Sodium/bicarbonate cotransporter NBCn1/slc4a7 increases cytotoxicity in magnesium depletion in primary cultures of hippocampal neurons.
Growing evidence suggests that pharmacological inhibition of Na/H exchange and Na/HCO(3) transport provides protection against damage or injury in cardiac ischemia. In this study, we examined the contribution of the sodium/bicarbonate cotransporter NBCn1 (slc4a7) to cytotoxicity in cultured hippocampal neurons of rats. In neurons exposed to extracellular pH (pH(o)) ranging from 6.2 to 8.3, NBCn1 protein expression increased by fivefold at pH < 6.5 compared to the expression at pH(o) 7.4. At pH(o) 6.5, the intracellular pH of neurons was approximately 1 unit lower than that at pH 7.4. Immunochemistry showed a marked increase in NBCn1 immunofluorescence in plasma membranes and cytosol of the soma as well as in dendrites, at pH(o) 6.5. NBCn1 expression also increased by 40% in a prolonged Mg(2+)-free incubation at normal pH(o). Knockdown of NBCn1 in neurons had negligible effect on cell viability. The effect of NBCn1 knockdown on cytotoxicity was then determined by exposing neurons to 0.5 mm glutamate for 10 min and measuring lactate dehydrogenase (LDH) release from neurons. Compared to normal incubation (pH(o) 7.2 for 6 h) after glutamate exposure, acidic incubation (pH(o) 6.3 for 6 h) reduced cytotoxicity by 75% for control neurons and 78% for NBCn1-knockdown neurons. Thus, both controls and knockdown neurons showed acidic protection from cytotoxicity. However, in Mg(2+)-free incubation after glutamate exposure, NBCn1 knockdown progressively attenuated cytotoxicity. This attenuation was unaffected by acidic preincubation before glutamate exposure. We conclude that NBCn1 has a dynamic upregulation in low pH(o) and Mg(2+) depletion. NBCn1 is not required for acidic protection, but increases cytotoxicity in Mg(2+)-free conditions.
Cooper
Deborah S
DS
Department of Physiology, Emory University School of Medicine, Atlanta, GA 30322, USA.
Yang
Han Soo
HS
He
Peijian
P
Kim
Eunjin
E
Rajbhandari
Ira
I
Yun
Chris C
CC
Choi
Inyeong
I
D000136:Acid-Base Equilibrium / Q000235:genetics; D000818:Animals; D002460:Cell Line; D002470:Cell Survival / Q000235:genetics; D002478:Cells, Cultured; D019610:Cytoprotection / Q000235:genetics*; D003603:Cytotoxins / Q000633:toxicity*; D005455:Fluorescent Antibody Technique; D018698:Glutamic Acid / Q000633:toxicity; D006624:Hippocampus / Q000378:metabolism*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008275:Magnesium Deficiency / Q000378:metabolism* / Q000503:physiopathology; D009474:Neurons / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D034622:RNA Interference / Q000502:physiology; D051381:Rats; D017207:Rats, Sprague-Dawley; D027982:Sodium-Bicarbonate Symporters / Q000235:genetics / Q000378:metabolism*; D015854:Up-Regulation / Q000235:genetics
D003603:Cytotoxins; C498902:Slc4a7 protein, rat; D027982:Sodium-Bicarbonate Symporters; D018698:Glutamic Acid; D007770:L-Lactate Dehydrogenase