Alanine: glyoxylate aminotransferase 1 is present in the peroxisomes of guinea pig kidney

The subcellular distribution of alanine: glyoxylate aminotransferase 1 in guinea pig and rabbit kidneys was examined by centrifugation in a sucrose density gradient. The enzyme was located in the peroxisomes of guinea pig kidney and cross-reactive with the antibody against rat liver alanine: glyoxylate aminotransferase 1. This is the first report on the presence of the enzyme in the peroxisomes of mammalian kidney. The enzyme was found to be located in the mitochondria but not in the peroxisomes in rabbit kidney.     

Message amplification phenotyping of an inherited delta-aminolevulinate dehydratase deficiency in a family with acute hepatic porphyria

The molecular basis of the enzymatic defect responsible for acute hepatic porphyria due to delta-aminolevulinate dehydratase (ALAD) deficiency was investigated in a family including a proband with the acute disease. In order to delineate the mutation in the proband, cDNA for deficient ALAD was synthesized from the proband's cells. The ALAD phenotype was studied by message amplification phenotyping with total RNA extracted from lymphoblastoid cells of the proband and his family members. Two independent mutant alleles of ALAD were identified in the proband's cells. One mutant allele was shown to result in an amino acid substitution at residue 274 (Ala274----Thr). Message amplification phenotyping studies have also permitted us to define the ALAD phenotype of each subject in the family. This is the first mutation to be recognized in the human ALAD gene.     

Inhibition of biological actions of big endothelin-1 by phosphoramidon

Endothelin (ET)-1 and big ET-1 both caused contraction of isolated porcine coronary arteries, but the potency of big ET-1 was 1/100-1/200 that of ET-1. These responses were independent of the vascular endothelium. Phosphoramidon blocked the vasoconstriction caused by 30 nM big ET-1, but was ineffective on the action of 0.3 nM ET-1. Also in vivo, phosphoramidon had no effect on the ET-1-induced pressor actions, but blocked the pressor and airway-contractile responses to big ET-1 in rats and/or guinea pigs. Thus, it is likely that the vascular responses to exogenous big ET-1 are at least in part due to its conversion to ET-1 by a phosphoramidon-sensitive ET converting enzyme(s) in the vascular smooth muscle in vitro and in vivo.     

Slalom chromatography: size-dependent separation of DNA molecules by a hydrodynamic phenomenon

Slalom chromatography, a size-dependent DNA fractionation method based on a new principle [Hirabayashi, J., & Kasai, K. (1989) Anal. Biochem. 178, 336-341], was systematically studied in detail. In this method, larger DNA fragments are eluted much later than smaller ones from columns packed with spherical microbeads. Elution of a series of DNA fragments was systematically examined by using columns packed with polymer-based packings of different diameter and different pore size for high-performance gel permeation chromatography. Packings of smaller diameter proved to be superior for resolving the smaller size range of DNA, while the reverse was the case for larger DNAs. Application of a faster flow rate led to larger retardation of every DNA fragment, while at the lowest flow rate applied (0.067 cm/min), all the fragments were eluted almost at the void volume. When the column temperature was lowered, retardation of DNA became larger. On the other hand, differences in the chemical nature and the pore size of packings, or in the hydrophobicity of the eluting solvent, had little effect on DNA retardation. Size-dependent fractionation of DNA was also achieved even on columns packed with nonporous packings having anionic groups (cation exchangers). In conclusion, these results confirmed the previous conclusion that slalom chromatography is not based on an adsorption or equilibrium phenomenon but should be attributed to a hydrodynamic phenomenon.     

Expression in Escherichia coli of chemically synthesized gene for a novel opiate peptide alpha-neo-endorphin

Chemically synthesized alpha-neo-endorphin gene was fused to the Escherichia coli beta-galactosidase gene on the plasmid pKO13. The resulting recombinant DNA was used to transform E. coli cells. Radioimmunoassay for alpha-neo-endorphin in CNBr-treated bacterial cells showed that alpha-neo-endorphin was synthesized at approximately 5 x 10(5) molecules per single E. coli cell. One of the transformants, WA802/p alpha NE2, was used for alpha-neo-endorphin purification. From 10.9 g of wet cells, we isolated 4 mg of chemically pure and biologically active alpha-neo-endorphin.     

Independent control of sexual and scent marking behaviors of male gerbils by cells in or near the medial preoptic area

This research studied the role of the medial preoptic area and adjacent cell populations in androgen control of scent marking and sexual behavior in male gerbils (Meriones unguiculatus). Experiment 1 replicated previous research showing that implants of testosterone propionate in or near the medial preoptic area reinstate marking behavior in castrates. Implant sites near the diagonal band of Broca or in the posterior part of the medial preoptic area, near the anterior hypothalamus, are more effective than other sites. Experiment 2 showed that medial preoptic area lesions permanently impair sexual behavior despite testosterone stimulation. Experiments 2–4 showed that lesions in or near the medial preoptic area can also disrupt scent marking; however, this behavior gradually recovered in many lesioned males, especially if they received testosterone. The data suggest that both scent marking and sexual behavior are controlled by androgens acting on cells in or near the medial preoptic area, but the cell populations involved in these two behaviors are probably not the same.     

