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S E Leeman

Publications and source records attributed to S E Leeman.

At least 145 records · Page 8Linked to original sources
PubMed1976

Radioimmunoassay for neurotensin, a hypothalamic peptide.

A highly sensitive and specific radioimmunoassay for neurotensin has been developed which utilizes 125I-labeled neurotensin and rabbit antisera raised toward synthetic neurotensin which has been coupled specifically through its lysine side chain to several proteins. The three antisera described have different specificities but are directed primarily towards the COOH-terminal region of neurotensin which is the biologically active portion of the molecule. Two of the antisera, poly(Glu60, Lys40) (from animal no. 4) and keyhole limpet hemocyanin (from animal no. 8), cross-react fully with COOH-terminal partial sequences of neurotensin while antiserum poly(Glu60, Lys40) (from animal no. 6) requires the entire molecule for full recognition. The assay can detect less than 3 fmol of neurotensin and the dose-response curves for synthetic and native neurotensin are superimposable, irrespective of the antiserum employed. Using these assay systems, the immunoactivity in acid/acetone extracts of 45 kg of bovine hypothalami was purified to homogeneity and shown to be attributable to intact neurotensin and not to fragments of neurotensin nor to related molecules. Radioimmunoassayable neurotensin (R-NT) obtained from bovine, rat, guinea pig, and rabbit hypothalami also gave dose-response curves which paralleled that of neurotensin and the neurotensin equivalents per g of wet tissue were in the range 45 to 70 pmol/g. Measurements with the three antisera were in agreement, especially after the extracts were chromatographed on Sephadex G-25; R-NT in these hypothalamic extracts has also been shown to be destroyed by treatment with various enzymes known to cleave neurotensin.

Animals↗
PubMed1976

Characterization of radioimmunoassayable neurotensin in the rat. Its differential distribution in the central nervous system, small intestine, and stomach.

Using radioimmunoassay, the distribution of radioimmunoassayable neurotensin (R-NT) has been determined in acid/acetone extracts of tissues from week-old and adult rats. Whereas only 10% of the R-NT in week-old rats was found in the head, 90% was found in extracts of the body, and 85% was present in the intestine. The distribution of R-NT in the central nervous system of adult male rats was as follows: hypothalamus (35%), brain stem (35%), cerebral cortex (17%), thalamus (11%), cerebellum (1%), and pituitary gland (1%). Concentrations of R-NT were highest in the hypothalamus (60 pmol/g) and lowest in the cerebellum (0.8 pmol/g). More than 10 times as much R-NT was found in extracts of adult rat small intestine than in brain. The concentration of R-NT was highest in the jejuno-ileal section of the intestine (approximately 50 pmol/g) but it was also detected in the esophogus, stomach, duodenum, and large intestine (approximately 1 to 8 pmol/g) than the remaining muscle layer (approximately 40 pmol/g). Rat jejuno-ileal R-NT co-chromatographed with neurotensin on Sephadex G-25, displayed equal immunological potency with the antisera utilized, and was destroyed by enzymes known to cleave neurotensin. High concentrations (approximately 400 pmol/g) of an R-NT of smaller molecular weight than neurotensin were found in rat stomach. This peptide reacted most strongly with a COOH-terminal-directed antiserum and appeared to be four to five amino acids in length; it might possibly be a breakdown product of neurotensin or a related peptide with biological activity. Acid/acetone extracts of rat and bovine plasma were found to contain an R-NT which displayed the properties of neurotensin (approximately 50 fmol/ml) as well as a substance which behaved similarly to the small molecular weight stomach R-NT (approximately 1000 fmol/ml).

Aging↗
PubMed1976

Isolation of a tridecapeptide from bovine intestinal tissue and its partial characterization as neurotensin.

