Choline acetyltransferase activity and total protein content in selected optic areas of the rat after complete light-deprivation during CNS development

Choline acetyltransferase (acetyl-CoA: choline O-acetyltransferase, I.U.B. 2.3.1.6) activity and total protein content in visual and extra-visual areas were compared in normal Long-Evans rats and in rats subjected to complete light-deprivation for 21 days from birth. The enzyme activity and the protein content in the superior colliculi, lateral geniculate bodies and visual cortex, as well as in the sensory-motor cortex, hypothalamus, brain stem and cerebellum, were measured in both mothers and progeny. By means of a radiochemical technique modified in this laboratory, a significant decline of ChAc activity was observed in the lateral geniculate bodies and superior colliculi, with no significant decline in the visual cortex of the experimental progeny. Total protein content, measured colorimetrically, was significantly decreased in the superior colliculi of the progeny. The biochemical data obtained from all other brain areas in the experimental animals (progeny and mothers) and controls demonstrated no marked differences. The enzymic alterations observed in the cholinergic system of progeny after complete light-deprivation during this critical period of CNS development can be specifically correlated with decreased functional maturation of the visual system. If it is accepted that ACh is a neuro-transmitter in some parts of the visual pathway, the data presented here suggest that complete light-deprivation from birth to 21 days of age in the rat has an effect on a biochemical system involved in synaptic transmission.     

Growth yields of bacteria on selected organic compounds

Cell yields were determined for two bacterial soil isolants grown aerobically in minimal media on a variety of synthetic organic compounds. 1-Dodecanol, benzoic acid, phenylacetic acid, phenylglyoxylic acid, and diethylene, triethylene, and tetraethylene glycols were tested. Two “biochemicals,” succinate and acetate, were also tested for comparison. Yields were calculated on the basis of grams of cells obtained per mole of substrate utilized, gram atom of carbon utilized, mole of oxygen consumed, and equivalent of “available electrons” in the substrates. This latter value appears to be nearly constant at 3 g of cells per equivalent of “available electrons.” Yields predicted on this basis for other bacteria and for yeasts on other substrates are in fair agreement with reported values.     

Metabolism of the reserve polysaccharide of Streptococcus mitis: Properties of a transglucosylase

1. A transglucosylase has been separated from cell extracts of Streptococcus mitis, and has been partially purified by chromatography on DEAE-cellulose. 2. The transglucosylase was present in the six strains of Streptococcus mitis that were examined, and the activity of the enzyme was the same whether the cells had grown on glucose or on maltose. Four of the strains could store intracellular iodophilic polysaccharide when grown on high concentrations of glucose or maltose (1%), but none of the strains stored polysaccharide during growth on 0·1% glucose. The activity of transglucosylase in cell extracts was the same whether or not the cells had stored polysaccharide. 3. The transglucosylase degrades amylose in the presence of a suitable acceptor, transferring one or more glucosyl residues from the non-reducing end of the donor to the non-reducing end of the acceptor. With [¹⁴C]glucose as acceptor the maltodextrins produced were labelled in the reducing glucose unit only. 4. The enzyme can synthesize higher maltodextrins from maltose and maltotriose. Maltotetraose is disproportionated to give products of sufficient chain length to give a stain with iodine. 5. The action pattern of S. mitis during the degradation of synthetic amylose was shown to be intermediate between the single-chain and multi-chain mechanism.     

Statistical evaluation of electronic and plate counts for estimating bacterial populations

Bacteria from cultures of Staphylococcus aureus, Pseudomonas geniculata, and Rhodotorula glutenis were counted by use of an electronic counter and by plate counts from broth cultures at selected periods from 0 to 24 hr. Variations in the two methods were noted, and the results were compared after calculating correlations, coefficients of variation, and nested analyses of variance. It was not possible to determine the absolute accuracy of the two methods; however, the precision of the results obtained with an electronic counter was better than that obtained with plate counting. It appears that electronic counting, because of ease and convenience, provides better comparative counts at a particular stage of growth, especially during the early stages of the growth period, whereas plate counting yields the best results in determining growth curves.     

The relationship between serum and cell type on the development of rat and pig cultured preadipocytes

Stromal-vascular cells from rats and pigs were isolated from adipose tissue and used to measure preadipocyte proliferation and differentiation. Cells from rats and pigs were grown in either 2.5% pig serum or 2.5% rat serum. Cells were either supplemented or unsupplemented with insulin after five days of growth in culture. In these cultures, pig fat cells developed as discrete clusters while rat fat cells developed as loose clusters or as individual cells. Rat cells had greater levels of sn-glycerol phosphate dehydrogenase activity compared to pig cells. Rat serum increased soluble protein in plated cells when compared to cells grown in pig serum. Pig serum increased glycerol phosphate dehydrogenase specific activity when compared to rat serum. In this system, there was no response to insulin. The cells grown in rat serum did not resemble adipocytes in regard to the presence of large lipid droplets (oil red 0 staining). These results demonstrate that rat and pig stromal-vascular cells in culture are morphologically different. Cells from both species, however, responded similarly to sera from either species showing that cells from rats and pigs responded to the growth and differentiation factors present in these sera.Mention of a trade name, proprietary product, or specific equipment does not constitute a guarantee or warranty by the U.S. Department of Agriculture and does not imply its approval to the exclusion of other products that may be suitable.     

