Vasopressin-enhanced urea transport by rat inner medullary collecting duct cells in culture

Abstract. The distal inner medullary collecting duct (IMCD) is critical in the urinary concentrating process, in part because it is the site of vasopressin (AVP)-regulated permeability to urea. The purpose of these experiments was to develop a cell culture model of the IMCD on permeable structure and to characterize the responsiveness to AVP. Rat IMCD cells were grown to confluence on collagen-coated Millipore filters glued onto plastic rings. To assess the time required to achieve confluence, the transepithelial resistance was measured periodically and was found to be stable after 2 weeks, at a maximal value of 595 ± 22 ω cm². In separate monolayers the effect of AVP on inulin and urea permeability was determined. While inulin permeability was unchanged after AVP, urea permeability increased from 6.0 ± 0–4 to peak values of 16.0 ± 3–8(10nM),23.1 ± 3–9(1 μM)and28 1 ± 4–9(10μM) X 10^-6cms^-1 ( n = 24). In 10 other monolayers, after the addition of 1 mM 8-Br-cAMP, urea permeability increased from 5.1 ±0–3 to 8.1 ± 1–6 times 10^-6 cm s^-1 and, after 8-Br-cAMP +3-isobutyl-l-methylxanthine, to 12.2 ± 0–7 times 10^-6 cms^-1. We conclude that rat IMCD cells grown in culture exhibit the characteristics of a 'tight' epithelium. Inulin and urea permeability are not different in the absence of AVP, consistent with high resistance junctional complexes. Furthermore, IMCD cells retain the capacity for AVP-regulated urea permeability, a characteristic feature of this nephron segment in vivo.     

Uptake of proline by brushborder vesicles isolated from human kidney cortex

Proline transport into renal brushborder membrane vesicles isolated from human kidney is mediated by two uptake systems. The high-affinity system is stimulated by a Na gradient and appears to be shared with glycine while the low-affinity system is not. Uptake curves of low concentrations of proline exhibit a Na-gradient-dependent overshoot indicative of electrogenic transport. The proline transport systems observed in isolated human renal brushborder membrane vesicles appear to have characteristics similar to those in rat kidney membranes.     

Idiotype regulation: a model for B cell tolerance

Immunological tolerance represents an individual's nonresponsiveness to self-antigens or foreign antigens. Inasmuch as the mechanism of self-nonself discrimination tolerance is unknown, it remains an area of active investigation because of its implied involvement in certain immune disorders. In this paper we discuss neonatal idiotype suppression and internal idiotope vaccination as models for B cell tolerance. The results demonstrate opposite aspects of idiotope regulation. Neonatal idiotype suppression represents an anti-idiotype-induced establishment of tolerance, whereas an internal antigen vaccine overcomes an established tolerance to self-antigens. New views on the functional importance of idiotype self-tolerance are discussed.     

Isolation and characterization of a large,neurite-associated glycoconjugate from neuroblastoma cells

A high molecular weight glycoconjugate has been isolated from neurite-producing neuronal tumor cells in culture and has been designated as I(0) based on its elution characteristics in gel filtration chromatography. This molecule cannot be found in a variety of nonneuronal cells. I(0) is found in the substratum-attached material or cell fraction of neurite-producing neuroblastoma cells, depending upon culture conditions. It is found in the substratum-bound fraction of B104 rat neuroblastoma cells during serum starvation and in the EGTA-detached cell fraction of B104 cells grown in chemically defined N2 medium. It occurs only in the cell fraction of the human neuroblastoma line Platt. Examination of behavioral variants of the B104 rat line further strengthens the association of I(0) with neurite production; the constitutive neurite-producing E(R)B9 variant contains I(0) while the non-neurite-producing E(R)A11 variant does not. I(0) is large, eluting in the void volume of sepharose-CL2B columns. Radioiodination of intact cells with lactoperoxidase shows I(0) to be a cell surface component. Metabolic radiolabeling studies show that it contains a high proportion of polysaccharide to protein, does not contain mannose, and is unsulfated. Alkaline borohydride reduction release two size classes of large polysaccharide chain. The alkaline reduction results, along with the mannose incorporation studies, show the presence of O-glycosidic linkages and few, if any, N-linkages. Resistance to nitrous acid deamination, insensitivity to glycosaminoglycan lyases, and the absence of sulfation, indicate that I(0) does not contain the glycosaminoglycans hyaluronic acid, chondroitin-, dermatan-, or heparin- sulfates. Affinity column chromatography reveals high binding affinity of I(0) to polyornithine and no binding to gelatin (collagen) or the glycosaminoglycans hyaluronate and heparin. These studies describe a unique high molecular weight glycoconjugate on the surface of neurite-producing neuroblastoma cell lines from two species.     

Intravenous versus inhaled atropine for inhibiting bronchoconstrictor responses in dogs

We studied whether the muscarinic antagonist, atropine, given intravenously or by inhalation, inhibits the bronchoconstrictor responses to inhaled acetylcholine and to acetylcholine released by electrical stimulation of the vagus nerves to the same degree. We assessed bronchoconstrictor responses in anesthetized dogs by determining the increase in total pulmonary resistance before and after increasing doses of atropine and then constructing inhibition dose-response curves. Before atropine the responses to the two stimuli were equal in magnitude. After intravenous atropine (initial dose 0.12 micrograms/kg, total dose 16 micrograms/kg) both responses were progressively inhibited to a similar degree. By contrast, after inhaled atropine (initial dose 0.02 micrograms/kg, total dose 2.4 micrograms/kg) the response to acetylcholine inhalation was inhibited to a much greater degree than the response to vagal stimulation. Thus, in studies designed to inhibit bronchoconstriction due to an inhaled muscarinic agonist to the same degree as bronchoconstriction due to a vagal reflex, atropine might better be given intravenously than by inhalation.     

Deep-level diagnostic value of the rDNA-ITS region

The similarity of certain reported angiosperm rDNA internal transcribed spacer (ITS) region sequences to those of green algae prompted our analysis of the deep-level phylogenetic signal in the highly conserved but short 5.8S and hypervariable ITS2 sequences. We found that 5.8S sequences yield phylogenetic trees similar to but less well supported than those generated by a ca. 10-fold longer alignment from rDNA-18S sequences, as well as independent evidence. We attribute this result to our finding that, compared to 18S, the 5.8S has a higher proportion of sites subject to vary and greater among-site substitution rate homogeneity. We also determined that our phylogenetic results are not likely affected by intramolecular compensatory mutation to maintain RNA secondary structure nor by evident systematic biases in base composition. Despite historical homology, there appears to be no ITS2 primary sequence similarity shared sufficient similarity to cluster correctly on the basis of alignability. Our results indicate that groups, however, share sufficient similarity to cluster correctly on the basis of alignability. Our results indicate that ITS region sequences can diagnose organismal origins and phylogenetic relationships at many phylogenetic levels and provide a useful paradigm for molecular evolutionary study.