Effects of ACTH(1-24) and ACTH/MSH(4-10) on isolation-induced distress vocalization in domestic chicks

The effects of centrally administered ACTH(1-24) and ACTH(4-10) on isolation-induced distress vocalizations (DVs) were assessed in the presence or absence of social cues (mirrored and plain environments). A dose-response analysis indicated that ACTH(1-24) at doses of 0.5 nM and above increased DVs relative to controls when the animals were tested in mirrored or social environments which reduce baseline levels of calling. This effect, however, was short-lived (approx. 15 min). When tested again 1 hr after injection, the treated animals did not differ from controls. ACTH/MSH(4-10) had no effect on vocalization when the animals were tested immediately after injection, but marginally increased calling when animals were tested an hour later. In addition to vocalization changes, ACTH(1-24) induced squatting when animals were isolated in the test boxes, and yawning, head shaking, wing flapping and preening when animals were reunited after testing. ACTH(1-24)-treated chicks also exhibited longer latencies to close their eyes when they were held in the cupped hands of the experimenter. Taken together, the results suggest that ACTH(1-24) induces a central state of arousal in chicks that resembles fear/anxiety.     

Intranasal administration of ACTH(1-24) stimulates catecholamine secretion.

Previously, we reported that intranasal (IN) ACTH(1-24) administration stimulates adrenocortical steroid secretion in normal subjects. To determine the efficiency of transmucosal absorption of ACTH into the adrenal medulla, we measured serum cortisol, aldosterone, epinephrine, norepinephrine and dopamine levels after IN vs. intravenous (IV) administration of 250 microg ACTH(1-24) in 7 healthy adult men (mean age 21.7 +/- 1.2 yr; range, 21 - 24 yr). Blood was collected at 0, 30, 60 and 120 min after administration of ACTH(1-24), and the levels of adrenocortical steroids and catecholamines were measured by specific RIA and HPLC methods, respectively. There were no side effects associated with IN or IV ACTH administration. Consistent with the previous study, serum cortisol and aldosterone increased after IN administration of ACTH(1-24), peaking 30 min after administration. Sixty minutes after IN and IV administration of ACTH, epinephrine levels increased by 41.9 +/- 13.1 % and 63.3 +/- 11.8 %, respectively, and remained elevated throughout the sampling period. Thirty minutes after IN or IV administration of ACTH(1-24), plasma norepinephrine levels increased by 55.9 +/- 13.4 % and 73.7 +/- 15.0 %, respectively, peaking 30 min after ACTH(1-24) administration, and decreasing to basal levels within 60 min. Plasma dopamine levels did not change after IN administration of ACTH(1-24). Adrenocortical steroid and catecholamine levels did not increase after IN administration of saline. These results demonstrate that IN administration of ACTH(1-24) not only stimulates adrenocortical steroids, but also epinephrine and norepinephrine.     

Effects of continuous alpha (1-24) ACTH infusion in the dog

Dogs chronically infused with alpha (1-24) ACTH for 2 weeks showed continuous elevations in plasma ACTH, cortisol, and progesterone levels. Haematologic changes included immediate increases in numbers of mature neutrophils and monocytes and reduced numbers of eosinophils and lymphocytes. Haematocrits were also reduced with ACTH infusion. Whereas serum potassium levels fell in association with ACTH, serum sodium was unchanged. Activities of two serum enzymes of probable liver origin, alkaline phosphatase and alanine aminotransferase, increased gradually with ACTH treatment. Histologic examination of liver tissue revealed prominent hepatocellular vacuolisation. The trophic action of ACTH infusion was manifested by an increased adrenal gland weight and an enhanced cortisol response to a bolus ACTH injection given 1 day after the infusion ceased. Long-term infusion of ACTH resulted in haematologic, biochemical and morphologic changes resembling those observed in dogs with spontaneous pituitary-dependent hyperadrenocorticism.     

