Insulin binding and effects on pyrimidine nucleoside uptake and incorporation in cultured mouse astrocytes.
Binding of 125I-insulin to primary cultures of differentiated mouse astrocytes was time-dependent, reaching equilibrium after 2 h at 22 degrees C, with equilibrium binding corresponding to 20.79 fmol/mg of protein, representing approximately 5,000 occupied binding sites/cell. The half-life of 125I-insulin dissociation at 22 degrees C was 2 min, with an initial dissociation rate constant of 4.12 X 10(-2) s-1. Dissociation of bound 125I-insulin was not accelerated significantly in the presence of unlabeled insulin (16.7 microM). Porcine and desoctapeptide insulins competed for specific 125I-insulin binding in a dose-dependent manner, whereas growth hormone, glucagon, and somatostatin did not. For porcine insulin, Scatchard analysis suggested multiple-affinity binding sites (high-affinity Ka = 4.92 X 10(8) M-1; low-affinity Ka = 0.95 X 10(7) M-1). After incubation with insulin (0.5 microM) for 2 h at 37 degrees C, increases above basal values of 254 +/- 23 and 189 +/- 34% for [3H]uridine uptake and incorporation, respectively, were observed. After incubation with insulin (0.5 microM) for 24 h at 37 degrees C, there were increases of 145 +/- 6% for [3H]thymidine uptake and 166 +/- 11% for thymidine incorporation. Basal and stimulated uridine and thymidine uptake and incorporation were inhibited by 50 microM dipyridamole. These studies confirm that mouse astrocytes in vitro possess specific insulin receptors and demonstrate an effect of insulin on pyrimidine nucleoside uptake and incorporation.
Kum
W
W
Cockram
C S
CS
Zhu
S Q
SQ
Teoh
R
R
Vallance-Owen
J
J
Young
J D
JD
D000818:Animals; D001253:Astrocytes / Q000187:drug effects / Q000378:metabolism*; D002478:Cells, Cultured; D007328:Insulin / Q000378:metabolism* / Q000494:pharmacology; D007457:Iodine Radioisotopes; D007700:Kinetics; D051379:Mice; D008813:Mice, Inbred ICR; D011972:Receptor, Insulin / Q000378:metabolism; D013936:Thymidine / Q000378:metabolism*; D014529:Uridine / Q000378:metabolism*
D007328:Insulin; D007457:Iodine Radioisotopes; D011972:Receptor, Insulin; D013936:Thymidine; D014529:Uridine