Selective survival of neurons from chick embryo sensory ganglionic dissociates utilizing serum-free supplemented medium

Bottenstein, Jane E.; Skaper, Stephen D.; Varon, Silvio; Sato, Gordon

doi:10.1016/0014-4827(80)90202-5

Cited by 426 publications

(158 citation statements)

References 11 publications

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“…For experiments, the cells were washed with serum-free DMEM/N1 [Dulbecco's modified Eagle's medium (DMEM; Gibco, Grand Island, NY); N1 ϭ 0.25% BSA; 6.25 ϫ 10 Ϫ8 M transferrin; 8.3 ϫ 10 Ϫ7 M insulin; 3 ϫ 10 Ϫ8 M selenium; 2 ϫ 10 Ϫ8 M progesteron; 1 ϫ 10 Ϫ4 M putrescine (all from Sigma Immunochemicals; Munich, Germany); 50 g/ml penicillin; 50 g/ml streptomycin; 100 g/ml neomycin; according to Bottenstein et al (1980)] and detached by incubating with a trypsin/EDTA solution [0.125%/0.05% (w/v); Biochrom KG] in PBS for 5 min at 37C. The cells were harvested with DMEM/N1 containing 1 mg/ml soybean trypsin inhibitor (Serva; Heidelberg, Germany) and centrifuged at 70 ϫ g for 5 min (Beckman TJ-6).…”

Section: Cell Culturementioning

confidence: 99%

Induction of Tubular Peroxisomes by UV Irradiation and Reactive Oxygen Species in HepG2 Cells

Schrader¹,

Wodopia

Hd³

1999

J Histochem Cytochem.

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SUMMARY Peroxisomes in the human hepatoblastoma cell line HepG2 exhibit a high degree of plasticity. Whereas in confluent cultures they appear as small (0.1-0.3 m) spherical particles, they undergo dramatic changes, forming elongated tubules measuring up to 5 m on separation of cells and cultivation at low density. We recently showed that several growth factors, including nerve growth factor (NGF), induce the formation of tubular peroxisomes and that this induction is sensitive to K 252b, a specific tyrosine kinase inhibitor, suggesting the involvement of this signal transduction pathway. Because tyrosine kinase is also involved in signal transduction via the reactive oxygen species (ROS), we have analyzed in this study the effects of UV irradiation, H 2 O 2 , and oxygen on tubulation of peroxisomes. UVC irradiation induced a significant increase in formation of tubular peroxisomes (40-50% of cells) and this effect was dose-dependently inhibited by pretreatment with N-acetyl cysteine, confirming the involvement of ROS in the UV effect. Furthermore, H 2 O 2 also directly induced the tubulation of peroxisomes, although to a lesser extent. Finally, cultivation under hypoxic conditions (1.5% O 2 ) drastically reduced the inducing effect of fetal calf serum on tubulation of peroxisomes, suggesting the involvement of oxygen-mediated signaling. Taken together, our observations indicate that ROS induce the tubulation of peroxisomes in HepG2 cells. Because peroxisomes harbor most of the enzymes for catabolism of ROS, the tubulation and expansion of the peroxisome compartment could have a cell rescue function against the destructive effects of ROS.

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Section: Cell Culturementioning

confidence: 99%

Induction of Tubular Peroxisomes by UV Irradiation and Reactive Oxygen Species in HepG2 Cells

Schrader¹,

Wodopia

Hd³

1999

J Histochem Cytochem.

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“…After 12 h, defined F14 medium was added, containing 2 mM-glutamine, penicillin (50 i.u. ml-') and streptomycin (50 ,ug ml-'), nerve growth factor (7 5 ,ug rhl-'), insulin (5 ,ug ml-'), transferrin (5 ,ug ml-'), putrescine (0-1 mM), progesterone (0-2 lAm) and selenium (30 nM); this was changed every 3-4 days (Bottenstein, Skaper, Varon & Sato, 1980). If cells were to be kept for more than 1 week, cytosine arabinoside (10-5 M) was added after 36 h in culture, for 48 h, to reduce non-neuronal cell proliferation, although this was slow in the defined medium used.…”

Section: Tissue Culturementioning

confidence: 99%

Ca2+ channel currents in rat sensory neurones: interaction between guanine nucleotides, cyclic AMP and Ca2+ channel ligands.

