An early transient increase of intracellular Na+ may be one of the first components of the mitogenic signal. Direct detection by 23Na-NMR spectroscopy in quiescent 3T3 mouse fibroblasts stimulated by growth factors.
23Na-NMR spectroscopy was designed to allow for continuous recording of intracellular Na+ in 3T3 fibroblasts stimulated by serum growth-factors in the presence of ion transport inhibitors. The metabolic state of cells at rest and following stimulation was monitored by 31P-NMR spectra of ATP and related high-energy phosphates. The study demonstrates that early activation of ion transporters by addition of serum is marked by the appearance of transient increase of the intracellular Na+, beginning 3 min after addition of serum to quiescent culture and lasting approx. 20 min. The initial rise in cellular Na+ results from an increased activity of the bumetanide-sensitive Na+/K+/Cl- cotransport and of the amiloride-sensitive Na+/H+ antiport. It is suppressed by any one of these inhibitors. Subsequent activation of the ouabain-sensitive Na+/K(+)-ATPase results in an increased Na+ efflux, leading to a return of intracellular Na+ to its initial baseline. Previous work had shown that the early activation of bumetanide-sensitive and amiloride sensitive ion-transporters by growth-factors was essential for induction of cell division, at least in some cell types. Preventing ion activation by adding ion-transport inhibitors lead to the inhibition of DNA synthesis 18 h later. This process was reversible upon elimination of these inhibitors. Even though alternative non-specific effects of these inhibitors cannot be ruled out, the observed transient peak in intracellular Na+ may be one of the earliest components of the mitogenic signal. On the basis of previous works, its effect seems to be related to the activation of Ca(2+)-dependent and cyclic AMP second messenger pathways. The different mechanisms whereby the activated Na+/K+/Cl- cotransport and the Na+/H+ antiport contribute to this signal need to be further investigated.
D016475:3T3 Cells; D000584:Amiloride / Q000494:pharmacology; D000818:Animals; D002034:Bumetanide / Q000494:pharmacology; D002352:Carrier Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D004734:Energy Metabolism; D009682:Magnetic Resonance Spectroscopy; D051379:Mice; D008807:Mice, Inbred BALB C; D008938:Mitosis / Q000502:physiology*; D010042:Ouabain / Q000494:pharmacology; D010477:Perfusion; D015398:Signal Transduction; D012964:Sodium / Q000378:metabolism*; D017923:Sodium-Hydrogen Exchangers / Q000037:antagonists & inhibitors / Q000378:metabolism; D028021:Sodium-Potassium-Chloride Symporters
D002352:Carrier Proteins; D017923:Sodium-Hydrogen Exchangers; D028021:Sodium-Potassium-Chloride Symporters; D002034:Bumetanide; D010042:Ouabain; D000584:Amiloride; D012964:Sodium