neuromedin-n and xenopsin

Cathepsin E (EC 3.4.23.34), an intracellular aspartic proteinase, was purified from monkey intestine by simple procedures that included affinity chromatography and fast protein liquid chromatography. Cathepsin E was very active at weakly acidic pH in the processing of chemically synthesized precursors such as the precursor to neurotensin/neuromedin, proopiomelanocortin, the precursor to xenopsin, and angiotensinogen. The processing sites were adjacent to a dibasic motif in the former two precursors and at hydrophobic recognition sites in the latter two. The common structural features that specified the processing sites were found in the carboxyl-terminal sequences of the active peptide moieties of these precursors; namely, the sequence Pro-Xaa-X'aa-hydrophobic amino acid was found at positions P4 through P1. Pro at the P4 position is thought to be important for directing the processing sites of the various precursor molecules to the active site of cathepsin E. Although the positions of Xaa and X'aa were occupied by various amino acids, including hydrophobic and aromatic amino acids, some of these had a negative effect, as typically observed when Glu/Arg and Pro were present at the P3 and P2 positions, respectively. Cathepsin D was much less active or was almost inactive in the processing of the precursors to neurotensin and related peptides as a result of the inability of the Pro-directed conformation of the precursor molecules to gain access to the active site of cathepsin D. Thus, the consensus sequence of precursors, Pro-Xaa-X'aa-hydrophobic amino acid, might not only generate the best conformation for cleavage by cathepsin E but might be responsible for the difference in specificities between cathepsins E and D.

Topics: Amino Acid Sequence; Animals; Cathepsin E; Cathepsins; Macaca; Molecular Sequence Data; Neurotensin; Oligopeptides; Peptide Fragments; Peptides; Protein Precursors; Xenopus Proteins

Intracerebral microinjections of neurotensin (NT) decrease intracranial self-stimulation (ICSS) of the medial prefrontal cortex (MPC) in the rat. This effect could be due to the ability of NT to bind dopamine. To test this hypothesis we studied the effects of intracerebral microinjections of neuromedin N, a natural NT analogue that does not bind dopamine, on ICSS of the rat MPC. Unilateral microinjections of neuromedin N into the MPC at doses of 2.5, 5, 10, 20 and 40 nmol produced a dose-related decrease in ICSS of the ipsilateral MPC. ICSS of the contralateral MPC, used as a control, was not affected by the microinjections. These results suggest that the inhibitory effect of NT on ICSS is independent of NT-dopamine binding. Because neuromedin N is also present in the MPC, these results also suggest a possible neuromodulatory role of this neuropeptide on ICSS of the prefrontal cortex.

Topics: Animals; Depression, Chemical; Dose-Response Relationship, Drug; Male; Microinjections; Neurotensin; Oligopeptides; Peptide Fragments; Peptides; Prefrontal Cortex; Rats; Rats, Wistar; Self Stimulation; Stereotaxic Techniques; Xenopus Proteins

Neurotensin is released from endocrine N cells of the ileum after meals, and might take part in the regulation of bicarbonate secretion by the pancreas and duodenum. The aim of this study was to define the effect of neurotensin on duodenal bicarbonate secretion, in comparison with its effect on gastric and pancreatic secretions. Neurotensin produced a dose-related increase of duodenal bicarbonate secretion with an ED50 of 60 pmol/kg.h. The maximal effect (about 2 times the basal level) was observed with 600 pmol/kg.h. Equimolar doses (600 pmol/kg.h) of xenopsin, neuromedin N, neurotensin 8-13 produced the same effect as neurotensin 1-13. Neurotensin fragments 1-11 and 9-13 (600 pmol/kg.h) had no significant effect on duodenal bicarbonate secretion. Indomethacin, atropine, naloxone, or the CCK antagonist L364,718 had no effect on neurotensin-stimulated bicarbonate secretion. Hexamethonium and vagotomy reduced the neurotensin effect by about 50 percent (P less than 0.05). Neurotensin produced a dose-related increase of pancreatic bicarbonate secretion with an ED50 of 150 pmol/kg.h, a decrease of gastric acid secretion with an ED50 of 2,400 pmol/kg.h, and a decrease of gastric pepsin secretion with an ED50 of 2,760 pmol/kg.h. This study shows that neurotensin stimulates duodenal bicarbonate secretion in doses which may be physiological. This biological activity depends on the presence of the C-terminal 8-13 fragment. The mechanism is complex, and depends, for approximately half, on vagal fibers (sensitive or motor), nicotinic synapses, and a non cholinergic effector. The other half of the effect, still unexplained, could be a direct effect on mucosal cells. Pancreatic and duodenal bicarbonate secretions were more sensitive to neurotensin than gastric secretion (acid and pepsin).

