Secondary Energy Failure After Cerebral Hypoxia???Ischemia in the Immature Rat (original) (raw)
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Brain, 2008
Following hypoxia-ischaemia (HI), an early biomarker of insult severity is desirable to target neuroprotective therapies to patients most likely to benefit; currently there are no biomarkers within the 'latent phase' period before the establishment of secondary energy failure. Brief transient phosphocreatine (PCr) recovery overshoot (measured absolutely or relative to nucleotide triphosphate, NTP) following HI has been observed in cardiac and skeletal muscle; its significance however is unclear. To investigate cerebral PCr recovery levels after HI in relation to (i) baseline metabolism, (ii) insult severity, (iii) energy metabolism at recovery and (iv) subsequent metabolic derangement, cerebral NTP, PCr and inorganic phosphate (relative to the exchangeable highenergy phosphate pool) were measured serially in an in vivo model of perinatal asphyxial encephalopathy using phosphorus-31 magnetic resonance spectroscopy. Measures were compared either in all piglets or between 3 subgroups with no (n = 5, favourable outcome), moderate (n = 8, intermediate outcome) or severe (n = 5, unfavourable outcome) secondary energy failure at 24 h after HI. Maximum NTP, PCr and inorganic phosphate recoveries were observed 2^8 h after HI. Following resuscitation, in subjects with favourable outcome PCr recovered to higher than its baseline level (overshoot); in subjects with unfavourable outcome maximum PCr recovery was lower than baseline and lower than in subjects with favourable and intermediate outcomes.
Journal of Neurochemistry, 2008
By combining immunohistochemical technique with microassay methods, we analyzed regional energy metabolism in vulnerable and tolerant areas of gerbil brains during evolution of neuronal damage after bilateral common carotid artery occlusion for 10 min with subsequent reperfusion. Four animals were used for each reperfusion period. Based on the information from the immunohistochemical examination, we dissected out vulnerable and tolerant subregions of the hippocampus, cerebral cortex, and thalamus from freeze-dried 20-mthick sections, and measured the levels of creatine phosphate (P-Cr), adenine nucleotides, guanine nucleotides, and purine bodies by HPLC, and the levels of glucose, glycogen, and lactate by an enzyme-immobilized column method. There were no significant differences in the levels of metabolites between vulnerable and tolerant subregions of control brains. After reperfusion, both vulnerable and tolerant subregions recovered preischemic metabolic profiles by 2 days. Although the regional differences between vulnerable and tolerant subregions were minimal at each reperfusion period, there were delays in the recovery of P-Cr, ATP, and/or total adenine nucleotides in all vulnerable subregions. A decline of P-Cr, ATP, and GTP levels without change in %ATP, AMP, or purine bodies occurred after reperfusion for 3 days, coinciding with the development of immunohistochemical damage by the immunoreaction for microtubule-associated protein 1A. The results supported the notion that subtle but sustained impairment of energy metabolism caused by mitochondrial dysfunction in the early reperfusion period might trigger delayed neuronal death in vulnerable subregions. Key Words: Immunohistochemistry-Energy metabolism-Selective vulnerability-Microtubule-associated protein-Cerebral ischemia.
Molecular Brain Research, 1994
A unilateral hypoxia-ischaemia (HI) 21-day-old rat preparation was used to assess the effects of HI on the expression of the immediate-early gene proteins (IEGPs) c-Fos/FRAs, Fos B, c-Jun, Jun B, Jun D, Krox 20, Krox 24, and on the mRNA for the neurotrophic factor, brain-derived neurotrophic factor (BDNF). Moderate HI (15 min hypoxia) produced delayed, selective neuronal death and was associated with a rapid induction of c-Fos, Fos B, Jun B, Jun D, and c-Jun proteins, but not Krox 20 protein or BDNF mRNA, in neurons on the side of HI and also a delayed expression of c-Jun (and to a lesser extent c-Fos/FRA's and Fos B) 24–48 h after HI in neurons that underwent delayed neuronal death. Krox 24 showed an initial induction followed by a long-lasting suppression of its expression in regions undergoing cell loss. Severe HI (60 min hypoxia) resulted in seizures and rapid neuronal loss and infarction (necrotic cell death) on the side of HI, and was associated with early induction of c-Fos, Fos B, c-Jun, Jun B, Jun D, Krox 20 and Krox 24 protein and BDNF mRNA in neurons on the non-ligated side of the brain. Fos, c-Jun, Jun B, Jun D and Krox 24, but not Krox 20, Fos B, or BDNF mRNA, were also induced in non-nerve cells on the damaged side of the brain after both moderate and severe HI, and many of these cells appeared to be dividing. Thus, moderate HI induces IEGP's in neurons and non-nerve cells in damaged regions, whereas severe HI induces IEGP's and BDNF in non-damaged regions. c-Jun (and to a lesser extent c-Fos/FRA's) showed a prolonged expression in neurons undergoing delayed, but not necrotic, cell death suggesting that they may be involved in the biochemical cascade that causes selective delayed neuronal death. BDNF was not induced by HI, and therefore, does not appear to play an endogenous neuroprotective role in the CNS.
