Relationship between enhancement of morphine analgesia and inhibition of enkephalinase by 2S, 3R 3-amino-2-hydroxy-4-phenylbutanoic acid derivatives

The effects of nineteen AHPA* derivatives were examined on morphine analgesia by tail-flick test in rats and on enkephalinase inhibition which was based on the formation of tyrosyl-glycyl-glycine from met-enkephalin. The correlation between the enhancement of morphine analgesia in vivo and enkephalinase inhibition in vitro was analyzed. The different analogs varied considerably in the degree of enhancement of morphine analgesia and inhibition of enkephalinase. A close relationship between enkephalinase inhibition expressed by IC50 in vitro and enhancement of morphine analgesia in vivo was observed in thirteen out of nineteen AHPA derivatives examined. One of other six AHPA derivatives which showed weak effectiveness in potentiating on morphine analgesia but was highly potent as an enkephalinase inhibitor, caused potent analgesic action when it was applied intracisternally indicating poor penetration of the blood brain barrier. The possibility was discussed that some of other compounds excluded from the linear relationship might act on other enkephalin degrading enzymes such as aminopeptidase.     

Duality of alpha-subunit of canine kidney sodium- and potassium-activated adenosinetriphophatase in electrophoresis

Sodium- and potassium-activated adenosinetriphosphatase (Na+, K+-ATPase) purified from dog kidney outer medulla was examined by polyacrylamide gel electrophoresis and by photoaffinity labeling with N-(ouabain)-N'-(2-nitro-4-azidophenyl)-ethylenediamine (NAP-ouabain). The large subunit band (alpha-band) split into two bands on the gel after the enzyme was heat-treated in the presence of 1% sodium dodecylsulfate (SDS). Of the two bands (alpha I and alpha II), alpha I had the same electrophoretic mobility as the original band, while alpha II moved slightly faster. Total conversion into alpha II was not observed, about half of the original remaining as alpha I. Below 60 degree C, heat treatment did not produce alpha II. Phenylmethylsulfonyl fluoride did not prevent the appearance of alpha II. Both alpha I and alpha II were labeled with [3H]NAP-ouabain. Nonspecific incorporation of [3H]NAP-ouabain also occurred irrespective of illumination, but it was removed either by diffusion during staining and destaining of the gel or by treatment of the enzyme with trichloroacetic acid. It is tentatively concluded that the splitting of the band reflects some intrinsic differences in situ of the alpha-subunit of dog kidney membrane Na+,K+-ATPase.     

Photochemical reaction of 9-cis-retro-γ-rhodopsin at low temperatures

9-cis-Retro-γ;rhodopsin (λ_max = 420 nm) was prepared from 9-cis-retro-γ-retinal and cattle opsin. After cooling to liquid nitrogen temperature (77 K), the pigment was irradiated with light at 380 nm. The spectrum shifted to the longer wavelengths, owing to formation of a batho product. This fact indicates that the conjugated double bond system from C-5 to C-8 of the chromophoric retinal in rhodopsin was not necessary for formation of bathorhodopsin. Reirradiation of the batho product with light at wavelengths longer than 520 nm yielded a mixture composed of presumably 9- or 11-cis forms of retro-γ-rhodopsin. These three isomers are interconvertible by light at liquid nitrogen temperature. Thus the retro-γ-rhodopsin system is similar in photochemical reaction at 77 K to cattle rhodopsin system. Each system has its own batho product. Based on these results, it was infered that the formation of bathorhodopsin is due to photoisomerization of the chromophoric retinal of rhodopsin and is not due to translocation of a proton on the ring or on the side chain from C-6 to C-8 of the chromophoric retinal to the Schiff-base nitrogen.     

