Relationship between cardiac output and oxygen uptake at the onset of exercise

The purpose of the present study was to assess the relationship between the rapidity of increased gas exchange (i.e. oxygen uptake ) and increased cardiac output ( ) during the transient phase following the onset of exercise. Five healthy male subjects performed multiple rest-exercise or light exercise (25 W)-exercise transitions on an electrically braked ergometer at exercise intensities of 50, 75, or 100 W for 6 min, respectively. Each transition was performed at least eight times for each load in random order. The was obtained by a breath-by-breath method, and was measured by an impedance method during normal breathing, using an ensemble average. On transitions from rest to exercise, rapidly increased during phase I with time constants of 6.8–7.3 s. The also showed a similar rapid increment with time constants of 6.0–6.8 s with an apparent increase in stroke volume (SV). In this phase I, increased to about 29.7%–34.1% of the steady-state value and increased to about 58.3%–87.0%. Thereafter, some 20 s after the onset of exercise a mono-exponential increase to steady-state occurred both in and with time constants of 26.7–32.3 and 23.7–34.4 s, respectively. The insignificant difference between and time constants in phase I and the abrupt increase in both and SV at the onset of exercise from rest provided further evidence for a cardiodynamic contribution to following the onset of exercise from rest.     

Simultaneous determination of nitrazepam and its metabolites in urine by micellar electrokinetic capillary chromatography

We applied micellar electrokinetic capillary chromatography to simultaneous separation and determination of nitrazepam and its major metabolites, 7-aminonitrazepam and 7-acetamidonitrazepam, in spiked urine. Prior to electrophoresis, the three compounds were successfully extracted from the spiked urine with commercial disposable solid-phase cartridges. The optimum running buffer for the separation was prepared by combining 85 parts of 60 mM sodium dodecyl sulphate—6 mM phosphate—borate, adjusted to pH 8.5, with 15 parts of methanol. The separation order, completed within 25 min, was 7-aminonitrazepam > 7-acetamidonitrazepam > nitrazepam, at an applied potential of 20 kV. We obtained reproducible electropherograms in successive repetitions, and few other peaks or interferences appeared in the electropherogram. The detection limits of the three compounds were 50–100 pg (0.1–0.2 μg/ml of analyte in spiked urine), and the recoveries were 78.9–100.8% for 1 μg/ml and 84.1–100.3% for 5 μg/ml. The application of this method to forensic or clinical samples is demonstrated.     

Tissue-Specific Isoforms of Catalase Subunits in Castor Bean Seedlings

Catalases purified from endosperm glyoxysomes and non-specializedmicrobodies from hypocotyls of castor bean seedlings differedin their specific activity [90–164 and 0.89–4.9kunits (mg protein)^–1, respectively] and in their constituentsubunits [two subunits of 54 and 56 kDa for the endosperm enzymeand only one of 56 kDa for the hypocotyl enzyme]. Immunoblotanalysis also showed that particulate fractions from the endospermsand from etiolated and green cotyledons contained two catalasesubunits of 54 and 56 kDa, whereas such fractions from the hypocotylsand roots contained only the 56-kDa subunit. Leaf peroxisomesfrom green leaves had two catalase subunits of around 55 kDaeach. Results of translation in vitro indicated that the 54-and 56-kDa subunits were translated from distinct mRNAs andlevels of both mRNAs increased in the endosperms during germination,prior to increases in levels of catalase proteins. In the hypocotyls,the 56-kDa subunit seemed to be synthesized constitutively. ¹Present addresses: YO, Toyota Central Institute, 31-9 Musashizuka,Nagabuchi, Nagakute, Aichi 480-11, Japan     

Phototropism in hypocotyls of radish. VI. No exchange of endogenous indole-3-acetic acid between peripheral and central cell layers during first and second positive phototropic curvatures

Using a physicochemical method, the distribution of endogenous indole-3-acetic acid (IAA) was measured in the peripheral and central cell layers, as well as in the illuminated and shaded sides of phototropically stimulated radish hypocotyls (Raphanus sativus var. hortensis f. gigantissimus Makino). The IAA was evenly distributed over the illuminated and shaded sides in the first and second positive phototropic curvatures induced by a pulse or a continuous unilateral illumination with blue light. Also, no net exchange of the IAA between the peripheral and central cell layers was observed during these curvatures. These results suggest that phototropism of radish hypocotyls cannot be described by the Cholodny-Went theory.     

