A stochastic afterhyperpolarization model of repetitive activity in vestibular afferents

A stochastic version of Kernell's (1968, 1972) model with cumulative afterhyperpolarization (AHP) was simulated. A characteristic of the model is that the AHP is the result of an increased potassium conductance (g _K) that is time-dependent but not voltage-dependent. Quantal synaptic inputs are assumed to be the only source of interspike interval variability. The model reproduces many features of the steady-state discharge of peripheral vestibular afferents, provided that firing rates are higher than 40 spikes/s. Among the results accounted for are the interspike interval statistics occurring during natural stimulation, their alteration by externally applied galvanic currents and the increase in the interspike interval following an interposed shock. Empirical studies show that some vestibular afferents have a regular spacing of action potentials, others an irregular spacing (Goldberg and Fernández 1971b; Fernández and Goldberg 1976). Irregularly discharging afferents have a higher sensitivity to externally applied galvanic currents than do regular afferents (Goldberg et al. 1984). To explain the relation between galvanic sensitivity and discharge regularity requires the assumption that neurons differ in both their synaptic noise (_v) and the slopes of their postspike voltage trajectories (d _v/dt). The more irregular the neuron's discharge at a given firing frequency, the greater is _v and the smaller is d _v/dt. Of the two factors, d _v/dt is estimated to be four times more influential in determining discharge regularity across the afferent population. The shortcomings of the model are considered, as are possible remedies. Our conclusions are compared to previous discussions of mechanisms responsible for differences in the discharge regularity of vestibular afferents.     

Activation of hepatic and inactivation of renal xanthine dehydrogenase activity by dexamethasone during the prenatal period in chick eggs

In the experiments reported here, the effect of dexamethasone on hepatic and renal xanthine dehydrogenase toward the end of incubation was studied. Dexamethasone injected on day 17 of incubation into the chick eggs increased the hepatic and decreased the renal activity of xanthine dehydrogenase. Furthermore, dexamethasone was found to accelerate fetal development as measured by the precocious decrease of thymidine uptake into nuclear DNA and by the increased fetus weight/egg weight ratio. The data suggest that an activation of xanthine dehydrogenase is caused by a hatching-independent factor and by a factor related to the hatching process.     

Use of Vital Dyes in Conjunction with the Semivitro Technic for the Detection of Pollen Tube Sperm Nuclei

The technique we describe here is a modification of that used by Hough et al. (1985), combined with “semivitro” pollen tube observations. With the semivitro technique, pollen tubes grow from the cut ends of pollinated styles (Brewbaker and Majumder 1961). Pollen of Nicotiana alata was presoaked for 15 min in simplified medium (Brewbaker and Kwack 1963) (10% sucrose, 300 ppm Ca(NO₃)₂, 100 ppm H₃BO₃ with the addition of 0.5 mg/ml of Hoechst 33258 stain from Serva Biochemicals, Heidelberg, Control H, purchased June 1983). (For germination of Nicotiana alata pollen in vitro, we use this same solution, except with 12% sucrose). After this prestaining, the pollen suspension was centrifuged for 5 min at 1200 × g, the pellet resuspended in control Brewbaker medium (i.e., no stain), recentrifuged and used to pollinate detached pistils. The pistils were then incubated at 25 C in a water-saturated atmosphere for 20 hr. At this time, the styles were cut just ahead of the front of the growing pollen tubes (Mulcahy and Mulcahy 1985) and the cut stylar ends each dipped in fresh control Brewbaker medium. Twelve to 24 hours later, tubes growing out of the cut styles were viewed by fluorescence microscopy (exciter filter, BG 12 + KV 418, beam splitter, 500 nm, and barrier filter OG 515). A distinct green fluorescence was seen in the generative and vegetative nuclei (Fig. 1).     

Measurement of lipoprotein lipase and hepatic triacylglycerol lipase in post-heparin plasma of the cynomolgus monkey

Conditions for measurement of the lipolytic activities, lipoprotein lipase and hepatic triacylglycerol lipase in cynomolgus monkey postheparin plasma are described. The two activities are separable by heparin-Sepharose chromatography. Goat anti-human hepatic triacylglycerol lipase serum inhibits monkey hepatic triacylglycerol lipase activity and allows direct measurement of lipoprotein lipase in post-heparin plasma. While both human and homologous serum can be used as a source of activator apolipoprotein, homologous serum produces a much greater activation.     

Regulation of nitrogen metabolism in glutamine auxotrophs of Klebsiella pneumoniae

We examined the regulation of nitrogen metabolism in four classes (glnA, glnB, glnF, and glnG) of Gln- auxotrophs of Klebsiella pneumoniae. These studies indicate that glutamine synthetase does not directly mediate the physiological response to NH4+ in this organism. We present evidence suggesting that the effect of NH4+ on the expression of genes involved in nitrogen metabolism involves the products of the glnF and glnG genes.     

In vivo inactivation of glycerol dehydrogenase in Klebsiella aerogenes: properties of active and inactivated proteins.

