Editorial Views  |   May 2003
Perinatal Brain Injury: The Role of Development in Vulnerability
Author Affiliations & Notes
  • Rona G. Giffard, M.D., Ph.D.
  • Gary Fiskum, Ph.D.
  • *Associate Professor of Anesthesia, Vice Chair for Research, Stanford University School of Medicine, Stanford, California, †Professor and Research Director of Anesthesiology, University of Maryland School of Medicine, Baltimore, Maryland.
Article Information
Editorial Views
Editorial Views   |   May 2003
Perinatal Brain Injury: The Role of Development in Vulnerability
Anesthesiology 5 2003, Vol.98, 1039-1041. doi:
Anesthesiology 5 2003, Vol.98, 1039-1041. doi:
THE article appearing in this issue of Anesthesiology by Ditsworth et al.  1 reports that cell death in the brains of piglets subjected to 90 min of deep hypothermic circulatory arrest (DHCA) is largely apoptotic, accompanied by activation of caspases 3 and 8, as well as early release of cytochrome c and the presence of Fas. This article raises several issues of interest to anesthesiologists. These include 1) the potential danger of DHCA to the developing brain and whether we can do anything to better protect the brains of these infants during surgery, 2) the role of development in determining the mechanisms of brain injury, and 3) the role of development in susceptibility to brain injury and sensitivity to neuroprotective interventions.
Although DHCA clearly provides significant protection to the brain and other organs from ischemia, the results of this article and prior reports 2–4 demonstrate that DHCA for a sufficient duration does result in brain injury. This finding is not surprising, because the extent of brain injury resulting from ischemia must in part depend on the duration of the ischemic insult, even in the presence of hypothermia. This work and that of others has shown that 60–90 min of DHCA is sufficient to cause brain cell death. Further investigation of the effects of deep hypothermia alone are needed, as well as investigation into the use of low-flow perfusion versus  intermittent perfusion as a way to protect the brain but still permit adequate surgical conditions.
The results reported by Ditsworth et al.  1 focus on the mechanism of brain cell death following DHCA. Their observations strengthen the argument that much of the cell death is apoptotic by demonstrating activation of caspases 3 and 8, as well as cytochrome c release from mitochondria, a step often necessary for activation of the caspase proteases that kill the cell. Making the distinction between different types of cell death is not a purely academic one, since different mechanisms of cell death may suggest different strategies for protection. Necrotic cell death involves swelling rather than condensation of the cell and internal organelles, random DNA fragmentation, early disruption of organelles without formation of apoptotic bodies, and early loss of plasma membrane integrity.
In contrast, apoptosis is a type of cell death with a distinct morphology consisting of nuclear condensation, early preservation of nuclear and cytoplasmic membranes, and relative preservation of cellular organelles. 5 Apoptotic cell death plays a key role in the normal development of the central nervous system. 6 As each region develops, the number of cells is reduced from the number initially generated so that the number of different types of neurons is appropriate, 7 and the number of astrocytes and oligodendrocytes is matched to the number of neurons and axons. This process results in developmentally determined vulnerable periods for specific cell populations. For example, cerebral white matter injury consisting of periventricular leukomalacia and hypomyelination are the anatomic correlates of cerebral palsy. These forms of brain injury are thought to be due to the specific vulnerability of premyelinating oligodendrocytes in the mid to late third trimester of human pregnancy when ischemia or infection/cytokine exposure may result in excessive loss of oligodendrocytes and a reduced number of mature myelinating oligodendrocytes. 8,9 This response to ischemia is not seen in older patients or animals, suggesting that the tendency of a cell to undergo apoptosis is developmentally regulated. This concept is further supported by the recent observation that expression of caspase 3 decreases during postnatal development, 10 but it increases in very old animals. 11 In addition, the pro-apoptotic Bax molecule and associated release of cytochrome c is increased in brain mitochondria from immature compared to mature animals, further indicating that early postnatal brain cells are primed to undergo apoptosis. 12 
