Effect of DN-1417, a thyrotropin releasing hormone analog, on dopaminergic neurons in rat brain

Acute and chronic effects of γ-butyrolactone-γ-carbonyl-histidyl-prolinamide (DN-1417) were investigated on motor activity, dopamine (DA) metabolites and DA receptors in various brain regions of rats. The motor activity, as measured with Automex recorder, was enhanced after a single injection with DN-1417 (20 mg/kg, IP), and the motor stimulating action persisted during 21 daily injections. Acute DN-1417 elevated both homovanillic acid (HVA) and 3,4-dihydroxyphenylacetic acid (DOPAC) levels in 7 brain regions, prefrontal cortex polar, medial and lateral fields, nucleus accumbens, olfactory tubercles, amygdala and striatum. After chronic treatment for 7 days, the acute effect of DN-1417 on DA metabolites disappeared in all regions except for the striatum in which DN-1417 still increased HVA and DOPAC. The response of striatal DA metabolites was also observed after chronic treatment for 21 days. Chronic DN-1417 produced no significant change in ³H-spiperone binding in the prefrontal cortex, nucleus accumbens, olfactory tubercles and striatum, while striatal ³H-DA binding displaced by 30 nM spiperone was enhanced after chronic treatment. These results indicate that DN-1417 interacts with mesocortical, mesolimbic and nigrostriatal DA systems in the different modes of action. The lack of tolerance to motor hyperactivity, however, raises the question as to whether DN-1417-induced hyperactivity may be mediated by the activation of mesolimbic DA neurons. The involvement of nigrostriatal neurons in DN-1417-induced motor hyperactivity is suggested.     

Analysis of apolipoproteins in high density lipoproteins by high performance liquid chromatography

A simple and rapid method for the analysis of apolipoproteins in high density lipoprotein (HDL) by high performance liquid chromatography (HPLC) has been developed (Kinoshita et al. (1983) J. Biochem. 94, 615-617). With this method, using a sodium phosphate buffer containing 0.1% sodium dodecyl sulfate (SDS) as an eluent, apolipoproteins can be analyzed from a very small amount of HDL fraction without delipidation using organic solvents. Separation profiles of apolipoproteins by this method were examined using several techniques. The elution pattern monitored by A280 can give precise quantitative as well as qualitative information about size-distribution of apolipoproteins, except for the apo C group. Moreover, separation of apo E from apo A-I was found to be improved by column elongation.     

Morphology of spines and spine joint in the crown-of-thorns starfish Acanthaster planci (Echinodermata,Asteroida)

Summary Morphology and movement of the spines of Acanthaster planci were studied. All surfaces of the animal are covered with spines. The spines on the aboral surface are cylindrical with sharp tips. The spines on the oral surface are flat; they bend over to cover the mouth and the ambulacral grooves when these soft parts are stimulated. Those on the side of the animal make a barrier of crossed spines. Thus the structure and movement of the spines are well-adapted for defense.The junction between the primary aboral spine and its pedicel makes a movable joint. The ultrastructure of the connective tissue at the joint was studied. The connective tissue is mainly composed of collagen fibers. Presumed neurosecretory cells with processes which are filled with electron-dense granules of 0.2 m diameter were found between collagen fibers. Muscle fibers are mainly found in the connective tissue at the central holes. These observations support the view that the joint connective tissue has catch properties.     

Intracistronic mapping of the defective site and the biochemical properties of beta subunit mutants of Escherichia coli H+-ATPase: correlation of structural domains with functions of the beta subunit

Sixteen mutants of Escherichia coli defective in H+-ATPase (proton-translocating ATPase) were tested for their ability to recombine with hybrid plasmids carrying various portions of the beta subunit cistron. Twelve mutations were mapped within the carboxyl half of the cistron corresponding to amino acid residues 279 to 459 (domain II), while four mutations were mapped within residues 17 to 278 (domain I). The biochemical properties of these mutants were analyzed in terms of the proton permeability of their membranes and the assembly properties of their F1F0 complex. The mutants were classified according to the properties into three types, I, II, and III. In 12 mutants of type I, proton conduction in membrane vesicles was blocked and little F1 was released from the membranes under conditions in which F1 could be released from wild-type membranes, suggesting that assembly of the F1F0 complex is structurally and functionally defective. F1 was partially purified with very low recovery from one of the type I mutants, KF16. ATPase activity was reconstituted from this F1 with the beta subunit of the wild type, confirming the genetic results. Only one mutant, KF38, was classified as type II. Its membranes were partially leaky to protons and its F1 was releasable, suggesting that the interaction of its F1 and F0 was unstable. Type III mutants, KF11 and KF43, had an F1F0 complex with very low activity, in which the structure of F1 was relatively similar to that of the wild type. F1 was purified as a single complex from KF43 in this study and from KF11 previously (H. Kanazawa, Y. Horiuchi, M. Takagi, Y. Ishino, and M. Futai (1980) J. Biochem. 88, 695-703). Reconstitution experiments in vitro showed that the F1's of both mutants were defective in the beta subunit. The properties of the altered F1 of KF43 differed from those of F1 of KF11, suggesting that the mutation sites of KF43 and KF11 were different. From the results of mapping mutation sites and the biochemical properties of the mutants, the correlation of structural domains with function of the beta subunit is discussed. Most type I and type II mutations except that of KF39 were mapped in domain II, while the type III mutations were mapped in domain I, suggesting that domain II is more important than domain I for the function of the beta subunit, especially in terms of proper assembly of the F1F0 complex.     