Upregulation of Fas-induced apoptosis by interferon-γ accompanied by increased ICE expression in renal cell cancer cells

The integration of Fas/Apo-1 (CD95) by Fas ligand or anti-Fas antibody induces apoptosis, and this system plays a pivotal role for the lysis of target cells by cytotoxic T lymphocytes. Fas-mediated apoptosis is also increased by a prior incubation of Fas-bearing cells with interferon(IFN)-. Interleukin-1- converting enzyme (ICE) and/or CPP32, or other members of ICE family act as direct cell death executors downstream of this mechanism, and a tetrapeptide inhibitor of these cysteine proteases blocks Fas-mediated apoptosis. In this study, we examined the effect of IFN- on Fas-mediated apoptosis in ACHN cells. IFN- augmented apoptosis in a dose dependent manner and reached a plateau at 400 U/ml when exposed for 48 h before the end of culture. The kinetics revealed a significant increase in apoptosis after 24 h. Exposing ACHN cells to IFN- increased pro-ICE expression accompanied with a decrease of pro-CPP32. These results suggest that direct enhancement of ICE expression and/or upregulation of conversion of pro-CPP32 to active form increases Fas-mediated apoptosis by IFN- in ACHN cells.     

Nuclear localization activity of phytochrome B

Phytochromes are soluble red/far-red-light photoreceptor proteins which mediate various photomorphogenic responses of plants. Despite much effort, the signal transduction mechanism of phytochrome has remained obscure. Phytochromes are encoded by a small multigene family in Arabidopsis . Among the members of the family, phytochrome A (phyA) and B (phyB) are the best characterized. PhyB contains putative nuclear localization signals within its C-terminal region. Transgenic Arabidopsis plants were produced which expressed a fusion protein consisting of GUS and C-terminal fragments of phyB. GUS staining from the fusion protein in these transgenic plants was observed in the nucleus, which suggests that the nuclear localization signal of the fragment is functional. Next, it was examined whether the endogenous phyB was detected in the nucleus. Nuclei were isolated from the light-grown wild-type Arabidopsis leaves and subjected to the immunoblot analysis. The result indicated that a substantial fraction of total phyB was recovered in the isolated nuclei. This result was further confirmed by the immunocytochemical analysis of the protoplasts. Finally, the effects of light treatments on the levels of phyB in the isolated nuclei were examined. Dark adaptation of the plants before the nuclear isolation reduced the levels of phyB. The reduction was accelerated by irradiation of plants with far-red light before the transfer to darkness. Thus, nuclear localization of phyB was suggested to be light-dependent.     

Purification and Characterization of a Ca2+-Dependent Protein Kinase from the Halotolerant Green Alga Dunaliella tertiolecta

A Ca²⁺-dependent protein kinase (CDPK) that has been partiallypurified and characterized previously [Yuasa and Muto (1992)Arch. Biochem. Biophys. 296: 175] was further purified to about20,000-fold from the soluble fraction of Dunaliella tertiolecta.The enzyme preparation contained 60- and 52-kDa polypeptidesboth of which phosphorylated casein as a substrate. Both polypeptidesshowed a Ca²⁺-dependent increase in mobility during SDS-PAGEand ⁴⁵Ca²⁺-binding activity after SDS-PAGE and electroblottingonto a nitrocellulose membrane, suggesting that both the 60-and 52-kDa CDPKs directly bind Ca²⁺. The protein kinase inhibitors,K-252a and staurosporine, inhibited the CDPK competitively withrespect to ATP. An antibody raised against the 60-kDa CDPK crossreactedwith both the 60- and 52-kDa polypeptides. Both molecular specieswere autophosphorylated in the presence of Ca²⁺, and a highlyphosphorylated 80-kDa band appeared in addition to these phosphorylatedbands at 60 and 52 kDa in SDS-PAGE. However, the specific activityof CDPK was not changed by prior autophosphorylation when theautophosphorylated enzyme was assayed as a mixture of thesephosphorylated molecular species. Only the 60-kDa polypeptidewas immunodetected in subcellular fractions of Dunaliella cells.The 52-kDa polypeptide increased during storage of the enzyme.These results suggest that the 52-kDa polypeptide is a proteolyticartifact produced during purification. Immunoreactive bandsof 60-kDa were detected in extracts of several green algae butnot in extracts of higher plants or a brown alga. ¹This research was partly supported by Grants-in-Aid from theMinistry of Education, Science and Culture, Japan (No. 06454013and 06304023) and Research Fellowship of the Japan Society forthe Promotion of Science for Young Sciencists. ²Research Fellow (PD) of the Japan Society for the Promotionof Science.     

Formation of a metallocycle by dimerization of acrylonitrile. Crystal structure of the hexakis{(1,4-dicyanobutane-1,4-diyl)-nickel(II)} complex

A novel hexanickel(II) complex [Ni₆(NCCHCH₂CH₂CHCN)₆] (2) with 1,4-dicyanobutane-1,4-diyl (L) which was produced by the metal-induced dimerization of acrylonitrile (AN) has been isolated and the structure has been determined crystallographically. Complex 2 is triclinic, space group

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. Each nickel atom is coordinated by two carbon atoms of L and two nitrogen atoms of the cyano group of two other L, providing a square-plenar geometry. The six nickel atoms are bridged by the cyano group and carbon atom to form the slightly distorted octahedral Ni₆ core.