Low-dose ouabain protects against excitotoxic apoptosis and up-regulates nuclear Bcl-2 in vivo.
Sodium-potassium ATPase (Na+,K+-ATPase) regulates the electrochemical gradient in cells, thereby providing fluid and ionic homeostasis. Additionally, interaction of the Na+,K+ pump with cardiac glycosides can activate intracellular signaling cascades (resulting in cell growth) and up-regulate transcription factors that promote cell survival. We used an in vivo excitotoxicity model to assess if Na+,K+-ATPase plays a role in neuronal apoptosis. After unilateral, intrastriatal injection of the glutamate receptor agonist kainic acid into postnatal day 7 rats, Na+,K+ pump function was increased at 12 h after excitotoxic challenge, and levels of neuron-specific enzyme subunits were preserved (up to 24 h after injection) in membrane-enriched striatal fractions. In addition, co-injection of kainic acid with a low-dose (0.01 nmol) of the cardiac glycoside ouabain significantly (P<0.05) reduced striatal apoptosis (at 24 h post-injection) without diminishing Na+,K+-ATPase activity. To evaluate the possible mechanisms for this neuroprotection, we examined the levels of nuclear factor kappa B and Bcl-2 after cardiac glycoside exposure. Low-dose ouabain increased nuclear Bcl-2 (but not nuclear factor kappa B) protein levels at 6 h post injection. Our results suggest that Na+,K+-ATPase allows for progression of apoptosis in excitotoxically-injured neurons, and that sublethal concentrations of ouabain provide neuroprotection against excitotoxicity. The mechanism for this ouabain neuroprotection could be intracellular cascades linked to the Na+,K+-ATPase-ouabain interaction that modulate subcellular Bcl-2 levels. Targeted, therapeutic inhibition of apoptosis through cardiac glycosides may represent an effective strategy against excitotoxicity-mediated neuronal injury.
Golden
W C
WC
Department of Pediatrics, Eudowood Neonatal Pulmonary Division, Johns Hopkins University School of Medicine, 600 North Wolfe Street, Nelson-Harvey 2-133, Baltimore, MD 21287, USA. cgolden@jhmi.edu
Martin
L J
LJ
D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D002467:Cell Nucleus / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D018690:Excitatory Amino Acid Agonists / Q000008:administration & dosage; D007150:Immunohistochemistry; D007276:Injections, Intraventricular; D007608:Kainic Acid / Q000008:administration & dosage; D010042:Ouabain / Q000494:pharmacology*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism*; D051381:Rats; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism*; D015854:Up-Regulation
D004791:Enzyme Inhibitors; D018690:Excitatory Amino Acid Agonists; D019253:Proto-Oncogene Proteins c-bcl-2; D010042:Ouabain; D000254:Sodium-Potassium-Exchanging ATPase; D007608:Kainic Acid