Neurotensin receptors on the rat liver plasma membranes

Neurotensin (NT) is now classified as a brain-gut peptide in the central nervous system and gastrointestinal tract. In the present study, we characterized the NT receptors on the rat liver plasma membranes. The specific binding of [3H]NT was time dependent, reversible, and saturable. Scatchard analysis of the specific binding data yielded two classes of binding sites, a high affinity site and a low affinity site. The average maximum number of binding sites (Bmax) amounted to 13.3 +/- 1.1 fmol/mg protein at high affinity site and 122.3 +/- 21.5 fmol/mg protein at low affinity site, respectively. The dissociation constant (Kd) had values of 0.39 +/- 0.01 nM at high affinity site and 8.1 +/- 1.1 nM at low affinity site, respectively. The amount of specifically bound [3H]NT was significantly reduced in the presence of mono and divalent cations, EDTA, EGTA and a peptidase inhibitor bacitracin, NT1-13 competed with [3H]NT for its binding site with an IC50 of 0.19 nM at high affinity site (0.2 nM concentration of [3H]NT) and 0.7 nM at low affinity site (4.0 nM concentration of [3H]NT). Xenopsin, a NT analogue separated from the skin of Xenopus laevis, was equipotent (IC50 0.75 nM) with NT1-13 at 4.0 nM concentration of [3H]NT. C-terminal sequence of NT contains the structure necessary for interaction with NT binding sites whereas N-terminal sequence had no binding activity. Since NT has a hyperglysemic and a hypercholesterolemic effects in rats, these NT receptors on the rat liver plasma membranes may be involved in the hyperglycemia and/or hypercholesteroremia induced by NT.     

Effects of bombesin and gastrin releasing peptide on catecholamine secretion from rat adrenal gland, in vitro

The effects of bombesin and gastrin releasing peptide (GRP) on the release of catecholamine were investigated by using isolated rat adrenal gland. Bombesin and GRP stimulated an epinephrine (E) release with dose-dependency. A half maximal effect of bombesin was observed at 1.2 X 10(-9) M, and a maximal release of E occurred at 1 X 10(-6) M of bombesin. The stimulatory effect of GRP on the E release was very similar to that of bombesin. Although both these peptides also stimulated a norepinephrine (NE) release, a significant effect was detected at concentrations of bombesin and GRP above 1 X 10(-7) M. Nicotine and pilocarpine stimulated both E and NE releases dose dependently, but the effect of pilocarpine on E and NE release was 1/100 or less potent than that of nicotine. Bombesin-induced catecholamine releases were not inhibited by hexamethonium or atropine that fully impeded the stimulatory effects of nicotine or pilocarpine. In addition, bombesin had additive effects on the nicotine- or pilocarpine-induced E and NE releases. These data strongly suggest that bombesin or GRP plays a physiological role as one of the important regulators in catecholamine secretion in the adrenal gland.     

Isolation and chemical characterization of glicentin C-terminal hexapeptide in porcine pancreas

Using a radioimmunoassay specific for porcine glicentin C-terminal hexapeptide, we isolated a peptide from porcine pancreas and characterized it as the C-terminal 64-69 sequence of glicentin: H-Asn-Lys-Asn-Asn-Ile-Ala-OH. The purification steps included gel filtration, ion-exchange chromatography and HPLC. In each step, the recovery of the desired peptide, radioimmunologically estimated from the respective elution profile, was 71.4-91.7%. The final yield of the hexapeptide was 22 micrograms (4.3%) from 800 g pancreas. The pancreatic content of this peptide was estimated to be approximately equimolar to that of pancreatic glucagon. No hexapeptide-like component was detected in porcine intestinal extracts. The data confirmed that the processing of pancreatic proglucagon liberates the C-terminal hexapeptide of the intramolecular glicentin sequence in a tissue-specific manner during the production of glucagon.     

Effects of synthetic peptides on giant neurons identified in the ganglia of an African giant snail (Achatina fulica Férussac)--II

Thirteen synthetic biologically-active peptides, which were classified into the peptides proposed as neurotransmitters in mammals and invertebrates and neural venom peptides, were investigated for their effects on the following six identifiable giant neurons of an African giant snail (Achatina fulica Férussac): RAPN (right anterior pallial neuron), INN (intestinal nerve neuron), RPeNLN (right pedal nerve large neuron), LPeNLN (left pedal nerve large neuron), d-LPeLN (dorsal-left pedal large neuron) and d-LPeCN (dorsal-left pedal constantly firing neuron). Oxytocin and proctolin at 10(-4)M excited the RAPN membrane potential, whereas FMRFamide at the same concentration inhibited the same neuron. FMRFamide at 10(-4)M markedly inhibited the d-LPeLN membrane potential, sometimes produced inhibition of RPeNLN and LPeNLN, showed varied effects (excitatory or inhibitory) on INN, and had no effect on d-LPeCN. The other peptides examined had almost no effect on any of the neurons tested.     

Effects of adrenergic blocking agents on lipolysis and adenyl cyclase activity induced by vasoactive intestinal polypeptide (VIP).

VIP stimulates lipolysis and adenyl cyclase activity in the rat adipose tissue. VIP-induced lipolysis and adenyl cyclase activity are not affected by phenoxybenzamine. VIP-induced lipolysis is inhibited by propranolol but VIP-induced adenyl cyclase activity is not.     

