Ovine Beta-lipotropin Research Articles

Multiple feedback regulatory loops upon rat hypothalamic corticotropin-releasing hormone secretion. Potential clinical implications.

To examine whether the hypothalamic corticotropin-releasing hormone (CRH) neuron is regulated by CRH, by products of the proopiomelanocortin (POMC) gene, and/or by glucocorticoids, we used a rat hypothalamic organ culture system in which rat CRH secretion from single explanted hypothalami was evaluated by an RIA (iCRH) specific for rat CRH. The effects of graded concentrations of ovine CRH (oCRH), adrenocorticotropin hormone (ACTH), beta-endorphin (beta-EP), alpha-melanocyte-stimulating hormone (alpha-MSH), corticotropin-like intermediate lobe peptide (CLIP), ovine beta-lipotropin (ovine beta-LPH), and dexamethasone (DEX) upon unstimulated and serotonin- (5HT), acetylcholine- (ACh), and norepinephrine-(NE) stimulated CRH secretion were determined. oCRH and DEX inhibited unstimulated iCRH secretion with ID50 at the 10(-8) M range. ACTH had no detectable suppressive effect at 10(-8) M. oCRH, ACTH, and DEX inhibited 5HT-, ACh-, and NE-stimulated iCRH secretion in a dose-dependent fashion. beta-EP, alpha-MSH, and CLIP also inhibited 5HT-induced iCRH secretion. Of the latter peptides, the strongest inhibitor was beta-EP and the weakest was CLIP. Ovine beta-LPH had only a weak inhibitory effect on 5HT-induced iCRH secretion. Generally, the concentrations required for 50% suppression of neurotransmitter-stimulated iCRH secretion were significantly lower than those required for a similar suppression of unstimulated iCRH secretion. In conclusion, these data suggest the presence of multiple negative feedback loops involved in the regulation of the hypothalamic CRH neuron: an ultrashort CRH-mediated loop, a short, hypothalamic POMC-derived peptide loop, and a long, glucocorticoid-mediated negative feedback loop. The potency of these negative feedback loops may be determined by the state of activation of the CRH neuron.

Detection from rat pituitary of beta-lipotropin and materials containing opiatelike activity by combined enzymatic radioreceptor assay.

Tonin, a proteolytic enzyme isolated from rat submaxillary gland, was allowed to react upon ovine beta-lipotropin (beta-LPH) at 37 degrees C at a variety of pH values and for different lengths of time. Opiatelike activity generated by the reaction was assessed using a radioreceptor assay for beta-endorphin with rat brain homogenate. [3H]naloxone, and beta-endorphin as receptors, tracer, and hormone standard, respectively. Cleavage of beta-LPH with tonin produced a 10-fold increase in opiatelike activity as compared with beta-LPH alone. Digestion of beta-LPH with other enzymes such as renin, cathepsin D, trypsin, and chymotrypsin produced much less opiatelike activity. beta-Endorphin and methionine-enkephalin were not cleaved by tonin. Using this new assay, we were able to detect beta-LPH and materials containing opiatelike activity from rat pituitary extracts after gel chromatography. It is more specific and more sensitive than trypsin digest.

Synthesis and biological activity of ovine beta-lipotropin-(41--91)-henkaipentekontapeptide.

The 51-residue peptide ovine beta-lipotropin-(41--91) has been synthesized by the solid-phase method in about 5% overall yield. The synthetic product was characterized by partition chromatography on agarose gel, thin-layer chromatography in two solvent systems, paper electrophoresis at two pH values, polyacrylamide gel electrophoresis, amino acid analyses of acid and enzymic hydrolysates, and bioassay for lipolytic and melanotropic activities. The synthetic peptide is about 5.4 times as active on a weight basis as ovine beta-lipotropin in the lipolytic assay. In the melanotropic assay, it was about 2.4 times more active than the beta-lipotropin but only 5% as active as bovine beta-melanotropin. It had negligible opiate activity in the guinea pig ileum assay.

Conformation of beta-endorphin and beta-lipotropin: formation of helical structure in methanol and sodium dodecyl sulfate solutions.

Circular dichroic spectra of camel beta-endorphin and ovine beta-lipotropin in water show little, if any, secondary structure. Intrinsic viscosities and sedimentation coefficients of the two peptides also suggest that the molecules are not compact and globular. Methanol or sodium dodecyl sulfate promotes the formation of helical structure to an extent as much as one-half of either peptide molecule. The conformation of the complex between camel beta-endorphin and dodecyl sulfate may be related to the opiate-like function of this peptide hormone.

IMMUNOCHEMICAL INVESTIGATIONS OF HUMAN PITUITARY β‐LIPOTROPIN

Antiserum to human beta-lipotropin has been raised in rabbits and characterized by gel double diffusion, quantitative precipitin, microcomplement fixation and biological neutralization tests. A sensitive radioimmunoassay for human beta-lipotropin has been developed. It was demonstrated that the antiserum shows specificity for this hormone. Human beta-melanotropin and human beta-endorphin showed only a minimal degree of cross-reaction, while ovine beta-lipotropin showed a partial but significant degree of cross-reactivity.

Isolation and structure of an untriakontapeptide with opiate activity from camel pituitary glands.

The isolation of an untriakontapeptide from camel pituitary extracts has been described. Its structure has been determined and shown to be identical to the sequence of carboxyl-terminal 31 amino acids of ovine beta-lipotropin. The peptide possesses very low lipotropic activity but significant opiate activity.

Effect of ACTH, α-MSH, and β-Lipotropin on Calcium and Phosphorus Metabolism in the Rabbit

Studies were undertaken to investigate the effects of synthetic 1-24 adrenocorticotropin (ACTH), bovine alpha melanocyte-stimulating hormone (alpha-MSH), and ovine beta lipotropin (beta-LPH) on plasma calcium and phosphate in rabbits. Equimolar concentrations of these hormones were infused intravenously in intact and thyroidectomized animals. In addition, ACTH was similarly administered to adrenalectomized rabbits. ACTH, alpha-MSH, and beta-LPH all lowered plasma calcium and raised plasma phosphate. These changes were not prevented by prior thyroidectomy. ACTH was equally effective in inducing hypocalcemia and hyperphosphatemia in the absence of the adrenal glands, while adrenalectomy alone raised plasma calcium. From these findings we have concluded that 1) ACTH, alpha-MSH, and betaLPH affect phosphate as well as calcium metabolism; 2) these hormones do not act by releasing calcitonin; and 3) ACTH exerts its hypocalcemic-hyperphosphatemic effect, at least in part, independently of its trophic action on the adrenal glands.

Action of plasmin on ovine beta-lipotropin: Revision of the car☐yl terminal sequence

Human plasmin was found to cleave the lysylasparagine bond specifically in positions 79 and 80 of the ovine beta-lipotropin molecule. Structural studies of the COOH-terminal fragment revealed the amino acid sequence to be: Asn-Ala-Ile-Ile-Lys-Asn-Ala-His-Lys-Lys-Gly-Gln which is slightly different from the structure originally proposed. Together with an earlier revision of the NH 2-terminal sequence, the corrected structure of ovine beta-lipotropin is shown in Figure 2.