Thus an important aspect of understanding brain injury due to cerebral ischemia requires understanding the role played by development. Although several investigators argue against a role for apoptosis in adult brain ischemia, there is much better agreement that apoptosis plays an important role in the response to ischemia in the perinatal period. Work from several laboratories studying normothermic ischemia clearly suggests that apoptotic cell death in the brain is developmentally regulated, with apoptosis being readily detected after models of perinatal hypoxia/ischemia, but less prominent in adult models of cerebral ischemia. 13–22 
Many genes involved in the cell death process have been identified. Many biochemical changes and specific signaling pathways have been shown to participate in this process. Apoptosis may result from imbalances in signaling pathways (such as lack of growth factors), may be initiated by activation of membrane receptors, and has several potential pathways for execution. These include (1) activation of proteases called caspases that carry out the cell death, 23–26 (2)participation of mitochondria in the release of proapoptotic proteins, 27,28 and (3) regulation by the bcl-2 family of proteins. 29 
Several steps in the apoptosis cascade have provided new ways to reduce ischemic cell death in models of cerebral ischemia. Caspase inhibitors and overexpression of antiapoptotic regulatory proteins (such as bcl-2) have been shown to be effective at reducing ischemic brain injury in animal models. 30–35 Despite this type of evidence, due to the heterogeneity of the morphologic picture in cerebral ischemia, there is still disagreement about the extent to which cell death during stroke involves apoptosis, necrosis, or a combination of both. 36–40 
Complicating our understanding of the mechanisms of cell death are recent findings that suggest that there are multiple methods of genetically controlled cell death and that the morphologic picture of apoptosis does not always correlate with activation of caspases nor does the appearance of a necrotic death rule out an active genetically determined type of cell death. Genetically determined types of cell death independent of caspase activation have been described, 41,42 which may still display the cellular morphology of apoptosis. Cell death in which activation of caspases is important but results in necrosis-like morphology has also been reported. 43 Recent data suggest that both caspase-dependent and caspase-independent forms of cell death are involved in cerebral ischemia. 44 
In addition to changes in the mechanisms of brain cell loss with development, the effect of the same insult changes with age. Vulnerability or the extent of injury observed in response to an ischemic insult increases as a function of age. Vulnerability to ischemia changes rapidly with age in the perinatal period as demonstrated in a study of combined focal ischemia–hypoxia in rat between the ages of postnatal days 1 and 7. 45 Brains of postnatal day 5 rats showed markedly less injury than did brains of postnatal day 7 animals. Similar changes in response to injury have also been seen in brain cell cultures. 46 Thus, although a given duration of ischemia may result in less severe injury in an infant, because this deficit will be present throughout life, it is still a matter of great concern.
Understanding differences in the mechanisms of brain injury provoked by ischemia in neonates compared to adults will lead to the development of age-specific protective strategies. At this time, a great deal remains to be learned about age-specific responses to cerebral ischemia, and the efficacy of potential protective strategies should be evaluated in both perinatal and adult models of cerebral ischemia.
This Editorial View accompanies the following article: Ditsworth D, Priestley MA, Loepke AW, Ramamoorthy C, McCann J, Staple L, Kurth D: Apoptotic neuronal death following deep hypothermic circulatory arrest in piglets. Anesthesiology 2003; 98:1119–27.