Somatostatin radioimmunoassay with 125I-Nalpha-tyrosyl-somatostatin

Nalpha-Tyrosyl-somatostatin was synthesized and proved to be homogeneous. Radioiodination of this tyrosine-containing somatostatin analogue by either the lactoperoxidase method or the chloramine T method led to the formation of crude iodinated compound, which was purified by ion exchange chromatography on CM-Sephadex C-25 using a linear ammonium acetate buffer gradient. This purification process was found to be satisfactorily reproducible and suitable for the preparation of 125I-Nalpha-tyrosyl-somatostatin. Using the purified 125I-somatostatin analogue, radioimmunoassay for somatostatin was performed and the assay system was proved to be sensitive and specific for somatostatin. Immunoassays of hot-water extracts of porcine and tupaia brain, pancreas, stomach and various regions of the intestine in the system revealed that those tissues contained immunoreactive somatostatin at various concentrations. Of the results, it was remarkable that somatostatin immunoreactivity was found in the ileum, middle colon and rectum in both animals, although the concentration were lower when compared with those in the stomach, duodenum and jejunum.     

Control of tomato bacterial wilt without disinfection using a new functional polymer that captures microbial cells alive on the surface and is highly biodegradable

This report describes a green chemical method for controlling soil-borne plant diseases without disinfection using an equimolar copolymer of N-benzyl-4-vinylpyridinium chloride with styrene (PBVP-co-ST) that captures microbial cells alive on the surface and is highly biodegradable. Tomato bacterial wilt caused by Ralstonia solanacearum was controlled by the addition of sawdust coated with PBVP-co-ST prior to transplantation. This effected 87% reduction in appearance and 89% reduction in the index of symptom under appropriate conditions. The coated sawdust did not exhibit bactericidal activity. The half-life of PBVP-co-ST was 5.6 d when treated with activated sludge in soil. The disease control was explained in terms of reduction of infectious contact between the roots of tomato and the cells of R. solanacearum due to coagulation-like interaction between microbial cells and the coated sawdust, in addition to capture of microbial cells by the coated sawdust.     

A new family of satellite DNA sequences as a major component of centromeric heterochromatin in owls (Strigiformes)

We isolated a new family of satellite DNA sequences from Hae III- and Eco RI-digested genomic DNA of the Blakistons fish owl ( Ketupa blakistoni). The repetitive sequences were organized in tandem arrays of the 174 bp element, and localized to the centromeric regions of all macrochromosomes, including the Z and W chromosomes, and microchromosomes. This hybridization pattern was consistent with the distribution of C-band-positive centromeric heterochromatin, and the satellite DNA sequences occupied 10% of the total genome as a major component of centromeric heterochromatin. The sequences were homogenized between macro- and microchromosomes in this species, and therefore intraspecific divergence of the nucleotide sequences was low. The 174 bp element cross-hybridized to the genomic DNA of six other Strigidae species, but not to that of the Tytonidae, suggesting that the satellite DNA sequences are conserved in the same family but fairly divergent between the different families in the Strigiformes. Secondly, the centromeric satellite DNAs were cloned from eight Strigidae species, and the nucleotide sequences of 41 monomer fragments were compared within and between species. Molecular phylogenetic relationships of the nucleotide sequences were highly correlated with both the taxonomy based on morphological traits and the phylogenetic tree constructed by DNA-DNA hybridization. These results suggest that the satellite DNA sequence has evolved by concerted evolution in the Strigidae and that it is a good taxonomic and phylogenetic marker to examine genetic diversity between Strigiformes species.An erratum to this article can be found at Communicated by Y. Hiraoka     

Do infant Japanese macaques (<Emphasis Type="Italic">Macaca fuscata</Emphasis>) categorize objects without specific training?

In the present study, we examined whether infant Japanese macaques categorize objects without any training, using a similar technique also used with human infants (the paired-preference method). During the familiarization phase, subjects were presented twice with two pairs of different objects from one global-level category. During the test phase, they were presented twice with a pair consisting of a novel familiar-category object and a novel global-level category object. The subjects were tested with three global-level categories (animal, furniture, and vehicle). It was found that they showed significant novelty preferences as a whole, indicating that they processed similarities between familiarization objects and novel familiar-category objects. These results suggest that subjects responded distinctively to objects without training, indicating the possibility that infant macaques possess the capacity for categorization.     

Glucagon, glicentin, proglucagon, PYY, PP and proPP-icosapeptide immunoreactivities of rectal carcinoid tumors and related non-tumor cells

Glucagon/PP-related peptides were detected immunohistochemically in 18 out of 22 cases of rectal tumors investigated. The reactive tumors showed prevalence of trabecular or mixed trabecular-acinar structure and moderate staining with Grimelius' silver and lead-hematoxylin. Three of the remaining 4 cases were characterized by reactivity for 5-hydroxytryptamine only, prevalence of a solid nest structural component and intense staining with Grimelius' silver technique and lead-hematoxylin. Fifteen of the 18 glucagon/PP-reactive cases were investigated immunohistochemically with a series of 6 sera directed against different sequences of glucagon, glicentin and proglucagon, and of 7 sera directed against PP, PYY and proPP-icosapeptide. A large spectrum of glucagon-related immunoreactivities, including C-terminus and mid-portion glucagon-immunoreactivity, N- and C-terminus glicentin-immunoreactivity, GLP1- and GLP2-immunoreactivity, were detected in human rectal L cells and most rectal carcinoids. With the exception of a few scattered cells in the rectal mucosa and in 3 tumors, C-terminus glucagon-immunoreactivity was obtained only after trypsin or subtilisin treatment of tissue sections. Both PYY and PP/proPP-like peptide(s) were detected in rectal L cells and carcinoids, with prevalence of PYY in normal cells and PP/proPP-like peptides in tumor cells. It is concluded that the same or closely related hormone/prohormone sequences are synthesized and stored in rectal endocrine cells and carcinoid tumors although differences of quantitative expression, post-translational cleavage or reactivity to antibodies may occur. The usefulness of protease treatments of tissue sections to unmask immunoreactivities of uncleaved propeptides or fixative-denatured peptides is outlined.