Isolation of ACTH1-39,ACTH1-38 and CLIP from the calf anterior pituitary

Calf anterior pituitaries were defatted and homogenized and peptides were adsorbed from the homogenate supernatant onto octadecylsilyl-silica. After elution, the resulting extract was subjected to gradient elution reversed-phase high pressure liquid chromatography (RP-HPLC) using aqueous acetonitrile containing 0.1% ($\text{v}\text{v}$) trifluoroacetic acid (TFA). Radioimmunoassay of column fractions for corticotropin (ACTH) revealed three major areas of immunoreactivity. Each was purified to homogeneity by gradient elution RP-HPLC employing aqueous acetonitrile containing either 0.13% heptafluorobutyric acid ($\text{v}\text{v}$) or 0.1% TFA ($\text{v}\text{v}$). Amino acid analysis and exopeptidase and trypsin digestions revealed the three forms of corticotropin to be ACTH_1–38, corticotropin-like intermediary lobe peptide, (CLIP, ACTH_18–39) and ACTH_1–39. ³H-labeled ACTH_1–39 did not give rise to either ³H-ACTH_1–38 or ³H-CLIP during isolation.     

Adrenocorticotropin radioimmunoassay: properties of antisera against synthetic ACTH(1-24) and its clinical application

Two antisera against synthetic ACTH(1-24) developed in rabbit showed strikingly different affinities toward the ACTH molecule. Both antisera (A-6 and A-7) were highly specific for the COOH-terminal region of ACTH(1-24). Antisera A-6 recognized ACTH(1-39) poorly. Radioimmunoassays (RIAs) using these antisera permitted the rapid (less than or equal to 18 h) quantitation of ACTH(1-24) (A-6) or ACTH(1-39) (A-7) at picogram levels. ACTH levels were determined on silicic acid extracts of rat and human plasma samples by the RIA specific for mid-region of ACTH(1-39) (A-7) and compared with that obtained by an ACTH(34-39) (C-terminal) RIA. In nearly all cases the C-terminal/mid-region ACTH ratios were less than 1.0, indicating that C-terminus of ACTH is more readily degraded by tissue or blood peptidases than are internal sequences. A solid-phase immunoadsorbent RIA specific for the extreme COOH-terminus of ACTH(1-24) was developed by coupling antiserum (A-6) to Sepharose 4B. This assay exhibited the same specificity as the soluble antiserum, yet tolerated relatively high concentrations of protein. Although the assay was suitable for rapid quantitation of ACTH(1-24), a decrease in sensitivity was observed in comparison to a conventional assay.     

Centrally administered N-terminal fragments of ACTH (1-10, 4-10, 4-9) display convulsant properties in rabbits

Electroencephalographic and behavioral effects of the following ACTH fragments: 1-4, 4-9, 4-11, 1-10, 4-10, 1-13, 1-17 and 1-24 were studied in rabbits. Sequences 4-9, 1-10 and 4-10 displayed some epileptic properties, i.e., they induced epileptic seizures (only electrographic or also behavioral) or increased hippocampal spiking. The 4-9 sequence seemed to be the common sequence responsible for these proconvulsant effects. The possible involvement of the enkephalinergic system is discussed.     

Influence of intracerebroventricular (ICV) injection of ACTH (1-24) on plasma gonadotropin in female rats: dose-response study

The intracerebroventricular (ICV) injection of ACTH (1-24) (0.1, 1.0 and 2.5 micrograms) to adult conscious ovariectomized (OVX) rats caused a dose-related inhibition of plasma LH at 10 min postinjection. The ICV injection of ACTH (1-24) (2.5 micrograms) to OVX rats in the absence or presence of a single dose of estradiol benzoate (OVX + EB): a) Decreased significantly plasma LH levels in OVX rats at 10 and 30 min postinjection. b) Decreased significantly plasma LH levels in (OVX + EB) rats at 10 min but not at 30 min postinjection. c) Did not change plasma FSH levels at 10 or 30 min postinjection in both (OVX) or (OVX + EB) rats. d) Did not change plasma ACTH levels at 10 or 30 min postinjection in (OVX) rats. Our observation suggest that ACTH (1-24) inhibited plasma LH, possibly through brain sites of action.     

