Effects of cycloheximide on delayed neuronal death in rat hippocampus

doi:10.1016/0006-8993(90)90144-Z

Brain Research

Volume 534, Issues 1–2, 26 November 1990, Pages 299-302

https://doi.org/10.1016/0006-8993(90)90144-Z Get rights and content

Abstract

The effect of cycloheximide, a protein synthesis inhibitor, on hippocampal selective neuronal death was morphologically studied in rats subjected to 10 min forebrain ischemia using a 4-vessel occlusion model. Neuronal damage in the hippocampal CA1 subfield 72 h after ischemic insult was dramatically decreased by the lasting inhibition of protein synthesis through consecutive administration of cycloheximide. Cycloheximide, which was administered once within the first 24 h of recirculation, showed protective action on ischemic cell necrosis and its most potent effect was observed when injected at 12 h of post-ischemia. After 36 h of recirculation, however, treatment with cycloheximide could no longer prevent cell death. The possibility is considered that hippocampal delayed neuronal death following transient ischemia is caused by abnormal protein(s).

Reference (18)

JørgensenM.B. et al.
Delayed c-fos proto-oncogene expression in the rat hippocampus induced by transient global cerebral ischemia: an in situ hybridization study
Brain Research
(1989)
KirinoT.
Delayed neuronal death in the gerbil hippocampus following ischemia
Brain Research
(1982)
SzekelyA.M. et al.
Activation of specific glutamate receptor subtypes increases c-fos protooncogene expression in primary cultures of neonatal rat cerebellar granule cells
Neuropharmacology
(1987)
AraiH. et al.
Energy metabolism in delayed neuronal death of CA1 neurons of the hippocampus following transient ischemia in the gerbil
Metab. Brain Dis.
(1986)
BrierleyJ.B.
Cerebral hypoxia
BustoR. et al.
The importance of brain temperature in cerebral ischemic injury
Stroke
(1989)
Ch'ihJ.J. et al.
Regulation of mammalian protein synthesis in vivo
Biochem. J.
(1977)
FahrbachS.E. et al.
Mechanisms for programmed cell death in the nervous system of a moth
JacewiczM. et al.
Selective gene expression in focal cerebral ischemia
J. Cereb. Blood Flow Metab.
(1986)

There are more references available in the full text version of this article.

Cited by (200)

