Microelectrode studies of the active Na transport pathway of frog skin

When the outer surface of short-circuited frog skin was penetrated with microelectrodes, stable negative potentials that averaged near -100 mV were recorded consistently, confirming the results of Nagel (W. Nagel. 1975. Abstracts of the 5th International Biophysics Congress, Copenhagen. P-147.). The appearance of these stable potentials, V(O), concurrent with the observations that (a) a high resistance outer barrier R(O) accounting for approximately 75 percent or more of the transcellular resistance of control skins had been penetrated and that (b) 10(-5) M amiloride and reduced [Na] outside caused the values of V(O) to increase towards means value near -130 mV while the values of percent R(O) increased to more than 90 percent. It was of relationships were the same as the values of E(1) observed in studies of the current-voltage relationships were the same as the values of E’(1) defined as the values of voltage at the inner barrier when the V(O) of the outer barrier was reduced to zero by voltage clamping of the skins. Accordingly, these data are interpreted to mean that the values of E(1), approximately 130 mV, represent the E(Na) of the sodium pump at the inner barrier. 2,4-DNP was observed to decrease the values of transepithelial voltage less than E(1) the V(O) was negative. These data can be interpreted with a simple electrical equivalent circuit of the active sodium transport pathway of the frog skin that includes the idea that the outer membrane behaves as an electrical rectifier for ion transport.     

I-A Mutation resulted in a selective loss of an antigen-specific Ir gene function

The immune responses to several antigens were compared in the I-A mutant mouse strain B6.C-H-2^bm12 and the wild-type strain C57BL/6. With a lymph node cell proliferation assay, the response to two of these antigens, beef insulin and (TG)A-L, was demonstrated to be controlled by a gene in the I-A^b region. B6.C-H-2^bm12 mice failed to respond to beef insulin, while their responses to (TG)A-L, DNP-OVA and PPD were comparable with those of the wild-type strain C57BL/6. Taken together with previous studies, these data suggest that the product of a single pleiotropic I-A gene, an la molecule, functions as a histocompatibility, la, and MLR antigen, as well as a necessary component for Ir gene function. Furthermore, the data reported here demonstrate that la molecules have multiple functional “Ir determinants,” one of which has been altered in the B6.C-H-2^bm12 mutant. The B6.C-H-2^bm12 mice, therefore, represent a powerful analytical tool for the understanding of the cellular and molecular basis for Ir gene control of the immune response.     

Opposition of the vasopressin-induced vasoconstriction in the isolated perfused rat kidney by some prostaglandins

PGE₂ can vasoconstrict or vasolidate the isolated Krebs-perfused rat kidney depending on the tone of the renal vasculature. Thus, it is weakly constrictor (threshold 5–10 ng bolus dose) in the perfused kidney whose perfusion pressure is 47 ± 2 SD mmHg (n = 6), but becomes a vasodilator (threshold ～ 10 pg) in the kidney whose perfusion pressure has been raised to 73 ± 6 SD mmHg (n = 6) or 121 ± 8 SD mmHg (n = 6) through constant infusion of Vasopressin (0.1 and 0.25 mU/ml respectively). PGE₁ was equally effective as PGE₂ while other PGs, I₂, I₁, and 6-keto E₁, were less effective in opposing vasoconstriction. PGF_2α was inactive up to a dose of 10 ng.     

Fibrinolytic activity associated with cultured human neoplastic and normal cells

Summary Fibrinolytic activity was studied in a number of different established as well as secondary human cell cultures derived from both malignant and normal tissues. The ability to degrade [¹²⁵I]-labeled fibrin was found to be characteristic of some malignant cultures as well as some normal cultures, and to be dependent upon the presence of serum. For the most part, this activity was detected in cultures with a relatively shortin vitro passage history (<30 passages). Low passaged colon and rectal carcinoma cells, HCT-8 and HRT-18, as well as normal rectal, colon and foreskin fibroblasts were positive for fibrinolytic activity, while long established (>100 passages) cultures of malignant cells (colon carcinoma, HeLa, Hep-2, KB) as well as normal cells (HEI, AV₃) were negative. It is proposed that although some normal cells synthesize plasminogen activators, the fibrinolytic capability of both malignant and normal cells may be lost on prolongedin vitro cultivation.     