Antioxidant activity and low toxicity of (E)-1-(1-(methylthio)-1-(selenopheny) hept-1-en-2-yl) pyrrolidin-2-one

The aim of the present study was to evaluate the potential pharmacological and toxicological properties of (E)-1-(1-(methylthio)-1-(selenopheny) hept-1-en-2-yl) pyrrolidin-2-one (compound 1), an organoselenium compound. In vitro experiments showed that compound 1 presented a reduction in the lipid peroxidation induced by Fe2? in thiobarbituric acid-reactive species (TBARS) production, and in the generation of reactive species caused by Fe2?/malonate in DCFH-DA oxidation. The high dose (500 mg/kg) induced an increase on ALT but not on AST activity. Hepatic, but not cerebral, δ-ALA-D activity from mice treated with 500 mg/kg presented a significant inhibition. Brain catalase activity was significantly inhibited by 100 mg/kg whereas hepatic catalase activity showed a significant increase at all doses. Hepatic lipid peroxidation was diminished only at lowest dose (100 mg/kg) whereas for brain tissue, all doses induced a significant reduction in TBARS levels. Brain and liver ascorbic acid contents were increased only at highest dose of compound 1. Urea and creatinine levels were not significantly altered by treatments. This is a promising compound with antioxidant activity and low toxicity, suggesting the potential beneficial activity of compound 1 against oxidative damage in many parameters studied in rats and mice.     

ACTH1-24 stimulates muscle cell glucose uptake

3H-2-deoxyglucose (2-DG) uptake was measured in L6A-1 rat skeletal muscle cells (a rapidly fusing subclone of L6), following addition of several concentrations (10(-16) to 10(-9)M) of the N-terminal fragment of ACTH1-24 to cells deprived of serum and insulin for 21 hours, but maintained in the presence of (5 micrograms/ml) insulin (stimulated state). There was a marked dose-dependent increase of 2-DG uptake at the various ACTH1-24 (P less than 0.001). There was no correlation between the time of exposure of the cells to serum-free conditions and the rate of uptake of 2-DG at the various ACTH1-24 concentrations both in the basal and insulin-stimulated states. Addition of catochalasin B (50 microM) to the cells, which inhibited both basal and insulin-stimulated uptake of 2-DG (by 70% and 91%, respectively) completely eliminated the enhancement of both of these uptake rates to 10(-12)M ACTH1-24. The results suggest that: 1) ACTH1-24 stimulates carrier-mediated uptake of glucose in skeletal muscle cells. 2) The site of action of ACTH1-24 is on the non-insulin mediated glucose uptake (NIMGU) system. 3) ACTH1-24 may be a useful probe to delineate some of the events associated with the NIMGU pathway.     

2(S)-(Cycloalk-1-enecarbonyl)-1-(4-phenyl-butanoyl)pyrrolidines and 2(S)-(aroyl)-1-(4-phenylbutanoyl)pyrrolidines as prolyl oligopeptidase inhibitors

In order to replace the P2-P1 amide group, different 1-cycloalkenyls and 2-aryls were studied in the place of the P1 pyrrolidine group of a 4-phenylbutanoyl-L-Pro-pyrrolidine structure, which is a well-known prolyl oligopeptidase inhibitor SUAM-1221. The 1-cyclopentenyl and the 2-thienyl groups gave novel compounds, which were equipotent with the corresponding pyrrolidine-analog SUAM-1221. It was shown that the P2-P1 amide group of POP inhibitors can be replaced by an alpha,beta-unsaturated carbonyl group or the aryl conjugated carbonyl group.     

Comparison between adrenal steroid secretion after stimulation with two synthetic analogues of corticotropin : ACTH 1-24 and ACTH 1-17

Chemical manipulations of the native ACTH (1-39) molecule led to the synthesis of a newly characterized compound which shows several interesting features, such as an enhanced half-life, an increased resistance against enzymic degradation and lower immunogenic activity. This study aimed at comparing the adrenal steroidogenetic response of the 1-17 analogue and of the other commercially available synthetic ACTH (1-24). Results suggest that ACTH 1-17 has a more pronounced and prolonged activity, which is confined to glico- and mineralocorticoid secretion.     

NMR structural characterization of cecropin A(1-8) - magainin 2(1-12) and cecropin A (1-8) - melittin (1-12) hybrid peptides.