Ditsworth D, Priestley M, Loepke AW, Ramamoorthy C, McCann J, Staple L, Kurth D: Apoptotic neuronal death following deep hypothermic circulatory arrest in piglets. A nesthesiology 2003; 98: 1119–27Ditsworth, D Priestley, M Loepke, AW Ramamoorthy, C McCann, J Staple, L Kurth, D
Nomura F, Forbess JM, Jonas RA, Hiramatsu T, du Plessis AJ, Walter G, Stromski ME, Holtzman DH: Influence of age on cerebral recovery after deep hypothermic circulatory arrest in piglets. Ann Thorac Surg 1996; 62: 115–22Nomura, F Forbess, JM Jonas, RA Hiramatsu, T du Plessis, AJ Walter, G Stromski, ME Holtzman, DH
Kurth CD, Priestley M, Golden J, McCann J, Raghupathi R: Regional patterns of neuronal death after deep hypothermic circulatory arrest in newborn pigs. J Thorac Cardiovasc Surg 1999; 118: 1068–77Kurth, CD Priestley, M Golden, J McCann, J Raghupathi, R
Langley SM, Chai PJ, Miller SE, Mault JR, Jaggers JJ, Tsui SS, Lodge AJ, Lefurgey A, Ungerleider RM: Intermittent perfusion protects the brain during deep hypothermic circulatory arrest. Ann Thorac Surg 1999; 68: 4–12; discussion 12–3Langley, SM Chai, PJ Miller, SE Mault, JR Jaggers, JJ Tsui, SS Lodge, AJ Lefurgey, A Ungerleider, RM
Kerr JF, Wyllie AH, Currie AR: Apoptosis: a basic biological phenomenon with wide-ranging implications in tissue kinetics. Br J Cancer 1972; 26: 239–57Kerr, JF Wyllie, AH Currie, AR
Oppenheim RW: Cell death during development of the nervous system. Annu Rev Neurosci 1991; 14: 453–501Oppenheim, RW
Thomaidou D, Mione MC, Cavanagh JF, Parnavelas JG: Apoptosis and its relation to the cell cycle in the developing cerebral cortex. J Neurosci 1997; 17: 1075–85Thomaidou, D Mione, MC Cavanagh, JF Parnavelas, JG
inney HC, Back SA: Human oligodendroglial development: relationship to periventricular leukomalacia. Semin Pediatr Neurol 1998; 5: 180–9inney, HC Back, SA
Back SA, Luo NL, Borenstein NS, Levine JM, Volpe JJ, Kinney HC: Late oligodendrocyte progenitors coincide with the developmental window of vulnerability for human perinatal white matter injury. J Neurosci 2001; 21: 1302–12Back, SA Luo, NL Borenstein, NS Levine, JM Volpe, JJ Kinney, HC
Hu BR, Liu CL, Ouyang Y, Blomgren K, Siesjo BK: Involvement of caspase-3 in cell death after hypoxia-ischemia declines during brain maturation. J Cereb Blood Flow Metab 2000; 20: 1294–300Hu, BR Liu, CL Ouyang, Y Blomgren, K Siesjo, BK
Martin DS, Lonergan PE, Boland B, Fogarty MP, Brady M, Horrobin DF, Campbell VA, Lynch MA: Apoptotic changes in the aged brain are triggered by interleukin-1beta- induced activation of p38 and reversed by treatment with eicosapentaenoic acid. J Biol Chem 2002; 277: 34239–46Martin, DS Lonergan, PE Boland, B Fogarty, MP Brady, M Horrobin, DF Campbell, VA Lynch, MA
Polster BM, Robertson CL, Bucci CJ, Suzuki M, Fiskum G: Postnatal brain development and neural cell differentiation modulate mitochondrial Bax and BH3 peptide-induced cytochrome c release. Cell Death Diff 2003; IN PRESS
eilharz EJ, Williams CE, Dragunow M, Sirimanne ES, Gluckman PD: Mechanisms of delayed cell death following hypoxic-ischemic injury in the immature rat: evidence for apoptosis during selective neuronal loss. Brain Res Mol Brain Res 1995; 29: 1–14eilharz, EJ Williams, CE Dragunow, M Sirimanne, ES Gluckman, PD
van Lookeren Campagne M, Gill R: Ultrastructural morphological changes are not characteristic of apoptotic cell death following focal cerebral ischaemia in the rat. Neurosci Lett 1996; 213: 111–4van Lookeren Campagne, M Gill, R