Stress granule induction after brain ischemia is independent of eukaryotic translation initiation factor (eIF) 2α phosphorylation and is correlated with a decrease in eIF4B and eIF4E proteins
2016, Journal of Biological Chemistry
Citation Excerpt :
In conclusion, IR stress during R3d decreased eIF4B and eIF4E levels in the hippocampal CA1 region concurrently with the induction of SGs. Cycloheximide has been previously described by our group and others as a neuroprotective agent in ischemia models (20, 30, 31). Moreover, cycloheximide (CHX) is known as an SG assembly inhibitor in cell lines (5).
Stress granules (SGs) are cytoplasmic ribonucleoprotein aggregates that are directly connected with the translation initiation arrest response to cellular stresses. Translation inhibition (TI) is observed in transient brain ischemia, a condition that induces persistent TI even after reperfusion, i.e. when blood flow is restored, and causes delayed neuronal death (DND) in selective vulnerable regions. We previously described a connection between TI and DND in the hippocampal cornu ammonis 1 (CA1) in an animal model of transient brain ischemia. To link the formation of SGs to TI and DND after brain ischemia, we investigated SG induction in brain regions with differential vulnerabilities to ischemia-reperfusion (IR) in this animal model. SG formation is triggered by both eukaryotic translation initiation factor (eIF) 2α phosphorylation and eIF4F complex dysfunction. We analyzed SGs by immunofluorescence colocalization of granule-associated protein T-cell internal antigen-1 with eIF3b, eIF4E, and ribosomal protein S6 and studied eIF2 and eIF4F complex. The results showed that IR stress induced SG formation in the CA1 region after 3-day reperfusion, consistent with TI and DND in CA1. SGs were formed independently of eIF2α phosphorylation, and their appearance was correlated with a decrease in the levels of eIF4F compounds, the cap-binding protein eIF4E, and eIF4B, suggesting that remodeling of the eIF4F complex was required for SG formation. Finally, pharmacological protection of CA1 ischemic neurons with cycloheximide decreased the formation of SGs and restored eIF4E and eIF4B levels in CA1. These findings link changes in eIF4B and eIF4E to SG induction in regions vulnerable to death after IR.
Electrophysiology of cerebral ischemia
2008, Neuropharmacology
Citation Excerpt :
Apparent recovery after a transient ischemic episode is often followed by cell death 24 or more hours later (Kirino, 1982; Petito et al., 1987; Urban et al., 1989; Hori and Carpenter, 1994). Various processes are involved, notably recurrent SD (Strong and Dardis, 2005) and continued activation of NMDA receptors (Rothman et al., 1987; Foster et al., 1988; Hori and Carpenter, 1994), persistent Ca2+ influx (Chinopoulos et al., 2004; MacDonald et al., 2006) – including influx through some glutamate, non-NMDA receptors (Tsubokawa et al., 1995) – formation of free radicals (McCord, 1985; Cao et al., 1988) and failure of normal protein synthesis (Goto et al., 1990; DeGracia and Hu, 2007). Ischemic damage of white matter is also caused by an excessive rise in cytoplasmic Ca2+.
Organized brain activity requires the coordinated firing of vast numbers of nerve cells. To maintain this, all these cells must be adequately polarized, their axons capable of conducting action potentials and releasing transmitters at an even greater numbers of synapses. Hence the often dire consequences of any interruption in the normal supply of O₂ and glucose. Initially, though both cognitive and synaptic functions are soon suppressed, membrane potentials in the brain change little – indeed, many neurons are hyperpolarized – and all these effects are fully reversible when glucose and/or O₂ supplies are restored. The early events, suppression of synaptic and cognitive function, sharply reduce the brain's needs of energy, enabling it to maintain the minimal metabolism required for survival. Even this minimum cannot be sustained for more than a few minutes: if ischemia is prolonged, a slowly progressive depolarization (mainly caused by glutamate release) suddenly accelerates owing to the activation of several inward currents. The resulting near-total depolarization and large increase in Ca²⁺ influx – as well as Ca²⁺ release from internal stores (including mitochondria) – leads to a rapid rise in cytoplasmic [Ca²⁺]. As long as this does not reach the critical level that triggers the irreversible processes leading to cell death, restoring energy supplies reactivates the membrane pumps that re-establish normal ionic gradients and membrane potentials, and thus make possible the return of synaptic and cognitive functions, Rapid advances in knowledge suggest a wide spectrum of agents potentially capable of delaying or even preventing irreversible outcomes of brain ischemia.
Expression and localization of Fas-associated proteins following focal cerebral ischemia in rats
2008, Brain Research
Citation Excerpt :
The tight correlation of TUNEL-positive cells with the level of the gene expression suggests the importance of these Fas-associated proteins in ischemic neuronal death. Our finding of FADD and caspase-8 induction in ischemic neurons suggests that, like the Fas ligand and Fas (Martin-Villalba et al., 1999; Matsuyama et al., 1994; Rosenbaum et al., 2000), the Fas-associated proteins may also participate in the process of delayed cell death in peri-infarct area after focal ischemia and supports the hypothesis that delayed neuronal cell death requires de novo protein synthesis (Goto et al., 1990). Oxidative stress induced by H2O2 can trigger rapid induction of FADD and caspase-8 in cultured cardiomyocytes (Eskes et al., 2000; Yaniv et al., 2005), which can be blocked by the FLICE-inhibitory protein (FLIP) (Scaffidi et al., 1999b).
