Multiple Forms of Choline-O-Acetyltransferase in Mouse and Rat Brain: Solubilization and Characterization

Three forms of acetyl coenzyme A: choline-O-acetyltransferase (EC 2.3.1.6, ChAT) have been isolated from mouse and rat forebrain synaptosomes with a 100 mM sodium phosphate (NaP) buffer of pH 7.4, a high-salt solution (500 mM NaCl), and a 2% Triton DN-65 solution, respectively. The Triton-solubilized form of ChAT differed from the other two forms in its capacity to acetylate homocholine, its pH profile, and its sensitivity to denaturation. NaCl-solubilized ChAT could be distinguished from the other two forms with respect to pH profile, sensitivity to inhibition by 4-(1-naphthylvinyl) pyridine (in the presence of Triton), and apparent Km value for choline acetylation. The caudate and putamen of rat brain contained the highest amount of ChAT activity, based on tissue wet weight, and the cerebellum contained the least of the brain regions examined; only the cerebellum had more membrane-bound than soluble ChAT. Septal lesion reduced ChAT activity in the NaP- and Triton-solubilized fractions prepared from hippocampus by 68% and 64%, respectively, whereas it reduced the activity of the NaCl-solubilized fraction by only 21%. These results suggest that three different forms of ChAT may exist in both mouse and rat brain.     

Rabbit kidney function in vitro: the effect of colloids, energy substrate, a vasodilator, perfusion pressure, and bovine serum albumin

An in vitro perfusion system at 37 degrees C for the assessment of rabbit kidney function is described. The purpose of this assay system is to evaluate the effects of cryobiological manipulation on kidney function. The effect of the colloids dextran (MW = 70,000, 80,000, and 180,000) in the perfusate at 110 mm Hg were compared to a reduced perfusion pressure, colloid-free perfusate. Better function was obtained at lower perfusion pressure with the colloid-free perfusate. Less damage was noted histologically on light and electron microscopy. Investigation of energy substrates on rabbit kidney function demonstrated that butyrate, or lactate, in addition to glucose resulted in increased sodium and glucose reabsorption over glucose alone. Substrate-free perfused kidneys exhibited depressed Na transport. Lactate, and to some extent butyrate, decreased net glucose utilization. An alpha-adrenergic blocking agent, isoxsuprine, in the initial flush solution did not appear to be beneficial. An increase of perfusion pressure from 50 to 75 mm Hg resulted in an increase in GFR. Tubular function was enhanced by inclusion of small amounts of BSA in the perfusate.     

Panagrellus ceylonensis n. sp. (Nematoda: Cephalobidae)

Panagrellus ceylonensis n. sp. is described from rubber latex in Ceylon. Its long, slender spicules are curved ventrally with a dorsal deflection at two-thirds of their length. The manubrium is thick and hooked, and the shaft is not widened at the shoulder. The terminal bifurcations are wide, with the dorsal one considerably longer and thicker than the ventral. The main ventral element is continuous with the terminal sheath with the ventral bifurcation arising within it.     

Synchronization of Mating Reactivity Rhythms in Populations of Paramecium bursaria

Cell populations of Paramecium bursaria show arhythmic mating reactivity after exposure to constant light (LL) for more than 2 wk. After this arhythmic population is exposed to darkness for 9 h, the mating reactivity rhythm of the cell population reappears. The phases of rhythms in individual cells are synchronized to each other. When the arhythmic population in constant light is exposed to dark pulses of various durations, the first peak of the recovered mating reactivity rhythm appears 6 h after the end of the dark pulse. Thus, in the case of dark pulses to cells in LL, the transition from dark to light sets the phase of the subsequent mating reactivity rhythm. When an arhythmic population in LL is transferred to constant darkness (DD), a rhythm of mating reactivity also appears and, in this case, the first peak of the rhythm occurs 18 h after the LL to DD transition. Therefore, arhythmic populations of cells in LL can be synchronized by either a dark pulse or by transition to continuous darkness. When the arhythmic populations in LL were transferred to various light/dark (LD) cycles, the mating reactivity rhythms entrained to LD cycles of 18 to 30 h in duration. Finally, mating rhythms can also be synchronized by treatment with puromycin (400 μg/ml for 6–18 h).     

