Tonic and phasic modifications of viscerosensory evoked potentials during sleep in cats

Modification of the viscerosensory evoked potentials (EPs) were studied during the sleep-wakefulness cycle of the rat. Electrical stimuli of various intensity were delivered either to the mucosal surface of a fistula of the small intestine or to the left splanchnic nerve during wakefulness (W), drowsiness (D), slow-wave-sleep (SWS), and paradoxical sleep (PS). The average EPs were recorded from the somatosensory (SI and SII) and associative (AS) areas of the cortex, the ventrobasal complex of the thalamus (VPL), the posterior hypothalamus (HPT) and the dorsal hippocampus (HPC). The amplitude of each component of the EPs in all explored structures were the largest in SWS and the smallest in W. A phasic increase in amplitude was observed in the EPs recorded immediately before the appearance of the spindles of SWS and during the REM episodes of PS. The peak latencies of the late components were the longest in SWS. These changes of the amplitudes and latencies were greater in the responses to weak stimulation than in EPs to strong ones. The possible synaptic events of the sleep-dependent control of viscerosensory activity are discussed.     

Transition-state stabilization at the oxyanion binding sites of serine and thiol proteinases: hydrolyses of thiono and oxygen esters

X-ray diffraction studies suggested that the tetrahedral intermediate formed during the catalysis by serine and thiol proteinases can be stabilized by hydrogen bonds from the protein to the oxyanion of the intermediate [cf. Kraut, J. (1977) Annu. Rev. Biochem. 46, 331-358; Drenth, J., Kalk, K.H., & Swen, H.M. (1976) Biochemistry 15, 3731-3738]. To obtain evidence in favor or against this hypothesis, we synthesized thiono substrates (the derivatives of N-benzoyl-glycine methyl ester and N-acetylphenylalanine ethyl ester) containing a sulfur in place of the carbonyl oxygen atom of the scissile ester bond. We anticipated that this relatively subtle structural change specifically directed to the oxyanion binding site should produce serious catalytic consequences owing to the different properties of oxygen and sulfur if transition-state stabilization in the oxyanion hole is indeed important. In fact, while in alkaline hydrolysis the chemical reactivities of oxygen esters and corresponding thiono esters proved to be similar, neither chymotrypsin nor subtilisin hydrolyzed the thiono esters at a measurable rate. This result substantiates the crucial role of the oxyanion binding site in serine proteinase catalysis. On the basis of the similar values of the binding constants found for oxygen esters and their thiono counterparts, it can be concluded that the substitution of sulfur for oxygen significantly influences transition state stabilization but not substrate binding. The thiol proteinases papain and chymopapain react with the oxygen and thiono esters of N-benzoylglycine at similar rates. Apparently, in these reactions the above stabilizing mechanism is absent or not important, which is a major mechanistic difference between the catalyses by serine and thiol proteinases.     

The chimpanzee M blood-group antigen is a variant of the human M-N glycoproteins

Chimpanzee erythrocytes express strong M but weak, occasional N blood-group activity, as detected by anti-M and anti-N reagents. We have found that the M activity is carried by a major membrane glycoprotein that is similar but not identical to the human MM glycoprotein (glycophorin A). We have isolated and characterized this glycoprotein from erythrocyte membranes of four individual chimpanzees. The purified glycoproteins strongly inhibited agglutination of M cells by rabbit anti-human M sera and only weakly inhibited the agglutination of N cells by rabbit anti-human N sera. They also displayed medium-to-strong inhibitory activity against chimpanzee iso- and crossimmune antisera tested with chimpanzee erythrocytes of various V-A-B-D and W^c specificities, which are known as chimpanzee extensions of the human type M-N system and the Miltenberger counterpart, respectively. Each glycoprotein was cleaved with CNBr into three fragments, whose size, solubility, and composition were analogous to those obtained by similar treatment of the human M-N antigens. The amino-terminal fragment was found to be a glycooctapeptide whose amino acid composition and partial sequence indicated that it is an intermediate form of the human M and N glycooctapeptides. Its carbohydrate content comprised two threonine-linked saccharide units that, although similar in composition to the human threonine-linked units, were fewer in number than the three units found in the corresponding human glycooctapeptides. Structural similarities to the human antigens strongly suggest that the amino terminus bears the major antigenic determinants of the molecule, and the occurrence in this region of numerous, albeit rare, variants among humans and in chimpanzees indicates that the corresponding coding sequence of the structural gene is particularly susceptible to mutational events. We conclude that the chimpanzee M gene product is a variant of the human type and that the chimpanzee gene is an allele of the human polymorphic M-N locus.This research was supported by National Institutes of Health Grants GM 16389 and HL 19011 and March of Dimes Grant 1-661.     

Identification of guanine and adenine adducts in DNA alkylated by dibromodulcitol in vitro and in vivo

DNA reacted with dibromodulcitol in neutral solution yielded 3- and 7-alkyl substituted purines after hydrolysis at neutral pH-value at 37°C. The alkylated products were identified by mass spectrometry and by comparison of their UV absorption spectra and chromatographic properties on thin-layer chromatography (TLC) and various columns with those of the corresponding galactitylpurine derivatives obtained by synthetic route from alkylation of the appropriate nucleic bases or nucleosides. The labelled alkylpurines occurring in DNA of Yoshida tumour cells treated with [³H]dibromodulcitol in vivo were also indentified by co-chromatography of labelled DNA hydrolysate with synthetic 3- and 7-alkyl substituted purines. On the basis of the same chromatographic behaviour 3-(1-deoxy-3,6-anhydrogalactit-1-yl)adenine, 7-(1-deoxygalactit-1-yl)guanine, 7-(1-deoxy-3,6-anhydrogalactit-1-yl)guanine and 1,6-di(guanin-7-yl)-1,6-dideoxygalactitol were identified as main alkylated products in tumor cell DNA after in vivo treatment with dibromodulcitol.     

