Mechanisms of secondary injury to spinal cord axons in vitro: role of Na+, Na(+)-K(+)-ATPase, the Na(+)-H+ exchanger, and the Na(+)-Ca2+ exchanger.
There is evidence that intracellular Na+ entry potentiates hypoxic-ischemic cell death by causing cytotoxic cell edema, intracellular acidosis, and gating of Ca2+ entry by reverse activation of the Na(+)-Ca2+ exchanger. In this study, we examined the role of Na+ in mediating traumatic injury to spinal cord axons. Dorsal column segments from adult rats (n = 87) were isolated and maintained in an in vitro recording chamber while being superfused with oxygenated Ringer's solution (95% O2/5% CO2, 25 degrees C). Selected experiments (n = 10) also were done at 33 degrees C. Compound action potentials (CAP) were recorded from microelectrodes. Injury was performed by compression of the dorsal column segment for 15 sec with a modified aneurysm clip exerting a closing force of 2 gm. With injury, the CAP decreased to 72.1 +/- 9.6% of baseline values. Removal of extracellular Na+ and replacement with the impermeant cation N-methyl-D-glucamine enhanced recovery of the CAP to 98.3 +/- 18.3% (p < 0.05) of baseline. The Na+ channel blockers tetrodotoxin and procaine also improved recovery of the CAP to 96.3 +/- 23.7% (p < 0.05) and 82.8 +/- 4.6% (p < 0.05) of baseline values, respectively. In contrast, increasing Na+ permeability with veratridine resulted in greater attenuation of CAP amplitude after 1 hr of trauma (60.1 +/- 8.4%, p < 0.05). Similarly, prevention of extrusion of Na+ from the intracellular compartment by inhibiting the Na(+)-K(+)-ATPase pump with ouabain resulted in greater attenuation of CAP amplitude at 1 hr after trauma (56.7 +/- 3.6%, p < 0.05). The Na(+)-H+ exchange blockers amiloride (100 microM) and harmaline (100 microM) significantly improved recovery after injury to 89.6 +/- 17.0% (p < 0.05) and 85.7 +/- 7.2% (p < 0.05) of baseline, respectively. However, administration of the Na(+)-Ca2+ exchange blockers benzamil (100 or 500 microM) and bepridil (50 microM) was ineffective. In summary, reduction of extracellular Na+ confers neuroprotection after spinal cord injury in vitro. Intracellular sodium rises appear to be mediated by voltage-gated Na+ channels. Blockade of the Na(+)-H+ exchanger also is neuroprotective, possibly by reducing intracellular acidosis. Furthermore, prevention of extrusion of intracellular Na+ by the Na(+)-K(+)-ATPase pump exacerbates the effects of compression trauma. However, reverse operation of the Na(+)-Ca2+ exchanger does not explain the injurious effects of Na+ in traumatically injured CNS white matter.
Agrawal
S K
SK
Spinal Cord Injury Neurophysiology Laboratory, Playfair Neuroscience Unit, Toronto Hospital Research Institute, University of Toronto, Ontario, Canada.
Fehlings
M G
MG
D000818:Animals; D001369:Axons / Q000502:physiology*; D002352:Carrier Proteins / Q000378:metabolism*; D066298:In Vitro Techniques; D017136:Ion Transport; D008297:Male; D008564:Membrane Potentials / Q000502:physiology; D051381:Rats; D017208:Rats, Wistar; D012964:Sodium / Q000494:pharmacology*; D019831:Sodium-Calcium Exchanger; D017923:Sodium-Hydrogen Exchangers; D000254:Sodium-Potassium-Exchanging ATPase; D013119:Spinal Cord Injuries / Q000503:physiopathology*; D013997:Time Factors
D002352:Carrier Proteins; D019831:Sodium-Calcium Exchanger; D017923:Sodium-Hydrogen Exchangers; D012964:Sodium; D000254:Sodium-Potassium-Exchanging ATPase