Brain tissue responses to ischemia (original) (raw)

Glutamate-induced neuronal death. The main excitatory neurotransmitter throughout the CNS is the dicarboxylic amino acid, glutamate. Reflecting this ubiquitous role in cell-cell signaling, average whole brain concentrations are on the order of 10 mM, with presumably much higher concentrations within synaptic vesicles. Under ischemic conditions, transmitter glutamate is massively released (initially mediated by vesicular release from nerve terminals, and later by reverse transport from astrocytes), reaching near-millimolar concentrations in the extracellular space. Unfortunately, such concentrations of glutamate are neurotoxic, and substantial evidence now implicates the toxicity of glutamate (excitotoxicity) in the pathogenesis of neuronal death after ischemia and other acute insults.

Extracellular glutamate accumulating under ischemic conditions overstimulates _N_-methyl-D-aspartate (NMDA), alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA), and kainate-type glutamate receptors, promoting Na+ influx and K+ efflux through glutamate receptor-activated membrane channels. NMDA receptor–gated ion channels are additionally highly permeable to Ca2+ and mediate Ca2+ influx into neurons. The gating of glutamate receptor–activated channels effectively achieves membrane shunting, which spreads in waves (spreading depression) from the ischemic core out toward the margins of the ischemic zone (ischemic penumbra). Spreading depression increases metabolic demand and energy failure, thus further enhancing glutamate release. Marked neuronal cell body swelling and dendrite swelling occur, hallmarks of necrosis death, as Na+ and Ca2+ entry is joined by the influx of Cl– and water. Elevations in neuronal intracellular free Ca2+ ([Ca2+]i), mediated both directly by NMDA receptors and indirectly via membrane depolarization–activated voltage-gated Ca2+ channels and reverse operation of the Na+-Ca2+ exchanger, bear particular responsibility for promoting spreading depression and triggering deleterious cytotoxic cascades.

In neuronal cell cultures, selective NMDA receptor blockade prevents most of the Ca2+ influx and cell death induced by brief intense glutamate exposures (1). NMDA antagonists also markedly attenuated the death of cultured neurons induced by oxygen and/or glucose deprivation, observations that fit well with studies conducted with selective agonists. Exposure to NMDA for as little as 3–5 minutes is sufficient to trigger widespread cultured cortical neuronal death (“rapidly triggered excitotoxicity”), whereas exposure to even saturating concentrations of kainate typically requires hours to do the same (“slowly triggered excitotoxicity”). This difference in time course fits with a higher rate of Ca2+ influx mediated directly by NMDA receptor–gated channels, compared with a slower rate of Ca2+ influx mediated by the voltage-gated channel and exchanger routes activated by AMPA or kainate receptors. NMDA receptor antagonists are also highly neuroprotective in animal models of focal brain ischemia, as well as hypoglycemia or trauma (2), although not transient global ischemia (3). In this latter setting, NMDA receptor–mediated excitotoxicity may be less prominent than AMPA receptor–facilitated Zn2+ entry in inducing lethal neuronal injury (see below). Reasons for this shift in prominence are presently not well-defined, but a contributing factor may be extracellular acidity due to accumulation of lactic acid during global ischemia, an event less prominent in the penumbra of focal ischemia where perfusion is partially maintained. This acid shift selectively downregulates NMDA receptors and NMDA receptor–mediated excitotoxicity but enhances AMPA receptor–mediated excitotoxicity (4); it may also enhance toxic Zn2+ entry through voltage-gated Ca2+ channels (5).

Other signaling messengers and growth factors. In addition to glutamate, other neurotransmitters released to the extracellular space during ischemia can significantly influence resultant brain injury. Dopamine, which increases 500-fold in the extracellular space following global ischemia, may contribute to striatal neuronal death. Moreover, experimental reduction of dopamine release, which can be accomplished by creating lesions in the dopaminergic neurons projecting from the substantia nigra or by using the tyrosine hydroxylase inhibitor alpha-methyl-_p_-tyrosine to deplete endogenous stores of dopamine, attenuates striatal injury in rodent global ischemia models (6). Contributing to dopamine-induced potentiation of ischemic injury may be its ability to enhance glutamate receptor currents.

