlate with the data obtained on MMCs by the patch-clamp method. It was established that individual MMCs in OSM can operate in different states of conductance (~350, 500-650, ~1000, and 1300 pS).…”

Section: Discussionsupporting

confidence: 88%

“…Besides, an elevation of Ca 2+ concentration increased the transparency to light in a part of mitochondria. 58 In our experiments, there was a rather weak correlation between the amplitude of mitochondrial swelling and the average PTP size in suspension (compare Figures 5 and S2). However, the increase in the mitochondrial transparency to light due to the MOM damage can occur (see Figure 3).…”

Section: Discussionmentioning

confidence: 57%

“…However, the increase in the mitochondrial transparency to light due to the MOM damage can occur (see Figure 3). Besides, the opening of several ~3-nm pores and/or their enlargement in 500-1000-nm organelles may contribute considerably to 6%-8% changes in mitochondrial transparency to light observed by Shibata et al 58 After the induction of the PTP, the rate of its transition to a higher conductance state (Da/min) almost linearly depended on the Ca 2+ concentration in the medium (Figure 5F): the higher Ca 2+ , the greater quantity of PTPs become bigger or better adapt to large PEG. Hence, Ca 2+ makes the structure of the PTP channel more elastic.…”

Section: Discussionmentioning

confidence: 94%

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Dynamics of the permeability transition pore size in isolated mitochondria and mitoplasts

et al. 2021

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Section: Discussionsupporting

confidence: 88%

Section: Discussionmentioning

confidence: 57%

Section: Discussionmentioning

confidence: 94%

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Dynamics of the permeability transition pore size in isolated mitochondria and mitoplasts

et al. 2021

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“…The current values seen in these experiments are consistent with very few pores, but further experiments will be needed to confirm this interpretation. Further, very recent imaging data measuring single mitochondrial swelling under conditions similar to those presented in this work demonstrated that swelling occurs in a steplike manner, suggesting that it is induced by the activation of a relatively small number of pores (Morikawa et al, 2014;Shibata et al, 2019). Interestingly, this study reported that despite the fact that all mitochondria undergo depolarization indicating PT activation, not all of them swell, suggesting an intriguing possibility that the high-conductance mode of PTP is not activated in every mitochondrion.…”

Section: Discussionsupporting

confidence: 61%

The very low number of calcium-induced permeability transition pores in the single mitochondrion

Neginskaya

Strubbe

Amodeo

et al. 2020

Journal of General Physiology

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Mitochondrial permeability transition (PT) is a phenomenon of stress-induced increase in nonspecific permeability of the mitochondrial inner membrane that leads to disruption of oxidative phosphorylation and cell death. Quantitative measurement of the membrane permeability increase during PT is critically important for understanding the PT’s impact on mitochondrial function. The elementary unit of PT is a PT pore (PTP), a single channel presumably formed by either ATP synthase or adenine nucleotide translocator (ANT). It is not known how many channels are open in a single mitochondrion during PT, which makes it difficult to quantitatively estimate the overall degree of membrane permeability. Here, we used wide-field microscopy to record mitochondrial swelling and quantitatively measure rates of single-mitochondrion volume increase during PT-induced high-amplitude swelling. PT was quantified by calculating the rates of water flux responsible for measured volume changes. The total water flux through the mitochondrial membrane of a single mitochondrion during PT was in the range of (2.5 ± 0.4) × 10−17 kg/s for swelling in 2 mM Ca2+ and (1.1 ± 0.2) × 10−17 kg/s for swelling in 200 µM Ca2+. Under these experimental conditions, a single PTP channel with ionic conductance of 1.5 nS could allow passage of water at the rate of 0.65 × 10−17 kg/s. Thus, we estimate the integral ionic conductance of the whole mitochondrion during PT to be 5.9 ± 0.9 nS for 2 mM concentration of Ca2+ and 2.6 ± 0.4 nS for 200 µM of Ca2+. The number of PTPs per mitochondrion ranged from one to nine. Due to the uncertainties in PTP structure and model parameters, PTP count results may be slightly underestimated. However, taking into account that each mitochondrion has ∼15,000 copies of ATP synthases and ANTs, our data imply that PTP activation is a rare event that occurs only in a small subpopulation of these proteins.

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“…Interestingly, in our study, we observed decreased autophagy-related protein expression in the shCTSS groups after silence of MCU. We hypothesize that knockdown of CTSS causes two kinds of effects: 1) because CTSS knockdown causes the malfunction of lysosomal function ( 34 ), autophagosomes accumulate in the cytoplasm, and damaged mitochondria cannot be eliminated in time and therefore accumulate in the cytoplasm; and 2) elevated cytoplasmic Ca 2+ and MCU levels contribute to mitochondrial Ca 2+ overload and cause the mitochondria to deteriorate ( 43 ), which induces mitochondrial apoptosis and exacerbates the burden of autophagy. The blockade of MCU complex partly broke this vicious circle, maintaining the balance of mitochondrial Ca 2+ levels and decreasing mitochondrial damage, thus decreasing autophagosome accumulation.…”

Section: Discussionmentioning

confidence: 99%

Inhibition of Cathepsin S Induces Mitochondrial Apoptosis in Glioblastoma Cell Lines Through Mitochondrial Stress and Autophagosome Accumulation

et al. 2020

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Cathepsin S (CTSS), a lysosomal cysteine protease, is overexpressed in various cancers, including glioblastoma (GB). A high level of CTSS is associated with tumor progression and poor outcome in GB. However, the underlying mechanisms of its role in the biological characteristics of G5B remain to be elucidated. Here, we uncovered a potential role of CTSS in the lysosomes and mitochondria of GB cells (GBCs). Downregulation of CTSS in GBCs could increase the expression of autophagy-related proteins; however, there was no significant change in p62, suggesting autophagy blockade. Moreover, inhibition of CTSS increased the expression of mitochondrial calcium uniporter (MCU) and enhanced mitochondrial Ca2+ uptake ability, causing mitochondrial Ca2+ overload, the generation of copious reactive oxygen species (ROS) and eventual mitochondrial apoptosis. Additionally, elevated damage to mitochondria exacerbated the burden of autophagy. Finally, we found that silence of MCU could alleviate the inhibition of CTSS-induced autophagosome accumulation and mitochondrial stress. Collectively, these results demonstrate that CTSS plays an important role in the process of autophagic flux and mitochondrial functions in GBCs.

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Time-lapse imaging of Ca2+-induced swelling and permeability transition: Single mitochondrion study

Cited by 14 publications

References 34 publications

Dynamics of the permeability transition pore size in isolated mitochondria and mitoplasts

Dynamics of the permeability transition pore size in isolated mitochondria and mitoplasts

The very low number of calcium-induced permeability transition pores in the single mitochondrion

Inhibition of Cathepsin S Induces Mitochondrial Apoptosis in Glioblastoma Cell Lines Through Mitochondrial Stress and Autophagosome Accumulation

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