Three types of alpha-melanocyte-stimulating hormone: bioactivities and half-lives.

Three types of alpha-melanocyte-stimulating hormone (alpha MSH) that differ in the acetyl status of the N-terminal serine have been found in the neurointermediate lobe of the pituitary gland and in the brain: desacetyl alpha MSH, which lacks an acetyl group; monoacetyl alpha MSH, in which the amino group of the serine is acetylated; and diacetyl alpha MSH, in which both amino and hydroxy groups of the serine are acetylated. We compared the lipolytic and melanotropic actions of these three peptides, and their rates of disappearance from plasma, both in vitro and in vivo. The following differences were found. a) For in vitro lipolytic actions on rabbit adipose tissue slices, the potencies differed according to the order diacetyl = monoacetyl greater than desacetyl. On rabbit isolated adipocytes, however, the three peptides were equipotent. b) For in vivo lipolytic action in the rabbit, not only potency but also kinetics differed. Diacetyl alpha MSH had the slowest onset, longest duration, and greatest potency. The desacetyl variant had the quickest onset, shortest duration, and least potency. c) The half-life for elimination from rabbit plasma both in vitro and in vivo was shortest for the desacetyl form and longest for the diacetyl peptide. d) For in vitro melanotropic effect on frog skin, kinetics of action were the same for all three peptides, but potency differed according to the order diacetyl = monoacetyl greater than desacetyl. Thus acetylation of alpha MSH alters lipolytic and melanotropic potencies in vitro and lipolytic potency and kinetics in vivo. These differences result in part from the fact that acetylation slows the degradation of the tridecapeptide both inside and outside the circulation.

Effects of opioid peptides and opiate alkaloids on insulin secretion in the rabbit.

In pentobarbital-anesthetized rabbits, iv injection of 9 nmol (31 micrograms) human beta-endorphin (beta h-endorphin)/kg BW caused a significant (P less than 0.05) increase in serum glucose and a significant decline in serum insulin during the subsequent 60 min. When 9 nmol/kg BW beta h-endorphin were injected simultaneously with 0.7 g glucose/kg BW, the clearance of serum glucose and the expected glucose-stimulated rise in serum insulin were both inhibited. The threshold dose for the insulinopenic effect of beta h-endorphin in the anesthetized, glucose-loaded rabbit was 0.09 nmol/kg BW. Threshold doses/kg BW were determined for six structurally related peptides found to possess insulinopenic activity: camel beta-endorphin, 0.09 nmol; N-arg-beta h-endorphin, 0.09 nmol; D-ala2-beta h-endorphin, 0.09 nmol; leu5-beta h-endorphin, 0.09 nmol; met-(O)5-beta h-endorphin, 0.9 nmol; and beta h-endorphin1-27, 0.9 nmol. Threshold dose/kg BW for somatostatin was 9 nmol. The following compounds were inactive at 9 nmol/kg BW: N-acetyl-beta h-endorphin; N-acetyl-arg-beta h-endorphin; beta h-endorphin2-31; beta h-endorphin6-31; beta h-endorphin(((1-5 + 6-31))); beta h-endorphin1-18 (gamma-endorphin); beta h-endorphin1-17 (alpha-endorphin); beta h-endorphin1-5 (met-enkephalin); leu5-beta h-endorphin (leu-enkephalin); met-NH2(5)-beta h-endorphin1-5 (met-enkephalinamide); D-ala2-leu5-beta h-endorphin1-5; D-ala2-N-me-phe4, met-(O)5-ol-beta h-endorphin1-5; and D-ala2-D-leu5-beta h-endorphin1-5. Ninety nmoles per kg BW of naloxone did not alter the insulinopenic effect of 0.9 nmol/kg BW beta h-endorphin. As little as 2.9 X 10(-10) molar beta h-endorphin inhibited glucose-stimulated release of insulin by rabbit pancreas slices in vitro. The capacities of the peptides and alkaloids to inhibit insulin secretion in vitro followed the same general order as the in vivo insulinopenic capacities. Naloxone at 2.9 X 10(-6) M did not reduce the antisecretagogue effect of 2.9 X 10(-8) M beta h-endorphin. These findings, when compared with previously described structure-activity relationships for opioid receptors, indicate the presence of a novel receptor for beta-endorphin in rabbit pancreas, the activation of which inhibits glucose-stimulated secretion of insulin.

Effects of melanotropic peptides on fetal adrenal gland.