Relationship between secondary constrictions and nucleolus organizing regions inAllium sativum chromosomes

Summary Allium sativum L. (2 n=16) had three types of clones with regard to the number of chromosomes carrying well-defined secondary constrictions: the first type had two secondary constricted chromosomes (type I), the second had three (type II) and the third had four (type III). Silver staining was applied to these three types of cells to determine the number of nucleolus organizing regions (NORs) per cell and to study the relationship between the morphological appearance of the secondary constrictions and the ability of the chromosomes to form nucleoli. Ag-positive regions appeared on two chromosomes in type I, on three in type II and on four in type III. The comparison of Giemsa and Feulgen stained chromosomes with the silver stained ones clearly indicated that the positive reaction with silver occurred exclusively on the secondary constricted regions that responded negatively to both Giemsa and Feulgen staining, indicating that the size of the achromatic secondary constrictions directly reflects the volume of the Ag-positive materials. However, all three types of clones had a maximum of four nucleoli at interphase. Of the four nucleoli, either two or one was extremely small (less than 1 m in diameter) in types I and II respectively. The size variations of the other nucleoli seemed to be positively correlated with those of the Ag-positive regions. This and the observation that the maximum number of nucleoli per cell did not coincide with the number of Ag-positive regions on the metaphase chromosome complement suggest strongly that the NORs responsible for the minute nucleoli cannot be detected on the metaphase chromosomes. The present observations indicate that not all NORs are indicated by the morphological appearance of secondary constrictions.     

A probable lack of function of c-type cytochrome in the photosynthetic system of the blue-green alga Anabaena variabilis

Quantitative study of the cytochrome c acting in the photosyntheticsystem of the blue-green alga Anabaena variabilis (M-2) wasdone with membrane fragments and intact cells. Membrane fragments highly active in the NADP⁺-Hill reaction(above 200 µmoles/mg chl.a;-hr) retained photoresponsivecytochrome c equal only one-tenth that of P700, while the plastocyanincontent was almost equal to that of P700. The cytochrome contentin intact cells was a little larger than that in membrane fragmentson the chlorophyll a basis. However, the values relative toP700 (1/9) and plastocyanin (1/10) were identical with thosein membrane fragments. The content was also far smaller thanthat of reaction center II's (1/6). If the cytochrome mediatesall electrons from reaction center II, the cytochrome oxidation-reductionshould have a rate constant of 2.4?102 sec^–1 which isone order above of the rate constant of the cytochrome reduction(2.3 to 3.5?10¹sec^–1). These quantitative relationshipsindicate that in Anabaena variabilis (M-2), c-type cytochrome,either cytochrome f or algal cytochrome c, cannot function inthe main electron flow between two reaction centers. (Received September 8, 1978; )     

Possible participation of glucocorticoid in elevation of 3',5'-cyclic AMP levels through inhibition of cyclic AMP phosphodiesterase.

Administration of prednisolone and cholate to rats elevated levels of cAMP (adenosine 3',5'-cyclic monophosphate) by 1.5- to 2.0-fold. Compounds such as prednisolone, hydrocortisone, cholate, and deoxycholate were found to be potent inhibitors of partially purified cAMP phosphodiesterase prepared from rat liver. Kinetic analysis showed that the prednisolone inhibition was noncompetitive with a Ki of 8.9 x 10(-4) M. These results suggest that in addition to increasing DNA-dependent RNA polymerase activity in vivo, a large application of glucocorticoid may incur elevation of intracellular cAMP levels.     

Molecular cloning of bovine leukemia virus DNA integrated into the bovine tumor cell genome

The bovine leukemia virus (BLV) DNA harbored in the bovine tumor cell genome was cloned in lambda Charon 4A phage. Using either representative or 3' half-enriched BLV cDNA as a blot hybridization probe, clone lambda BLV-1 was shown to carry 9 kb of the BLV genome, flanked by cellular sequences at both ends. Restriction mapping with twelve endonucleases and hybridization of the DNA fragments to BLV cDNA representing a 3'-end portion of the viral genome revealed the presence and precise location of two long terminal repeats (LTRs) and virus-cell junctions. Thus, lambda BLV-1 appears to contain the complete BLV genome and flanking tumor cellular sequences. The restriction map of the cloned BLV proviral DNA closely resembles that previously reported for unintegrated linear proviral DNA, but differs significantly from that of the integrated provirus of another BLV isolate, the difference occurring preferentially in the putative gag and pol genes.