Enhanced expression of Ca2+ channels by nerve growth factor and the v-src oncogene in rat phaeochromocytoma cells.
1. Rat phaeochromocytoma (PC12) cells were used to investigate the expression of Ca2+ channel types during neuronal differentiation. Neuronal differentiation was induced by treatment with nerve growth factor (NGF) or by activation of a temperature-sensitive tyrosine kinase (pp60v-src) in genetically modified PC12 (PC12/v-src) cells. PC12 cells differentiated morphologically in the presence of NGF. When grown at the permissive temperature of 37 degrees C which activates the kinase activity of pp60v-src, PC12/v-src cells differentiated morphologically with the extension of neurites. In contrast, PC12/v-src cells grown at the non-permissive temperature of 40 degrees C continued to divide and were morphologically indistinguishable from control PC12 cells. 2. Whole-cell Ca2+ currents were measured in PC12 cells using Ba2+ as the charge carrier. Ba2+ currents measured at the peak of the current-voltage curve from a holding potential of -80 mV were -0.28 +/- 0.04 nA (mean +/- S.E.M.) in control PC12 cells compared to -1.25 +/- 0.16 nA in NGF-differentiated cells. The current density increased from 9.4 +/- 0.7 pA/pF in control PC12 cells to 22.8 +/- 2.4 pA/pF in NGF-differentiated PC12 cells. Ba2+ currents were -0.24 +/- 0.04 nA in undifferentiated PC12/v-src cells grown at the non-permissive temperature of 40 degrees C compared to -0.95 +/- 0.16 nA in differentiated PC12/v-src cells grown at the permissive temperature of 37 degrees C. The current density increased from 4.5 +/- 0.5 pA/pF in PC12/v-src cells grown at the non-permissive temperature of 40 degrees C to 13.3 +/- 2.4 pA/pF in PC12/v-src cells grown at the permissive temperature of 37 degrees C. 3. The sensitivity of Ba2+ currents to omega-conotoxin GVIA (omega-CgTX) was determined for currents measured at the peak of the current-voltage curve (0 mV in 10 mM Ba2+) from a holding potential of -80 mV. In NGF-differentiated PC12 cells, 10 microM omega-CgTx inhibited 68.1 +/- 3.2% of the total Ba2+ current compared to 35.9 +/- 4.1% in control cells. The density of the omega-CgTX-sensitive current increased from 3.3 +/- 0.4 pA/pF in control cells to 15.7 +/- 2.0 pA/pF in NGF-differentiated cells. In differentiated PC12/v-src cells grown at 37 degrees C, omega-CgTX inhibited 52.2 +/- 4.2% of total Ba2+ current compared to 41.1 +/- 3.8% in PC12/v-src cells grown at 40 degrees C. The density of the omega-CgTX-sensitive current increased from 1.9 +/- 0.3 to 7.4 +/- 2.0 pA/pF with v-src-mediated differentiation.(ABSTRACT TRUNCATED AT 400 WORDS)
Lewis
D L
DL
Department of Pharmacology and Toxicology, Medical College of Georgia, Augusta 30912.
De Aizpurua
H J
HJ
Rausch
D M
DM
D000818:Animals; D001464:Barium / Q000378:metabolism; D001667:Binding, Competitive / Q000187:drug effects; D002121:Calcium Channel Blockers / Q000494:pharmacology; D015220:Calcium Channels / Q000187:drug effects / Q000378:metabolism*; D002454:Cell Differentiation / Q000187:drug effects; D004594:Electrophysiology; D007457:Iodine Radioisotopes; D009414:Nerve Growth Factors / Q000494:pharmacology*; D009543:Nifedipine / Q000494:pharmacology; D009857:Oncogenes / Q000502:physiology*; D016716:PC12 Cells; D010455:Peptides / Q000494:pharmacology; D011869:Radioligand Assay; D051381:Rats; D020866:omega-Conotoxin GVIA
D002121:Calcium Channel Blockers; D015220:Calcium Channels; D007457:Iodine Radioisotopes; D009414:Nerve Growth Factors; D010455:Peptides; D001464:Barium; D020866:omega-Conotoxin GVIA; D009543:Nifedipine