scholarly journals Comparison of rapid changes in surface morphology and coated pit formation of PC12 cells in response to nerve growth factor, epidermal growth factor, and dibutyryl cyclic AMP.

1984 ◽  
Vol 98 (2) ◽  
pp. 457-465 ◽  
Author(s):  
J L Connolly ◽  
S A Green ◽  
L A Greene
Keyword(s):  
Nerve Growth Factor ◽  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Cyclic Amp ◽  
Cell Surface ◽  
Dibutyryl Cyclic Amp ◽  
Coated Pits ◽  
Pit Formation ◽  
Epidermal Growth ◽  
Surface Changes

Scanning and transmission electron microscopic studies were carried out on the rapid cell surface response of PC12 pheochromocytoma cells to treatment with nerve growth factor (NGF), epidermal growth factor (EGF), and dibutyryl cyclic AMP. EGF induced a rapidly initiated series of surface changes identical to those previously observed with NGF. Ruffles appear over the dorsal surface of the cells by 30 s, are prominent at 3 min, and are absent by 7 min. Microvilli disappear as dorsal ruffles become prominent. Peripheral ruffles are seen by 3 min, are prominent on most of the cells by 7 min, and are virtually absent by 15 min. Large blebs are present on 50% of the cells by 2 h and are markedly decreased by 4 h. Within 30 s after NGF or EGF addition, an increase in the density of 60-130-nm coated pits per unit membrane is detectable. This reaches a maximum of two- to threefold in from 1 to 3 min and gradually decreases. Combined treatment with NGF and EGF increases surface ruffling and, after an early peak in coated pits which at 3 min is similar in magnitude to that observed for the separately administered factors, maintains a greater number of pits per unit area than either treatment alone. 3-d pretreatment with NGF greatly reduces the response of the cells to EGF both with respect to surface ruffling and coated pit formation while 4-h NGF pretreatment has no effect on the EGF response. Dibutyryl cyclic AMP induced none of the rapidly onsetting changes caused by NGF or EGF, and therefore it seems unlikely that cyclic AMP mediates these surface changes. Changes in cell surface architecture induced by NGF and EGF on PC12 cells and by NGF in normal sympathetic neurons (as previously described) indicates that such responses may be a widespread phenomenon associated with the interaction of at least some peptide growth factors/hormones with their receptors. These responses may represent or reflect primary events in the mechanism by which these factors act.

scholarly journals Differential inhibition of nerve growth factor and epidermal growth factor effects on the PC12 pheochromocytoma line.

1984 ◽  
Vol 98 (2) ◽  
pp. 417-426 ◽  
Author(s):  
P J Seeley ◽  
A Rukenstein ◽  
J L Connolly ◽  
L A Greene
Keyword(s):  
Nerve Growth Factor ◽  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Cell Surface ◽  
Surface Morphology ◽  
Neurite Outgrowth ◽  
Decarboxylase Activity ◽  
Differential Inhibition ◽  
Epidermal Growth ◽  
Cell Surface Morphology

Tests have been made of the action of the methyltransferase inhibitors 5'-S-methyl adenosine, 5'-S-(2-methyl-propyl)-adenosine, and 3-deaza-adenosine +/- L-homocysteine thiolactone, on nerve growth factor (NGF)-dependent events in the rat pheochromocytoma line PC12. Each of these agents inhibited NGF-dependent neurite outgrowth at concentrations of the order of millimolar. Slow initiation of neurite outgrowth over several days and more rapid regeneration of neurites (congruent to 1 d) were blocked, as was the priming mechanism necessary for genesis of neurites. The inhibitions were reversible in that PC12 cells maintained for several days in the presence of inhibitors grew neurites normally after washout of these agents. Other NGF-dependent responses of the PC12 line (i.e., induction of ornithine decarboxylase activity [over 4 h], enhancement of tyrosine hydroxylase phosphorylation [over 1 h], and rapid changes in cell surface morphology [30 s onward]) were inhibited by each of the agents. In contrast, corresponding epidermal growth factor-dependent responses in ornithine decarboxylase activity, phosphorylation, and cell surface morphology were not blocked, but instead either unaffected or enhanced, by the methylation inhibitors. These inhibitors did not act by blockade of binding of NGF to high- or low-affinity cell surface receptors, though they partially inhibited internalization of [125I]NGF. The inhibition of rapidly-induced NGF-dependent events and the differential inhibition of responses to NGF and epidermal growth factor imply that the methyltransferase inhibitors specifically block one of the first steps in the mechanistic pathway for NGF.

scholarly journals Pit formation and rapid changes in surface morphology of sympathetic neurons in response to nerve growth factor.

