GTP gamma S increases Nav1.8 current in small-diameter dorsal root ganglia neurons.
Tetrodotoxin-resistant (TTX-R) sodium current in small-size dorsal root ganglia (DRG) neurons is upregulated by prostaglandin E(2) and serotonin through a protein kinase A (PKA)/protein kinase (PKC) pathway, suggesting G protein modulation of one or more TTX-R channels in these neurons. Recently, GTP(gammaS), a hydrolysis-resistant analogue of GTP, was shown to increase the persistent current produced by the TTX-R Na(v)1.9. In this study, we investigated the modulation of another TTX-R channel, Na(v)1.8, by GTP(gammaS) in small-diameter DRG neurons from rats using whole-cell voltage clamp recordings. Because it has been suggested that fluoride, often used in intracellular recording solutions, may bind to trace amounts of aluminum and activate G proteins, we recorded Na(v)1.8 currents with and without intracellular fluoride, and with the addition of deferoxamine, an aluminum chelator, to prevent fluoride-aluminum binding. Our results show that GTP(gammaS) (100 micro M) caused a significant increase in Na(v)1.8 current (67%) with a chloride-based intracellular solution. Although the inclusion of fluoride instead of chloride in the pipette solution increased the Na(v)1.8 current by 177%, GTP(gammaS) further increased Na(v)1.8 current by 67% under these conditions. While the effect of GTP(gammaS) was prevented by pretreatment with H7 (100 micro M), a non-selective PKA/PKC inhibitor, the fluoride-induced increase in Na(v)1.8 current was not sensitive to H7 (100 micro M), or to inclusion of deferoxamine (1 mM) in the intracellular solution. We conclude that G protein activation by GTP(gammaS) increases Na(v)1.8 current through a PKA/PKC mechanism and that addition of fluoride to the pipette solution further enhances the current, but is not a confounding variable in the study of Na(v)1.8 channel modulation by G proteins independent of a PKA/PKC pathway or binding to aluminum.
Saab
Carl Y
CY
Department of Neurology and PVA/Eastern Paralyzed Veterans Association Neuroscience Research Center, Yale Medical School, New Haven, CT 06510, USA.
Cummins
Theodore R
TR
Waxman
Stephen G
SG
D000200:Action Potentials / Q000187:drug effects / Q000502:physiology; D000818:Animals; D005727:Ganglia, Spinal / Q000187:drug effects* / Q000502:physiology; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology*; D008297:Male; D062559:NAV1.8 Voltage-Gated Sodium Channel; D009419:Nerve Tissue Proteins / Q000502:physiology*; D009474:Neurons / Q000187:drug effects* / Q000502:physiology; D051381:Rats; D017207:Rats, Sprague-Dawley; D015222:Sodium Channels / Q000502:physiology*
D062559:NAV1.8 Voltage-Gated Sodium Channel; D009419:Nerve Tissue Proteins; C568319:Scn10a protein, rat; D015222:Sodium Channels; D016244:Guanosine 5'-O-(3-Thiotriphosphate)