Characterization of the binding and internalization of tetanus toxin in a neuroblastoma hybrid cell line.

Tetanus toxin is known to bind neuronal tissue selectively. To study the interactions of this potent neurotoxin in an intact cell system, the binding of 125I-tetanus toxin was characterized in a neuroblastoma retina hybrid cell line, N18-RE-105. The binding of 125I-tetanus toxin to membranes prepared from N18-RE-105 cells showed many similarities to the interactions of 125I-toxin with rat synaptic membranes. The binding was decreased with increasing temperature, ionic strength, and pH. 125I-Toxin bound to membranes with high affinity: KD = 0.62 +/- 0.05 nM; Bmax = 196 +/- 45 pmol/mg protein. Quantitative thin-layer chromatography and acid-degradation analysis revealed that N18-RE-105 cells contained polysialogangliosides GD1a and GT1b in high concentrations. An assay was developed to quantitate surface-bound and internalized 125I-tetanus toxin by exploiting the observation that surface-bound 125I-toxin is susceptible to pronase digestion. When cells were incubated with 125I-tetanus toxin at 0 degree C, all of the bound 125I-toxin could be degraded with pronase. In contrast, when the incubations were performed at 37 degrees C, within 10 min about 50% of the total cell-associated 125I-toxin was pronase-resistant. Temperature pulse experiments demonstrated that 125I-tetanus toxin that was bound to cells at 0 degree C rapidly disappeared from the surface when the cells were warmed to 37 degrees C, as revealed by the appearance of pronase-resistant radioactivity. This internalization was sensitive to metabolic inhibitors.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of naphthalene on color changes of the sand fiddler crab, Uca pugilator.

The polycyclic aromatic hydrocarbon, naphthalene, inhibited the melanin dispersion, which normally occurs in the fiddler crab, Uca pugilator, during the dark phase of its circadian rhythm of color change. Naphthalene produced this inhibition no matter whether the melanin was fully aggregated or more or less dispersed at the time of initial exposure. Inhibition of the circadian melanin dispersion does not appear to be due to a direct action of naphthalene on the melanophores. This inhibition was concentration-dependent with acute daily exposure. Naphthalene does not, however, inhibit melanin dispersion in response to a black background, indicating thereby that naphthalene-exposed crabs continue to synthesize melanin-dispersing hormone and norepinephrine.

A mechanism of action for the inhibition of black pigment dispersion in the fiddler crab, Uca pugilator, by naphthalene.

The polycyclic aromatic hydrocarbon, naphthalene, inhibits the circadian dispersion of epidermal black pigment in the fiddler crab, Uca pugilator, by inhibiting the release of black pigment dispersing hormone. Naphthalene caused no permanent neural damage in Uca pugilator. Naphthalene did not cause a chemically-induced phase shift in the circadian rhythm of black pigment dispersion but reduced the daytime peak of that dispersion. Black pigment concentration, which occurs at night, was not affected by exposure to naphthalene. Black pigment dispersing hormone in naphthalene-exposed crabs can be released by an injection of norepinephrine. Given the points above, and previously published data, it is concluded that naphthalene inhibits circadian black pigment dispersion in Uca pugilator by inhibiting the release of the neurotransmitter, norepinephrine.