Bayesian Nonparametric Modeling for Comparison of Single-Neuron Firing Intensities

Summary .  We propose a fully inferential model-based approach to the problem of comparing the firing patterns of a neuron recorded under two distinct experimental conditions. The methodology is based on nonhomogeneous Poisson process models for the firing times of each condition with flexible nonparametric mixture prior models for the corresponding intensity functions. We demonstrate posterior inferences from a global analysis, which may be used to compare the two conditions over the entire experimental time window, as well as from a pointwise analysis at selected time points to detect local deviations of firing patterns from one condition to another. We apply our method on two neurons recorded from the primary motor cortex area of a monkey's brain while performing a sequence of reaching tasks.     

Ca- and Mg-dependent association of phosphorylase kinase with human erythrocyte membranes

The interaction of rabbit muscle phosphorylase kinase (EC 2.7.1.38) with human erythrocyte membranes was investigated. It was found that at pH 7.0 the kinase binds to the inner face of the erythrocyte membrane (inside-out vesicles) and that this binding is Ca²⁺- and Mg²⁺-dependent. The sharpest increase in the binding reaction occurs at concentrations between 70 and 550 nM free Ca²⁺. Erythrocyte ghost or right-side out erythrocyte vesicles showed a significantly lower capacity to interact with phosphorylase kinase. Autophosphorylated phosphorylase kinase shows a similar Ca²⁺-dependent binding profile, while trypsin activation of the kinase and calmodulin decrease the original binding capacity by about 50%. Heparin (200 μg/ml) and high ionic strength (50 mM NaCl) almost completely blocks enzyme-membrane interaction; glycogen does not affect the interaction.     

Effect of anoxia on the kinetic properties of pyruvate kinase and phosphofructokinase,and on glycogen phosphorylase activity in marine worms and earth worms

Summary The kinetic properties of PK and PFK were studied in aerobic versus 12-hours anoxic marine worms Hedistae(=Nereis) diversicolor and Diopatra neapolitana and earth worms Allolobophora calliginosa and Eisenia foetida. The total glycogen phosphorylase (a+b) activity and the percentage of active a form were also measured in the marine and earth worms under the same conditions. Anoxia exposure did not result in any significant changes of kinetic parameters of PK and total activities of glycogen phosphorylase from marine worms, but it altered the kinetic characteristics of PFK from H. diversicolor. Chromatographical studies showed that PK from both aerobic and anoxic marine worms is eluted from DEAE-cellulose as a single peak at 50 mM KCl. In contrast to marine worms, however, anoxia caused a marked change in kinetic properties of PK from both earth worms, resulting in a reduction of enzyme affinity for its substrate PEP. In addition, the enzyme existed in both earth worms in two distinct variants eluted from DEAE-cellulose column as peak I and peak II at 50 mM and 150 mM KCl, respectively. The ratio of enzyme units (peak I/peak II) was reduced significantly after 12 h of anoxia, indicating that these two peaks are interconvertible. Anoxia also caused a reduction of total glycogen phosphorylase activity in E. foetida and lowered the percentage of active a form of the enzyme by approximately 50% in both earth worms. Kinetic properties of PFK from both earth worms were not significantly affected by anoxia. However, their low Ka values for F-2,6-P₂ imply that this effector may play an important role in PFK control in earth worms under anoxia.Abbreviations F6P fructose-6-phosphate - FBP fructose-1,6-bisphosphate - F-2,6-P fructose-2,6-bisphosphate - PEP phosphoenoylpruvate - PFK 6-phosphofructo-1-kinase (E.C.2.7.1.11) - PK pyruvate kinase (E.C.2.7.1.40) - Pi inorganic phosphate - PMSF phenyl methylsulfonyl fluoride     

Methods of assessment of transfer of a gentamycin-resistance gene (aacC1) from genetically engineered microorganisms into agriculturally important soil bacteria

