Calcium green-5N, a novel fluorescent probe for monitoring high intracellular free Ca2+ concentrations associated with glutamate excitotoxicity in cultured rat brain neurons

Rajdev, Sunita; Reynolds, Ian J.

doi:10.1016/0304-3940(93)90582-6

Cited by 104 publications

(51 citation statements)

References 18 publications

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“…CG5N is canonically used as an extracellular dye in eukaryotic cell biology (34); however, we observed that extended incubation time (≥1 h) of M. testarum strain BC008 in CG5N resulted in intracellular CG5N signal. Actively excavated mineral chip fragments were placed in 10 mL PES-30 with 1 μM CG5N and incubated for at least 1 h under standard culture conditions before imaging.…”

Section: Methodsmentioning

confidence: 40%

Extreme cellular adaptations and cell differentiation required by a cyanobacterium for carbonate excavation

Guida

García-Pichel

2016

Proc. Natl. Acad. Sci. U.S.A.

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Some cyanobacteria, known as euendoliths, excavate and grow into calcium carbonates, with their activity leading to significant marine and terrestrial carbonate erosion and to deleterious effects on coral reef and bivalve ecology. Despite their environmental relevance, the mechanisms by which they can bore have remained elusive and paradoxical, in that, as oxygenic phototrophs, cyanobacteria tend to alkalinize their surroundings, which will encourage carbonate precipitation, not dissolution. Therefore, cyanobacteria must rely on unique adaptations to bore. Studies with the filamentous euendolith, Mastigocoleus testarum, indicated that excavation requires both cellular energy and transcellular calcium transport, mediated by P-type ATPases, but the cellular basis for this phenomenon remains obscure. We present evidence that excavation in M. testarum involves two unique cellular adaptations. Long-range calcium transport is based on active pumping at multiple cells along boring filaments, orchestrated by the preferential localization of calcium ATPases at one cell pole, in a ring pattern, facing the cross-walls, and by repeating this placement and polarity, a pattern that breaks at branching and apical cells. In addition, M. testarum differentiates specialized cells we call calcicytes, that which accumulate calcium at concentrations more than 500-fold those found in other cyanobacteria, concomitantly and drastically lowering photosynthetic pigments and enduring severe cytoplasmatic alkalinization. Calcicytes occur commonly, but not exclusively, in apical parts of the filaments distal to the excavation front. We suggest that calcicytes allow for fast calcium flow at low, nontoxic concentrations through undifferentiated cells by providing buffering storage for excess calcium before final excretion to the outside medium.

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Section: Methodsmentioning

confidence: 40%

Extreme cellular adaptations and cell differentiation required by a cyanobacterium for carbonate excavation

Guida

García-Pichel

2016

Proc. Natl. Acad. Sci. U.S.A.

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“…Mitochondrial Ca 2+ fluxes were measured by monitoring the changes in Ca 2+ concentration in the reaction medium using the hexapotassium salt of the fluorescence probe Calcium Green 5-N [20]. Mitochondria (0.8 mg) were resuspended in 2 ml of reaction medium supplemented with 100 nM Calcium Green 5-N followed by addition of Ca 2+ (45 nmol) and energization with 5 mM succinate.…”

Section: Measurement Of Ca 2+ Fluxesmentioning

confidence: 99%

Doxorubicin increases the susceptibility of brain mitochondria to Ca2+-induced permeability transition and oxidative damage

