Antipyretic doses of α-MSH do not alter afebrile body temperature in the cold

1. 1.|α-MSH (1/2–13) was injected centrally and peripherally in rabbits exposed to cold.

2. 2.|Doses of the peptide previously shown to reduce fever had no effect on afebrile body temperature in the cold; larger doses did lower temperature.

3. 3.|We conclude that the antipyretic effect of small doses of α-MSH does not depend upon inhibition of central heat productioin and heat conservatioin pathways.

4. 4.|The antipyretic and hypthermic activities of α-MSH are independent and may represent different actions within the CNS.

Identification and characterization of beta1-adrenergic receptors in rat parotid membranes

• Beta-adrenergic receptor identification and properties are probed in rat parotid membranes utilizing the high affinity β-adrenergic antagonist(−)-[³H]dihydroalprenolol.

• The binding of (−)-[³H]dihydroalprenolol to membrane preparations of parotid is rapid, equilibrium being reached in 5 min. Strict stereospecificity is observed, (−)-propanolol being 100 times more potent than (+)-propranolol in competing with (−)-[³H]dihydroalprenolol for binding sites. Beta-adrenergic agonists compete for the binding sites with (−)-[³H]dihydroalprenolol with the same characteristics, i.e., much higher concentrations of the (+)-stereoisomers than the (−)-stereoisomers are required to produce 50% inhibition, the range varies from 14-fold for epinephrine to 300-fold for isoproterenol. Among the (−)-stereoisomers, the relative potency of inhibitory action is (−)-propranolol > (−)-isoproterenol > (−)-epinephrine ≡ (−)-norepinephrine. (−)-Isoproterenol is about 20 times as potent as norepinephrine, the least potent agonist among all the catecholamine (−)-stereoisomers.

• The binding of (−)-[³H]dihydroalprenolol is saturable, with a maximum number of binding sites equalling 450 fmol/mg protein and a dissociation constant of 7.9 nM. The Scatchard plots show no significant curvilinear character. Hill plots consistently give a Hill coefficient close to unity (0.92–1.05). Both pieces of evidence suggest a single-component system with no significant cooperativity.

• Dissociation kinetics study after the method of De Metys et al. (1973) Biochem. Biophys. Res. Commun. 155, 154, indicates a lack of site-to-site interactions among the binding sites. The rate of dissociation of bound (−)-[³H]dihydroalprenolol is the same in the presence and absence of 1 · 10⁻⁵ M (±)-alprenolol.

• Based on the binding of (−)-[³H]dihydroalprenolol, it is concluded that the beta-adrenergic receptors can be identified in rat parotid and that these binding sites display β₁ character. Results of the study indicate a one-component system with no observable site-to-site interactions.

Chemical characterization of sialyl oligosaccharides in milk of the Japanese black bear, Ursus thibetanus japonicus

Sialyl oligosaccharides were separated from two samples of Japanese black bear milk by extraction with chloroform/methanol, gel filtration on Bio Gel P-2, ion exchange chromatography on DEAE-Sephadex A-50 and high-performance liquid chromatography (HPLC) on a TSK gel Amido-80 column. They were characterized by ¹H-NMR spectroscopy. The structures of four sialyl oligosaccharides separated from the milk were the following:

Neu5Ac(α2-3)Gal(β1-4)Glc

Neu5Ac(α2-6)Gal(β1-4)GlcNAc(β1-3) Gal(α1-3)Gal(β1-4)[Fuc(α1-3)]GlcNAc(β1-6) Gal(β1-4)Glc

Neu5Ac(α2-6)Gal(β1-4)GlcNAc(β1-3) Gal(α1-3)[Fuc(α1-2)]Gal(β1-4)[Fuc(α1-3)]GlcNAc(β1-6) Gal(β1-4)Glc

Neu5Ac(α2-6)Gal(β1-4)GlcNAc(β1-3)[Neu5Ac(α2-6)Gal(β1-4)GlcNAc(β1-6)]Gal(β1-4)Glc

Determination of calcium transport and phosphoprotein phosphatase activity in microsomes from respiratory and vascular smooth muscle

1. 1. Calcium transport into microsomal vesicles of respiratory (tracheal) smooth muscle was characterized. This calcium transport was ATP dependent and stimulated by the presence of the oxalate ion. The magnitude of transport was similar to that reported for microsomes from other types of smooth muscle.

2. 2. Bovine and rabbit, heavy and light microsomes were isolated from respiratory (tracheal) and vascualar (aortic) smooth muscle. Preincubation of these vesicles with cyclic AMP and protein kinase did not alter the transport of calcium into the vesicles. There was no evidence of phosphate incorporatio into microsomal membrane proteins. Similar results were obtained if phosphorylase b kinase replaced the combination of cyclic AMP and protein kinase during the preincubation.