In order to elucidate the structure-antibiotic activity relationships of the peptides, the three-dimensional structures of two hybrid peptides, CA(1-8) - MA(1-12) and CA(1-8) - ME(1-12) in trifluoroethanol-containing aqueous solution were investigated by NMR spectroscopy. Both CA(1-8) - MA(1-12) and CA(1-8) - ME(1-12) have strong antibacterial activity but only CA(1-8) - ME(1-12) has hemolytic activity against human erythrocytes. CA(1-8) - MA(1-12) has a hydrophobic 310-helix of only two turns combined with one short helix in the N-terminus with a flexible hinge section in between. CA(1-8) - MA(1-12) has a severely bent structure in the middle of the peptide. These structural features as well as the low hydrophobicity of CA(1-8) - MA(1-12) seem to be crucial for the selective lysis against the membrane of prokaryotic cells. CA(1-8) - ME(1-12) has an alpha-helical structure of about three turns in the melittin domain and a flexible structure with one turn in the cecropin domain connected with a flexible hinge section in between, and these might be the structural features required for membrane disruption against prokaryotic and eukaryotic cells. The central hinge region (Gly9-Ile10-Gly11) in an amphipathic antibacterial peptide is considered to play an important role in providing the conformational flexibility required for ion channel formation of the C-terminal hydrophobic alpha-helix on cell membrane.     

Tetracosactide (ACTH 1-24) stimulates the external pancreatic secretion by a vagal mechanism in the rat

The use of ACTH has been recently proposed in the therapy of the withdrawal syndrome in opiate addicts. By analogy with previous results obtained with opiates and clonidine, we looked for a possible central effect of ACTH 1-24 (tetracosactide) on the nervous pathways controlling the external pancreatic secretion in the rat. Tetracosactide displayed a central, vagally mediated, stimulant effect on the external pancreatic secretion, at variance with opiates and clonidine, which displayed central inhibitory effects in the same model. These results suggest that tetracosactide has a mechanism of action different from clonidine.     

Solution conformation of asparagine-linked oligosaccharides: alpha(1-2)-, alpha(1-3)-, beta(1-2)-, and beta(1-4)-linked units

The solution conformation is presented for representatives of each of the major classes of asparaginyl oligosaccharides. In this report the conformation of alpha(1-3)-, alpha(1-2)-, beta(1-2)-, and beta(1-4)-linked units is described. The conformational properties of these glycopeptides were determined by high-resolution 1H nuclear magnetic resonance in conjunction with potential energy calculations. The NMR parameters that were used in this analysis were chemical shifts and nuclear Overhauser enhancements. Potential energy calculations were used to evaluate the preferred conformers available for the different linkages in glycopeptides and to draw conclusions about the behavior in solution of these molecules. It was found that the linkage conformation of the Man alpha 1-3 residues was not affected by substitution either at the 2-position by alpha Man or beta GlcNAc or at the 4-position by beta GlcNAc or by the presence of a bisecting GlcNAc on the adjacent beta Man residue.     

Structural and functional organization of ACTH: synthesis and properties of analogs of ACTH-(11-24)-tetradeca- and ACTH-(1-24)-tetracosapeptides containing hexa-amino acids instead of a natural sequence of ACTH 19-24 amino acids

To assess the role of amino acid sequence ACTH 19-24 in the corticotropin structure and steroidogenic activity, the analogues of ACTH-(11-24)-tetradeca- and ACTH-(1-24)-tetracosapeptides containing hexaglycine, hexaphenylalanine, hexaglutamic acid or hexalysine instead of the natural 19-24 sequence have been synthesized by conventional methods. All these compounds in water have the CD curves characteristic of random coil, CD spectra of analogue ACTH-(1-24)-tetracosapeptide and hexalysine-containing analogue ACTH-(11-24)-tetradecapeptide in trifluoroethanol indicate the presence of alpha-helices. The latter compound manifested higher steroidogenic activity than ACTH-(11-24)-tetradecapeptide. All the other analogues were either less active than ACTH-(1-24)-tetracosapeptide or inactive over the concentration range 10(-5)-10(2) mg/ml, thereby testifying to functional importance of the 19-24 sequence for manifesting full steroidogenic activity.