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Biomedical subjects

R Radcliffe

Publications and source records attributed to R Radcliffe.

14 recordsLinked to original sources
PubMed2001

Neurotensin levels in specific brain regions and hypnotic sensitivity to ethanol and pentobarbital as a function of time after haloperidol administration in selectively bred rat lines.

Evidence indicates that sensitivity to ethanol is a good predictor of the development of alcoholism. Thus, identification of neuronal processes that regulate ethanol sensitivity has been the subject of much recent research. The present studies were designed to further test the hypothesis that neurotensinergic processes mediate, in part, hypnotic sensitivity to ethanol. Single doses of haloperidol were administered to lines of rats [selectively bred for high and low sensitivity (HAS and LAS, respectively) to hypnotic effects of ethanol] to produce increases in neurotensin (NT) levels in brain regions. At 20 h after administration, haloperidol produced dose-dependent increases in NT immunoreactivity levels in nucleus accumbens (NA) and caudate putamen (CP) in both HAS and LAS lines. Levels of NT in NA and CP returned to control values at 48 h after 4 mg/kg haloperidol. These studies used two measures of hypnotic sensitivity to ethanol: duration of loss of righting reflex (sleep time) and blood ethanol concentration at regain of righting reflex (BECRR). At 20 h, but not 48 h, after haloperidol treatment, both HAS and LAS rats displayed increases in ethanol-induced sleep time with concomitant decreases in BECRR. Pentobarbital-induced sleep time was not increased 20 h after administration of 4 mg/kg haloperidol; however, hypnotic sensitivity to both pentobarbital and ethanol was increased by acute (30-min) pretreatment with 1 mg/kg. These results suggest that NT levels in NA, acting via NT receptors, enhance hypnotic sensitivity to ethanol, but not pentobarbital.

Alcoholic Intoxication↗
PubMed1994

Thrombus imaging with technetium-99m synthetic peptides based upon the binding domain of a monoclonal antibody to activated platelets.

UNLABELLED: Monoclonal antibodies which recognize fibrin or platelets have enabled imaging of vascular thrombi, however, early imaging has been difficult because of the slow blood disappearance of even small antibody fragments. It was theorized that it might be possible to synthesize peptides which possess the same thrombus affinity as monoclonal antibodies, but which would leave the blood pool much more rapidly. METHODS: In this study, peptides were synthesized with amino acid sequences based on the primary binding region of the platelet glycoprotein IIb/IIIa-directed monoclonal antibody PAC1. Both termini of the peptides were blocked to prevent rapid proteolysis and a metallothionein-derived sequence was incorporated as a chelating agent for reduced technetium. RESULTS: Technetium-99m-labeled peptides produced images of fresh clots in the jugular veins of rabbits and day-old thrombi in the femoral veins of dogs within 2 hr after injection. In control experiments, a 99mTc-labeled nonspecific peptide failed to produce focal images of thrombus. Another control compound, 99mTc-glucoheptonate, did produce images of fresh clots in rabbits but failed to produce focal images of day-old thrombi. As was hoped, blood clearance of the 99mTc peptides was rapid, with excretion through the kidneys, however, none of the peptides studied had better thrombus-to-blood ratios than iodinated fibrinogen and all had significantly lower deposition in the thrombus. CONCLUSION: Using labeled synthetic peptides appears to be technically feasible but the absolute binding to thrombus is not yet sufficient for reliable imaging of pre-existing thrombi.

Amino Acid Sequence↗
PubMed1990

Low doses of ethanol reduce neurotensin levels in discrete brain regions from LS/Ibg and SS/Ibg mice.

