Endothelin-3 stimulates the hypothalamic-pituitary-adrenal axis.

Endothelin-3 (ET-3) is a member of the novel vasoconstrictive peptide family, identified in porcine central nervous system. Intravenous bolus injection of 1000 pmol/kg of ET-3 in freely moving rats caused significant increases in plasma ACTH and corticosterone levels, almost equivalent to those of 100 pmol/kg of rat corticotropin-releasing hormone (rCRH). The action of ET-3 was virtually abolished by pretreatment of CRH-antagonist, alpha-helical CRH. When ET-3 was added to cultured anterior pituitary cells, neither direct stimulation of ACTH release nor potentiation of rCRH action was noted. The results indicate that ET-3 may function as a neuropeptide and stimulation of the CRH-neurons, direct or inderect, is mainly responsible for activation of ACTH and corticosterone release.     

Characterization and Intracellular Distribution of Pea Phytochrome I Polypeptides Expressed in E. coli

The pea phytochrome I (PI) cDNA clone, pPP1001, was expressedin E. coli. The plasmid pPP1001 contains pea PI cDNA which coversthe entire coding region with the Shine-Dalgarno consensus sequencejoined upstream of the cDNA in an expression vector pNUT6. ThepPP1001 transformants formed typical inclusion bodies when culturedat 32?C. However, when cultured at 37?C or in the presence ofisopropyl-ß-D-thiogalactopyranoside (IPTG) at 32?C,the bacteria lysed before inclusion body formation. Immuno-stainingwith anti-PI monoclonal antibody, mAP5, of transformants fixedby cold methanol showed that stainable materials were distributedin whole cytoplasmic region. When the inclusion bodies wereobserved clearly, the regions corresponding to the inclusionbodies became difficult to stain. Western blot analysis, however,showed that a ca. 100 kDa PI polypeptide was detected in thefraction from inclusion bodies and a ca. 90 kDa PI polypeptidefrom the soluble fraction. The amino acid sequence analysisof purified 100 kDa PI sample indicated that its amino terminusis blocked. However, minor signals in one experiment yieldeda sequence corresponding to the expected amino terminus of peaPI except for the initiation methionine. One of the anti-peaPI monoclonal antibodies, mAP9, that recognizes the near N-terminusof pea phytochrome was reactive to the 100 kDa polypeptide. (Received June 22, 1990; Accepted November 18, 1990)     

Cytoskeletal changes visualized by fluorescence microscopy during amoeba-to-flagellate and flagellate-to-amoeba transformations inPhysarum polycephalum

Summary Changes in the intracellular distribution of microtubules and microfilaments during amoeba-to-flagellate and flagellate-to-amoeba transformations inPhysarum polycephalum were examined by fluorescence microscopy using anti-tubulin antibody and NBD-phallacidin, respectively. Amoebae contained an extensive microtubular cytoskeleton, which was converted to a flagellar cone structure during transformation to flagellates in liquid medium. When flagellates reverted back to amoebae, this conical structure disintegrated prior to flagella resorption. Amoebae showed some microfilament-enriched domains along the periphery, from which numerous filamentous extrusions, probably pseudopods and filopods, emanated. Flagellates contained a ridge, a sheet-like structure, along their dorsal axis, especially in the earlier stages of flagellation. Another microfilament-enriched thick filamentous structure ran along the dorsal axis, starting from the anterior tip of the cell. This structure apparently coincided spatially with one of the bundles of microtubules. During the reversion to amoebae, other localized microfilaments were transiently observed at the posterior end. A model of cytoskeletal changes in the transformations between these two cell types was proposed.     

Characterization of antisera to 2-hydroxyestradiol and 4-hydroxyestradiol using 6-(O-carboxymethyl)oxime- and 17-hemisuccinate-bovine serum albumin conjugates and radioimmunoassay

For radioimmunoassay of the catechol estrogens, four hapten-bovine serum albumin (BSA) conjugates were prepared from 6-oxo-2-hydroxyestradiol 6-(O-carboxymethyl)oxime, 2-hydroxyestradiol 17-hemisuccinate, 6-oxo-4-hydroxyestradiol 6-(O-carboxymethyl)oxime and 4-hydroxyestradiol 17-hemisuccinate by coupling with BSA, employing the mixed anhydride method. The antisera elicited in rabbits by immunization with these antigens showed high affinity and specificity for 2-hydroxyestradiol or 4-hydroxyestradiol with cross-reactivities to a few structurally related estrogens. The specificity of antisera obtained is discussed in relation to the site of attachment of the hapten to BSA.     

On the relationship between two indices (“Bulk density” and “dry-matter content”) of dry-matter accumulation in plant organs

“Bulk density” and “dry-matter content” are useful indices of dry-matter accumulation in plant organs. A theoretical equation describing the relationship between these two indices was put forward. To examine the reliability of this equation, the seasonal changes of these two indices were investigated in the leaves and stems of different ages ofAucuba japonica. In each organ both indices varied seasonally almost parallel to each other, but the seasonal changes of dry-matter content were less obvious than those of bulk density. The observed bulk density was always larger than that calculated from the observed dry-matter content by the theoretical equation. A drying experiment showed that this discrepancy was caused by the decrement of the volume of the plant material by water loss during the period from the weight measurement to the volume measurement. When the water loss was negligible, the equation described well the relationships between the two indices of the experimental materials. It was also shown that this equation was useful for the estimation of the amount of air space in plant materials.     

Sensitivity of mice to systemic hyperthermia: effect of the transplantation of ascites tumor cells modified by preheating, and by an injection of CDDP or interferon

Effects of preheating and injection of cis-DDP (CDDP) or interferon on tumor-induced sensitization to systemic hyperthermia (SH) was investigated in mice. LD50 of SH at 42.0 +/- 0.2 degrees C (core body temperature) was 43 min in normal mice and 8 min in mice which were i.p. transplanted with FMA3 cells at a dose of 10(5) one day before. In mice which had received the SH for 10 min one hour before, one hour after or one day before the transplantation, LD50 of the SH one day after the transplantation was 41, 35 and 22 min, respectively. An injection of CDDP given i.p. at a dose of 4 mg/kg one day after the transplantation, which was effective to kill about 99% of the tumor cells, did not change the course of thermosensitization after the transplantation. An i.p. injection of mouse interferon did not change the thermosensitivity of normal mice, but greatly suppressed the thermosensitizing effect of tumor cells when it was given one day before the transplantation.