Neurotransmitters do not all act to promote injury; several, including serotonin, gamma-aminobutyric acid (GABA; see below), and adenosine, are neuroprotective. Adenosine, which accumulates rapidly during ischemia due to breakdown of ATP, has beneficial effects in many tissues. The activation of adenosine A2a receptors on vascular smooth muscle cells and neutrophils enhances blood flow and decreases inflammation, respectively (7). Adenosine also has nervous system–specific beneficial effects, mediated by the ability of neuronal adenosine A1-receptors to reduce neurotransmitter release and membrane excitability. In addition, the expression of several growth factors increases in ischemic tissues, likely as a protective response. Exogenous administration of growth factors has shown therapeutic promise in several experimental models of organ ischemia, including in liver, kidney, heart, and brain. Examples of growth factors whose administration reduces brain damage in rats subjected to cerebral ischemia are nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), neurotrophins 4/5 (NT-4/5), basic fibroblast growth factor, and IGF-1, which apparently blocks neuronal apoptosis (see below). Some growth factors may also enhance nerve fiber sprouting and synapse formation after ischemic injury, thereby promoting functional recovery.

Despite their overall salutary effects, certain growth factors may also enhance neuronal vulnerability to excitotoxic and free radical–induced death. Acute exposure to BDNF, NT-3, or NT-4/5 reduces the vulnerability of cultured neocortical neurons to apoptosis, but exacerbates the cellular necrosis of the same cells after exposure to oxygen-glucose deprivation or NMDA. The deleterious consequences of the neurotrophins as well as IGF-1 may be explained in part by enhanced NMDA receptor–mediated Ca2+ influx, enhanced production of free radicals, or possibly acute proexcitatory effects that could increase excitotoxicity (8). These deleterious consequences are not restricted to embryonic or in vitro systems, as free radical–mediated tissue damage induced by direct intraparenchymal injection of iron into the adult rat spinal cord was increased by pretreatment with BDNF, NT-3, or NT-4/5 (J. McDonald et al., unpublished data). If these growth factors have an injury-enhancing component effect in the ischemic brain, perhaps masked by other survival-promoting effects, interventions aimed specifically at blocking this component may uncover higher levels of net neuroprotective effects.

Zinc toxicity. Zinc, the second most abundant transition metal in the human body, is present in all cells, for the most part tightly bound to proteins, such as metalloenzymes and transcription factors, where it serves catalytic and structural roles. In the brain, there is an additional substantial pool of chelatable Zn2+ localized to synaptic vesicles in excitatory (glutamatergic) nerve terminals, which is released in a Ca2+-dependent fashion with depolarization and can alter the behavior of several transmitter receptors and voltage-gated channels (9). While the normal functional significance of this presumptive signaling Zn2+ pool is not presently understood, growing evidence suggests that it contributes to nerve cell death under pathological conditions such as ischemia or seizures or following head trauma (10).

Following transient global ischemia, chelatable Zn2+ translocates from nerve terminals into the cell bodies of vulnerable neurons (11). This translocation precedes neuronal degeneration, and its interruption by the intracerebroventricular (icv) injection of a chelator, ethylenediaminetetraacetic acid saturated with equimolar Ca2+ (CaEDTA), reduces subsequent neuronal death. Furthermore, exposure to the high micromolar concentrations of Zn2+ likely to occur in brain extracellular space after synchronous cellular depolarization is sufficient to kill cultured neurons, especially if the neurons are depolarized, which facilitates entry of Zn2+ across the plasma membrane through voltage-gated Ca2+ channels (10). Recent observations from our laboratory suggest that Zn2+ toxicity may also contribute to the development of cerebral infarction following mild transient focal ischemia (G.J. Zipfel et al., unpublished observations).