Abstract
The long-term modulation of calcium (Ca2+) currents (I Ca) was studied in 108CC15 neuroblastoma × glioma hybrid (N×G) cells grown under various culture conditions. The following results were obtained:
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1.
Addition of 1 mM dibutyryl cyclic adenosine monophosphate (db-cAMP) or 0.1 μM forskolin to the culture medium increased a transient component of I Ca two-fold within 3 days, from 21.0±1.6 pA/pF (n=22) to a maximum of 40.0±2.6 pA/pF (n=28). Under these conditions, cells also expressed a slowly inactivating I Ca component (maximum after 3 days, 20.5±1.6 pA/pF, n=28).
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2.
The fast inactivating I Ca as well as the db-cAMP-induced slowly inactivating I Ca were completely down-regulated during incubation of NxG cells with the inorganic Ca2+ channel blocker, nickel (Ni2+, 100 μM). The suppressing effect was reversed within 3 days of incubation in db-cAMP-containing medium lacking Ni2+.
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3.
Binding studies on membrane preparations of control and Ni2+-pretreated NxG cells revealed a marked difference in the maximal (+)3H-PN200-110 binding. The difference was seen in undifferentiated as well as in dbc-AMP-incubated cells.
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4.
The protein synthesis blocker, cycloheximide, suppressed both the db-cAMP-induced increase and the reappearance of I Ca following Ni2+ pretreatment.
It is suggested that chronic application of db-cAMP or Ni2+ to NxG cells increases and decreases the number of Ca2+ channel proteins, respectively.
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References
Barish ME (1986) Differentiation of voltage-gated potassium current and modulation of excitability in cultured amphibian spinal neurons. J Physiol (Lond) 375:229–250
Bodewei R, Hering S, Schubert B, Wollenberger A (1985) Sodium and calcium currents in neuroblastoma x glioma hybrid cells before and after morphological differentiation by dibutyryl cyclic AMP. Gen Physiol Biophys 4:113–127
Brown D, Higashida H (1988) Voltage- and calcium-activated potassium currents in mouse neuroblastoma × rat glioma hybrid cells. J Physiol (Lond) 397:149–165
Brown D, Docherty RJ, McFadzean I (1989) Calcium channels in vertebrate neurons. Ann NY Acad Sci 560:358–372
De Lean A, Munson PJ, Rodbard D (1978) Simultaneous analysis of families of sigmoidal curves: application to bioassay, radioligand assay, and physiological dose-response curves. Am J Physiol 235:E97-E102
Docherty RJ (1988) Gadolinium selectively blocks a component of calcium current in rodent neuroblastoma x glioma hybrid (NG108-15) cells. J Physiol (Lond) 398:33–47
Glossmann H, Ferry DR (1985) Assay for calcium channels. Methods Enzymol 109:513–550
Hamill OP, Marty A, Neher E, Sakmann B, Sigworth FJ (1981) Improved patch-clamp techniques for high-resolution current recording from cells and cell-free membrane patches. Pflügers Arch 391:85–100
Hamprecht B, Glaser T, Reiser G, Bayer E, Probst F (1985) Culture and characteristics of hormone-responsive neuroblastoma × glioma hybrid cells. Methods Enzymol 109:316–341
Hescheler J, Rosenthal W, Hinsch KD, Wulfern M, Trautwein W, Schultz G (1988) Angiotensin II-induced stimulation of voltage dependent Ca2+ currents in an adrenal cortical cell line. EMBO J 7:619–624
Klee WA, Nirenberg M (1974) A neuroblastoma × glioma hybrid cell line with morphine receptors. Proc Natl Acad Sci USA 71:3474–3477
Maurer R (1989) Both isoforms of the cAMP-dependent protein kinase catalytic subunit can activate transcription of the prolactin gene. J Biol Chem 264:6870–6873
McGee R, Simpson P, Nirenberg M (1978) Regulation of acetylcholine release from neuroblastoma x glioma hybrid cells. Proc Natl Acad Sci USA 75:1314–1318
Montminy M, Bilezikjian L (1987) Binding of a nuclear protein to the cyclic-AMP response element of the somatostatin gene. Nature 328:175–178
Mullaney I, Magee AI, Unson CG, Milligan G (1988) Differential regulation of amounts of the guanine-nucleotide-binding proteins Gi and G0 in neuroblastoma × glioma hybrid cells in response to dibutyryl cyclic AMP. Biochem J 256:649–656
Nirenberg M, Wilson S, Higashida H, Rotter A, Krueger K, Busis N, Ray R, Kenimer JG, Adler M (1983) Modulation of synapse formation by cyclic adenosine monophosphate. Science 222:794–799
Peterson GL (1983) Determination of total protein. Methods Enzymol 91:95–119
Riabowol KT, Fink JS, Gilman MZ, Walsh DA, Goodman RH, Feramisco JR (1988) The catalytic subunit of cAMP-dependent protein kinase induces expression of genes containing cAMP-responsive enhancer elements. Nature 336:83–86
Rosenthal W, Koesling D, Rudolph U, Kleuss C, Pallast M, Schultz G (1986) Identification and characterization of the 35-kDa β-subunit of guanine nucleotide-binding proteins by an antiserum raised against transducin. Eur J Biochem 158:255–263
Seamon K, Daly J (1986) Forskolin: its biological and chemical properties. Adv Cyclic Nucleotide Protein Phosphorylation Res 20:1–150
Trautwein W, Hescheler J (1990) Regulation of cardiac L-type calcium current by phosphorylation and G proteins. Annu Rev Physiol 52:257–274
Usowicz MM, Porzig H, Becker C, Reuter H (1990) Differential expression by nerve growth factor of two types of Ca channels in rat phaeochromocytoma (PC12) cell lines. J Physiol (Lond) 426:95–116
Waterman M, Murdoch GH, Evans RD, Rosenfeld MG (1985) Cyclic AMP regulation of eukaryotic gene transcription by two discrete molecular mechanisms. Science 229:267–269
Vazquez D (1974) Inhibitors of protein synthesis. FEBS Lett 40:S63-S84
Yaari Y, Hamon B, Lux HD (1987) Development of two types of calcium channels in cultured mammalian hippocampal neurons. Science 235:680–682
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Eckert, R., Hescheler, J., Krautwurst, D. et al. Calcium currents of neuroblastoma × glioma hybrid cells after cultivation with dibutyryl cyclic AMP and nickel. Pflugers Arch. 417, 329–335 (1990). https://doi.org/10.1007/BF00371000
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DOI: https://doi.org/10.1007/BF00371000