The effect of nerve growth factor on the development of sodium channels in PC12 cells

We have studied the development of the action potential Na+ channels in PC12 cells, an established line that has been useful as a model for neuronal differentiation. In continuous culture PC12 cells, although electrically inexcitable, nevertheless have a low level of Na+ channels as judged by the increase in 22Na+ uptake in the presence of veratridine and scorpion toxin. These two neurotoxins have been shown to promote activation of Na+ channels in a variety of electrically excitable cells. Following treatment with nerve growth factor (NGF), conditions which induce differentiation to an electrically excitably neuronal-cell type, the neurotoxin-activated 22Na+ uptake increases approximately 12-fold, on a per cell basis, reaching a maximum in 12-16 days. The dose-response curves for veratridine and scorpion toxin are unchanged by NGF treatment (K0.5 for veratridine, 18-14 microM; K0.5 for scorpion toxin, 120-96 nM). Na+ channels in both undifferentiated and differentiated cells are tetrodotoxin sensitive and NGF treatment has no effect on the inhibition constant (Ki, 10-12 nM). Na+ channel sites were measured directly by the specific binding of [3H]saxitoxin. In NGF-treated cells, the saxitoxin receptor density reaches 154 fmol/mg protein (Kd, 1.3 nM), a level comparable to other excitable cells. Levels in control cells were too low to measure accurately. These findings show that NGF treatment of PC12 cells leads to a substantial increase in the expression of neurotoxin-sensitive Na+ channels. Furthermore, these channels are pharmacologically similar, if not identical, to those which exist in undifferentiated cells and therefore do not appear to result from the conversion of preexisting channels.     

Isolation and elucidation of some functional properties of the "mute" catalytic subunit of cAMP-dependent protein kinase

A mute isoenzyme of type II cAMP-dependent protein kinase from rat muscle has been reported that is released from the regulatory subunit by cAMP but remains inactive until combination with heat- and acid-stable modulator has occurred. This enzyme has now been obtained in isolation free of the normal catalytic subunit using affinity chromatography with both an ATP analog (Blue Dextran/Sepharose) and a protein substrate analog (Kemptide/CH-Sepharose). Separation can be effected in both cases before activation of the mute enzyme. Affinity of the mute enzyme for Blue Dextran--a ligand specific for the dinucleotide fold in this kinase--is somewhat higher than that of the normal enzyme. Conversely, before reaction with the modulatory protein the mute enzyme will not bind at all to Kemptide/CH-Sepharose, where the normal enzyme elutes at 50 mM KCl. When pretreated with the modulatory protein and so activated, mute enzyme binds to Kemptide with a very high affinity and can only be eluted using a natural substrate (phosphorylase kinase), up to 500 mM salt being ineffective. The modulator thus appears to act through alteration of the protein substrate binding site on the enzyme.     

Copper toxicity and copper tolerance in Enteromorphacompressa (L.) Grev.

The responses of ship-fouling and non-fouling isolates of Enteromorpha compressa (L.) Grev. have been compared in media containing copper at 0.0?9.6 μmol · dm^?3. The responses of each isolate were found to vary, according to the conditions of the original habitat. Thus ship-fouling E. compressa was found to be tolerant of copper concentrations up to 9.6 μmol · dm^?3 showing a maintenance of all of the physiological processes studied during the present research (cell viability, net photosynthesis, intracellular K⁺ and dimethylsulphoniopropionate content). Non-fouling plant material showed symptoms of copper toxicity at all levels of copper from 1.8 μmol · dm^?3 to 9.6 μmol · dm^?3. Copper tolerance in ship-fouling E. compressa appears to be genetically determined, since the progeny from ship-fouling plants are also tolerant to copper concentrations within the range 1.8 to 9.6 μmol · dm^?3. The rate of accumulation of copper in ship-fouling thalli is, however, almost identical to that of non-fouling thalli, suggesting that tolerance may be due primarily to internal detoxification, rather than an exclusion mechanism.     

Anti-idiotypic antibodies to HLA-DR4 and DR2

The aim of our study was to determine whether antibodies recognizing epitopes of HLA-DR antigens (idiotypic antibodies or Ab1) induce the production of anti-idiotypic antibodies (Ab2). We tested the capacity of the F(ab')2 fragment obtained from two sera, one with no anti-HLA antibodies (serum ES) and one depleted by absorption of anti-HLA lymphocytotoxins (serum FH), to block the anti-DR antibodies reacting with the HLA-DR antigens of the immunizing donor. The F(ab')2 fragment obtained from serum ES inhibited the anti-DR2 activity of an earlier post-delivery bleeding obtained from the same woman. The anti-idiotypic antibodies contained by this serum also inhibited the anti-DR2 activity of a reference anti-DR2 antiserum 8W907 and of an anti-MT1 antiserum 8W1231. Similarly, the F(ab')2 fragment obtained from serum FH, after absorption of her anti-DR4 antibody, inhibited the anti-DR4 activity of autologous and homologous antisera. These data suggest that sera of parous women contain anti-idiotypic antibodies directed against regulatory idiotypes of anti-DR antibodies.     