In order to assess the risk associated with the deliberate release of genetically engineered microorganisms (GEMs) into the agricultural environment, the transfer of plasmids between bacterial strains was investigated under laboratory conditions. Genetically modified Rhizobium leguminosarum and Agrobacterium tumefaciens strains carrying the gentamycin acetyltransferase resistance gene (aacC1) on various plasmids were investigated for their ability to transfer the aacC1 gene to their wild-type (w.t.) counterparts, as well as to Pseudomonas syringae. Conjugation experiments between the various strains, were carried out after the relevant characteristics and conditions for selective growth of each bacterial strain had been ascertained. After conjugations on filters had been completed, the putative transconjugants were grown in media containing antibiotics and assessed for the presence of aacC1 gene by: (a) DNA plasmid profile; (b) expression of AAC(3)-I enzyme activity; (c) colony hybridization using a ³²P-labelled DNA probe complementary to the aacC1 gene. The results obtained indicate that transfer of the aacC1 gene from genetically modified strains of R. leguminosarum into a plasmid-free strain of A. tumefaciens occurred via self-transmissible plasmids. Alternatively, genetically modified A. tumefaciens bearing the aacC1 gene on plasmids acquired from R. leguminosarum strains, transferred it ineffectively to a hardly detectable frequency. No transfer of the aacC1 gene from genetically modified R. leguminosarum or A. tumefaciens strains into P. syringae has been observed. These data indicate that in the absence of the RP4 element, genetically modified A. tumefaciens is not able to efficiently transfer aacC1 into w.t. R. leguminosarum and P. syringae. Correspondence to: A. S. Tsiftsoglou     

Cyanide-resistant Respiration in Fresh and Aged Sweet Potato Slices

The respiration of fresh sweet potato (Ipomoea batatas) slices is resistant to, and often stimulated by, cyanide and antimycin A. m-Chlorobenzhydroxamic acid (CLAM), a selective inhibitor of the alternate path, inhibits respiration in the presence of cyanide and has a limited inhibitory effect in the presence of antimycin A. Thus, a partial bypass of the antimycinsensitive site is indicated. Respiration rises 2-fold at best with slice aging, the increment being cytochrome-mediated. The cyanide-resistant pathway contributes neither to coupled fresh slice respiration nor to the induced respiration in the absence of inhibitors of the cytochrome path. In the presence of uncoupler, however, the alternate path is engaged both in fresh and aged slices. V_cyt, the maximal capacity of the cytochrome path, remains essentially the same with slice aging, whereas V_alt decreases from 20 to 60 per cent. The induced respiration is readily accommodated by the potential cytochrome path capacity of fresh slices, which is realized on aging. Accordingly, there is no need to invoke mitochondrial proliferation in explanation of the development of the induced respiration. The engagement of the alternate path in response to uncoupler reflects substrate mobilization to a degree that substrate oxidation exceeds the electron transport capacity of the cytochrome path.

Fresh slices do not utilize exogenous substrates, whereas aged slices do so readily. Cerulenin, a specific inhibitor of fatty acid synthesis, prevents the development of the induced respiration as well as the capacity to oxidize exogenous substrates. It is suggested that lipid, and ultimately membrane, biosynthesis is central to the development of the induced respiration and the ability to use exogenous substrates, much as in potato.

     

Wound-induced membrane lipid breakdown in potato tuber

Freshly cut slices of potato tuber show an extensive loss of membrane lipid components which may be as great as 35% for phospholipids and 30% for glycolipids, in less than 15 minutes at 3 C. Phosphatidyl-choline, phosphatidyl-ethanolamine and mono- and di-galactosyl diglycerides comprise the bulk of the lipids that are degraded. Concomitantly, there is a pronounced loss of linoleic and linolenic acids. Whereas degradative events elicited by slicing proceed to a depth of at least 10 millimeters from the surface, phospholipid biosynthesis, as well as the development of the wound induced respiration and cyanide resistance on aging, are restricted to the superficial 1 millimeter.     

A sequential test for assessing observed agreement between raters

Assessing the agreement between two or more raters is an important topic in medical practice. Existing techniques, which deal with categorical data, are based on contingency tables. This is often an obstacle in practice as we have to wait for a long time to collect the appropriate sample size of subjects to construct the contingency table. In this paper, we introduce a nonparametric sequential test for assessing agreement, which can be applied as data accrues, does not require a contingency table, facilitating a rapid assessment of the agreement. The proposed test is based on the cumulative sum of the number of disagreements between the two raters and a suitable statistic representing the waiting time until the cumulative sum exceeds a predefined threshold. We treat the cases of testing two raters' agreement with respect to one or more characteristics and using two or more classification categories, the case where the two raters extremely disagree, and finally the case of testing more than two raters' agreement. The numerical investigation shows that the proposed test has excellent performance. Compared to the existing methods, the proposed method appears to require significantly smaller sample size with equivalent power. Moreover, the proposed method is easily generalizable and brings the problem of assessing the agreement between two or more raters and one or more characteristics under a unified framework, thus providing an easy to use tool to medical practitioners.