Radioimmunoassayable neurotensin (R-NT) has been isolated from acid/acetone extracts of 50 kg of calf small intestine with an overall yield of approximately 15%. The concentration of R-NT in calf intestinal tissue was approximately 35 pmol/g wet weight. Throughout the purification procedures which involved adsorption onto sulfopropyl (SP)-Sephadex, chromatography on Sephadex G-25 and SP-Sephadex, immunoadsorption on neurotensin-antibody Sepharose and high voltage paper electrophoresis, R-NT displayed the chromatographic and electrophoretic properties of neurotensin. R-NT was found to contain a tridecapeptide with the same amino acid composition as neurotensin. This peptide yielded the same products as neurotensin when submitted to digestion by carboxypeptidase A or papain. Its immunological properties were indistinguishable from those of neurotensin and its potency in stimulating hypotension in anesthetized rats was comparable to that of synthetic neurotensin. If the amino acid sequence of this peptide proves to be the same as that of neurotensin, then neurotensin is another biologically active peptide isolated from both brain and intestinal tissues.

Amino Acids↗
PubMed1976

Regional distribution of substance P in the brain of the rat.

Using a sensitive radioimmunoassay we have studied the regional distribution of substance P. The level of substance P is higher in the mesencephalon, hypothalamus and preoptic area than in other regions of the brain. Substance P is found in especially high concentrations in the reticular part of the substantia nigra and the interpeduncular nucleus. It is present in large amounts in several septal, preoptic and hypothalamic nuclei as well.

Animals↗
PubMed1976

Synthesis and activities of neurotensin, and its acid and amide analogs: possible natural occurrence of [Gln4]-neurotensin.

It was considered, a priori, that the isolation of the tridecapeptide, neurotensin, might have inadvertently sllowed the hydrolysis of either the [Gln4]- or the [Leu13-NH2]-moieties. Neurotensin and its three acid and amide analogs, i.e., [Gln4]-neurotensin, neurotensin-NH2, and [Gln4]-neurotensin-NH2 were synthesized. Neurotensin and [Gln4]-neurotensin were indistinguishable by the hypotensive assay, hyperglycemic assay, contraction of the ileum, and radioimmunoassay. Neurotensin-NH2 and [Gln4]-neurotensin-NH2 showed less than 1% of these neurotensin activities. Present information does not elucidate whether the glutamic acid residue in position 4 of neurotensin in situ is present as Glu4 or as Gln4. At high levels, neurotensin released the luteinizing hormone, follicle stimulating hormone, and thyrotropin; [Gln4]-neurotensin-NH2 released thyrotropin, and [Gln4]-neurotensin released luteinizing hormone and follicle stimulating hormone, but these activities do not appear biologically significant.

Animals↗
PubMed1976

Hyperglycemic effect of neurotensin, a hypothalamic peptide.

A hypotensive, gut-contracting peptide neurotensin (NT), recently isolated from bovine hypothalami, has been found to produce hyperglycemia within minutes after iv injection into anesthetized rats. The dose-response relationship (deltaglucose, 15 min after injection) was linear over the range 30-200 pmol/100 g BW. NT did not alter the disappearance rate of [14C]glucose from plasma during the development of the hyperglycemia. However, the peptide caused a fall in liver glycogen (52 +/- 6.5 to 41 +/- 3.3 mg/g) and a 7-fold increase in the activity of the 5'-AMP independent form of liver glycogen phosphorylase. Activation of liver glycogen phosphorylase did not occur in vitro under conditions found suitable for demonstrating the effectiveness of glucagon, suggesting the possible involvement of an intermediary substanc(s) in vivo. Acute adrenalectomy did not prevent the response. Hypophysectomized rats (4 days post-operative) were less sensitive to NT, perhaps as a consequence of their diminished liver glycogen levels (normal, 52 +/- 6.5 mg/g; hypophysectomized, 23 +/- 1.8 mg/g); however, the presence of the pituitary was not essential for this response. NT was also effective in rats with hereditary diabetes insipidus (Brattleboro strain). At the time intervals sampled, radioimmunoassayable plasma levels of growth hormone, glucagon, and insulin were not significantly changed after injection of NT into normal rats. Pretreatment of rats with reserpine (7 mg/kg), morphine sulfate (10 mg/kg), propranolol (5mg/kg), or phenoxybenzamine (10 mg/kg) did not prevent the response. These findings characterize the action of NT on liver glycogen metabolism and blood glucose levels, but a physiological role for NT in this regard remains to be demonstrated.

Adrenal Glands↗
PubMed1975

The synthesis of neurotensin.