Further observations on the effect of air ions on influenza in the mouse

In earlier papers we reported that the course of respiratory infections in mice produced by intranasal instillation of measured amounts of: (1) a fungus, COCCIDIOIDES IMMITIS, (2) a bacterium, KLEBSIELLA PNEUMONIAE and (3) a virus, PR8 strain of influenza virus was materially affected by the air ion environment. When the challenge dose of influenza virus was administered as an aerosol, as described here, the cumulative mortality rate was completely uninfluenced by shifts in the concentration of positive and negative air ions in the ambient atmosphere and by the accompanying electrical fields. A hypothetical mechanism accounting for the different results obtained with intranasal and aerosol challenge is presented.

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Zusammenfassung In früheren Arbeiten wurde über den Verlauf von Infektionen der Atemwege von Mäusen nach intranasaler Instillation bekannter Mengen (1) des Fungus COCCIDIOIDES IMMITIS, (2) KLEBSIELLA PNEUMONIAE und (3) PR8 Influenzavirus berichtet, die durch Luftionen beeinflusst waren. Hier wurde die Dosis von Influenzavirus als Aerosol verabreicht. Die kumulative Mortalitätsrate wurde durch den Wechsel der Konzentrationen von positiven und negativen Luftionen in der Umgebungsatmosphäre und begleitenden elektrischen Feldern nicht beeinflusst. Eine Erklärung für die unterschiedlichen Ergebnisse bei intranasaler und Aerosol-Virusapplikation wird diskutiert.

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Resume Dans des travaux précédents, on a montré que l'évolution de maladies du système respiratoire provoquées chez des souris par l'instillation intranasale de doses déterminées d'agents pathogènes était affectée de façon significative par le taux d'ionisation de l'air ambiant. Il s'agissait d'agents cryptogamiques (COCCIDIOIDES IMMITIS), bactériens (KLEBSIELLA PNEUMONIAE) et de virus (lignée PR8 du virus de la grippe). Lorsque la dose minimum de virus grippal est appliquée sous forme d'aérosole — selon la méthode décrite ici — le taux cumulatif de mortalité n'est aucunement influencé par les variations de concentration d'ions positifs ou négatifs de l'air ambiant ni par les champs électriques qui les accompagnent. On développe une hypothèse pour expliquer la diversité des résultats obtenus au moyen des infectations intranasales ou par aérosoles.

     

“Killer Character” of Saccharomyces cerevisiae: Curing by Growth at Elevated Temperature

Normal "killer" strains of Saccharomyces cerevisiae, when grown at 37 to 40 C, produce almost exclusively nonkiller cells due to loss or mutation of at least part of the non-chromosomal killer genome.     

Mutants of Saccharomyces cerevisiae That Incorporate Deoxythymidine-5′-Monophosphate Into Deoxyribonucleic Acid In Vivo

Spontaneous mutants of Saccharomyces cerevisiae able to incorporate deoxythymidine-5′-monophosphate (dTMP) into deoxyribonucleic acid (DNA) have been selected based on their ability to grow in the presence of aminopterin and sulfanilamide if dTMP is present. Essentially all mutants (called tup) selected in this way required dTMP for growth in the presence of the two drugs, but none required dTMP in the absence of the drugs. Neither thymine nor thymidine would satisfy this requirement. Equimolar amounts of ³²P- and ³H-base-labeled dTMP were incorporated by the mutants into alkali-stable, deoxyribonuclease-sensitive material. In the presence of aminopterin and sulfanilamide, this incorporation was sufficient to account for a substantial proportion of the thymine residues in the cellular DNA, whereas in the absence of the drugs only about 40% as much of the thymine residues originated from the medium. Of 29 mutants examined, all were recessive and 17 showed 2:2 segregation in crosses with a wild-type strain. The lesions in these mutants fell into four complementation groups: one (tup1) occurs on chromosome III; another (tup3) is on chromosome II; and a third (tup4) was centromere linked. Strains of the genotype α tup1 mated with lower than normal efficiency with a strains, but with higher than normal efficiency with α strains. Strains of genotype a/α tup1/tup1 failed to sporulate, whereas homozygous diploids for tup2, tup3, or tup4 sporulated normally, as did a/α tup1/+ strains.