Role of Na(+)/H(+) exchanger during O(2) deprivation in mouse CA1 neurons.
To determine the role of membrane transporters in intracellular pH (pH(i)) regulation under conditions of low microenvironmental O(2), we monitored pH(i) in isolated single CA1 neurons using the fluorescent indicator carboxyseminaphthorhodafluor-1 and confocal microscopy. After total O(2) deprivation or anoxia (PO(2) approximately equal to 0 Torr), a large increase in pH(i) was seen in CA1 neurons in HEPES buffer, but a drop in pH(i), albeit small, was observed in the presence of HCO(3)(-). Ionic substitution and pharmacological experiments showed that the large anoxia-induced pH(i) increase in HEPES buffer was totally Na(+) dependent and was blocked by HOE-694, strongly suggesting the activation of the Na(+)/H(+) exchanger (NHE). Also, this pH(i) increase in HEPES buffer was significantly smaller in Na(+)/H(+) exchanger isoform 1 (NHE1) null mutant CA1 neurons than in wild-type neurons, demonstrating that NHE1 is responsible for part of the pH(i) increase following anoxia. Both chelerythrine and H-89 partly blocked, and H-7 totally eliminated, this anoxia-induced pH(i) increase in the absence of HCO. We conclude that 1) O(2) deprivation activates Na(+)/H(+) exchange by enhancing protein kinase activity and 2) membrane proteins, such as NHE, actively participate in regulating pH(i) during low-O(2) states in neurons.
Yao
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Section of Respiratory Medicine, Department of Pediatrics, Yale University School of Medicine, New Haven, Connecticut 06520, USA.
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Douglas
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D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine / Q000494:pharmacology; D000143:Acids / Q000378:metabolism; D000470:Alkaloids; D000818:Animals; D053119:Benzophenanthridines; D015687:Cell Hypoxia / Q000502:physiology; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D006146:Guanidines / Q000494:pharmacology; D006624:Hippocampus / Q000166:cytology*; D006863:Hydrogen-Ion Concentration; D002534:Hypoxia, Brain / Q000378:metabolism*; D066298:In Vitro Techniques; D007546:Isoquinolines / Q000494:pharmacology; D008297:Male; D051379:Mice; D008818:Mice, Neurologic Mutants; D009474:Neurons / Q000378:metabolism*; D010100:Oxygen / Q000494:pharmacology*; D010617:Phenanthridines / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism; D012964:Sodium / Q000494:pharmacology; D017923:Sodium-Hydrogen Exchangers / Q000037:antagonists & inhibitors / Q000378:metabolism*; D013449:Sulfonamides; D013450:Sulfones / Q000494:pharmacology
D000143:Acids; D000470:Alkaloids; D053119:Benzophenanthridines; D004791:Enzyme Inhibitors; D006146:Guanidines; D007546:Isoquinolines; D010617:Phenanthridines; D017923:Sodium-Hydrogen Exchangers; D013449:Sulfonamides; D013450:Sulfones; C069139:growth factor-activatable Na-H exchanger NHE-1; C074107:3-methylsulfonyl-4-piperidinobenzoyl guanidine; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D012964:Sodium; C016299:chelerythrine; D017868:Cyclic AMP-Dependent Protein Kinases; D011493:Protein Kinase C; C063509:N-(2-(4-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide; D010100:Oxygen