Metabolic stages, mitochondria and calcium in hypoxic/ischemic brain damage
Cell Calcium, 2004
Cerebral hypoxia/ischemia leads to mitochondrial dysfunction due to lack of oxygen leaving the glycolytic metabolism as a main pathway for ATP production. Inhibition of mitochondrial respiration thus triggers generation of lactate and hydrogen ions (H + ), and furthermore dramatically reduces ATP generation leading to disregulation of cellular ion metabolism with subsequent intracellular calcium accumulation. Upon reperfusion, when mitochondrial dysfunction is (at least partially) reversed by restoring cerebral oxygen supply, bioenergetic metabolism recovers and brain cells are able to re-institute their normal ionic homeostatic mechanisms. However, the initial restoration of normal mitochondrial function may be only transient and followed by a secondary, delayed perturbation of mitochondrial respiratory performance seen as a decrease in cellular ATP levels and known as "secondary energy failure". There have been several mechanisms considered responsible for delayed post-ischemic mitochondrial failure, the mitochondrial permeability transition (MPT) being one that is considered important. Although the amount of calcium available during early reperfusion in vivo is limited, relative to the amount needed to trigger the MPT in vitro; the additional intracellular conditions (of acidosis, high phosphate, and low adenine nucleotideae levels) prevailing during reperfusion, favor MPT pore opening in vivo. Furthermore, the cellular redistribution and/or changes in the intracellular levels of pro-apoptotic proteins can alter mitochondrial function and initiate apoptotic cell death. Thus, mitochondria seem play an important role in orchestrating cell death mechanisms following hypoxia/ischemia. However, it is still not clear which are the key mechanisms that cause mitochondrial dysfunction and lead ultimately to cell death, and which have more secondary nature to brain damage acting as aggravating factors.
Neuronal death and survival in two models of hypoxic-ischemic brain damage
Brain research …, 1999
Ž . Two unilateral hypoxic-ischemia HI models moderate and severe in immature rat brain have been used to investigate the role of various transcription factors and related proteins in delayed neuronal death and survival. The moderate HI model results in an apoptotic-like neuronal death in selectively vulnerable regions of the brain while the more severe HI injury consistently produces widespread necrosis resulting in infarction, with some necrosis resistant cell populations showing evidence of an apoptotic type death. In susceptible regions undergoing an apoptotic-like death there was not only a prolonged induction of the immediate early genes, c-jun, c-fos Ž . and nur77, but also of possible target genes amyloid precursor protein APP and CPP32. In contrast, increased levels of BDNF, 751 phosphorylated CREB and PGHS-2 were found in cells resistant to the moderate HI insult suggesting that these proteins either alone or in combination may be of importance in the process of neuroprotection. An additional feature of both the moderate and severe brain insults Ž . was the rapid activation andror proliferation of glial cells microglia and astrocytes in and around the site of damage. The glial response Ž . following HI was associated with an upregulation of both the CCAAT-enhancer binding protein a microglia only and NFk B transcription factors. q
Brain ischemia and reperfusion: molecular mechanisms of neuronal injury
Journal of the Neurological Sciences, 2000
Brain ischemia and reperfusion engage multiple independently-fatal terminal pathways involving loss of membrane integrity in partitioning ions, progressive proteolysis, and inability to check these processes because of loss of general translation competence and reduced survival signal-transduction. Ischemia results in rapid loss of high-energy phosphate compounds and generalized depolarization, which induces release of glutamate and, in selectively vulnerable neurons (SVNs), opening of both voltage-dependent and glutamate-21 regulated calcium channels. This allows a large increase in cytosolic Ca associated with activation of m-calpain, calcineurin, and phospholipases with consequent proteolysis of calpain substrates (including spectrin and eIF4G), activation of NOS and potentially of 21 Bad, and accumulation of free arachidonic acid, which can induce depletion of Ca from the ER lumen. A kinase that shuts off translation initiation by phosphorylating the a-subunit of eukaryotic initiation factor-2 (eIF2a) is activated either by adenosine