Petito CK, Torres-Munoz J, Roberts B, Olarte JP, Nowak TS, Jr, Pulsinelli WA: DNA fragmentation follows delayed neuronal death in CA1 neurons exposed to transient global ischemia in the rat. J Cereb Blood Flow Metab 1997; 17: 967–76Petito, CK Torres-Munoz, J Roberts, B Olarte, JP Nowak, TS Pulsinelli, WA
Portera-Cailliau C, Price DL, Martin LJ: Excitotoxic neuronal death in the immature brain is an apoptosis- necrosis morphological continuum. J Comp Neurol 1997; 378: 70–87Portera-Cailliau, C Price, DL Martin, LJ
Sidhu RS, Tuor UI, Del Bigio MR: Nuclear condensation and fragmentation following cerebral hypoxia- ischemia occurs more frequently in immature than older rats. Neurosci Lett 1997; 223: 129–32Sidhu, RS Tuor, UI Del Bigio, MR
Colbourne F, Sutherland GR, Auer RN: Electron microscopic evidence against apoptosis as the mechanism of neuronal death in global ischemia. J Neurosci 1999; 19: 4200–10Colbourne, F Sutherland, GR Auer, RN
Ishimaru MJ, Ikonomidou C, Tenkova TI, Der TC, Dikranian K, Sesma MA, Olney JW: Distinguishing excitotoxic from apoptotic neurodegeneration in the developing rat brain. J Comp Neurol 1999; 408: 461–76Ishimaru, MJ Ikonomidou, C Tenkova, TI Der, TC Dikranian, K Sesma, MA Olney, JW
Cheng Y, Deshmukh M, D'Costa A, Demaro JA, Gidday JM, Shah A, Sun Y, Jacquin MF, Johnson EM, Holtzman DM: Caspase inhibitor affords neuroprotection with delayed administration in a rat model of neonatal hypoxic-ischemic brain injury. J Clin Invest 1998; 101: 1992–99Cheng, Y Deshmukh, M D'Costa, A Demaro, JA Gidday, JM Shah, A Sun, Y Jacquin, MF Johnson, EM Holtzman, DM
Nakajima W, Ishida A, Lange MS, Gabrielson KL, Wilson MA, Martin LJ, Blue ME, Johnston MV: Apoptosis has a prolonged role in the neurodegeneration after hypoxic ischemia in the newborn rat. J Neurosci 2000; 20: 7994–8004Nakajima, W Ishida, A Lange, MS Gabrielson, KL Wilson, MA Martin, LJ Blue, ME Johnston, MV
Gill R, Soriano M, Blomgren K, Hagberg H, Wybrecht R, Miss MT, Hoefer S, Adam G, Niederhauser O, Kemp JA, Loetscher H: Role of caspase-3 activation in cerebral ischemia-induced neurodegeneration in adult and neonatal brain. J Cereb Blood Flow Metab 2002; 22: 420–30Gill, R Soriano, M Blomgren, K Hagberg, H Wybrecht, R Miss, MT Hoefer, S Adam, G Niederhauser, O Kemp, JA Loetscher, H
Alnemri ES: Mammalian cell death proteases: a family of highly conserved aspartate specific cysteine proteases. J Cell Biochem 1997; 64: 33–42Alnemri, ES
Thornberry NA, Lazebnik Y: Caspases: enemies within. Science 1998; 281: 1312–16Thornberry, NA Lazebnik, Y
Honig LS, Rosenberg RN: Apoptosis and neurologic disease. Am J Med 2000; 108: 317–30Honig, LS Rosenberg, RN
Yuan J, Yankner BA: Apoptosis in the nervous system. Nature 2000; 407: 802–9Yuan, J Yankner, BA
Mignotte B, Vayssiere JL: Mitochondria and apoptosis. Eur J Biochem 1998; 252: 1–15Mignotte, B Vayssiere, JL
Kroemer G, Reed JC: Mitochondrial control of cell death. Nat Med 2000; 6: 513–9Kroemer, G Reed, JC
Reed JC, Jurgensmeier JM, Matsuyama S: Bcl-2 family proteins and mitochondria. Biochim Biophys Acta 1998; 1366: 127–37Reed, JC Jurgensmeier, JM Matsuyama, S
Linnik MD, Zahos P, Geschwind MD, Federoff HJ: Expression of bcl-2 from a defective herpes simplex virus-1 vector limits neuronal death in focal cerebral ischemia. Stroke 1995; 26: 1670–74Linnik, MD Zahos, P Geschwind, MD Federoff, HJ
Lawrence MS, Ho DY, Sun GH, Steinberg GK, Sapolsky RM: Overexpression of Bcl-2 with herpes simplex virus vectors protects CNS neurons against neurological insults in vitro and in vivo. J Neurosci 1996; 16: 486–96Lawrence, MS Ho, DY Sun, GH Steinberg, GK Sapolsky, RM
