Guanine nucleotide stimulation of norepinephrine secretion from permeabilized PC12 cells: effects of Mg2+, other nucleotide triphosphates and N-ethylmaleimide.
The addition of either Ca2+ or guanosine 5'-O-3-(thiotriphosphate), GTP gamma S, to digitonin-permeabilized rat pheochromocytoma PC12 cells stimulates norepinephrine release. Unlike Ca(2+)-stimulated release, there is a delay between the time of addition of GTP gamma S to digitonin-permeabilized PC12 cells and stimulation of norepinephrine release. Preincubation of the permeabilized cells in the absence of Mg2+ eliminates this lag and increases the initial rate of GTP-gamma S-stimulated norepinephrine secretion. This suggests that the rate of GDP dissociation from the GTP-binding protein responsible for this stimulation is faster in the absence of Mg2+ than in its presence. While an equimolar concentration of GTP gives 50% inhibition of GTP gamma S-stimulated release, 100-fold excesses of ITP, ATP, UTP and CTP gave no inhibition of GTP gamma S-stimulated release. Both the inability of ITP to inhibit GTP gamma S-stimulated secretion and the increase in GTP gamma S-stimulated secretion caused by preincubation in the absence of Mg2+ indicate that some of the properties of the GTP-binding protein responsible for this stimulation are more like those of the low molecular weight GTP-binding proteins rap1 and ras than those of a heterotrimeric G-protein. Low concentrations of N-ethylmaleimide gave more inhibition of GTP gamma S-stimulated release than Ca(2+)-stimulated release which suggests that the mechanisms by which Ca2+ and GTP gamma S stimulate norepinephrine release are at least in part distinct.
Yang
Y C
YC
Laboratory of Biochemistry, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892.
Vu
N D
ND
Wagner
P D
PD
D000818:Animals; D004072:Digitonin / Q000494:pharmacology; D005033:Ethylmaleimide / Q000494:pharmacology*; D019204:GTP-Binding Proteins / Q000378:metabolism; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology*; D008274:Magnesium / Q000494:pharmacology*; D009638:Norepinephrine / Q000378:metabolism*; D009711:Nucleotides / Q000494:pharmacology*; D016716:PC12 Cells; D010738:Type C Phospholipases / Q000378:metabolism
D009711:Nucleotides; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D010738:Type C Phospholipases; D019204:GTP-Binding Proteins; D008274:Magnesium; D004072:Digitonin; D005033:Ethylmaleimide; D009638:Norepinephrine