Ionic mechanisms of anoxic injury in mammalian CNS white matter: role of Na+ channels and Na(+)-Ca2+ exchanger

Abstract

White matter of the mammalian CNS suffers irreversible injury when subjected to anoxia/ischemia. However, the mechanisms of anoxic injury in central myelinated tracts are not well understood. Although white matter injury depends on the presence of extracellular Ca2+, the mode of entry of Ca2+ into cells has not been fully characterized. We studied the mechanisms of anoxic injury using the in vitro rat optic nerve, a representative central white matter tract. Functional integrity of the nerves was monitored electrophysiologically by quantitatively measuring the area under the compound action potential, which recovered to 33.5 +/- 9.3% of control after a standard 60 min anoxic insult. Reducing Na+ influx through voltage-gated Na+ channels during anoxia by applying Na+ channel blockers (TTX, saxitoxin) substantially improved recovery; TTX was protective even at concentrations that had little effect on the control compound action potential. Conversely, increasing Na+ channel permeability during anoxia with veratridine resulted in greater injury. Manipulating the transmembrane Na+ gradient at various times before or during anoxia greatly affected the degree of resulting injury; applying zero-Na+ solution (choline or Li+ substituted) before anoxia significantly improved recovery; paradoxically, the same solution applied after the start of anoxia resulted in more injury than control. Thus, ionic conditions that favored reversal of the normal transmembrane Na+ gradient during anoxia promoted injury, suggesting that Ca2+ loading might occur via reverse operation of the Na+)-Ca2+ exchanger. Na(+)-Ca2+ exchanger blockers (bepridil, benzamil, dichlorobenzamil) significantly protected the optic nerve from anoxic injury. Together, these results suggest the following sequence of events leading to anoxic injury in the rat optic nerve: anoxia causes rapid depletion of ATP and membrane depolarization leading to Na+ influx through incompletely inactivated Na+ channels. The resulting rise in the intracellular [Na+], coupled with membrane depolarization, causes damaging levels of Ca2+ to be admitted into the intracellular compartment through reverse operation of the Na(+)-Ca2+ exchanger. These observations emphasize that differences in the pathophysiology of gray and white matter anoxic injury are likely to necessitate multiple strategies for optimal CNS protection.

A recoverable state of axon injury persists for hours after spinal cord contusion in vivo

“…The immediate rise in calcium after injury and our EGTA experiments suggested an extracellular source of calcium. Therefore, we performed a series of pilot experiments that pharmacologically queried established molecularly defined entry pathways (for example, voltage-gated calcium channels [17][18][19] ; reverse action of the sodium-calcium exchanger [20][21][22] ; glutamate receptors 23 ). These experiments failed to show strong effects on initial calcium rises (proportion of calcium-elevated axons at initial p.i.…”

“…Given the critical role of calcium accumulations in contused axons, it is important to consider where the intra-axonal calcium is coming from and which downstream mechanisms it initiates. In vitro and in a multitude of neurological conditions, including trauma, ischaemia, inflammation and degeneration 12,26 , a broad range of mechanisms have been implicated as sources for elevated axonal calcium, including extracellular sources (for example, voltage-gated ion channels [17][18][19] , the plasma membrane sodiumcalcium exchanger [20][21][22] , the acid sensing ion channel 27 or nodal glutamate receptors 23 ), as well as intracellular stores (such as mitochondria 28 or the endoplasmic reticulum 29 ). Our in vivo analysis now indicates that mechanoporation, that is, the formation of pores in the plasma membrane as first described after diffused brain injury 24 , is a dominant source of calcium influx at least in the first hours after contusion.…”

Disruption of ionic and cell volume homeostasis in cerebral ischemia: The perfect storm

“…Of special interest is the involvement of NCX in white matter injury (reviewed in [14]). Rat optic nerve and spinal cord white matter tracks are functionally protected against anoxic injury by removal of extracellular Ca 2+ or by NCX inhibitors benzamil, bepridil, DCB, and KN-R7943 [117][118][119]. These data are of particular importance, as ischemic white matter injury is less understood and may contribute substantially to neurological deficits in human stroke [120].…”

Incorporation of sodium channel blocking and free radical scavenging activities into a single drug, AM‐36, results in profound inhibition of neuronal apoptosis

“…Pharmacological modulation of voltage-sensitive sodium channels is considered to be a rational and e ective therapeutic strategy against neuronal damage in cerebral ischaemia (Carter, 1998;Rataud et al, 1994). Sodium in¯ux is an important initiating event leading to anoxic damage and a cascade of cellular events (Stys et al, 1992). Blockade of Na + channels during periods of reduced oxygen supply has therefore been proposed as an e ective way of limiting energy expenditure since a large part of the energy consumed by excitable cells is used to maintain ionic gradients across the cellular membrane (Carter, 1998;Urenjak et al, 1996).…”

“…Loss of ATP results in a drop in membrane potential to the point where voltage-dependent Na + channels open, causing Na + in¯ux, continual depolarization and initiation of a cascade of cellular events (Stys et al, 1992). Down-regulation of Na + channels has been proposed as a potential way of reducing energy expenditure in cerebral ischaemia since much of the energy required by excitable cells is used to maintain Na + and K + gradients across the cellular membrane (Carter, 1998;Urenjak et al, 1996).…”