The aim of this study was to investigate the changes of expression of Fas-associated proteins and its cellular localization in the peri-infarct region following transient focal cerebral ischemia. Adult male Sprague–Dawley rats underwent right middle cerebral artery occlusion (MCAo) for 2 h and reperfusion for 1, 3, 6, 12 and 24 h. The expression of Fas-associated death domain protein (FADD), Fas-associated phosphatase-1 (FAP-1) caspase-8 and death-associated protein (Daxx), the pro-apoptotic genes, were examined by methods of RT-PCR, immunohistochemistry and Western blot. The results showed that the expression levels of mRNA and protein for FADD and caspase-8 increased significantly at 1–3 h after reperfusion, peaked at 12 h, then declined markedly at 24 h. The time course change of FAP-1 was consistent with that of FADD. The expression level of mRNA and protein for death-associated protein (Daxx) increased significantly at 3 h after reperfusion and persisted for 24 h at a high level. Immunofluorescence double-staining laser scanning showed that the immunoreactivity of FADD was localized in cytoplasm, and Daxx immunoreactivity was translocated from nucleus to cytoplasm at 3 h after reperfusion. The TUNEL-positive cells could be found in peri-infarct region at 3 h and increased with time after reperfusion. Our findings suggest a possible association between expression of FADD, caspase-8, Daxx and FAP-1 genes and apoptosis following ischemia.
Catalpol modulates the expressions of Bcl-2 and Bax and attenuates apoptosis in gerbils after ischemic injury
2006, Brain Research
Our previous study described the neuroprotective effects of catalpol in gerbil ischemic model, in which catalpol was shown to prevent hippocampal neurons from death and ameliorate the cognitive ability of the animals. In the study, we focused on investigating the neuroprotective mechanism of catalpol. Animals were randomly assigned three groups as sham-operated, ischemia-treated with saline and ischemia-treated with catalpol. Transient global ischemia was produced by a 5 min occlusion of the bilateral common carotid arteries. Catalpol was intraperitoneally injected at the dose of 5 mg/kg immediately after reperfusion and repeatedly at 12, 24, 48 and 72 h. Histology as well as immunohistochemistry and TUNEL (the terminal deoxynucleotidyl transferase-mediated UTP nick end label) analysis were performed on serial slices through the dorsal hippocampus after gerbils were sacrificed. The results showed that 5 min transient global ischemia followed by 4 days reperfusion caused significant increases in TUNEL-positive and Bax-positive cells in hippocampal CA1 subfield. Catalpol not only significantly reduced TUNEL-positive and Bax-positive cells but also significantly increased Bcl-2-positive cells. All these suggested that catalpol could effectively inhibit apoptosis by modulating the expressions of Bcl-2 and Bax genes.
The specific FKBP38 inhibitor N-(N′,N′- dimethylcarboxamidomethyl)cycloheximide has potent neuroprotective and neurotrophic properties in brain ischemia
2006, Journal of Biological Chemistry
FK506 and FK506-derived inhibitors of the FK506-binding protein (FKBP)-type peptidylprolyl cis/trans-isomerases (PPIase) display potent neuroprotective and neuroregenerative properties in various neurodegeneration models, showing the importance of neuroimmunophilins as targets for the treatment of acute and chronic neurodegenerative diseases. However, the PPIase activity targeted by active site-directed ligands remainsed unknown so far. Here we show that neurotrophic FKBP ligands, such as GPI1046 and N-[methyl(ethoxycarbonyl)]cycloheximide, inhibit the calmodulin/Ca²⁺ (CaM/Ca²⁺)-regulated FKBP38 with up to 80-fold higher affinity than FKBP12. In contrast, the non-neurotrophic rapamycin inhibits FKBP38·CaM/Ca²⁺ 500-fold less affine than other neuroimmunophillins. In the context of the high expression of FKBP38 in neuroblastoma cells, these data suggest that FKBP38·CaM/Ca²⁺ inhibition can mediate neurotrophic properties of FKBP ligands. The FKBP38-specific cycloheximide derivative, N-(N′,N′-dimethylcarboxamidomethyl)cycloheximide (DM-CHX) was synthesized and used in a rat model of transient focal cerebral ischemia. Accordingly, DM-CHX caused neuronal protection as well as neural stem cell proliferation and neuronal differentiation at a dosage of 27.2 μg/kg. These effects were still dominant, if DM-CHX was applied 2-6 h post-insult. In parallel, sustained motor behavior deficits of diseased animals were improved by drug administration, revealing a potential therapeutic relevance. Thus, our results demonstrate that FKBP38 inhibition by DM-CHX regulates neuronal cell death and proliferation, providing a promising strategy for the treatment of acute and/or chronic neurodegenerative diseases.
Effects of β-mercaptoethanol and cycloheximide on survival and DNA damage of bovine embryos stored at 4°C for 72 h
2006, Theriogenology
The objectives of this study were to determine the effects of cycloheximide (CHX) and β-mercaptoethanol (β-ME) during storage of in vitro-produced (IVP) bovine blastocysts for 72 h at 4 °C on their survival, hatching capacity and DNA damage. In Experiment 1, when blastocysts were stored in a medium supplemented with 25, 50 or 100 μg/mL of CHX, or 25, 50 or 100 μM of β-ME, the blastocysts stored with 25 μg/mL of CHX had a significantly higher survival rate than that of the blastocysts stored without CHX (79.5% versus 54.2%). In contrast, β-ME had no apparent effects on the survival and hatching capacity of stored embryos. In Experiment 2, to investigate synergistic effects of CHX and β-ME during storage of blastocysts on their developmental parameters and DNA damage, they were stored in the medium with CHX (25 μg/mL) and β-ME (50 μM). The combination of CHX and β-ME had no significant effects on the survival of blastocysts. The proportion (6.8%) of DNA-fragmented cells in the blastocysts stored with CHX was similar to that (5.4%) in the non-stored blastocysts (positive control) and significantly lower than that (9.7%) in the blastocysts stored without CHX and β-ME (negative control). However, there were no significant differences among the proportions of dead cells of blastocysts in the storage groups. Therefore, the supplementation of CHX in the storage medium had a beneficial effect on the proportions of survival and DNA-fragmented cells in the stored embryos, whereas the β-ME alone or in combination with CHX had no positive effects on either of these proportions.