Topics: Animals; Bicarbonates; Duodenum; Ganglionic Blockers; Gastric Mucosa; Hexamethonium; Hexamethonium Compounds; Male; Neurotensin; Oligopeptides; Pancreas; Peptide Fragments; Peptides; Rats; Rats, Inbred Strains; Stimulation, Chemical; Xenopus Proteins

The tridecapeptide neurotensin (NT) is present at high concentrations within the mammalian gut, although its physiological function is undefined. In this study, the actions of NT and structurally related peptides neuromedin N (NM-N) and xenopsin were characterized on ion transport in the porcine distal jejunal mucosa in vitro. The serosal-side administration of these peptides elicited rapid changes in transmural potential difference and short-circuit current (Isc) which were greater in mesenteric than in antimesenteric segments. NT-induced elevations in Isc were dependent upon external permeant anions and were associated with net Cl transport in both segments. In mesenteric segments, NT and its homologs increased Isc with the order of potency: NT greater than NM-N greater than xenopsin. NT produced tachyphylaxis to its own actions and to those of NM-N; NM-N was ineffective in producing tachyphylaxis. Isc elevations produced by NT were inhibited by the neuronal conduction blocker tetrodotoxin (0.1 microM) or the Ca++-channel blocker dl-verapamil (100 microM) and reduced in Ca++-free media. Antagonists to the enteric transmitters acetylcholine, ATP, 5-hydroxytryptamine, substance P and histamine did not alter Isc responses of mesenteric segments to NT. Serosal administration of vasoactive intestinal peptide (10 nM) did not resemble the effects of NT. The magnitude of Isc responses to these agonists was smaller in antimesenteric segments. These results indicate that NT-related peptides present in mucosal endocrine cells or nerves of the porcine jejunum may modulate Cl transport through mechanisms that involve the Ca++-dependent release of unknown enteric neurotransmitters. Moreover, there appear to exist within the distal jejunum circumferential differences in mucosal responses to NT and other neurohumoral factors.

Topics: Animals; Bicarbonates; Biological Transport; Calcium; Chlorides; Electrolytes; Female; Jejunum; Male; Neurotensin; Oligopeptides; Peptide Fragments; Peptides; Substance P; Vasoactive Intestinal Peptide; Xenopus Proteins

Neurotensin, xenopsin and neuromedin N had a biphasic effect (initial small relaxation followed by a sustained contraction) on the motility of longitudinal muscle strips from the guinea pig gastric corpus. Kinetensin was without effect. Tetrodotoxin, adrenaline, histamine, 5-hydroxytryptamine, opioid peptides, angiotensin II and antagonists of acetylcholine, and desensitization of the strips to bradykinin did not modify the action of neurotensin. The contraction phase was inhibited in a concentration-dependent manner by indomethacin and acetylsalicylate, demonstrating that this activity of neurotensin was dependent on prostaglandin synthesis. The preparation responded to exogenous prostaglandins E1, E2, F1 alpha and F2 alpha with concentration-dependent contractions. The relaxation phase was abolished by verapamil and apamin, indicating the presence of inhibitory neurotensin receptors on smooth muscle cells that are linked to ionic channels.

Topics: Animals; Apamin; Aspirin; Drug Interactions; Guinea Pigs; Muscle Contraction; Muscle Relaxation; Muscle, Smooth; Neurotensin; Oligopeptides; Peptide Fragments; Peptides; Prostaglandins; Receptors, Neurotensin; Receptors, Neurotransmitter; Stomach; Verapamil; Xenopus Proteins

Neurotensin, neuromedin N and xenopsin induced a monophasic and concentration-dependent contraction of the intact guinea pig oesophagus but kinetensin was without effect. The responses were completely abolished by tetrodotoxin and atropine but were unaffected by hexamethonium. The maximum response induced by neurotensin was reduced (20-30%) by somatostatin and by dynorphin-(1-13) in a naloxone-reversible manner. Neurotensin did not contract the isolated muscularis mucosae. The effects of neurotensin-related peptides on the motility of the oesophagus are mediated exclusively through the release of acetylcholine.

