Re-examination of the putative roles of insulin and prolactin in the regulation of lipid deposition and lipogenesis in vivo in mammary gland and white and brown adipose tissue of lactating rats and litter-removed rats.

1. The effects of various treatments to alter either plasma prolactin (bromocryptine administration or removal of litter) or the metabolic activity of the mammary gland (unilateral or complete teat sealing) on the disposal of oral [14C]lipid between 14CO2 production and [14C]lipid accumulation in tissues of lactating rats were studied. In addition, the rates of lipogenesis in vivo were measured in mammary gland, brown and white adipose tissue and liver. 2. Bromocryptine administration lowered plasma prolactin, but did not alter [14C]lipid accumulation in mammary gland or in white and brown adipose tissue. 3. In contrast, complete sealing of teats results in no change in plasma prolactin, but a 90% decrease in [14C]lipid accumulation in mammary gland and a 4-fold increase in white and brown adipose tissue. The rate of lipogenesis in mammary gland was decreased by 95%, but there was no change in the rate in white and brown adipose tissue. Unilateral sealing of teats resulted in a decrease in [14C]lipid accumulation in white adipose tissue. 4. Removal of the litter for 24 h (low prolactin) produced a similar pattern to complete teat sealing, except that there was a 6-fold increase in lipogenesis in white adipose tissue. Re-suckling for 5 h increased plasma prolactin, but did not alter the response seen in litter-removed lactating rats. 5. Changes in lipoprotein lipase activity and in plasma insulin paralleled the reciprocal changes in [14C]lipid accumulation in white and brown adipose tissue and in mammary gland. 6. It is concluded that the plasma insulin is more important than prolactin in regulating lipid deposition in adipose tissue during lactation, and that any effects of prolactin must be indirect.     

An ESR study of nitrosyl-Aplysia brasiliana myoglobin and nitrosyl annelidae Glossoscolex paulistus erythrocruorin

The nitrosyl derivatives of Annelidae Glossoscolex paulistus hemoglobin (an earth worm erythrocruorin (Ec AGp)) and Aplysia brasiliana myoglobin (Mb Apb) are studied using ESR spectroscopy. These two proteins have a quite similar ESR spectra at 100 K, but a different temperature behaviour. The temperature dependence of the nitrosyl Mb Apb spectrum is in good agreement with the Boltzmann distribution. In the case of nitrosyl-Ec AGp, the results are explained by the existence of two types of spectrum in thermodynamic equilibrium, with delta H = 9.08 kJ/mol, delta S = 47.15 J/mol and T1/2 = 193 K. There is a great similarity of the nitrosyl-Ec AGp spectra with those reported for elephant myoglobin, suggesting the presence of the same heme environment with a glutamine residue in the distal site. The pH dependence of the spectrum of nitrosyl-Mb Apb shows that the affinity of nitrosyl binding is higher at high pH (7.3) than at low pH (4.6). The ESR parameters are the same for these two pH values.     

The acid-alkaline transition of a sea turtle myoglobin: coexistence at high pH of high- and low-spin forms

The microenvironment of the iron in a sea turtle Dermochelys coriacea myoglobin is studied using the spectroscopic techniques EPR and optical absorption. Optical absorption spectra in the visible region suggest a great homology between turtle Mb and other myoglobins, such as those from whale, human and elephant. The pK of the acid-alkaline transition is 8.4 slightly lower than the pK of whale and equal to that of elephant myoglobin. The EPR spectrum at pH 7.0 is characteristic of a high-spin configuration with axial symmetry (gx = gy = 5.95). At higher pH, this signal changes in a way different from that observed for whale myoglobin. We observe for turtle Mb both the formation of a low-spin configuration with rhombic symmetry (gx = 2.56, gy = 2.20, gz = 1.90) and of a high-spin species with rhombic distortion (gx = 6.79, gy = 5.18, gz = 2.12). This suggests a lowering of symmetry at the haem, so that now the x and y directions are no more equivalent. This can be explained by amino acid substitution at the distal positions of haem or to off-axial positioning of distal residues. The coexistence at high pH (pH 11.0) of these two spin forms could be explained by the existence of two protein conformations, in which the crystal field splitting factor, delta, and the electron exchange energy are of the same order, allowing the presence of different configurations simultaneously. The presence of different kinds of haem is ruled out by the experiments with nitrosyl turtle Mb and turtle Mb-F showing spectra very similar to those of whale myoglobin. The pk of the acid-alkaline transition, 8.5, obtained from EPR spectra, agrees very well with results from optical absorption.     