A tridecapeptide having the amino acid sequence, less than Glu-Leu-Tyr-Glu-Asn-Pro-Arg-Arg-Pro-Tyr-Iie-Leu-OH, (The nomenclature and symbols follow the suggestions of the IUPAC-IUB Commission on Biochemical Nomenclature (1972) J. Biol. Chem. 247, 977).) has been synthesized by the Merrifield solid-phase procedure. The synthetic scheme chosen involved synthesis of the peptide in the (Gln) form and cyclization to the less than Glu) form. After purification, the (Gln) peptide was obtained in a 7% yield and the (greater than Glu) peptide was obtained in a 35% yield. The (greater than Glu) was found to be chemically and biologically indistinguishable from the tridecapeptide, neurotensin, recently isolated from bovine hypothalami.

Animals↗
PubMed1975

The amino acid sequence of a hypothalamic peptide, neurotensin.

The amino acid sequence of neurotensin, a hypotensive peptide isolated from acid-acetone extracts of bovine hypothalami, has been established as less than Glu-Leu-Tyr-Glu-Asn-Lys-Pro-Arg-Arg-Pro-Tyr-Ile-Leu-Oh. (The nomenclature and symbols follow the suggestions of the IUPAC-IUB Commission on Biochemical Nomenclature (1972) J. Biol. Chem. 247, 977). This was accomplished by sequence analyses performed on the intact peptide as well as its isolated tryptic, chymotryptic, and papain-generated fragments. The results of enzymic hydrolyses were consistent with the specificities of the enzymes used and indicated that all of the amino acids are unsubstituted and in the L configuration. The absence of non-amino acid constituents was further supported by analyses of electrophoretic mobility-molecular weight relationships of neurotensin and its fragments.

Amino Acid Sequence↗
PubMed1974

Substance P.

Explore the source record for details and available documents.

Amino Acid Sequence↗
PubMedDate not supplied

Studies on the role of catecholamines in the regulation of the developmental pattern of hypothalamic aromatase.

Experiments were conducted to study the regulation of the developmental pattern of aromatase in the forebrain of the perinatal rat. Two experimental designs were used: aromatase measured in primary cultures of fetal hypothalamic cells and in cell-free preparations of forebrain tissue excised at varying ages. In cultured cells, aromatase decreased logarithmically at a slow rate (t1/2 = 7.8 days). Norepinephrine caused a pronounced dose (4 x 10(-6) M) and time-dependent (2-6 days) drop in aromatase without affecting the levels of 5 alpha-reductase or substance P. In isolated tissue, aromatase activity was compared with the concentrations of norepinephrine and dopamine in the forebrain of males vs females at different perinatal ages and in discrete forebrain areas at postnatal day 4. In no case was a sex difference in catecholamines seen. An overall developmental decline in aromatase was associated with developmental increases in catecholamine levels. Acute treatment with the beta-agonist, isoproterenol, had no effect on brain aromatase activity.

Animals↗
PubMedDate not supplied

The stability and metabolism of intravenously administered neurotensin in the rat.

The clearance and metabolism of synthetic and tritiated (3H) neurotensin (NT) were studied following its intravenous injection in a pharmacologic dose (500 pmol/kg) into anesthesized rats. Immunoreactive NT (iNT), measured in a radioimmunoassay (RIA) with use of a carboxyl-(C)-terminal directed antiserum, displayed an apparent half-life (t 1/2) of 0.55 min, while that measured by an amino-(N)-terminal directed antiserum had a t 1/2 of 5 min. The radiolabel from injected 3H-NT (3H on Tyr3,11) had a t 1/2 of 6.5 min. High-pressure liquid chromatography of extracts of plasma obtained from the circulation 0.5-3 min after injection of NT and 3H-NT showed the presence of NT and the generation mainly of the fragments NT1-8, NT1-11, and NT9-13, as well as free 3H-labeled tyrosine. The apparent half-lives of intravenously injected synthetic NT1-8, NT1-11 and NT1-12 measured with the N-terminal RIA were 9, 5 and 5 min, respectively, while that for NT9-13 was less than 0.5 min. These results indicate that exogenously injected NT is rapidly metabolized to form N-terminal fragments which are cleared more slowly than NT. These findings suggest that use of N-terminal antisera to detect the release of endogenous NT into the circulation is likely to yield measurements of the fragments NT1-8 and NT1-11 which thus far have been found to be biologically inactive.

Animals↗