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B F Dickey

Publications and source records attributed to B F Dickey.

43 records · Page 3Linked to original sources
PubMed1987

Reconstitution of the rat liver vasopressin receptor coupled to guanine nucleotide-binding proteins.

The V1 vasopressin receptor has been solubilized from rat liver membranes with the zwitterionic detergent 3-[(3-cholamidopropyl)dimethylammoniol]-1-propanesulfonate (CHAPS) and reconstituted into phospholipid vesicles. There is essentially complete solubilization of the receptor by 3% CHAPS at a protein concentration of 15 mg/ml. Reconstitution into soybean phospholipid vesicles is readily achieved either by gel filtration chromatography or by membrane dialysis. The binding of [3H]vasopressin to proteoliposomes is specific, saturable, reversible, and magnesium-dependent. In contrast, the detergent-soluble vasopressin receptor does not display specific binding. The apparent affinity of the reconstituted receptor for [3H]vasopressin is approximately 4-fold lower than that of the receptor in native membranes. In addition, the binding of [3H]vasopressin to reconstituted vesicles is not sensitive to 100 microM guanosine 5'-O-thiotriphosphate (GTP gamma S) as it is in native membranes. However, the apparent affinity of the reconstituted receptor for ligand approximates that of native membranes when membranes are prebound with vasopressin prior to solubilization and reconstitution into vesicles. Furthermore, vesicles reconstituted from membranes prebound with vasopressin show GTP gamma S sensitivity of [3H] vasopressin binding. This finding strongly suggests that vasopressin stabilizes a receptor-G-protein complex during solubilization. The rat liver vasopressin receptor is a glycoprotein, as shown by its specific binding to the lectin "wheat germ agglutinin." The vasopressin receptor can be reconstituted from the N-acetylglucosamine-eluted peak of a wheat germ agglutinin-Sepharose column, and [3H] vasopressin binding activity is purified 5-6-fold from membranes by this chromatographic procedure. The functionality of the partially purified receptor is indicated by its ability to bind ligand with high affinity and by its ability to functionally interact with a G-protein when vasopressin is bound prior to solubilization.

Animals↗
PubMed1986

Reversible inactivation of vasopressin and angiotensin II binding to hepatocyte membranes by a calcium-dependent, cytosolic protein.

A cytosolic protein is described which inhibits the binding of vasopressin and angiotensin to their rat liver receptors in the presence of calcium. The binding of insulin and transferrin was unaffected. Inhibition was temperature-dependent; it was maximal in 10 min at 37 degrees C, but required longer incubation times at lower temperatures. The pH optimum was 7.4. Inhibition also required the presence of calcium, with half-maximal inhibition at 6-8 microM calcium, but did not require any other low molecular weight cofactors. Inhibition could be reversed by washing the membranes at pH 5.5, but not by incubation with EGTA. Sephacryl S-300 chromatography showed that activity eluted in two peaks with approximate molecular weights of 70,000 and 150,000. In the presence of calcium, the inhibitory activity eluted at 150,000; in the absence of calcium, most of the inhibitory activity eluted at 70,000. A radiolabeled cytosolic protein with a molecular weight of 70,000 was eluted from inhibited rat liver membranes at pH 5.5 as analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. We propose that vasopressin and angiotensin II, which both mobilize calcium in hepatocytes via phosphatidylinositol turnover, can, by this same mechanism, activate a protein(s) which reduces further binding to their receptors.

Angiotensin II↗
PubMed1985

Internalization, recycling, and redistribution of vasopressin receptors in rat hepatocytes.

Three hours after isolation, cultured hepatocytes have approximately 150,000 surface vasopressin receptors/cell, and these exhibit a Kd for 125I-vasopressin of 6 nM based on calculation of Koff/Kon, or a Kd of 9.5 nM based on Scatchard plot analysis. After the binding of 125I-vasopressin to its receptor on the hepatocyte surface, this complex is internalized with a t1/2 of 3-6 min. Following this internalization, the number of vasopressin receptors on the cell surface is restored both in vitro and in the isolated perfused liver with a t1/2 of 8-10 min. This restoration is blocked in vitro by incubation of the hepatocytes at 18 degrees C, but not by cycloheximide, suggesting that internalized vasopressin receptors recycle back to the cell surface. Prolonged incubation of hepatocytes with vasopressin results in the loss of greater than 75% of the vasopressin surface binding at concentrations of vasopressin approximately equivalent to its Kd. The binding of vasopressin to cultured hepatocytes 3-5 h after isolation resembles binding to the isolated perfused whole liver with respect to receptor dynamics. During culture for 48 h, however, we observe a progressive loss of hepatocyte surface vasopressin receptors. Concomitant with this reduction in surface receptors with time in culture, there appears to be a marked elevation in intracellular receptors.

Animals↗
PubMed1983

Polymorphonuclear leukocyte participation in acute oleic-acid-induced lung injury.

The purpose of this study was twofold: (1) to analyze the cellular components of bronchoalveolar lavage fluid throughout the development of oleic-acid-induced lung injury in the rat and (2) to investigate the role of polymorphonuclear leukocytes (PMN) in the pathogenesis of this disease. Animals were killed and lavaged at various times after a single intravenously administered injection of oleic acid. The results demonstrate that a significant influx of inflammatory cells appear in the lavage fluid as early as 4 h after the administration of oleic acid. The PMN are the first cells to appear, and significant levels persist through Day 5 after injection. There is a transient yet significant influx of lymphocytes between 3 and 7 days after treatment. Rats treated with oleic acid displayed significant increases in lung vascular permeability over control animals at 1 and 4 h after injection. Depletion of PMN by anti-PMN serum significantly decreased the permeability changes induced by oleic acid. Treatment of oleic-acid-injected animals with catalase, superoxide dismutase, or dimethyl sulfoxide failed to inhibit lung permeability changes induced in this model.

Acute Disease↗
PubMed1981

Oleic-acid-induced lung injury in the rat. Failure of indomethacin treatment or complement depletion to ablate lung injury.

The purpose of this study was to establish a rat animal model of acute respiratory distress syndrome using the intravenous injection of oleic acid. Further, we attempted to inhibit the development of lung injury by pretreatment of the rats with indomethacin or cobra venom factor. Histologic evidence of lung injury was apparent within hours after the administration of a single intravenous injection of oleic acid. By 24 hours, interstitial and intraalveolar edema and hemorrhage were noted with vascular congestion and an extensive interstitial infiltrate. The lungs appeared virtually normal by 12 days, with no evidence of chronic lung injury. Multiple injections of oleic acid also did not progress into chronic pulmonary inflammation. Treatment of the rats with indomethacin or cobra venom factor had no effect on ablating acute lung injury. An animal model of adult respiratory distress syndrome is presented which does not progress to chronic lung injury.

Animals↗