3. 3. The phosphoprotein phosphatase activity of cardiac sarcoplasmic reticulum and smooth muscle microsomes was determined. The activity of this enzyme was found to be several-fold less in the cardiac sarcoplasmic reticulum than in various smooth muscle microsome preparations.

EFFECT OF ULCERATIVE COLITIS ACTIVITY ON PLASMA CONCENTRATION OF TRANSFORMING GROWTH FACTOR β1

Enhanced expression of transforming growth factor-β₁(TGF-β₁) demonstrated in human colonic mucosa of patients with ulcerative colitis (UC), indicates its possible significance in the pathogenesis of this disease. The aim of this study was to evaluate plasma TGF-β₁concentration in patients with different degrees of colonic mucosal injury, as a possible indicator of ulcerative colitis activity. TGF-β₁concentration was measured with an enzyme immunoassay (EIA) in plasma of 45 patients with endoscopically confirmed UC. Values observed in UC patients (40.5±15.9 ng/ml) were significantly higher than in healthy people (18.3±11.6 ng/ml) and higher than in patients with irritable colon syndrome (ICS), (20.5±13.6 ng/ml). The highest plasma TGF-β₁(58.6±112.1 ng/ml) was in patients with the severe UC course. TGF-β₁level analysed in all UC patients revealed significant positive correlation with scored degree of mucosal injury (r=0.396;P<0.01). Among other possible laboratory markers of the disease activity, only C-reactive protein concentration demonstrated significant correlation. Enhanced production of TGF-β₁can be related to inflammation activity. Measurement of plasma TGF-β₁may be considered as a biomarker of the disease activity.     

Fluorimetric determination of the resting potential changes associated with the chemotactic response in Paramecium

1. (1) Evidence is presented which indicates that the carbocyanine dye (3,3′ dipropyl thiadicarbocyanine) can be used as a spectroscopic probe for monitoring the resting potential across the plasma membrane of the ciliated protozoan Paramecium.

2. (2) The dye at low concentrations ( 1 μM) does not affect either the viability or the motility of the cells, nor does it induce a chemotactic response.

3. (3) The fluorescence of the dye bound to the cells alters as the potential across the membrane is changed by increasing the external cation concentration.

4. (4) The absorbance of the bound dye also changes in response to an alteration of the membrane potential.

5. (5) The membrane potential changes as measured by the fluorescence method have been correlated with the measurements of the potential estimated by microelectrode methods.

6. (6) Both cations which induce a negative chemotactic response in Paramecium (K⁺, Na⁺, Ba²⁺) and several non-toxic cations bring about a rapid depolarization of the plasma membrane. The significance of these rapid changes in relation to the swimming behaviour of the ciliate is discussed.

alpha-Galactoside Binding Proteins from Plant Membranes: Isolation, Characterization, and Relation to Helminthosporoside Binding Proteins of Sugarcane

α-Galactoside binding proteins were isolated from cellular membranes of mint and tobacco as well as two clones of sugarcane which differ in their sensitivity to helminthosporoside, a toxic galactoside. Sodium trichloroacetate was used to disrupt membranes after which the proteins were purified using a melibiose-Sepharose-6B affinity column. Proteins from mint, tobacco, and susceptible sugarcane had equal electrophoretic mobilities, whereas resistant sugarcane protein migrated more slowly. Pretreatment of the proteins with fluorescamine caused them to migrate with the tracking dye. Each of the proteins had molecular weights of about 100,000 and each was shown to be oligomeric. Gel filtration revealed that aqueous solutions of these membrane proteins contained a mixture of size species which included a high molecular weight multimer and lower molecular weight oligomers. The relative abundance of the oligomers was dependent upon protein concentration: the lower concentrations yielded higher relative amounts of oligomers (Kenfield and Strobel 1980 Biochim Biophys Acta 600: 705-712). Also, the binding activity of these receptors was inversely proportional to protein concentration. At low protein concentration (4 micrograms per milliliter), the K_d's of each of the proteins for galactinol, raffinose, and helminthosporoside was about 10 micromolar. At high protein concentrations (100 micrograms per milliliter), mint and resistant sugarcane proteins failed to bind α-galactosyl ligands, whereas proteins from tobacco and susceptible sugarcane exhibited a markedly decreased binding activity compared to that at lower protein concentrations. Binding proteins from susceptible sugarcane were mixed with receptors from either resistant sugarcane or mint at low protein concentrations, then assayed for binding activity. Such mixtures showed a concentration-dependent decrease in binding activity analogous to the activity of homogeneous protein solutions. Bovine serum albumin, a nonsubunit protein, had no effect on the binding activity of the protein from susceptible sugarcane. Thus, receptors from diverse plants can associate in vitro and form functional oligomers. The amino acid composition of each of the binding proteins was similar but not identical. The significance of these results is discussed in regard to regulation of carbohydrate transport and sensitivity to phytotoxins.