Possible physiological role of epidermal growth factor in the development of the mouse mammary gland during pregnancy

Epidermal growth factor stimulated cell proliferation in a primary mammary epithelial cell culture derived from mice at different stages of pregnancy. Moreover, the peptide hormone inhibited casein production induced by the synergistic actions of insulin, cortisol and prolactin. The inhibitory effect of epidermal growth factor was influenced by the gestational stages of the mammary gland. These effects of epidermal growth factor were exerted at physiological concentrations. The dual actions of epidermal growth factor on mammary cells implicate its participation in regulation of the growth and differentiation of the mammary gland during pregnancy.     

The large intracellular pool of asialoglycoprotein receptors functions during the endocytosis of asialoglycoproteins by isolated rat hepatocytes

The function of intracellular asialoglycoprotein receptors during the endocytosis of asialo-orosomucoid in isolated hepatocytes was assessed by following changes in the occupancy of intracellular receptors. Unoccupied total cellular (inside and surface) or surface receptors were quantified at 0 degrees C by the binding of 125I-asialo-orosomucoid in the presence or absence, respectively, of digitonin. Freshly isolated cells had about 17% of their total receptors on the surface. After incubation at 37 degrees C, the receptor distribution changed to 25 to 50% on the cell surface and 50 to 75% inside the cell. At 37 degrees C, the average total number of receptors/cell was 4.5 x 10(5). Dissociation constants, determined from equilibrium binding studies in the presence or absence of digitonin to assess total or surface receptors, were identical (5.4 +/- 1.4 and 5.6 +/- 1.1 x 10(-9) M, respectively). In the presence of asialo-orosomucoid at 37 degrees C, there was both a time- and a concentration-dependent decrease in surface and intracellular receptor activity. This receptor activity decrease was reversed by removing asialo-orosomucoid from the medium or by washing the digitonin-permeabilized cells with ethylene glycol bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid prior to quantification of receptor activity. Within 1 to 2 h in the presence of excess asialo-orosomucoid, a steady state was attained in which approximately 70% of the intracellular receptors were occupied. The kinetics of receptor activity recovery on the cell surface after internalization of a pulse of ligand is different than the rate of recovery of internal receptor activity. The results suggest that all of the internal asialoglycoprotein receptors are functional and participate during endocytosis. Internal receptors may be functionally equivalent to those on the surface or they may serve a reservoir or routing function for internalized ligand.     

Adsorption of mutagens to cotton bearing covalently bound trisulfo-copper-phthalocyanine

A method for separating nonpolar mutagens from their dilute aqueous solutions is described. It utilizes the affinity of the mutagens to a phthalocyanine derivative attached to cotton through a covalent bond. For mutagens having 3 or more fused aromatic rings in their structures, efficient adsorption took place on soaking the cotton in their solutions. The mutagens adsorbed can be recovered by elution with ammoniacal methanol. Mutagenicity in smoker's urine, cooked beef, and river water was detected by use of this method.     

Characterization of tyrosine hydroxylase from bovine adrenal medulla

Tyrosine hydroxylase purified to apparent homogeneity from the soluble fraction of bovine adrenal medulla had an apparent Mr of about 280,000 by Bio-Gel A-1.5m chromatography, and gave a single band with a Mr of 60,000 by sodium dodesyl sulfate polyacrylamide gel electrophoresis. The enzyme is considered to be composed of four identical subunits. Isoelectric point of purified enzyme was pH 6.0. The amino acid composition of the enzyme was characterized by fairly high contents of glutamic acid and alanine residues. The N-terminal amino acid was determined to be glutamic acid.     

Dichroism of bacteriochlorophyll in cells of the photosynthetic bacterium Rhodopseudomonas palustris

1. The dichroism was measured at varied wavelengths of polarized light in “intact” cells of Rhodopseudomonas palustris and in films of air-dried lamellar fragments of the bacterium.