Dolphin

1991

The Journal of Physiology

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SUMMARY1. The characteristics have been examined of the high threshold calcium channel current in cultured rat dorsal root ganglion (DRG) neurones recorded in the presence of guanosine-5'-O-(3-thiotriphosphate) (GTPyS; 200 ,UM in the patch pipette). This current, termed IBa, GTPyS' was slowly activating and showed little inactivation over 100 ms.2. External application of forskolin (10 ,UM) to elevate internal cyclic AMP levels increased the amplitude of IBa, GTPyS whereas it had no effect on the control IBa. This effect was prevented by inclusion in the patch pipette of the peptide inhibitor of cyclic AMP-dependent protein kinase (PKI; 25 ,tM). 5. The 'agonist' response of IBa, GTPyS to D600 (10 fM) was also occluded by application of forskolin (10 /,M) in the patch pipette. Forskolin alone, applied in this manner, increased IBa, GTPyS to a similar extent to D600 applied alone.6. The agonist effect of ( + )-202-791 (5 /tM) on IBa, GTPyS was not prevented by prior enhancement with forskolin, nor was it prevented by PKI.7. In conclusion, internal GTPyS activates G proteins which may interact directly with calcium channels to influence the kinetics of activation and to reduce steadystate inactivation of the channels. There is also an indirect effect on the generation

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“…Chick embryos [stage 17-18;Hamburger and Hamilton (1951)] were dissected as described previously (Oakley and Tosney, 1993). Small explants were aspirated from either the anterior or posterior somite with a flame-polished electrode (tip diameter, 50 m), washed in neuron media (NM) composed of Ham's F12 (Life Technologies, Grand Island, N Y) supplemented with 10% horse serum, antibiotics, and hormone additives (Bottenstein et al, 1980), plated on polyornithine/ laminin-coated (Life Technologies) glass coverslips, and maintained at 37°C, 5% C O 2 overnight. Culture purity was verified by lectin binding and immunocytochemistry as in Oakley and Tosney (1993).…”

Section: Cell Culturementioning

confidence: 99%

Contact with Isolated Sclerotome Cells Steers Sensory Growth Cones by Altering Distinct Elements of Extension

Steketee¹,

Tosney

1999

J. Neurosci.

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During pathfinding, growth cones respond to guidance cues by altering their motility. This study shows that motile responses can be highly specific: filopodial contact with two different, physiologically relevant cells differentially alters discrete elements of motility. With each cell type, the responses to contact are invariant. Each cell induces a distinct response in sensory growth cones with every filopodial contact. Contact with an inhibitory cell, posterior sclerotome, alters a discrete motile characteristic; contact locally inhibits the ability of veils to extend down contacting filopodia. The inhibition is precise. Contact fails to alter other individual veil characteristics such as initiation frequency or extension rate. Moreover, despite local veil inhibition, the general level of extension across the growth cone is retained, as though protrusive activity is regulated to some set point. Contact with a stimulatory cell, anterior sclerotome, elicits a biphasic response. First, contact stimulates extension generally, altering the set point of protrusion. Contact increases veils and filopodia throughout the growth cone persistently. Then contacting processes consolidate, forming neurite. Filopodia contacting either cell type have similar lifetimes but different fates. Filopodia contacting posterior cells show morphological indications of structural instability, likely related to their inability to support veil extension. Filopodia contacting anterior cells branch, become morphologically complex, and ultimately consolidate into neurite. The invariance and precision of these responses suggests they are the steering components elicited by contact. These steering components, when integrated with other motile events, modulate growth cone trajectory. The discreteness of these responses suggests that guidance cues affect equally discrete elements in signaling cascades.

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Selective survival of neurons from chick embryo sensory ganglionic dissociates utilizing serum-free supplemented medium

Cited by 426 publications

References 11 publications

Induction of Tubular Peroxisomes by UV Irradiation and Reactive Oxygen Species in HepG2 Cells

Induction of Tubular Peroxisomes by UV Irradiation and Reactive Oxygen Species in HepG2 Cells

Ca2+ channel currents in rat sensory neurones: interaction between guanine nucleotides, cyclic AMP and Ca2+ channel ligands.

Contact with Isolated Sclerotome Cells Steers Sensory Growth Cones by Altering Distinct Elements of Extension

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