Studies were designed to examine the previously proposed hypothesis that some of the pharmacological actions of ethanol are mediated by neurotensinergic processes. Neurotensin-immunoreactivity (NT-ir) was extracted from various brain regions and shown by high performance liquid chromatography to possess the same retention time as authentic bovine NT1-13. The highest levels of NT-ir were observed in the hypothalamus with intermediate levels in the midbrain and striatum and lowest levels in the frontal cortex. Levels of NT-ir were higher in hypothalamus and midbrain from long-sleep (LS) than from short-sleep (SS) mice. Ethanol, in vivo, produced a dose-dependent decrease in NT-ir in several brain regions; low doses, 1.5 to 3.0 g/kg, but not high doses, 4.1 g/kg, of ethanol significantly decreased NT-ir in hypothalamus, midbrain, and striatum of LS and SS mice. Levels of NT-ir in the frontal cortex were not altered by ethanol administration. Ethanol-induced decreases in NT-ir were of rapid onset with a maximum decrease in 5 min after intraperitoneal (i.p.) injection, and they were of long duration with levels remaining depressed for 4 hr. These findings show that subhypnotic, intoxicating doses of ethanol enhance NT release, in vivo, and support the hypothesis that some of ethanol's actions are mediated by neurotensinergic systems.

Animals↗
PubMed1990

Further characterization of LSxSS recombinant inbred strains of mice: activating and hypothermic effects of ethanol.

Lines of mice selected for differential initial sensitivity to the anesthetic effects of ethanol also differ in their locomotor responses to lower doses of ethanol. Sixteen recombinant inbred strains of mice derived from long-sleep (LS) and short-sleep (SS) selected lines as well as inbred LS and SS mice were used in a genetic correlational study to investigate possible associations between high-dose and low-dose indices of initial sensitivity to ethanol. Measurements of high-dose (4.1 g/kg) effects of ethanol were hypothermia, sleep time, and blood ethanol content at regaining of righting response, and the index of low-dose (1.875 g/kg) sensitivity was distance traveled during a 5-min period immediately following intraperitoneal injection with ethanol. The results indicated wide genetic variation in hypothermia and ethanol-induced locomotor activation in a manner consistent with polygenic influence. Furthermore, correlations between low-dose locomotor activity and hypnotic dose effects tended to be low and nonsignificant, indicating independence of inherited mechanisms underlying high- and low-dose ethanol sensitivity.

Alcoholism↗
PubMed1983

A critical role of lysine residues in the stimulation of tissue plasminogen activator by denatured proteins and fibrin clots.

Two-chain 70 000-dalton plasminogen activator of tissue origin displays only weak activity toward plasminogen in a two-component system. The rate of activation is enhanced a minimum of 50-fold by the presence of fibrin clots or denatured proteins. The stimulation must depend on both chemical determinants and spatial configuration, since native proteins, including fibrinogen, lack significant stimulatory activity. These studies employed chemical modifications of four stimulatory proteins (fibrin, denatured fibrinogen, denatured IgG and denatured ovalbumin) to identify a critical role for lysine residues. Arginine, aspartic acid, cysteine, cystine, glutamic acid, histidine, methionine, tyrosine and tryptophan were found not to be essential. The critical spatial determinant(s) remain(s) unknown.

Animals↗
PubMed1981

A lysine-absorbable plasminogen activator is elevated in conditions associated with increased fibrinolytic activity.

A plasminogen activator, or class of activators, that absorbs to lysine-agarose is present in human plasma. We have developed a quantitative assay for this plasminogen activator. The assay involves removal of the activator from plasma with lysine-agarose affinity columns and subsequent measurement of the activity by the conversion of plasminogen to plasmin on standardized fibrin agar plates. Using this assay, we investigated three physiologic conditions that have in the past been associated with increased fibrinolytic activity to determine whether elevation of the LAPA was involved. Normal individuals undergoing strenuous physical exercise and others subjected to venous occlusion as well as patients with cirrhosis of the liver were examined. Treadmill exercise to maximal exertion produced up to 15-fold increases in the level of LAPA; venous occlusion produced similar elevation. Certain individuals did not show increase fibrinolytic activity in response to exercise or venous occlusion, as indicated by unchanged euglobulin lysis times. These fibrinolytic hyporesponders did not show an elevation of their LAPA levels. In the third group examined, patients with cirrhosis, 24 of 62 had elevated levels of LAPA. Supplementation of plasma from normal individuals with this plasminogen activator from exercised individuals and cirrhotics resulted in increased rates of clot lysis.

Adsorption↗
PubMed1977

Activation of bovine factor VII by hageman factor fragments.

