Characteristics of an amiloride-sensitive sodium entry pathway in cultured rodent glial and neuroblastoma cells.
We have studied the induction of an amiloride-sensitive sodium influx into C6 glioma, NIE, and NB2A neuroblastoma cell lines. In late log phase, cells grown continuously in the presence of 10% fetal calf serum showed Na+ influxes of approximately 25-30 nmol/mg protein min; less than 5% of this flux was inhibited by amiloride. Removal of serum for 24 h caused a decrease in the total Na+ influx to 15-20 nmol/mg protein/min. Upon readdition of serum to the incubation medium, there was an increase in total Na+ influx, depending on the cell type, of 20-400% within 2 min. This increment in Na+ influx represented an increase in amiloride-sensitive Na+ transport with an apparent K'1 of 0.4 mM. By adding serum back at various times after serum deprivation, it was determined that 4 h was required to observe a detectable increase in the amiloride-sensitive Na+ flux. Thus, serum removal results in the induction of the amiloride transport system which, however, remains latent until the reintroduction of serum to the medium. Addition of 5 micrograms/ml of cycloheximide blocked the increase in Na+ transport, indicating that de novo protein synthesis mediated this serum deprivation-induced increase in Na+ transport. Moreover, inhibition of de novo lipid synthesis by 0.1 mM fenfluramine also blocked the induction of this transport activity, suggesting that a coordinated synthesis of lipid and protein is required for the expression of this sodium transport site. We have also found that this serum stimulated Na+ influx did not saturate with Na+ concentration, up to 140 mM. Also, among commonly used inhibitors of passive Na+ entry into epithelial tissues, only amiloride was capable of inhibiting this transport system in these neural cell lines.
D000584:Amiloride / Q000031:analogs & derivatives / Q000494:pharmacology*; D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D001769:Blood; D000001:Calcimycin / Q000494:pharmacology; D002455:Cell Division; D002460:Cell Line; D003513:Cycloheximide / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D005277:Fenfluramine / Q000494:pharmacology; D005910:Glioma; D007700:Kinetics; D051379:Mice; D009447:Neuroblastoma; D009474:Neurons / Q000378:metabolism*; D011719:Pyrazines / Q000494:pharmacology*; D051381:Rats; D012964:Sodium / Q000378:metabolism*
D011719:Pyrazines; D005277:Fenfluramine; D000001:Calcimycin; D000584:Amiloride; D003513:Cycloheximide; D012964:Sodium