Effects of the developmental state of the tissue on the competence for flower bud regeneration in pedicel explants of tobacco

The competence of pedicel explants of tobacco (Nicotiana tabacum L. cv Samsun) to regenerate flower buds in response to auxin was manipulated by preincubating excised tissues in the absence of auxin. When exposed to 1 micromolar 1-naphthaleneacetic acid, these tissues formed fewer buds than controls that were not preincubated. The number of buds eventually formed correlated with the 1-naphthaleneacetic acid concentration in the tissue 6 hours after the start of hormone application. The internal concentrations in pretreated explants were lower than in tissues that were not pretreated due to diminished uptake per milligram fresh weight and increased hormone conjugation. The change in the developmental state induced by auxin deprivation had a dual effect on bud regeneration: (a) the pretreatment caused fewer buds to be formed at any 1-naphthaleneacetic acid concentration tested, and (b) a higher auxin concentration in the medium was required to get a maximum bud number on precultured explants. An increase of the 1-naphthaleneacetic acid concentration in the medium led to an elevated hormone level in freshly cut as well as in preincubated tissues. It was concluded that the developmental state of the tissue directly affects the maximum number of buds that can be regenerated. Apart from that there is an indirect effect exerted via modulation of the ratio between external and internal auxin concentration. The change in this ratio can be compensated for by an adjustment of the auxin concentration in the medium.     

Immobilization of Candida cylindracea lipase on colloidal liquid aphrons (CLAs) and development of a continuous CLA-membrane reactor

A novel technique is described for the immobilization of Candida cylindracea lipase in the soapy-shell of colloidal liquid aphrons (CLAs). CLAs consist of a micron-sized solvent droplet surrounded by a thin, aqueous, soapy-film and are stabilized by a mixture of nonionic and ionic surfactants. Retention of lipase within the CLAs is primarily determined by electrostatic interactions between the surface charges on the protein and those of the anionic surfactant used (SDS) because leakage of the lipase from dispersed CLAs was reduced at low continuous phase pH (相似文献   

Calcineurin,a Type 2B Protein Phosphatase,Modulates the Ca2+-Permeable Slow Vacuolar Ion Channel of Stomatal Guard Cells

The slowly activating vacuolar (SV) channel of plant vacuoles is gated open by cytosolic free Ca2+ and by cytosol-positive potentials. Using vacuoles isolated from broad bean guard cell protoplasts, SV-mediated currents could be measured in the whole-vacuole configuration of a patch clamp as the time-dependent increase in current at cytosol-positive voltages. Time-dependent deactivation of the SV currents when changing from activating to nonactivating voltages (tail currents) was used to calculate the selectivity of the channel to Ca2+ and Cl- with respect to K+. Changing the equilibrium potential for each permeant ion (Ca2+, Cl-, and K+) at least once for individual vacuoles allowed the relative permeabilities (P) of each of these ions to be calculated in a single experiment. The resulting Pca:Pcl:Pk ratio was close to 3:0.1:1. In accord with its characterization as a weakly selective Ca2+ channel, the SV-mediated current density decreased with increasing Ca2+ activity in the vacuole lumen. SV currents were potently modulated by the Ca2+-dependent, calmodulin-stimulated protein phosphatase 2B (calcineurin). At low concentrations ([less than or equal to]0.4 units per mL), calcineurin stimulated SV currents by ~60%, whereas at higher concentrations the phosphatase was inhibitory, reaching ~90% inhibition at 3 units per mL. Bovine calmodulin had no direct effect on SV-mediated currents, although calcineurin stimulated by exogenous calmodulin inhibited SV currents at all concentrations tested with half-maximal inhibition for calcineurin at 0.16 units per mL. The inhibitory effect of calcineurin could be blocked by the pyrethroid deltamethrin, indicating inhibition of SV channels by calcineurin via dephosphorylation. A model is discussed in which vacuolar Ca2+ release through SV channels is subject to both positive feedforward and negative feedback control through cytosolic Ca2+ and dephosphorylation, respectively.     

Inelastic neutron-scattering study of the proton transfer dynamics in polyglycine I at 20 K

Inelastic neutron-scattering (INS) spectra of three isotopic derivatives of polyglycine I (-COCH2NH-)n, (-COCD2NH-)n, and (-COCH2ND-)n at 20 K are presented from 30 to 4000 cm(-1). The band frequencies are compared to those observed in the infrared and Raman. Assignments in terms of group vibrations are proposed. These mostly resemble previous assignment schemes, except for the amide bands. The INS intensities reveal that the proton dynamics for the (N)H proton are totally different from those proposed previously. They are independent of the molecular frame and the valence bond approach is not consistent with observation. A phenomenological approach is proposed in terms of localized modes. The calculated intensities reveal that the (N)H stretching mode has two components at approximately 1377 and 1553 cm(-1). This is a dramatic change compared to all former assignments at approximately 3280 cm(-1) based on infrared and Raman data. These proton-dynamics are associated with a weakening of the NH bond due to the ionic character of the hydrogen bond (N(delta-)...H+...O(delta'-)) and proton transfer. The infrared and Raman spectra are re-examined and a new assignment scheme is proposed for the amide bands; the amide A and B bands are re-assigned to the overtones of the stretching modes. A symmetric double-minimum potential for the proton is consistent with all the observations.