During the early events of coagulation of human blood by the intrinsic pathway, factor XII is activated to a form which can activate factor XI, and is proteolytically fragmented to smaller species (30,000 daltons and 70,000 daltons) which have lost most of the ability to activate factor XI but which can activate prekallikrein rapidly. The effect of these fragments on factor VII was studied. It was found that these Hageman factor fragments promoted rapid proteolysis of one-chain factor VII to a more active two-chain form. The amino-terminal sequences of the chains of activated factor VII were found to be Ala-Asx-Gly- and Ile-Val-Gly-, the same as were earlier observed after activation of factor VII by activated factor X. This finding indicates that initiation of coagulation by the intrinsic pathway also primes the extrinsic pathway.

Amino Acid Sequence↗
PubMed1976

Mechanism of activation of bovine factor VII. Products of cleavage by factor Xa.

Coagulation Factor VII from bovine plasma is a glycoprotein containing a single peptide chain. The NH2-terminal sequence of Ala-Asx-Gly-Phe-Leu- is homologous with the NH2 termini of prothrombin, Factor IX, and the light chain of Factor X. Factor Xa in the presence of calcium ions and phospholipid cleaves Factor VII at an Arg-Ile bond in the sequence Arg-Ile-Val-Gly-Gly-, producing a two-chain molecule with at least 85 times the coagulant activity of single-chain Factor VII and a new NH2-terminal sequence homologous with the corresponding chains of thrombin, Factor IXa and Factor Xa. A second slower cleavage at an Arg-Gly bond destroys Factor VII activity. Bovine Factor VII, unlike prothrombin, Factor IX, and Factor X, is rapidly inhibited by diisopropylphosphorofluoridate (iPr2PF). [3H]iPr2PF is readily incorporated into one-chain, two-chain, and three-chain forms of Factor VII up to ratios of approximately 0.9 moles of [3H]diisopropylphosphate per mole of protein. The radioactive peptides generated from each form of [32P]iPr2PF-inhibited Factor VII by tryptic and thermolytic digestion were found to migrate together on paper electrophoresis. This indicates that the iPr2PF is incorporated stoichiometrically into the same specific site in each form.

Amino Acid Sequence↗
PubMed1975

Activation and control of factor VII by activated factor X and thrombin. Isolation and characterization of a single chain form of factor VII.

Factor VII purified as previously described, was found to consist of two polypeptide chains joined by disulfide bridges. We now report the isolation and 200,000-fold purification of a single chain form of Factor VII. This was accomplished by protecting the molecule against proteolysis by including benzamidine during the entire purification. The purification was essentially as previously reported except that barium cirtate was substituted for barium sulfate as an absorbant for Factor VII as it resulted in a 4-fold increase in yield. Single chain Factor VII is rapidly hydrolyzed by Factor Xa in the presence of calcium ions and phospholipids, and by thrombin, to a two-chain form which possesses at least 85 times the Factor VII clotting activity of the single chain species. The two-chain form of the enzyme requires tissue factor in order to activate Factor X. From the observed rates of activation of Factor VII by Xa in the presence of calcium ions and phospholipids, it was calculated that at approximately physiological concentration, Factor VII activity would increase at an initial rate of 20-fold per min; this reaction is sufficiently rapid to constitute a feedback control mechanism. The action of thrombin is approximately 40-fold slower under these conditions. Diisopropylphosphorofluoridate inactivates the single chain and two-chain forms of Factor VII at approximately equal rates. After inhibition, the single chain species could be cleaved but not activated by proteolysis.

Animals↗
PubMedDate not supplied

Genetic-based differences in neurotensin levels and receptors in brains of LS x SS mice.

Levels of endogenous neurotensin (NT-IR) in the LS x SS RI strains differed by 3.0-, 4.7-, 5.4-, and 6.9-fold in the ventral midbrain (VMB), hypothalamus (HY), nucleus accumbens (NA), and caudate putamen (CP), respectively. Frequency distributions and estimates of the number of genes indicate that differences in NT-IR are polygenically influenced. The NT-IR levels in NA and CP were significantly correlated, but levels in the VMB did not correlate with those in the NA or CP. Specific binding to either low (NTL)- or high (NTH)-affinity receptors as measured in the absence or presence of levocabastine differed significantly in brain regions from among LS X SS mouse strains. Results indicate a polygenic influence mediating the differences in receptor densities and suggest differences in genetic regulation of NTL and NTH receptors.

Animals↗