View all citing articles on Scopus

: This work was supported by the Science and Technology Agency.

View full text

Article preview

Brain Research

Abstract

Reference (18)

Delayed c-fos proto-oncogene expression in the rat hippocampus induced by transient global cerebral ischemia: an in situ hybridization study

Brain Research

Delayed neuronal death in the gerbil hippocampus following ischemia

Brain Research

Activation of specific glutamate receptor subtypes increases c-fos protooncogene expression in primary cultures of neonatal rat cerebellar granule cells

Neuropharmacology

Energy metabolism in delayed neuronal death of CA1 neurons of the hippocampus following transient ischemia in the gerbil

Metab. Brain Dis.

Cerebral hypoxia

The importance of brain temperature in cerebral ischemic injury

Stroke

Regulation of mammalian protein synthesis in vivo

Biochem. J.

Mechanisms for programmed cell death in the nervous system of a moth

Selective gene expression in focal cerebral ischemia

J. Cereb. Blood Flow Metab.

Cited by (200)

Stress granule induction after brain ischemia is independent of eukaryotic translation initiation factor (eIF) 2α phosphorylation and is correlated with a decrease in eIF4B and eIF4E proteins

Electrophysiology of cerebral ischemia

Expression and localization of Fas-associated proteins following focal cerebral ischemia in rats

Catalpol modulates the expressions of Bcl-2 and Bax and attenuates apoptosis in gerbils after ischemic injury

The specific FKBP38 inhibitor N-(N′,N′- dimethylcarboxamidomethyl)cycloheximide has potent neuroprotective and neurotrophic properties in brain ischemia

Effects of β-mercaptoethanol and cycloheximide on survival and DNA damage of bovine embryos stored at 4°C for 72 h