A study of the metabolism of amphibian neural crest cells during their migration and pigmentation in vitro

Flickinger, R.A.

doi:10.1002/jez.1401120306

Cited by 111 publications

(8 citation statements)

References 12 publications

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“…After removal of the jelly coat and vitelline membrane with fine forceps, pairs of blastomeres were isolated from axolotl (Ambystoma rnexicanum) or anuran (Xenopus laevis and Rana pipiens) mid-cleavage stage embryos (stages 6-8;Harrison, 1969) with glass dissecting needles. Cell pairs were placed in physiological saline (Niu-Twitty solution; Flickinger, 1949) containing 58.2 mM NaCl, 0.7 mM KCI, 0.3 mM Ca(NO3)2, 0.8 mM MgSO4, 0.8 mM Na2HPO4, 0.1 mM KH2PO4, and 0.4 mM NaHCO2 adjusted to pH 7.8 with NaOH, to which was added up to 0.1% colchicine to inhibit mitosis. After isolation cell pairs were allowed to stabilize for at least 0.5 h before experimentation was started.…”

Section: Methodsmentioning

confidence: 99%

Equilibrium properties of a voltage-dependent junctional conductance.

Spray¹,

Harris²,

Bennett³

1981

The Journal of General Physiology

378

207

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The conductance of junctions between amphibian blastomeres is strongly voltage dependent. Isolated pairs of blastomeres from embryos of Ambystoma mexicanum, Xenopus laevis, and Rana pipiens were voltage clamped, and junctional current was measured during transjunctional voltage steps. The steady-state junctional conductance decreases as a steep function of transjunctional voltage of either polarity. A voltage-insensitive conductance <5% of the maximum remains at large transjunctional voltages. Equal transjunctional voltages of opposite polarities produce equal conductance changes. The conductance is half maximal at a transjunctional voltage of -15 inV. Thej unctional conductance is insensitive to the potential between the inside and outside of the cells. The changes in steady-state junctional conductance may be accurately modeled for voltages of each polarity as arising from a reversible two-state system in which voltage linearly affects the energy difference between states. The voltage sensitivity can be accounted for by the movement of about six electron charges through the transjunctional voltage. The changes in junctional conductance are not consistent with a current-controlled or ionic accumulation mechanism. We propose that the intramembrane particles that comprise gap junctions in early amphibian embryos are voltage-sensitive channels.

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Section: Methodsmentioning

confidence: 99%

Equilibrium properties of a voltage-dependent junctional conductance.

Spray¹,

Harris²,

Bennett³

1981

The Journal of General Physiology

378

207

View full text Add to dashboard Cite

show abstract

“…Pairs of blastomeres from early cleavage stages (stages 6 to 8; Harrison, 1969) of Ambystoma mexicanum embryos were isolated mechanically and maintained in physiological saline (Flickinger, 1949; Niu-Twitty solution: 58.2 mM NaCl, 0.7 mM KCl, 0.3 mM Ca(NO&, 0.8 mM MgS04, 0.8 mM Na2HP04, 0.1 mM KH%POd, and 0.4 mM NaHC02, adjusted to pH 7.8 with NaOH) as previously described (Spray et al, 1981a). Mitosis was blocked with up to 0.05% colchicine.…”

Section: Methodsmentioning

confidence: 99%

Control of intercellular communication by voltage dependence of gap junctional conductance

Harris

Spray²,

Bennett³

1983

J. Neurosci.

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The junctional conductance between coupled amphibian blastomeres exhibits a high degree of voltage dependence, as previously described in voltage clamp studies (Spray, D. C., A. L. Harris, and M. V. L. Bennett (1981) J. Gen. Physiol. 77: 77-95; Harris, A. L., D. C. Spray, and M. V. L. Bennett (1981) J. Gen. Physiol. 77: 95-117). The present study examines the properties which this voltage dependence confers on electrotonic coupling between cells. The effects of applied pulses and ramps of current are studied experimentally and are modeled by calculation. During sufficiently large current pulses applied to one cell of a pair, the cells uncouple and then recouple after termination of the pulses. Ramps of current applied to one of the cells can give voltage-current (V-I) relations with a region of hysteresis within which the cells are stably coupled or stably uncoupled depending on previous history. Intrinsically generated currents are able to cause bistability of coupling in the absence of externally applied current. Calculations from the parameters of junctional conductance defined under voltage clamp fully account for these findings and illustrate how junctional and nonjunctional conductances affect the V-I relations in the region of bistability. Recordings from several cells within a small group show that boundaries of intercellular communication can be altered by applied current, a finding that also can be accounted for by voltage dependence of junctional conductance. The "Appendix" examines quantitatively the criteria required for bistability of coupling and the relevance of bistability for intercellular signaling. The plasticity of coupling which the voltage dependence of junctional conductance confers on cells offers an intriguing mechanism by which patterns of intercellular communication could be determined and changed in developing tissues.

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“…At the early blastula stage, embryos were transferred into 20% MMR and cultured overnight at 14°C to stage 10. The resulting early gastrulae were placed in Flickinger medium (Flickinger, 1949) containing 25 µg/ml gentamycin, the vitelline membranes were removed by using sharpened forceps, and animal caps were dissected out by using electrolytically sharpened tungsten needles. Explants were cultured individually in 1% agarose-coated wells of 96-well plates, in 100 µl Flickinger medium containing a range of concentrations of retinoic acid, at 20°C until control embryos reached stage 23-27.…”

Section: Explantsmentioning

confidence: 99%