Crop/Weed gene flow:Cucurbita argyrosperma Huber andC. fraterna L. H. Bailey (Cucurbitaceae)

A mixed population of Cucurbita at Vado El Mow in northern Tamaulipas, Mexico showed an anomalous pattern of fruit bitterness. Some domesticated plants (C. argyrosperma andC. moschata) expressed cucurbitacin bitterness whereas some sympatric free-living plants produced non-bitter fruits. This reversal of typical cucurbitacin expression suggested gene flow between crop and weed at the site. Isozyme analysis provided little insight as to taxa involved in gene exchange, although progeny from a single free-living plant carried IDH allozymes that are associated with Mexican landraces ofC. pepo. Synthetic hybridization revealed that fertile F, hybrids are produced from crosses involvingC. fraterna as the pistillate parent andC. argyrosperma as the staminate parent. Interspecific crop/weed hybrids can produce viable progeny upon self-pollination or backcrossingto either parent, andF₂ families display normal allozyme segregation. Hybrid fertility, as indicated by pollen stainability, increases in progeny produced by backcrossingfrom theC. argyrosperma parent. Interspecific hybridfertility represents a potential for crop/weed gene flow that would be realized under natural conditions if pollen flow occurs betweenC. fraterna andC. argyrosperma in the fields of Tamaulipas. Oligolectic “squash bees” (Teponapis), efficientCucurbita pollen vectors, are present at the site. Thus, it is likely that natural interspecific crop/weed hybridization has occurred at Vado El Moro and this might at least partially explain the anomalous distribution of fruit bitterness among extant populations at the site.     

Flavonoids of Derris araripensis

Nineflavonoids: a dihydrochalcone,a flavone,four 3-methylflavonols,a flavanone, a 3-methylflavanonol and a flavan were isolated from the roots of Derris araripensis. Eight of these compounds are reported for the first time. Structures were established by spectral analysis and chemical degradation.     

Chemically defined media for growing anaerobic bacteria of the genus Veillonella

A chemically defined medium for Veillonella parvula and V. alcalescens is described. Some nutritional aspects of the two strains used were examined: the optimum concentration of reducing agents, the requirement for amino acids, diamines, vitamins and other growth factors, and the conditions needed for well balanced nutrition.No specific requirements for single amino acids were observed. A combination of l-cysteine, dl-aspartic acid, l-glutamic acid, l-serine and l-tyrosine, promoted growth. In V. alcalescens, serine could substitute both arginine and tryptophan (or histidine). No growth was obtained with ammonium salts as the sole N source.Decarboxylation of l-ornithine, l-lysine and l-arginine was not demonstrated in the Veillonella parvula strain, which required putrescine or cadaverine for growth. Spermine, spermidine, l-lysine, l-ornithine and l-arginine, could not substitute putrescine in Veillonella parvula. Veillonella alcalescens, which does not require putrescine in the medium, was able to decarboxylate l-ornithine while forming putrescine.     

Kinetics of reaction of (nitrotyrosyl)cytochrome c with ligands.

The reaction of [nitrotyrosyl]cytochrome c with ligands was studied by stopped-flow techniques. At pH 7.0 the reaction with imidazole shows two distinct phases, one fast phase being concentration-dependent and a slow phase being concentration-independent. The results are consistent with the existence of two forms of [nitrotyrosyl]cytochrome c in solutions [Schejter et al. (1970) Biochemistry 9, 5118-5122]; form I, the smaller fraction, seems to be responsible for the slow first-order process.     

Characterization of protein spin labeling by maleimide: evidence for nitroxide reduction

A quantitative determination of maleimide spin label (MAL) binding in oxi and met hemoglobin (Hb) and bovine serum albumin are investigated using double integration to the ESR signal. This determination permitted the observation that a considerable fraction of MAL is reduced, losing its paramagnetism. Experiments using the same spin label with myoglobin and Hb with blocked-SH groups, where reduction was not observed, indicate the involvement of SH groups in the process. The 4-hydroxy-2,2,6,6-tetramethylpiperidino-1-oxyl spin label (which is not able to bind in the SH group) is reduced too, but the dependence on the molar ratio is different in comparison with the MAL case. In both cases the reduction percentage depends on the molar ratio spin label to protein and to the protein concentration. In order to obtain the total SH groups labeled (two in the Hb case) it is necessary to use an excessive amount of label (around 18:1) in the 0.5 mM Hb concentration.