High oxygen tension leads to acute cell death in organotypic hippocampal slice cultures

Pomper, Jörn K.; Graulich, Johannes; Kovacs, Richard J.; Hoffmann, Ulrich; Gabriel, Siegrun; Heinemann, Uwe

doi:10.1016/s0165-3806(00)00132-2

Cited by 29 publications

(16 citation statements)

References 23 publications

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“…Sixty-percent O 2 caused a modest but significant increase in cell death (P Ͻ 0.05), and no time-dependent increase in cell death was observed using 40 and 20% O 2 . This pattern of hyperoxiainduced cell death is similar to that which others have reported using the organotypic hippocampal slice preparation (Graulich et al 2002;Pomper et al 2001) and suggests that hyperoxia (60 and 95% O 2 ) and increased ⅐O 2 Ϫ may increase oxidative stress-induced cell death.…”

Section: Oxygen Dose and Cell Death During Two-sided Superfusionsupporting

confidence: 88%

“…To the contrary, the highest level of EH-1 staining and thus cell death was observed after 4-h incubation in 95% O 2 . These data suggest that CA1 neurons in hippocampal slices are vulnerable to oxidative stress from prolonged exposure (Յ4 h) to hyperoxic control conditions (Fessel et al 2002;Rice 1999) This finding supports the earlier findings of Pomper et al (2001), who showed that CA1 cell death in organotypic slice cultures, prepared from younger rats, was greatest in 95% O 2 and less in 20% O 2 . Likewise, cell death was greatest in hippocampal and cerebral cortical neuronal cultures incubated in 95 and 21% O 2 and less in 9% O 2 (Brewer and Cotman 1989;Kaplan et al 1986).…”

Section: Cell Death and Cellular Viability As A Function Of O 2 Tensionsupporting

confidence: 86%

“…Similar observations have been made previously in our lab when comparing CA1 neuronal excitability of hippocampal slices maintained in 95, 60, and 40% O 2 . It is our opinion and others (Bingmann and Kolde 1979;Bingmann et al 1982;Fessel et al 2002;Pomper et al 2001;Shibata and Blatteis 1991) that neuronal activity that is traditionally characterized as "normal" in 95% O 2 has in fact been altered by an underlying level of oxidative stress due to hyperoxia. We propose that the brain slice preparation has increased excitability from the general stimulatory effects of hyperoxia Garcia et al 2005; this study, Fig.…”

Section: Cell Death and Cellular Viability As A Function Of O 2 Tensionmentioning

confidence: 75%

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Superoxide (·O₂⁻) Production in CA1 Neurons of Rat Hippocampal Slices Exposed to Graded Levels of Oxygen

D’Agostino¹,

Putnam²,

Dean³

2007

Journal of Neurophysiology

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Neuronal signaling, plasticity, and pathologies in CA1 hippocampal neurons are all intimately related to the redox environment and, thus tissue oxygenation. This study tests the hypothesis that hyperoxic superfusate (95% O(2)) causes a time-dependent increase in superoxide anion (*O(2)(-)) production in CA1 neurons in slices, which will decrease as oxygen concentration is decreased. Hippocampal slices (400 microm) from weaned rats were incubated with the fluorescent probe dihydroethidium (DHE), which detects intracellular *O(2)(-) production. Slices were loaded for 30 min using 10 microM DHE and maintained using one-sided superfusion or continuously loaded using 2.5 microM DHE and maintained using two-sided superfusion (36 degrees C). Continuous loading of DHE and two-sided superfusion gave the highest temporal resolution measurements of *O(2)(-) production, which was estimated by the increase in fluorescence intensity units (FIUs) per minute (FIU/min +/- SE) over 4 h. Superoxide production (2.5 microM DHE, 2-sided superfusion) was greatest in 95% O(2) (6.6 +/- 0.4 FIU/min) and decreased significantly during co-exposure with antioxidants (100 microM melatonin, 25 microM MnTMPyP) and lower levels of O(2) (60, 40, and 20% O(2) at 5.3 +/- 0.3, 3.3 +/- 0.1, and 1.6 +/- 0.2 FIU/min, respectively). CA1 cell death after 4 h (ethidium homodimer-1 staining) was greatest in 95% O(2) and lowest in 40 and 20% O(2). CA1 neurons generated evoked action potentials in 20% O(2) for >4 h, indicating viability at lower levels of oxygenation. We conclude that .O(2)(-) production and cell death in CA1 neurons increases in response to increasing oxygen concentration product (= PO(2) x time). Additionally, lower levels of oxygen (20-40%) and antioxidants should be considered to minimize superoxide-induced oxidative stress in brain slices.

