The effects of axotomy on electrophysiological properties of B cells of bullfrog sympathetic ganglia conditioned by a previous lesion

In bullfrog sympathetic B cells, axotomy decreases the amplitude and decay time of membrane afterhyperpolarization (AHP) and increases action potential (AP) duration. A second (test) axotomy, 7 days after an initial (conditioning) axotomy, did not amplify these changes. No recovery of AHP amplitude or AP duration occurred by 56 days post-axotomy, but AHP decay time recovered 21 days earlier than after test axotomy alone. Conditioning, previously shown to accelerate regeneration, speeds the return to normal of those membrane properties previously shown to recover after axotomy.     

Suppressors of the ras2 Mutation of SACCHAROMYCES CEREVISIAE

Saccharomyces cerevisiae contains two members of the ras gene family. Strains with disruptions of the RAS2 gene fail to grow efficiently on nonfermentable carbon sources. This growth defect can be suppressed by extragenic mutations called sra. We have isolated 79 independent suppressor mutations, 68 of which have been assigned to one of five loci. Eleven additional dominant mutations have not been assigned to a specific locus. Some sra1 and SRA4 and all SRA3 mutations were RAS independent, allowing growth of yeast cells that lack a functional RAS gene. Mutations in sra1, SRA3, SRA4 and sra6 are linked to his6, ino1, met3 and ade6, respectively. Some sra mutants have pleiotropic phenotypes that affect glycogen accumulation, sporulation, viability, respiratory capacity and suppression of two cell-division-cycle mutations, cdc25 and cdc35. The proposed functions of many of the suppressor genes are consistent with the model in which RAS activates adenylate cyclase.     

alpha-Difluoromethylornithine induces differentiation of a human embryonal carcinoma cell line in vitro

Human embryonal carcinoma cells could serve as a useful model system for analysis of early human development. A limited number of human embryonal carcinoma cell lines have been generated from in vivo tumors. We report here that alpha-difluoromethylornithine, a specific enzyme-activated inhibitor of ornithine decarboxylase activity, can induce differentiation in human embryonal carcinoma cells. The differentiated phenotype could be distinguished from undifferentiated cells by altered cellular morphology, biochemical and cell surface antigenic properties. These results suggest that alterations in the intracellular levels of polyamines may play a role in human embryonal carcinoma cell differentiation, and possibly human embryogenesis.     

Production of steroids and release of prostaglandins by spherical pig blastocysts in vitro

Levels of pregnenolone and progesterone in spherical pig blastocysts (near 4 and 15 microM respectively) exceeded respective levels in histotroph by about 400-fold. When blastocysts were cultured for 5 days in a synthetic medium containing pregnenolone sulfate (1 microM), daily rates of release of pregnenolone, progesterone, androstenedione, testosterone, oestrone and oestradiol were determined to be near 320, 45, 26, 27, 0.8 and 9.2 fmol per blastocyst respectively. Daily outputs of progesterone and testosterone (fmol per blastocyst) diminished (P less than 0.05) to 1.3 and undetectable levels (less than 2) respectively in the presence of Trilostane (94 microM). Increasing the content of pregnenolone sulfate in the culture medium (to 4.5 microM) resulted in higher daily rates of release of pregnenolone and progesterone (to near 1740 and 380 fmol per blastocyst respectively), verifying activity of 3 beta-hydroxy-delta 5-steroid dehydrogenase, and of arylsulfatase, in tissues of intact spherical pig blastocysts. Prostaglandin E2 was the predominant prostaglandin (PG) released by cultured blastocysts (about 1 fmol per blastocyst per hour), hourly rates of release of PGH2 (derived) and PGF2 alpha being near 0.1 and less than 0.06 fmol per blastocyst respectively. The data establish a capacity for spherical pig blastocysts to release a range of steroids and PGs of possible significance to embryonic growth and development in vivo.     

Spontaneous interstitial nephritis in kdkd mice. II. Characterization of a tubular antigen-specific, H-2K-restricted Lyt-2+ effector T cell that mediates destructive tubulointerstitial injury