1981 ◽  
Vol 90 (1) ◽  
pp. 176-180 ◽  
Author(s):  
J L Connolly ◽  
S A Green ◽  
L A Greene
Keyword(s):  
Nerve Growth Factor ◽  
Growth Factor ◽  
Cell Surface ◽  
Sympathetic Neurons ◽  
Cell Types ◽  
Coated Pits ◽  
Nerve Growth ◽  
Pheochromocytoma Cells ◽  
Pit Formation ◽  
Transmission Electron

Scanning and transmission electron microscope studies were carried out on the rapid cell surface responses of cultured newborn rat sympathetic neurons to nerve growth factor (NGF), a substance that promotes their survival and differentiation. The somas of sympathetic neurons continuously exposed to NGF or deprived of the factor for 4-5 h have a very smooth surface. After readdition of NGF to the latter type of cultures, there is rapidly initiated a transient, sequential change in the cell surface. Microvilli and small ruffles appear within 30 s and are most prominent by 1 min. By 3 min of exposure, the microvilli and ruffles decrease in prominence, and by 7 min the somal surface is again smooth. By 30 s after NGF readdition, as increase in the number of 60- tp 130-nm coated pits is also detectable. This increase reaches a maximum of about threefold from 0.5 to 3 min and then gradually decreases. Alterations in the surface did not occur on the nonneuronal cell types present in the cultures and were not observed in response to another basic protein (cytochrome c) or to physical manipulation. Changes in cell surface architecture induced by NGF in normal sympathetic neurons and, as previously described, in PC12 pheochromocytoma cells indicate that such responses may present or reflect primary events in the mechanism of the factor's action.

Epidermal growth factor increases inhibin synthesis by isolated segments of rat seminiferous tubules

1989 ◽  
Vol 123 (2) ◽  
pp. 213-219 ◽  
Author(s):  
G. F. Gonzales ◽  
G. P. Risbridger ◽  
D. M. de Kretser
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Cyclic Amp ◽  
Phorbol Esters ◽  
Seminiferous Tubules ◽  
Dibutyryl Cyclic Amp ◽  
Adult Rats ◽  
Adult Rat ◽  
Epidermal Growth

ABSTRACT The effect of epidermal growth factor (EGF) on the production of immunoreactive inhibin by adult rat isolated seminiferous tubules in vitro has been investigated. EGF (0·1–1000 ng/ml) added to cultures of seminiferous tubules from adult rats caused a dose-dependent increase in inhibin content in the tubules without changing the amount secreted into the media. However, after continuous stimulation with EGF for periods in excess of 5 days, an increase in inhibin secretion was observed. In the presence of 10 and 100 ng FSH/ml, EGF (10 ng/ml) produced a further increment in the inhibin content of the tubules, but this effect was not found with FSH concentrations of 500 or 1000 ng/ml. EGF also increased the tubule content of inhibin after the addition of 100 μg dibutyryl cyclic AMP/ml but no effect of EGF was observed on the FSH- or dibutyryl cyclic AMP-induced secretion of inhibin into the medium. The effect of EGF on inhibin content in the tubules was partially suppressed by the addition of 4β-phorbol-12β-myristate-13α-acetate (20 ng/ml). Insulin (1–100 ng/ml) decreased basal inhibin secretion without changing the inhibin content of tubules and this effect was antagonized by EGF (10 ng/ml) with insulin doses of 1–50 ng/ml whereas, at 100 ng/ml, the effect of EGF on tubule inhibin content was reversed. The addition of EDTA (2 mmol/l) resulted in an inhibition of basal and EGF-induced inhibin production. These data demonstrate a stimulatory effect of EGF on inhibin production by isolated seminiferous tubules which is inhibited by insulin and phorbol esters, both stimulators of protein kinase C activity. Journal of Endocrinology (1989) 123, 213–219

scholarly journals Structure of the gene encoding VGF, a nervous system-specific mRNA that is rapidly and selectively induced by nerve growth factor in PC12 cells.

