New PCR-RFLP analysis for the mouse Tnfsf6gld gene caused by a point mutation in the Tnfsf6 [tumor necrosis factor (ligand) superfamily, member 6] locus.

A new polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis was developed for genetic typing of the mouse Tnfsf6gld mutation. An artificial restriction site was introduced to the mouse Tnfsf6gld mutation by PCR amplification using a modified primer. The three genotypes of the Tnfsf6 locus (Tnfsf6gld/Tnfsf6gld, Tnfsf6gld/+, and +Tnfsf6-gld/+Tnfsf6-gld) could be distinguished clearly and easily. This PCR-RFLP analysis was found to be useful for the identification of the mouse Tnfsf6gld mutation.     

New segment synthesis of alpha-inhibin-92 by the acyl disulfide method

The thiocarboxyl group reacts with diaryl disulfides to give an unsymmetrical acyl disulfide in dimethylformamide (DMF) and a symmetrical diacyl disulfide in aqueous DMF. Both acyl disulfides react with the alpha-amino group to form the peptide bond. The method was used in a new segment synthesis of alpha-inhibin-92 (alpha-IB-92) with use of 2,2'-dipyridyl disulfide as activator. Thiocarboxyl peptides were synthesized by the solid-phase method on 4-[alpha-(Boc-Gly-S)benzyl]phenoxyacetamidomethyl-resin. The segments alpha-IB-92-(1-34)SH (I), Msc-alpha-IB-92-(35-65)SH (II), Msc-alpha-IB-92(66-92)OH (III), and Msc-alpha-IB-92-(35-92)OH (VI) were prepared in yields of 33, 36, 41, and 25%, respectively, with use of crystalline symmetrical anhydrides in double and triple coupling protocols. Segments I, II, and III were used in a 3-segment synthesis of alpha-IB-92 with an overall yield based on starting resin of about 8% while a 2-segment synthesis using I and IV gave 11%. An all stepwise synthesis of alpha-IB-92 gave 4.5%.     

Succession of sessile organisms on experimental plates immersed in Nabeta Bay,Izu Peninsula,Japan V. An integrated consideration on the definition and prediction of succession

Successional changes of sessile organisms over about 3 years on concrete plates immersed at depths of 1.0, 2.5, 4.0 and 5.5 m in Nabeta Bay, Shimoda, Pacific coast of Japan, were investigated from their species composition, percentage cover, and the structure, metabolism, biomass and maturity of the community for the purpose of verifying the hypothesis of succession by Margalef (1968) and Odum (1969). The process of succession was divided into five stages from a cluster analysis based on Horn's (1966) similarity in community structure. The five stages were characterized from dominant, species and 18 community attributes in terms of structure, metabolism, biomass and maturity. With the advance of stages, two large-sized, long-lived species, the kelpEisenia bicyclis and the oysterCrassostrea nippona, dominated; 18 community attributes showed a directional succession ending in a constant community structure, low community metabolism, accumulation of community biomass except chlorophylla and an enhanced degree of community maturity. This directionality in succession of community attributes was valid for the prediction of Margalef and Odum, despite a tendency for estimated daily energy budget to be more heterotrophic in such an open system. From these results, it was suggested that the definition of succession by Odum (1969) should be interpreted as the regularity in shift of dominant species, not individual species, and as the control of system by feedback mechanism developing with succession rather than community-controlled mechanism.     

The receptors responsible for heat production in brown adipose tissue in the young rabbit

It is known that adrenergic agonists stimulate thermogenesis in the brown fat of the young rabbit but the receptors responsible for mediating the response have not been identified. The infusion of either noradrenaline or isoproterenol (1-2 micrograms . kg-1 X min-1) produced an increase in subcutaneous temperature (0.93 +/- 0.15 and 1.22 +/- 0.10 degrees C, respectively over the interscapular brown fat. At low doses (0.4 microgram . kg-1 X min-1) only isoproterenol was effective. The thermogenic response to isoproterenol was blocked by atenolol, a beta 1-adrenergic antagonist. Neither salbutamol or terbutaline, both beta 2-agonists, produced a temperature increase. Collectively, these data suggest that stimulation of beta 1-adrenoceptor is primarily responsible for the thermogenic activity of brown fat in the rabbit. However, it was found that 53% of the increase in temperature could be blocked by prazosin, an alpha 1-antagonist. Phentolamine was not effective as a blocker. Although a maximal brown fat thermogenic response can be achieved by stimulating the beta-adrenoceptors, the alpha-adrenoceptors appears to play at least an auxiliary role in young rabbit.     

