Edited by: Steve Bunk
M. Ankarcrona, J.M. Dypbukt, E. Bonfoco, B. Zhivotovsky, S. Orrenius, S.A. Lipton, P. Nicotera, "Glutamate-induced neuronal death: A succession of necrosis or apoptosis depending on mitochondrial function," Neuron, 15:961-73, 1995. (Cited in 120 papers through October 1997)
Comments by Stuart A. Lipton, Cerebrovascular and NeuroScience Research Institute, Brigham and Women's Hospital and Program in Neuroscience, Harvard Medical School; and Pierluigi Nicotera, Molecular Toxicology Program, University of Konstanz, Germany
When a region of an injured brain is deprived of blood and oxygen, there are two ways the cells can die, by necrosis or by apoptosis. The mode of death makes a big difference to surrounding tissue. In the study that prompted this paper, a multinational team discovered that the functioning of a brain neuron's mitochondria-the energy-producing organelles outside the nucleus-is the critical determinant of how the cell dies during blood-deprived or ischemic brain injury, as well as in several neurodegenerative diseases. This knowledge, in turn, is essential to devising ways of preventing neuronal cell death and of combating neurodegenerative disease.
|WAYS TO DIE: Determining how and why a neuron dies was the goal of Stuart Lipton of Harvard, at TK, and Pierluigi Nicotera, then of the Karolinska Institute. |
Apoptosis, or programmed cell death, is a genetically regulated process characterized by cell shrinkage, nuclear condensation, and production of membrane-enclosed particles that are digested by other cells. "I call it death with dignity," Lipton quips, "because the cells don't spill their guts and damage other tissue." In contrast, necrosis is messy; the cells enlarge and then release cytoplasmic material into the surrounding area.
Both forms of cell death can happen during stroke, but apoptosis may predominate in neurodegenerative disorders such as Huntington's disease, AIDS dementia, Lou Gehrig's disease (amyotrophic lateral sclerosis), and even glaucoma. To analyze the two types of cell deaths as they developed, the team studied model cultures of cerebellar granule cells and cerebrocortical neurons. These cells have N-methyl-D-aspartate (NMDA) receptor subtypes that are known to become overstimulated by too much glutamate, a natural neurotransmitter in the brain and one of the excitatory amino acids. When overstimulated by too much glutamate, "the NMDA channels are exquisitely poised on the threshold of exciting the cells to death by allowing excessive calcium ion influx and the generation of free radicals," says Lipton. Such excitotoxicity occurs during a variety of neurodegenerative diseases.
The team exposed the granule cell neurons to varying concentrations of glutamate. Relying on Nicotera's expertise in confocal microscopy and fluorescence staining, they used a complex succession of different dyes to assess the cells' functions. "With dyes, you can look at the living state of the cells," Nicotera notes. For example, they saw that mitochondria of neurons undergoing necrosis after exposure to toxic amounts of glutamate became permeable and unable to hold dye.
During ischemic brain injury, cells near the center of the blood-deprived region are exposed to excessive amounts of glutamate and die by necrosis, which usually causes inflammation and degeneration of surrounding tissue. More distant or penumbral neurons may survive and recover their energy, only to die later by apoptosis, which does not injure their neighbors.
Lipton says the significance of this paper is that it was the first to show that the energy state of the mitochondria determines whether an insult will cause necrosis or apoptosis in a neuron. "What we found is that as the mitochondria depolarize, neurons lose their energy charge," he says. "When this occurs, apoptosis cannot proceed and the cells swell, lyse, and thus die by necrosis." The paper also suggests that some cells may die by necrosis because they are too damaged to summon the energy to enter apoptosis.
"Apoptosis is an active process," says Nicotera, "and sufficient generation of ATP [adenosine triphosphate, a nucleotide essential to the release of energy] is required for apoptosis to proceed."
Current work is focused on developing a safe antagonist to NMDA receptors or to downstream signaling pathways, to prevent either form of cell death. A leading candidate is memantine, a compound being tested to help treat AIDS dementia complex (ADC), stroke, and neuropathic pain.