黑腹果蝇P转座因子研究 Ⅰ.我国黑腹果蝇的P-M测定及其地理分布

采用性腺败育(GD不育)作为标准检定方法。对我国20个地方的黑腹果蝇的P因子活性和细胞型进行了测定。结果表明我国北部沿海城市为Q型;南部沿海和内地皆为M型。各地的M品系所产生的GD不育能力各不相同,但表现出与地理位置相关的梯度变化。这一变化规律为研究我国黑腹果蝇的P因子起源及P和M品系的形成提供了重要的理论依据。     

Distribution,properties, and functions of midgut carboxypeptidases and dipeptidases from Musca domestica larvae

Dipeptidase and carboxypeptidase A activities were determined in cells and luminal contents of the fore-, mid-, and hind-midgut of Musca domestica larvae. Dipeptidase activity was found mainly in hind-midgut cells, whereas carboxy-peptidase activity was recovered in major amounts in both cells and in luminal contents of hind-midguts. The subcellular distribution of dipeptidase and part of the carboxypeptidase A activities is similar to that of a plasma membrane enzyme marker (aminopeptidase), suggesting that these activities are bound to the microvillar membranes. Soluble carboxypeptidase A seems to occur both bound to secretory vesicles and trapped in the cell glycocalyx. Based on density-gradient ultracentrifugation and thermal inactivation, there seems to be only one molecular species of each of the following enzymes (soluble in water or solubilized in Triton X-100): membrane-bound dipeptidase (pH optimum 8.0; Km 3.7 mM GlyLeu, M_r 111,000), soluble carboxypeptidase (pH optimum 8.0; Km 1.22 mM N-carbobenzoxy-glycyl-L-phenylalanine (ZGlyPhe), M_r45,000) and membrane-bound carboxypeptidase (pH optimum 7.5, Km 2.3 mM ZGlyPhe, M_r58,000). The results suggest that protein digestion is accomplished sequentially by luminal trypsin and luminal carboxypeptidase, by membrane-bound carboxypeptidase and aminopeptidase, and finally by membrane-bound dipeptidase.     

Vanadate-stimulated oxidation of NAD(P)H

Vanadate stimulates the oxidation of NAD(P)H by biological membranes because such membranes contain NAD(P)H oxidases which are capable of reducing dioxygen to O₂⁻ and because vanadate catalyzes the oxidation of NAD(P)H by O₂⁻, by a free radical chain mechanism. Dihydropyridines, such as reduced nicotinamide mononucleotide (NMNH), which are not substrates for membrane-associated NAD(P)H oxidases, are not oxidized by membranes plus vanadate unless NAD(P)H is present to serve as a source of O₂⁻. When [NMNH] greatly exceeds [NAD(P)H], in such reaction mixtures, one can observe the oxidation of many molecules of NMNH per NAD(P)H consumed. This reflects the chain length of the free radical chain mechanism. We have discussed the mechanism and significance of this process and have tried to clarify the pertinent but confusing literature.     

Monoclonal Antibodies Allow Precipitation of Esterasic but Not Peptidasic Activities Associated with Butyrylcholinesterase