Adrenal glands from early, mid, and late fetuses of rabbit, guinea pig, and rat, and from newborn animals of each species, were incubated for 1-4 h with and without 0.1 nM-1 microM ACTH, alpha- or beta-melanocyte-stimulating hormone (alpha MSH or beta MSH). The effects of the peptides were measured on production of glucocorticoids, and on incorporation of labeled thymidine or leucine into DNA or protein, respectively. The findings were similar in all three species. ACTH stimulated synthesis of glucocorticoids throughout fetal life. Potency increased progressively, as reflected by declining minimal effective dose and rising maximal response. In early and mid fetus alpha MSH and beta MSH caused a modest glucocorticoid steroidogenic effect. ACTH and alpha MSH stimulated DNA and protein synthesis in the early and mid fetal gland. alpha MSH was more potent than ACTH in these respects, minimal effective dose being generally 10 times less and maximal response 25-200% greater. The effects diminished or disappeared in the late fetal and newborn gland. These data indicate that alpha- and beta MSH possess steroidogenic or growth-promoting properties, or both, for the fetal adrenal gland.

Effects of hormones on 3', 5' -cyclic adenosine monophosphate in choroid plexus.

Choroid plexus of rabbit and rat was incubated for 2-30 min at 37 degrees C under 95% O2-5% CO2 in Tyrode solution containing 10 mM glucose and 1 mM theophylline with these agents: epinephrine, norepinephrine, isoproterenol, dopamine, histamine, serotonin, arginine, and lysine vasopressins, oxytocin, angiotensin, adrenocorticotropin (ACTH), beta-melanocyte-stimulating hormone, and choroid plexus peptide IIF. After incubation, tissue and medium were analyzed for 3', 5' -cyclic adenosine monophosphate (cAMP) content. Each amine or peptide was tested initially at 1,000 microng/ml. Only ACTH and serotonin affected cAMP content of rabbit choroid plexus. At 1,000 microng/ml, these agents caused a 10 and 4 times (respectively) increase in cAMP content of tissue + medium at 2-10 min with decline in content at 10-30 min. More than 90% of the increment was located in tissue, less than 10% in medium. Minimal effective dose (MED) to cause a significant (P less than .05) accumulation of cAMP was 0.1 microng/ml (2.2 x 10(-8) M) for ACTH and 10 microng/ml (5.7 x10(-3) M) for serotonin. Only isoproterenol, epinephrine, and norepinephrine influenced cAMP content of rat choroid plexus. MED's for this effect by isoproterenol, epinephrine, and norepinephrine were .001, .01, and 10 microng/ml (4.7 x 10(-9), 5.5 x 10(-8), and 5.9 x 10(-5) M), respectively.

Orosomucoid content of pleural and peritoneal effusions.

22 nonneoplastic, noninflammatory effusions (cirrhosis and congestive heart failure), 12 non-neoplastic inflammatory effusions (tuberculosis, lupus erythematosus, rheumatoid arthritis, and idiopathic pleuropericarditis), and 58 neoplastic effusions (cancer of lung, breast, ovary, and pancreas, and lymphoma) were analyzed by radial immunodiffusion for orosomucoid concentration. The average concentration +/-SE was 35+/-4, 65+/-17, and 130+/-13 mg/100 ml in the three types of effusion, respectively. By gel filtration and ion exchange chromatography, orosomucoid was isolated from 12 nonmalignant and 14 malignant fluids. The orosomucoid preparations reacted as single components in acrylamide gel electrophoresis at pH 9.0, and in immunodiffusion and immunoelectrophoresis against antisera to human serum and to human plasma orosomucoid. In radial immunodiffusion, the slope of the line relating concentration to the square of the diameter of the precipitate area was identical for orosomucoid isolated from normal human plasma and from nonneoplastic effusions, but was subnormal for orosomucoid isolated from neoplastic fluids. All orosomucoid preparations had normal amino acid composition. Orosomucoid from the nonmalignant effusions had normal carbohydrate content. 11 of 14 samples of orosomucoid isolated from neoplastic fluids had abnormalities in carbohydrate composition, consisting of subnormal content of sialic acid (11 of 14), hexose (10 of 14), and hexosamine (3 of 14), and abnormally high content of hexosamine (4 of 14). Discriminant analysis showed that concentration of orosomucoid distinguished between neoplastic and nonneoplastic noninflammatory effusions more effectively than concentration of total protein, albumin, alpha(1), alpha(2), beta, or gamma-globulin.