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Section: Oxygen Dose and Cell Death During Two-sided Superfusionsupporting

confidence: 88%

Section: Cell Death and Cellular Viability As A Function Of O 2 Tensionsupporting

confidence: 86%

Section: Cell Death and Cellular Viability As A Function Of O 2 Tensionmentioning

confidence: 75%

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Superoxide (·O₂⁻) Production in CA1 Neurons of Rat Hippocampal Slices Exposed to Graded Levels of Oxygen

D’Agostino¹,

Putnam²,

Dean³

2007

Journal of Neurophysiology

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“…The atmosphere inside the plate was kept at 60% O 2 , 35% N 2 , and 5% CO 2 by directing a flow of gas into a water filled container inside the plate and performing the incubation with the lid on. All of the solutions were equilibrated with a gas mixture of 60% O 2 , 35% N 2 , and 5% CO 2 (19). The efflux experiments were carried out by incubating the slices with ACSF (400 l) on top of the membrane for 5 min.…”

Section: Methodsmentioning

confidence: 99%

Stimulated Efflux of Amino Acids and Glutathione from Cultured Hippocampal Slices by Omission of Extracellular Calcium

Stridh

Tranberg

Weber

et al. 2008

Journal of Biological Chemistry

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Omission of extracellular Ca2؉ for 15 min from the incubation medium of cultured hippocampal slices stimulated the efflux of glutathione, phosphoethanolamine, hypotaurine, and taurine. The efflux was reduced by several blockers of gap junctions, i.e. carbenoxolone, flufenamic acid, and endothelin-1, and by the connexin43 hemichannel blocking peptide Gap26 but was unchanged by the P2X 7 receptor inhibitor oxidized ATP, a pannexin1 hemichannel blocking peptide and an inactive analogue of carbenoxolone. Pretreatment of the slices with the neurotoxin N-methyl-D-aspartate left the efflux by Ca 2؉ omission unchanged, indicating that the stimulated efflux primarily originated from glia. Elevated glutamate efflux was detected when Ca 2؉ omission was combined with the glutamate uptake blocker L-trans-pyrrolidine-2,4-dicarboxylate and when both Ca 2؉ and Mg 2؉ were omitted from the medium. Omission of Ca 2؉ for 15 min alone did not induce delayed toxicity, but in combination with blocked glutamate uptake, significant cell death was observed 24 h later. Our results indicate that omission of extracellular Ca 2؉ stimulates efflux of glutathione and specific amino acids including glutamate via opening of glial hemichannels.

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“…A critical step is rapid removal of the brain from the skull, taking care to not penetrate the brain during removal and then rapid immersion in ice cold solution. Ideally the time between decapitation and removal of the brain from the skull should be no more than 30 -60 sec to preserve slice health 21 .…”

Section: Discussionmentioning

confidence: 99%

Generation of Local CA1 γ Oscillations by Tetanic Stimulation

Hatch

Reid

Petrou

2015

JoVE

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Neuronal network oscillations are important features of brain activity in health and disease and can be modulated by a range of clinically used drugs. A protocol is provided to generate a model for studying CA1 γ oscillations (20 -80 Hz). These γ oscillations are stable for at least 30 min and depend upon excitatory and inhibitory synaptic activity in addition to activation of pacemaker currents. Tetanically stimulated oscillations have a number of reproducible and easily quantifiable characteristics including spike count, oscillation duration, latency and frequency that report upon the network state. The advantages of the electrically stimulated oscillations include stability, reproducibility and episodic acquisition enabling robust characterization of network function. This model of CA1 γ oscillations can be used to study cellular mechanisms and to systematically investigate how neuronal network activity is altered in disease and by drugs. Disease state pharmacology can be readily incorporated by the use of brain slices from genetically modified or interventional animal models to enable selection of drugs that specifically target disease mechanisms. Video LinkThe video component of this article can be found at

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High oxygen tension leads to acute cell death in organotypic hippocampal slice cultures

Cited by 29 publications

References 23 publications

Superoxide (·O₂⁻) Production in CA1 Neurons of Rat Hippocampal Slices Exposed to Graded Levels of Oxygen

Superoxide (·O₂⁻) Production in CA1 Neurons of Rat Hippocampal Slices Exposed to Graded Levels of Oxygen

Stimulated Efflux of Amino Acids and Glutathione from Cultured Hippocampal Slices by Omission of Extracellular Calcium

Generation of Local CA1 γ Oscillations by Tetanic Stimulation

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High oxygen tension leads to acute cell death in organotypic hippocampal slice cultures

Cited by 29 publications

References 23 publications

Superoxide (·O2−) Production in CA1 Neurons of Rat Hippocampal Slices Exposed to Graded Levels of Oxygen

Superoxide (·O2−) Production in CA1 Neurons of Rat Hippocampal Slices Exposed to Graded Levels of Oxygen

Stimulated Efflux of Amino Acids and Glutathione from Cultured Hippocampal Slices by Omission of Extracellular Calcium

Generation of Local CA1 &#947; Oscillations by Tetanic Stimulation

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Superoxide (·O₂⁻) Production in CA1 Neurons of Rat Hippocampal Slices Exposed to Graded Levels of Oxygen

Superoxide (·O₂⁻) Production in CA1 Neurons of Rat Hippocampal Slices Exposed to Graded Levels of Oxygen

Generation of Local CA1 γ Oscillations by Tetanic Stimulation