Papadopoulos MC, Koumenis IL, Xu L, Giffard RG: Potentiation of murine astrocyte antioxidant defence by bcl-2: protection in part reflects elevated glutathione levels. Eur J Neurosci 1998; 10: 1252–60Papadopoulos, MC Koumenis, IL Xu, L Giffard, RG
Endres M, Namura S, Shimizu-Sasamata M, Waeber C, Zhang L, Gomez-Isla T, Hyman BT, Moskowitz MA: Attenuation of delayed neuronal death after mild focal ischemia in mice by inhibition of the caspase family. J Cereb Blood Flow Metab 1998; 18: 238–47Endres, M Namura, S Shimizu-Sasamata, M Waeber, C Zhang, L Gomez-Isla, T Hyman, BT Moskowitz, MA
Cao G, Pei W, Ge H, Liang Q, Luo Y, Sharp FR, Lu A, Ran R, Graham SH, Chen J: In Vivo Delivery of a Bcl-xL Fusion Protein Containing the TAT Protein Transduction Domain Protects against Ischemic Brain Injury and Neuronal Apoptosis. J Neurosci 2002; 22: 5423–31Cao, G Pei, W Ge, H Liang, Q Luo, Y Sharp, FR Lu, A Ran, R Graham, SH Chen, J
Mouw G, Zechel JL, Zhou Y, Lust WD, Selman WR, Ratcheson RA: Caspase-9 inhibition after focal cerebral ischemia improves outcome following reversible focal ischemia. Metab Brain Dis 2002; 17: 143–51Mouw, G Zechel, JL Zhou, Y Lust, WD Selman, WR Ratcheson, RA
Martin LJ, Al-Abdulla NA, Brambrink AM, Kirsch JR, Sieber FE, Portera-Cailliau C: Neurodegeneration in excitotoxicity, global cerebral ischemia, and target deprivation: A perspective on the contributions of apoptosis and necrosis. Brain Res Bull 1998; 46: 281–309Martin, LJ Al-Abdulla, NA Brambrink, AM Kirsch, JR Sieber, FE Portera-Cailliau, C
McConkey DJ: Biochemical determinants of apoptosis and necrosis. Toxicol Lett 1998; 99: 157–68McConkey, DJ
Roy M, Sapolsky R: Neuronal apoptosis in acute necrotic insults: why is this subject such a mess? Trends Neurosci 1999; 22: 419–22Roy, M Sapolsky, R
Fujikawa DG: Confusion between neuronal apoptosis and activation of programmed cell death mechanisms in acute necrotic insults. Trends Neurosci 2000; 23: 410–1Fujikawa, DG
Northington FJ, Ferriero DM, Graham EM, Traystman RJ, Martin LJ: Early Neurodegeneration after Hypoxia-Ischemia in Neonatal Rat Is Necrosis while Delayed Neuronal Death Is Apoptosis. Neurobiol Dis 2001; 8: 207–19Northington, FJ Ferriero, DM Graham, EM Traystman, RJ Martin, LJ
Sperandio S, de Belle I, Bredesen DE: An alternative, nonapoptotic form of programmed cell death. Proc Natl Acad Sci U S A 2000; 97: 14376–81Sperandio, S de Belle, I Bredesen, DE
Mathiasen IS, Jaattela M: Triggering caspase-independent cell death to combat cancer. Trends Mol Med 2002; 8: 212–20Mathiasen, IS Jaattela, M
Leist M, Jaattela M: Four deaths and a funeral: from caspases to alternative mechanisms. Nat Rev Mol Cell Biol 2001; 2: 589–98Leist, M Jaattela, M
Zhan RZ, Wu C, Fujihara H, Taga K, Qi S, Naito M, Shimoji K: Both caspase-dependent and caspase-independent pathways may be involved in hippocampal CA1 neuronal death because of loss of cytochrome c From mitochondria in a rat forebrain ischemia model. J Cereb Blood Flow Metab 2001; 21: 529–40Zhan, RZ Wu, C Fujihara, H Taga, K Qi, S Naito, M Shimoji, K
Grafe MR: Developmental changes in the sensitivity of the neonatal rat brain to hypoxic/ischemic injury. Brain Res 1994; 653: 161–6Grafe, MR
Papadopoulos MC, Koumenis IL, Yuan TY, Giffard RG: Increasing vulnerability of astrocytes to oxidative injury with age despite constant antioxidant defenses. Neuroscience 1998; 82: 915–25Papadopoulos, MC Koumenis, IL Yuan, TY Giffard, RG