Glycerol:oxidized nicotinamide adenine dinucleotide (NAD+) 2-oxidoreductase (EC 1.1.1.6), an inducible enzyme for anaerobic glycerol catabolism in Klebsiella aerogenes, was purified and found to have a molecular weight of 79,000 by gel electrophoresis. The protein seemed to be enzymatically active either as a dimer of a 40,000-dalton peptide at pH 8.6 or as a tetramer of 160,000 molecular weight at pH 7.0. The enzyme activity was present at high levels in cells growing anaerobically on glycerol, but disappeared with a half-life of about 45 min if molecular oxygen was introduced to the culture. In contrast, no such phenomenon occurred with dihydroxyacetone kinase activity, the second enzyme in the pathway. Immunochemical analysis showed that the inactivation of the oxidoreductase did not involve degradation of the protein. Furthermore, subunits of the active and inactive forms of the enzyme were indistinguishable in size on polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate and had similar isoelectric points (pH 4.7). Inactivation did, however, alter the gel filtration properties of the enzyme protein and, more importantly, reduced its affinity for the dye Cibacron F3GA and the coenzyme NAD+.     

Mapping of the substrate-binding site of the human granulocyte elastase by the aid of tripeptidyl-p-nitroanilide substrates

The kinetic properties of the human granulocyte elastase /EC 3.4.21.11/ were investigated with 24 tripeptidyl-pNA substrates. By the regression analysis of the kinetic data obtained with 15 substrates a relatively hydrophobic compound, Boc-D-Phe-Ala-Nle-pNA, was predicted as the optimal substrate sequence. The compound was synthesized, assayed and the predicted K_m = 4.2 uM was confirmed experimentally. The substrate-binding site of granulocyte elastase appeared to be hydrophobic and very much similar to that of the pancreatic enzyme at the S₂–S₄ subsites, but the S₁ subsite, which determines the primary specificity, could accomodate bulkier residues and it was less selective than that in the pancreatic enzyme.     

Reversible modification of arginine residues in neocarzinostatin. Isolation of a biologically active 89-residue fragment from the tryptic hydrolysate

Reaction of the antitumor protein neocarzinostatin with 1,2-cyclohexanedione in 0.25 M borate buffer, pH 9.0, resulted in complete modification of arginine residues in positions 66, 67, and 78. The arginine-modified protein lost its native structure and was biologically inactive in the inhibition of growth of HeLa cells, inhibition of DNA synthesis, and in vitro DNA strand scissions. Trypsin hydrolysis of 1,2-cyclohexanedione-modified neocarzinostatin resulted in selective cleavage of the Lys-Val (positions 20 and 21) bond of the primary structure yielding NH2-terminal 1-20 and the COOH-terminal 21-109 residue fragments. The latter contained modified arginine residues. Both peptide fragments were biologically inactive. Treatment of the arginine-modified neocarzinostatin and the arginine-protected 89-residue fragment with 0.25 M Tris-acetate buffer, pH 9.0, for 15 h resulted in the release of 1,2-cyclohexanedione, regenerating all three arginine residues. The regenerated protein and the 89-residue fragment were fully active biologically. Further, the regenerated 89-residue fragment possessed 70% of the reactivity of neocarzinostatin with antibody raised against the native protein. The conformation of the 89-residue fragment was almost identical with that of the native protein in CD spectral properties.     

Formation of Short-Chain Fatty Acids from H2 and CO2 by a Mixed Culture of Bacteria

The biological utilization of CO₂ and H₂ for the formation of short-chain fatty acids was studied by using a mixed culture of bacteria. Optimization of a medium was carried out in continuous culture to identify limiting factors which controlled growth and production of organic acids. The optimal pH for growth and acid production was 7.0 at 37°C; the maximal cell concentration obtained was 5.9 g of cells per liter (dry weight), and the maximal amount of volatile acids formed was 4.7 g/liter, with acetic acid as the predominant acid. With the optimized medium, it was found that the rate of transfer of hydrogen or carbon dioxide, or both, from gas to liquid was the limiting factor which controlled growth and production of acids.     

Characterization and measurement of human apolipoprotein A-II by radioimmunoassay

The development of a radioimmunoassay for apolipoprotein A-II (apo A-II) is described. Initial studies revealed a lack of immunological identity between purified apo A-II used as the standard and serum or HDL. Extensive testing of different buffers, standards, antisera, tracers, utilization of a detergent, and heating of sera failed to resolve the problem. Gel filtration of iodinated and non-iodinated apo A-II on Sephadex G-100 columns showed that apo A-II, in dilute solution, elutes in a higher molecular zone than expected with a broad, assymetrical profile. The use of a subfraction of the tracer in the assay resulted in parallelism in the serum and standard dilution curves. The apo A-II assay was sensitive, specific, and reproducible. Apo A-II added to sera was fully recovered and delipidation did not affect the immunoreactivity of either serum or HDL. Apo A-II contributed approximately 20% to the protein mass of HDL. Comparison of these results with those obtained by radial immunodiffusion, and with previously reported data, indicates that the reactivity of apo A-II in its native and delipidated forms may be markedly influenced by different immunologic methodologies and their specific reagents. Caution should thus be shown at present in assigning absolute concentrations to apo A-II in serum or HDL.