Cardoso

Santos

Carvalho

et al. 2008

Free Radical Biology and Medicine

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This study was aimed at investigating the effects of subchronic administration of doxorubicin (DOX) on brain mitochondrial bioenergetics and oxidative status. Rats were treated with seven weekly injections of vehicle (sc, saline solution) or DOX (sc, 2 mg kg −1 ), and 1 week after the last administration of the drug the animals were sacrificed and brain mitochondrial fractions were obtained. Several parameters were analyzed: respiratory chain, phosphorylation system, induction of the permeability transition pore (PTP), mitochondrial aconitase activity, lipid peroxidation markers, and nonenzymatic antioxidant defenses. DOX treatment induced an increase in thiobarbituric acid-reactive substances and vitamin E levels and a decrease in reduced glutathione content and aconitase activity. Furthermore, DOX potentiated PTP induced by Ca 2+ . No statistical differences were observed in the other parameters analyzed. Altogether our results show that DOX treatment increases the susceptibility of brain mitochondria to Ca 2+ -induced PTP opening and oxidative stress, predisposing brain cells to degeneration and death.© 2008 Elsevier Inc. All rights reserved. IntroductionDoxorubicin (DOX) is a potent broad-spectrum antineoplastic agent effective in the treatment of a wide variety of cancers, including both solid tumors and leukemias [1]. However, the chronic administration of this drug can induce toxicity to nontarget tissues, cardiotoxicity being the best known side effect [2]. DOX-induced cardiotoxicity has been attributed to a number of effects, including the direct inhibition of key transporters involved in ion homeostasis, alterations in cellular iron and calcium metabolism, disruption of sarcoplasmic reticulum function, mitochondrial dysfunction, and apoptotic cell loss [3]. The mechanisms underlying these events seem to be linked to an increased production of reactive oxygen species (ROS) and oxidative damage [3]. Oxidative stress results from an imbalance between the generation of ROS and reactive nitrogen species and their removal by the cellular antioxidant system [4] and has been implicated in many neurodegenerative disorders [5,6].Several studies in breast cancer survivors and other patients undergoing DOX-based chemotherapy have reported persistent changes in cognitive functions, including memory loss and difficulty in the performance of daily life tasks [4]. Despite the well-known side effects of DOX treatment in the heart, little is known about its effects in the brain. Park et al. [7] showed that DOX generates free radicals in cultured astrocytes and decreases cell viability in a concentration-dependent manner. Similarly, in a study made in primary neuronal cultures, Lopes et al. [2] observed that DOX can induce neuronal cell death by necrosis and apoptosis in a concentration-dependent manner.Mitochondria play a central role in both cell life and cell death [8]. These organelles are essential for the production of ATP through oxidative phosphorylation and regulation of intracellular Ca 2+ homeostasis and are t...

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“…Mitochondrial Ca 2+ fluxes were measured by monitoring the changes in Ca 2+ concentration in the reaction medium using the hexapotassium salt of the fluorescence probe Calcium Green 5-N (Rajdev and Reynolds, 1993). Mitochondria (0.5 mg) were resuspended in 2 ml of reaction medium supplemented with 100 nM of Calcium Green 5-N followed by addition of Ca 2+ (70 nmol/mg protein) and energization with succinate (5 mM).…”

Section: Mitochondrial Respirationmentioning

confidence: 99%

Estradiol affects liver mitochondrial function in ovariectomized and tamoxifen-treated ovariectomized female rats

Moreira

Custódio

Nunes

et al. 2007

Toxicology and Applied Pharmacology

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Given the tremendous importance of mitochondria to basic cellular functions as well as the critical role of mitochondrial impairment in a vast number of disorders, a compelling question is whether 17β-estradiol (E2) modulates mitochondrial function. To answer this question we exposed isolated liver mitochondria to E2. Three groups of rat females were used: control, ovariectomized and ovariectomized treated with tamoxifen. Tamoxifen has antiestrogenic effects in the breast tissue and is the standard endocrine treatment for women with breast cancer. However, under certain circumstances and in certain tissues, tamoxifen can also exert estrogenic agonist properties. We observed that at basal conditions, ovariectomy and tamoxifen treatment do not induce any statistical alteration in oxidative phosphorylation system and respiratory chain parameters. Furthermore, tamoxifen treatment increases the capacity of mitochondria to accumulate Ca 2+ delaying the opening of the permeability transition pore. The presence of 25 μM E2 impairs respiration and oxidative phosphorylation system these effects being similar in all groups of animals studied. Curiously, E2 protects against lipid peroxidation and increases the production of H 2 O 2 in energized mitochondria of control females. Our results indicate that E2 has in general deleterious effects that lead to mitochondrial impairment. Since mitochondrial dysfunction is a triggering event of cell degeneration and death, the use of exogenous E2 must be carefully considered.

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Calcium green-5N, a novel fluorescent probe for monitoring high intracellular free Ca2+ concentrations associated with glutamate excitotoxicity in cultured rat brain neurons

Cited by 104 publications

References 18 publications

Extreme cellular adaptations and cell differentiation required by a cyanobacterium for carbonate excavation

Extreme cellular adaptations and cell differentiation required by a cyanobacterium for carbonate excavation

Doxorubicin increases the susceptibility of brain mitochondria to Ca2+-induced permeability transition and oxidative damage

Estradiol affects liver mitochondrial function in ovariectomized and tamoxifen-treated ovariectomized female rats

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