Commercially available and affinity-purified butyrylcholinesterases isolated from human serum were examined for their esterasic activity and their ability to hydrolyze various neuropeptides, including neurotensin, substance P, and leucine-enkephalin. The three pools that displayed the lowest esterasic activities were shown to hydrolyze neurotensin with the same HPLC degradative pattern. By contrast, noticeable qualitative and quantitative discrepancies were observed when hydrolyses of substance P and leucine-enkephalin by these three butyrylcholinesterase pools were studied. The pool that exhibited the highest esterasic activity appeared to be homogeneously constituted by 90- and 180-kDa protein bands by sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis and was totally unable to hydrolyze these three neuropeptides. This suggested that the three other butyrylcholinesterase preparations could be contaminated by exogenous peptidases. This was confirmed by means of three distinct monoclonal antibodies directed toward human serum butyrylcholinesterase. The three IgG-purified fractions precipitated the esterasic activity, whereas they failed to precipitate the neuropeptide-hydrolyzing activities whatever the substrate examined. Altogether, these results demonstrate that peptidases associated with butyrylcholinesterase are contaminating enzymes that cannot be considered as intrinsic activities of this enzyme.     

Nutrients and mass in litter and top soil of ten tropical tree plantations

The importance of litter to nutrient and organic matter storage and the possible influence of species selection on soil fertility in ten stands each consisting of a separate tree species were examined in this study. The plantations had been grown under similar conditions in an arboretum in the Luquillo Experimental Forest, Puerto Rico. The species involved were: Anthocephalus chinensis, Eucalyptus × patentinervis, E. saligna, Hernandia sonora, Hibiscus elatus, Khaya nyasica, Pinus caribaea var. hondurensis, P. elliottii var. densa, Swietenia macrophylla, and Terminalia ivorensis. After 26 yr, litter mass ranged from 5 mg ha^-1 in the H. sonora stand to 27.2 Mg ha^-1 in the P. caribaea stand. Nutrients in the litter (N, P, K, Ca, and Mg) also varied widely, but stands were ranked in different order when ranked by nutrients in the litter than then ranked according to accumulation of mass. Only E. saligna and A. chinensis stands were ranked similarly in accumulation of both nutrients and mass, and the stand of H. elatus was ranked higher with respect to nutrient accumulation than to accumulation of mass. The nutrient concentration in standing leaf litter generally increased in the order of recently fallen <old intact< fragmented. Nutrient concentration of standing leaf litter appears to increase with age and depth in the litter layer. The amount of nutrients stored in the litter compartment of these plantations was in the same order of magnitude as the quantity of available nutrients in the top 10-cm of mineral soil. Total litter mass was negatively correlated with the mass-weighted concentration of N, K, and Mg. The same relationship was found for Ca in the leaf litter and N in the fine wood litter compartments. In some stands (notably P. caribaea, P. elliottii, and E. saligna), leaf litter derived from species other than the species planted in that particular stand had higher nutrient concentration than leaf litter from the planted species. Soils of the 10 stands were classified in the same soil series and had similar texture (clay soils). However, significantly different chemical characteristics were found. Results obtained by analysis of covariance and by limiting comparisons to adjacent stands with similar soil texture, indicate that different species have had different influences on the concentration of available nutrients in soil.     

Phosphorus concentration in barley (Hordeum vulgare L.) seed: Influence on seedling growth and dry matter production

The effect of phosphorus (P) concentration in barley seed on seedling growth has not been much investigated. Consequently, two experiments were conducted in the greenhouse to determine the effect of P concentration in barley seed (Hordeum vulgare L., cv. Empress) on the seedlings grown in sand-filled boxes receiving a culture solution without P. Seeds were selected with three P concentrations: high-P (113.0 mmol P kg⁻¹), medium-P (80.7 mmol P kg⁻¹) and low-P (54.9 mmol P kg⁻¹). At 21 days after sowing, the shoot and root yield or shoot height was the least with seedlings from low-P seed. In the other experiment, high-P and low-P seeds were wetted with distilled water or with a solution of 25.8 cmol L⁻¹ of NaH₂PO₄ for 24 h, and then grown for 31 days. Solution P had been imbibed by seeds whether low or high in native P, but only the imbibed P held by low native P seed benefited seedling dry matter accumulation and shoot elongation. The lack of benefit from seed-imbibed P on seedlings grown from high-P barley seed was associated with low recovery of the imbibed P in those seedlings.