Protection of foreign DNA against host-controlled restriction in bacterial cells

Summary Foreign Flac plasmid DNA which is introduced into potentially restricting E. coli recipient cells can be protected from restriction by preinfecting the recipient cells with UV-inactivated T3 or T7 bacteriophages which express the ocr gene function. The recipient cells survive and are able to replicate themselves as well as the newly acquired plasmid.     

Studies of L-canavanine incorporation into insectan lysozyme.

L-Canavanine is incorporated into the lysozyme synthesized, in response to administration of bacterial cell wall materials, by canavanine-treated larvae of the tobacco hornworm Manduca sexta (Sphingidae). Maximum canavanine incorporation into M. sexta lysozyme occurs when the larvae are provided 1 mg of canavanine g-1 fresh body weight. Analysis of canavanine-containing lysozyme purified from these insects reveals that 21% of the arginine residues are replaced by canavanine; this residue substitution results in a loss of 49.5% of the catalytic activity. When the larvae are provided 0.5 mg of canavanine g-1, 16.5% of the arginine residues are substituted by canavanine and 39.5% of the catalytic activity is lost. Canavanine is also incorporated into the lysozyme induced by canavanine-treated pupae of the giant silk moth Hyalophora cecropia (Saturnidae). In contrast, replacement of 17% of the arginine in H. cecropia lysozyme by canavanine fails to affect the catalytic activity. We have determined the primary structure of M. sexta lysozyme and compared it with the primary structure of H. cecropia lysozyme which has been described elsewhere. M. sexta lysozyme has an arginine at positions 23, 42, and 107. H. cecropia contains serine, lysine, and lysine, respectively, at these locations. The ability of incorporated canavanine to inhibit M. sexta lysozyme activity selectively may result from the fact that replacement of any one of the 3 arginine residues at position 23, 42, or 107 by canavanine causes the loss of catalytic activity.     

Anoxic survival of the isolated cerebellum of the turtle Pseudemis scripta elegans

Intact turtle brain provides a useful model for the study of anoxia and potential survival strategies, since this tissue maintains transmembrane ion gradients and ATP levels during prolonged anoxia and recovers functional activity afterwards. Since isolated tissues offer experimental advantages, the present study sought to determine effects of anoxia on the isolated turtle cerebellum and to define relationships between anoxia survival and glucose supply. In normoxia, the extracellular potassium ([K+]o) activity and evoked potentials were maintained with 5 mM glucose, but 20 mM glucose was required to maintain adenosine triphosphate (ATP) levels and prevent significant increases in [K+]o during anoxia. Inhibition of glycolysis by iodoacetic acid (IAA) during anoxia provoked large increases in [K+]o at all glucose levels. These results demonstrate the usefulness of the isolated turtle cerebellum for studies of anoxic survival since this tissue can maintain ATP levels and [K+]o during prolonged anoxia with 20 mM glucose in the artificial cerebrospinal fluid medium. They also suggest the presence of a Pasteur effect at least during the transition to a hypometabolic state.     

Progestins induce down-regulation of insulin-like growth factor-I (IGF-I) receptors in human breast cancer cells: potential autocrine role of IGF-II.

Insulin-like growth factor-I (IGF-I) receptors are present in breast cancer cells and may play a role in breast cancer cell growth. We have studied the effect of progestins on IGF-I receptors in T47D human breast cancer cells. T47D cells constitutively express high levels of progesterone receptors and are a model for studying the regulation of cellular functions by progestins. Treatment of T47D cells with either progesterone or the synthetic progestin promegestone (R5020) decreased IGF-I receptor content by approximately 50%, as measured by Scatchard analysis and receptor biosynthesis studies. In contrast to progestins, estradiol, dexamethasone, and dihydrotestosterone did not influence IGF-I receptor content. No effect of R5020 was seen after 12 h of incubation, a near-maximal effect was seen after 24 h, and greatest effects were seen after 72 h. R5020 decreased IGF-I receptor mRNA abundance, indicating that progestins acted at the level of gene expression. However, progestins also increased the secretion of IGF-II, a ligand for the IGF-I receptor. In contrast to IGF-II, T47D cells did not express IGF-I. The addition of exogenous IGF-II to T47D cells down-regulated both IGF-I receptor binding and IGF-I receptor mRNA abundance. This study indicates, therefore, that progestins regulate IGF-I receptors in breast cancer cells and suggests that this regulation occurs via an autocrine pathway involving enhanced IGF-II secretion.