degradation 21 products or depletion of ER lumenal Ca . Early during reperfusion, oxidative metabolism of arachidonate causes a burst of excess oxygen radicals, iron is released from storage proteins by superoxide-mediated reduction, and NO is generated. These events result in peroxynitrite generation, inappropriate protein nitrosylation, and lipid peroxidation, which ultrastructurally appears to principally damage the plasmalemma of SVNs. The initial recovery of ATP supports very rapid eIF2a phosphorylation that in SVNs is prolonged and associated with a major reduction in protein synthesis. High catecholamine levels induced by the ischemic episode itself and / or drug administration down-regulate insulin secretion and induce inhibition of growth-factor receptor tyrosine kinase activity, effects associated with down-regulation of survival signal-transduction through the Ras pathway. Caspase activation occurs during the early hours of reperfusion following mitochondrial release of caspase 9 and cytochrome c. The SVNs find themselves with substantial membrane damage, calpain-mediated proteolytic degradation of eIF4G and cytoskeletal proteins, altered translation initiation mechanisms that substantially reduce total protein synthesis and impose major alterations in message selection, down-regulated survival signaltransduction, and caspase activation. This picture argues powerfully that, for therapy of brain ischemia and reperfusion, the concept of single drug intervention (which has characterized the approaches of basic research, the pharmaceutical industry, and clinical trials) cannot be effective. Although rigorous study of multi-drug protocols is very demanding, effective therapy is likely to require (1) peptide growth factors for early activation of survival-signaling pathways and recovery of translation competence, (2) inhibition of lipid peroxidation, (3) inhibition of calpain, and (4) caspase inhibition. Examination of such protocols will require not only characterization of functional and histopathologic outcome, but also study of biochemical markers of the injury processes to establish the role of each drug.
Journal of Cerebral Blood Flow & Metabolism, 1994
The cerebral metabolic rate for glucose (CMRg1) and cerebral energy utilization (CEU) were assessed in immature rats during recovery from cerebral hypoxia–ischemia. CMRg1was determined using a modification of the Sokoloff technique with 2-deoxy-[14C]glucose (2-DG) as the radioactive tracer. CEU was determined using the Lowry decapitation technique. Seven-day postnatal rats underwent unilateral common carotid artery ligation, followed 4 h thereafter by exposure to 8% oxygen at 37°C for 3 h. At 1, 4, or 24 h of recovery, the rat pups underwent those procedures necessary for the measurement of either CMRg1or CEU. At 1 h of recovery, the CMRg1of the cerebral hemisphere ipsilateral to the carotid artery occlusion was 97% of the control rate (8.7 μmol 100 g−1min−1) but was only 48% of the control in the contralateral hemisphere. At 4 h of recovery, the CMRg1was increased 49% above baseline in the ipsilateral hemisphere, decreasing thereafter to 84% of the control at 24 h. The CMRg1of the ...
Bilateral Molecular Changes in a Neonatal Rat Model of Unilateral Hypoxic-Ischemic Brain Damage
Pediatric Research, 2006
Perinatal hypoxia ischemia (HI) is a frequent cause of neonatal brain injury. This study aimed at describing molecular changes during the first 48 h after exposure of the neonatal rat brain to HI. Twelve-day-old rats were subjected to unilateral carotid artery occlusion and 90 min of 8% O 2 , leading to neuronal damage in the ipsilateral hemisphere only. Phosphorylated-Akt levels were decreased from 0.5 to 6 h post-HI, whereas the level of phosphorylated extracellular signal-related kinases (ERK)1/2 increased during this time frame. Hypoxia-inducible factor (HIF)-1␣ protein increased with a peak at 3 h after HI. mRNA expression for IL- and tumor necrosis factor-␣ and - started to increase at 6 h with a peak at 24 h post-HI. Expression of heat shock protein 70 was increased from 12 h after HI onwards in the ipsilateral hemisphere only. Surprisingly, HI changed the expression of cytokines, HIF1-␣ ,and P-Akt to the same extent in both the ipsi-as well as the contralateral hemisphere, although neuronal damage was unilateral. Exposure of animals to hypoxia without carotid artery occlusion induced similar changes in cytokines, HIF-1␣, and P-Akt. We conclude that during HI, hypoxia is sufficient to regulate multiple molecular mediators that may contribute, but are not sufficient, to induce long-term neuronal damage.