Topics: Animals; Dynorphins; Esophagus; Female; Gastrointestinal Motility; Guinea Pigs; In Vitro Techniques; Male; Neurotensin; Oligopeptides; Peptide Fragments; Peptides; Somatostatin; Xenopus Proteins

The naturally occurring analogs of neurotensin-(8-13), xenopsin, [Lys8,Asn9]neurotensin-(8-13) (LANT-6) and neuromedin N stimulated the production of intracellular cyclic GMP in murine neuroblastoma clone N1E-115, an adrenergic neuronal cell type. The order of potency was neurotensin-(8-13) greater than neurotensin greater than xenopsin greater than neuromedin N greater than LANT-6. Furthermore, xenopsin, LANT-6 and neuromedin N each inhibited the specific binding of [3H]neurotensin to intact N1E-115 cells in a dose-related fashion. The order of affinity of the peptides for the neurotensin receptor was neurotensin-(8-13) greater than xenopsin greater than neurotensin greater than neuromedin N greater than LANT-6.

Topics: Animals; Clone Cells; Cyclic GMP; Mice; Neuroblastoma; Neurotensin; Oligopeptides; Peptide Fragments; Peptides; Receptors, Neurotensin; Receptors, Neurotransmitter; Xenopus Proteins

A novel nonapeptide with neurotensin-like immunoreactivity was isolated from pepsin-treated human plasma by dialysis, ion-exchange chromatography and high performance reversed-phase liquid chromatography. The amino acid sequence was determined by automated gas-phase sequence analysis as Ile-Ala-Arg-Arg-His-Pro-Tyr-Phe-Leu. Sequence homology with human serum albumin and with the biologically active peptides neurotensin and angiotensin is demonstrated. The name proposed for this peptide is kinetensin.

Topics: Amino Acid Sequence; Angiotensin I; Angiotensins; Animals; Cattle; Chickens; Chromatography, High Pressure Liquid; Humans; Microchemistry; Neurotensin; Oligopeptides; Pepsin A; Peptide Fragments; Peptides; Serum Albumin; Swine; Xenopus; Xenopus Proteins

A peptidase that cleaved neurotensin at the Pro10-Tyr11 peptide bond, leading to the formation of neurotensin-(1-10) and neurotensin-(11-13), was purified nearly to homogeneity from rat brain synaptic membranes. The enzyme appeared to be monomeric with a molecular weight of about 70,000-75,000 as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and high pressure liquid chromatography filtration. Isoelectrofocusing indicated a pI of 5.9-6. The purified peptidase could be classified as a neutral metallopeptidase with respect to its sensitivity to pH and metal chelators. Thiol-blocking agents and acidic and serine protease inhibitors had no effect. Studies with specific peptidase inhibitors clearly indicated that the purified enzyme was distinct from enzymes capable of cleaving neurotensin at the Pro10-Tyr11 bond such as proline endopeptidase and endopeptidase 24-11. The enzyme was also distinct from other neurotensin-degrading peptidases such as angiotensin-converting enzyme and a recently purified rat brain soluble metalloendopeptidase. The peptidase displayed a high affinity for neurotensin (Km = 2.6 microM). Studies on its specificity revealed that neurotensin-(9-13) was the shortest neurotensin partial sequence that was able to fully inhibit [3H]neurotensin degradation. Shortening the C-terminal end of the neurotensin molecule as well as substitutions in positions 8, 9, and 11 by D-amino acids strongly decreased the inhibitory potency of neurotensin. Among 20 natural peptides, only angiotensin I and the neurotensin-related peptides (xenopsin and neuromedin N) were found as potent as unlabeled neurotensin.

Topics: Animals; Brain; Chromatography, High Pressure Liquid; Chromatography, Ion Exchange; Electrophoresis, Polyacrylamide Gel; Isoelectric Focusing; Kinetics; Molecular Weight; Neurotensin; Octoxynol; Oligopeptides; Peptide Fragments; Peptide Hydrolases; Peptides; Polyethylene Glycols; Protease Inhibitors; Rats; Synaptic Membranes; Xenopus Proteins