The noradrenaline content and innervation of brown adipose tissue in the young rabbit

This work examined the noradrenaline content of brown adipose tissue, the metabolic response to endogenous noradrenaline released during tyramine infusion, and the innervation of brown fat at the electron microscopic level in the young rabbit. The noradrenaline content (ng/g) of the interscapular and cervical fat deposits ranged from 256 +/- 51 to 343 +/- 59 and 399 +/- 18 to 694 +/- 92, respectively, in four groups of rabbits (1-2, 7-8, 12-13, and 25-27 days of age). There was considerable variation amongst animals in each age group, but no evidence of a major increase or decrease in noradrenaline content during the first 4 weeks of life. Intravenous infusion of tyramine (100 micrograms X kg-1 X min-1) increased plasma noradrenaline concentration, oxygen consumption, and blood flow to brown fat. Thus noradrenaline released from endogenous sites, as well as injected noradrenaline, will initiate the thermogenic response of brown fat. Ultrastructurally, unmyelinated axons that were not organized in a fascicle were observed adjacent to the adipocytes in the late gestation fetus. By 1 week of age of axons were surrounded by Schwann cell cytoplasm which formed a fascicle. However, no evidence of myelination was found up to 21 days of age. Collectively, the data indicate that the brown adipocyte is fully responsive at 1-2 days of age even though myelination of the nerves is incomplete, and that the incomplete development of the sympathetic nerves at birth is not a factor in the synthesis of noradrenaline in the very young rabbit. In addition, brown fat of the newborn rabbit is not as thermogenically active as the brown fat of the cold-acclimated rat.     

Synthesis of a peptide with full somatostatin activity

A peptide has been synthesized according to the structure proposed for somatostatin by the solid phase method. The synthetic product was assayed and found to possess full somatostatin activity as compared with the natural material.     

Applications of fractal analysis to physiology

This review describes approaches to the analysis of fractal properties of physiological observations. Fractals are useful to describe the natural irregularity of physiological systems because their irregularity is not truly random and can be demonstrated to have spatial or temporal correlation. The concepts of fractal analysis are introduced from intuitive, visual, and mathematical perspectives. The regional heterogeneities of pulmonary and myocardial flows are discussed as applications of spatial fractal analysis, and methods for estimating a fractal dimension from physiological data are presented. Although the methods used for fractal analyses of physiological data are still under development and will require additional validation, they appear to have great potential for the study of physiology at scales of resolution ranging from the microcirculation to the intact organism.     

Kinetics of alpha 2-macroglobulin endocytosis and degradation in mutant and wild-type Chinese hamster ovary cells

The production of Chinese hamster ovary (CHO) cell mutants which are defective in endocytosis has led to a greater understanding of the process by which cells sort ligands and their receptors. Robbins and coworkers have obtained CHO mutants which are resistant to diphtheria toxin, defective in the delivery of endocytosed lysosomal enzymes to lysosomes, and have a decreased uptake of iron from transferrin (Robbins et al.: J. Cell Biol. 96:1064-1071, 1983). We have previously shown that these CHO mutants are markedly deficient in the acidification of early endocytic compartments (Yamashiro and Maxfield: J. Cell Biol. 105:2713-2721, 1987). In this study we examined the endocytosis of alpha 2-macroglobulin (alpha 2M) to determine whether the defects in early endosome acidification would alter the processing of this ligand. We found that the CHO mutants DTG 1-5-4 and DTF 1-5-1 bind, internalize, and degrade 125I-alpha 2M in a manner similar to the wild-type cells. We also found that the CHO mutants retain the ability to recycle the receptors for alpha 2M. Since the binding of alpha 2M is greatly reduced at mildly acidic pH (approximately 6.8), only slight acidification of the endosomal compartment should be sufficient to achieve sorting of alpha 2M from its receptor. In contrast, lysosomal enzymes require more acidic conditions (pH less than 6.0) for dissociation. The different behavior of the two ligands provides biochemical evidence for a partial (but not complete) defect in early endosome acidification in the mutants. The data also indicate that pH regulation in a relatively narrow range can achieve differential sorting of various ligands.