Evidence for a dual effect of dibutyryl cyclic AMP on the synthesis of tyrosine aminotransferase in rat liver

A single injection of dibutyryl cyclic AMP (Bt2cAMP) into adrenalectomized rats results in rapid and proportionate increases in hepatic tyrosine aminotransferase catalytic activity and in the amount of functional mRNA coding for this enzyme. This effect is transient in that mRNATAT peaks at 0.065% of total poly(A)+RNA activity at 1 h and is back to the basal level of 0.012% in 2.5 h. Enzyme activity peaks at 2.5 h and is back to the basal level by 5 h. If Bt2cAMP is repeatedly injected (0, 1, 2.5, and 4 h), enzyme activity remains at maximal levels for 4 to 5 h, whereas changes in mRNATAT activity are identical with those observed in the single injected rats. The rate of tyrosine aminotransferase synthesis at 5.5 h in the multiply injected rats, a time when mRNATAT has already returned to the basal level, is 3 to 4 times greater than that in either control or singly injected rats at the same time (0.3% of total protein versus 0.07%) and is equivalent to the maximal rate seen 1 h after the initial injection of the cyclic nucleotide. Since the rate of synthesis is increased in proportion to the increase in enzyme catalytic activity, stabilization of the enzyme against degradation is excluded as an induction mechanism at this late time point. These responses are not due to differences in the metabolism of Bt2cAMP, and the effect depends on the presence of metabolically active derivatives of this nucleotide. It thus appears that Bt2cAMP induces the synthesis of tyrosine aminotransferase in rat liver in two distinct ways. One is pretranslational and involves a transient and rapid increase in mRNATAT activity. The second appears to involve a delayed but sustained increase in translation of a basal level of mRNATAT.     

Factors Contributing to the Poor Myelination in the Brain of the Snell Dwarf Mouse

Abstract: Conventional histological examination of the pituitary does not distinguish Snell dwarf mutants (dw/dw) from their normal littermates (+/?) in the neonatal stage. However, immunohistochemical examination of pituitaries of litters born to heterozygous Snell parents revealed that in approximately 25% of the glands examined, the number of positive cells was very low in the neonatal stage. We attempted to delineate the events resulting in the poor myelination in the brain of the Snell dwarf mouse, and to devise an immunohistochemical method for identifying the mutant neonate. Differences in the brain weights of the dw/dw and +/? mice first became apparent on the 10th day of age, and from this time on no further increase in the weight of the dwarf mouse brain was recorded. Increase in CNPase activity was found to be suppressed in the cerebrum and brain stem throughout the developmental stage, but not in the other parts of the brain. The yield of isolated myelin decreased by 58% in the mutant mouse, but CNPase activity was equivalent to that of control myelin. Differences in DNA content per cerebrum from the dw/dw and +/? mice first became apparent on the 10th day of age. Henceforth, the dw/dw mice showed no further increase, although the +/? mice continued to increase. [³H]Thymidine incorporation into the DNA fraction in vivo on the 7th day of age, when glial cell proliferation in the cerebrum is most active, was suppressed to about 50% of the control level in all parts of the dwarf brain. These findings indicate that the poor myelination found in the mutant cerebrum is a hypomyelination due to reduced oligodendroglial proliferation caused by lack of circulating growth hormone.     

Co-operative response of chemically excitable membrane III. Three-state model

The general three-state model is formulated first, which is the direct extension of the unified two-state model previously formulated (Kijima & Kijima, 1978). In this model, each protomer in a symmetrically interacting system (oligomers or lattices) can take three states, S, R and Q, where S and R states are the same as in the two-state model and Q state is another state either corresponding to a different open-state of ionophore from R open-state or corresponding to another closed state of ionophore. The model has no restriction on the value of Hill coefficient at the midpoint of the dose-response curves in contrast to two-state models. It is applied on GABA sensitive inhibitory synapse of crayfish muscle to account for anomalous behaviour of the membrane in I^? solution.The simplified versions of the above general three-state model are also formulated (simplified three-state model), in which it is assumed that R and Q state are equivalent in regard to the nearest neighbor interaction. By this assumption, R and Q state are collectively treated as state A and mathematical formula obtained on Ising model are applicable on this model. This model is applied on the insect sugar receptor which was shown to be incompatible with the two-state models (Kijima & Kijima, 1980). Further simplification of the above simplified model results in two convenient models: three-state KNF model and three-state MWC model, which have minimum parameters but sufficient to account for most experiments. They give plausible physico-chemical base on the “classical model” in which the existence of both inactive and active ligand-receptor complex is assumed.     

Inhibition of thyroxine-induced metamorphosis by prolactin in tadpoles, Rana catesbeiana.

Thyroxine (T4)-prolactin interactions on hepatic arginase and ornithine transcarbamylase (OTC) as well as hind legs, tail, digestive tract and median eminence were investigated in tadpoles, Rana catesbeiana. Prolactin completely blocked T4-induced tail resorption, but failed to suppress hind-leg growth, shortening of digestive tract and promotion by T4 of the median eminence development. Prolactin blocked T4-induced increase in hepatic arginase activity but not in hepatic OTC activity. A possibility that T4 and prolactin are regulating the hepatic arginase indirectly is discussed.