1991 ◽  
Vol 11 (5) ◽  
pp. 2335-2349 ◽  
Author(s):  
S R Salton ◽  
D J Fischberg ◽  
K W Dong
Keyword(s):  
Nervous System ◽  
Nerve Growth Factor ◽  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Cyclic Amp ◽  
Pc12 Cells ◽  
Cdna Clone ◽  
Neural Differentiation ◽  
Epidermal Growth ◽  
Specific Mrna

Nerve growth factor (NGF) plays a critical role in the development and survival of neurons in the peripheral nervous system. Following treatment with NGF but not epidermal growth factor, rat pheochromocytoma (PC12) cells undergo neural differentiation. We have cloned a nervous system-specific mRNA, NGF33.1, that is rapidly and relatively selectively induced by treatment of PC12 cells with NGF and basic fibroblast growth factor in comparison with epidermal growth factor. Analysis of the nucleic acid and predicted amino acid sequences of the NGF33.1 cDNA clone suggested that this clone corresponded to the NGF-inducible mRNA called VGF (A. Levi, J. D. Eldridge, and B. M. Paterson, Science 229:393-395, 1985; R. Possenti, J. D. Eldridge, B. M. Paterson, A. Grasso, and A. Levi, EMBO J. 8:2217-2223, 1989). We have used the NGF33.1 cDNA clone to isolate and characterize the VGF gene, and in this paper we report the complete sequence of the VGF gene, including 853 bases of 5' flank revealed TATAA and CCAAT elements, several GC boxes, and a consensus cyclic AMP response element-binding protein binding site. The VGF promoter contains sequences homologous to other NGF-inducible, neuronal promoters. We further show that VGF mRNA is induced in PC12 cells to a greater extent by depolarization and by phorbol-12-myristate-13-acetate treatment than by 8-bromo-cyclic AMP treatment. By Northern (RNA) and RNase protection analysis, VGF mRNA is detectable in embryonic and postnatal central and peripheral nervous tissues but not in a number of nonneural tissues. In the cascade of events which ultimately leads to the neural differentiation of NGF-treated PC12 cells, the VGF gene encodes the most rapidly and selectively regulated, nervous-system specific mRNA yet identified.

Developing Concepts in Receptor Research

1983 ◽  
Vol 17 (5) ◽  
pp. 357-366 ◽  
Author(s):  
Pedro Cuatrecasas
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Cell Surface ◽  
Cell Biology ◽  
Protein Chemistry ◽  
Coated Pits ◽  
Surface Receptors ◽  
Epidermal Growth ◽  
Receptor Research ◽  
Receptor Assays

Recent advances in molecular biology and protein chemistry have permitted spectacular progress in understanding the chemistry and cell biology of cell surface receptors for hormones and drugs. Methodological approaches, such as the use of radiolabeled ligands and direct receptor assays, have permitted the characterization, categorization, and purification of many receptors. The knowledge gained in understanding fundamental hormone-receptor interactions can form the basis of future, rational new drug design. Another important advance relates to the dynamic nature of receptors in their membrane environment. Upon binding of hormones such as insulin and epidermal growth factor, the complexes rearrange topographically on the cell surface, forming microclusters that are internalized into “receptosomes” via receptor-mediated endocytosis (which utilizes coated pits). The internalized receptor-hormone complex can have various fates, including the generation of selective signals for controlling cell growth and differentiation. In at least one case (epidermal growth factor), the latter may be dependent on the processes that occur in an acidic endosomal compartment within the cell.

Effect of epidermal growth factor on the membrane potential of cultured porcine thyroid cells

1986 ◽  
Vol 109 (3) ◽  
pp. 321-324 ◽  
Author(s):  
J. R. Bourke ◽  
P. A. McGrath ◽  
G. J. Huxham ◽  
M. J. Waters ◽  
S. W. Manley
Keyword(s):  
Membrane Potential ◽  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Cyclic Amp ◽  
Follicle Cells ◽  
Dibutyryl Cyclic Amp ◽  
Thyroid Cells ◽  
Epidermal Growth

ABSTRACT Cultured porcine thyroid cells maintained in media containing TSH exhibited a membrane potential of −50 mV, and hyperpolarized by about 10 mV within 1 h of the addition of epidermal growth factor (EGF; 10 ng/ml). Follicle cells had depolarized to −45 mV after 4 h of exposure to EGF. Cells maintained in dibutyryl cyclic AMP (dbcAMP) did not alter their membrane potential when exposed to EGF for up to 4 h. Cultures washed to remove the TSH or dbcAMP hyperpolarized to − 75 mV within 30 min, and a reversible depolarization to − 60 mV was observed on addition of EGF. It was concluded that EGF acts as a physiological antagonist of TSH and also exerts a separate depolarizing influence on cultured thyroid cells. J. Endocr. (1986) 109, 321–324

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