Fractionation and characterization of rapidly phosphorylated nuclear proteins in salivary gland cells of Chironomus tentans

Rapidly phosphorylated nuclear proteins were investigated in explanted salivary gland cells of Chironomus tentans after labeling with ³²P_i. After sonication nuclei were fractionated by centrifugation at 18,000 g into sedimentable (80% of ³²P) and not sedimentable (supernatant) material. About 90% of ³²P in the supernatant fraction was sedimentable at 100,000 g (disperse chromatin). The disperse chromatin contained 20%–40% of the total nuclear DNA but only 5%–20% of ³²P. The ³²P-labeled phosphoproteins in the material pelleted at 20,000 g were further fractionated by differential solubility in lysis buffer. Electrophoretic analyses on SDS polyacrylamide gels resolved the ³²P-labeled nuclear proteins into 12 major bands in the M_r range of 12,000–120,000. The incorporation of ³²P into most bands reached a steady-state within 5–10 min of incubation with ³²P_i and was not measurably influenced by cycloheximide, an inhibitor of protein synthesis. The phosphate groups are linked to polypeptide chains by bonds vulnerable to pronase and alkaline phosphatase. All major bands in the pelleted chromatin were also present in the disperse chromatin except for an M_r 95,000 phosphoprotein. Two of the fastest moving ³²P-bands comigrated with the core histones H2A and H4. Both possessed a high pI value and were insoluble in 0.35 M NaCl. The H2A-like protein was partially soluble in lysis buffer while the H4-like one was not. The two fast moving ³²P-labeled bands with rapidly turned over phosphates may be fractions or variants of the core histones H2A and H4.     

Free radical signal of bile pigment in paraffin embedded liver tissue

A linear correlation have been found between the amplitude of the free radical signal of the electron spin resonance (ESR) spectrum of paraffin embedded liver blocks and the number of bile casts in the histological sections made from these blocks. It has been suggested that the major part of the ESR free radical signal arises from bile pigment, but the contribution of "age pigment" cannot be excluded. On the basis of model experiments the majority of these radical centers was assigned to protein bound bilirubin. In the course of histological processing more than 80% of various free radical centers arising in air dried liver tissue is extracted.     

Crystal and molecular structure of (E)-5-(2-bromovinyl-2''-deoxyuridine)

(E)-5-(2-bromovinyl-2'-deoxyuridine) crystallizes in the space group P2(1) with a = 12.976(1), b = 4.800(1), c = 20.385(2) A, beta = 96.88(1) degrees, Z = (two molecules a and b in the asymmetric unit). The structure has been determined by the use of 2400 diffractometer reflexions and refined by least-squares to R of 0.053. Conformational features of both molecules a and b resemble those of thymidine. The ribofuranose rings assume the rare C(3')-exo form observed also in thymidine. Similarly, the torsion angles around the glycosidic bonds (mean = 40(1) and 56(1) degrees fall in the anti range. In each molecule the best plane of the 2-bromovinyl moiety is bent out of the least-squares plane of the pyrimidine base by 6 degrees, so that the positively charged C(8)-H(8) group can donate an intramolecular hydrogen bond to 0(4) atom. Eight strong and weak intermolecular hydrogen bridges are built up between the symmetry independent and related molecules forming a complicated three dimensional hydrogen bond network.     

Annotated chromosome counts of czechoslovak plants (31–60) (Materials for “Flóra ČSSR”—3)

Chromosome counts of the following 30 taxa (106 populations) are given:Betonica officinalis (2n=16);Bidens frondosus (2n=48);Calamagrostis arundinacea (2n=28+0–2B);Dianthus carthusianorum subsp.latifolius (2n=30);Festuca gigantea (2n=42, 42+2B);Hypericum perforatum (2n=32);Koeleria macrantha (2n=28);Kohlrauschia prolifera (2n=30);Lilium martagon (2n=24+0–2B);Melica ciliata (2n=18);Poa remota (2n=14);Ranunculus polyanthemos (2n=16);R. sardous subsp.sardous (2n=16);Roegneria canina (2n=28+0–1B);Rudbeckia laciniata (2n=76);Scabiosa canescens (2n=16);Serratula tinctoria (2n=22);Seseli elatum subsp.heterophyllum var.beckii (2n=18);S. hippomarathrum (2n=20);Thlaspicaerulescens caerulescens subsp.tatrense (2n=14);Trifolium alpestre (2n=16);T. avense (2n=14);T. medium (2n=79, 80+0–2B, 82);T. rubens (2n=16);Veronica officinalis subsp. alpestris (2n=36);Vincetoxicum hirundinaria (2n=22);Vulpia bromoides (2n=14);Zerna benekenii (2n=28)Z. monoclada (2n=28+0–8B);Z. ramosa (2n=42). Remarks on taxonomy, nomenclature and chorology for some of these taxa are given.