In this report, we have examined the effector T cell repertoire in the spontaneous interstitial nephritis of kdkd mice. Lymph node cells from nephritic kdkd mice are capable of transferring this disease into thymectomized, irradiated, and bone marrow-reconstituted CBA/Ca recipients. CBA/Ca mice do not spontaneously develop interstitial nephritis and are normally resistant to the adoptive transfer of nephritic cells, a resistance that in the short term can be attenuated with low-dose cyclophosphamide. We therefore used delayed-type hypersensitivity responses and direct transfer of immune cells under the renal capsule to characterize nephritogenic effector cells from kdkd donor mice. Lyt-2+, L3T4- T cells from the peripheral lymphoid organs of nephritic kdkd mice, after adoptive transfer into cyclophosphamide-pretreated CBA/Ca recipients, mediate an antigen-specific delayed-type hypersensitivity response to renal tubular basement membrane antigens. These cells are restricted by gene products in H-2Kk; they are also present in nephritic, but not in control kidneys. We have also observed this same phenotypic subpopulation of kdkd lymphocytes mediate a destructive interstitial renal lesion within 7 days of being placed under the kidney capsule of CBA/Ca mice. These findings suggest that T lymphocytes reactive to a parenchymal tubular antigen are of substantial importance in the development of spontaneous interstitial nephritis in kdkd mice.     

Recognition of vesicular lipoproteins by the apolipoprotein B,E receptor of cultured fibroblasts

Vesicular lipoproteins (e.g., lipoprotein-X) are found in plasma in cholestasis or following infusion of Intralipid or phospholipid. To investigate the metabolism of vesicular lipoproteins, we isolated them from the plasma of subjects with cholestasis or following chronic or single Intralipid infusion. Cholestasis and chronic Intralipid therapy were found to be associated with elevated plasma concentrations of apoE, as determined by radioimmunoassay. Vesicular lipoproteins purified from each of the three types of plasma contained apoE, as well as other proteins. In cholestasis, in which levels of apoE were up to five times normal, a major portion of the plasma apoE was on vesicular lipoproteins. Normalized for apoE content, all preparations of vesicular lipoproteins displaced 125I-labeled LDL from apoB,E receptors of cultured fibroblasts identically. This displacement was inhibited by monoclonal antibodies that block receptor binding of apoE. Vesicular lipoproteins containing 125I-labeled apoE were internalized and degraded by fibroblasts. Different preparations caused small losses or gains of cellular cholesterol, with appropriate stimulation or suppression of apoB,E receptors. Thus, vesicular lipoproteins contain apoE, and apoE mediates their interaction with the apoB,E receptor. Our results suggest that the catabolism of cholesterol-rich vesicular lipoproteins, formed during cholestasis or following infusions of Intralipid or phospholipid, may be receptor-mediated.     

Dietary effects on hematologic and serum chemical values in gray short-tailed opossums (Monodelphis domestica)

At the time of weaning (8 weeks), 57 gray short-tailed opossums (Monodelphis domestica) were placed in four dietary groups. One group was fed a horsemeat-based diet that had been used for 2.5 years in our colony, and three groups were fed three different commercial fox food diets. After the animals had reached sexual maturity (6 months), blood samples were collected and subjected to standard hematologic and serum chemical assays. Significant differences were observed among the dietary groups and between sexes in several values, but all animals appeared to be healthy and robust. The ranges, means, and standard deviations for the values presented here can be used as reference values for healthy young adult animals being fed these particular diets.     

Reversion and immobilization of phage Mud1 cts (Apr lac) insertion mutations in Salmonella typhimurium

Summary Phage Mud1 cts (Ap^r lac), or Mud1, insertion mutations may be accompanied by adjacent deletion formation which can complicate use of lac fusions generated with this phage for gene regulatory studies. As for phage Mu insertion mutations, phage Mud1 insertions fail to revert at significant frequency (whether or not accompanied by an adjacent deletion). We describe isolation of revertible (X mutant) derivatives of phage Mud1 in Salmonella typhimurium. The X mutant derivatives allow use of reversion as a simple test to determine whether a Mud1 insertion has occurred precisely without an adjacent deletion that may have fused the lac genes to a promoter outside of the gene of interest. In addition, a simple method for stabilizing Mud1 generated lac fusions against subsequent transposition is described.     

Purification and characterization of actophorin, a new 15,000-dalton actin-binding protein from Acanthamoeba castellanii

Actophorin is a new actin-binding protein from Acanthamoeba castellanii that consists of a single polypeptide with a molecular weight of 15,000. The isoelectric point is 6.1, and amino acid analysis shows an excess of acidic residues over basic residues. The phosphate content is less than 0.2 mol/mol. There is 0.4 +/- 0.1 mg of actophorin/g of cells, so that the molar ratio of actin to actophorin is about 10:1 in the cell. Unique two-dimensional maps of tryptic and chymotryptic peptides and complete absence of antibody cross-reactivity show that Acanthamoeba actophorin, profilin, capping protein, and actin are separate gene products with minimal homology. Actophorin has features of both an actin monomer-binding protein and an actin filament-severing protein. Actophorin reduces the extent of actin polymerization at steady state in a concentration-dependent fashion and forms a complex with pyrene-labeled actin that has spectral properties of unpolymerized actin. During ultracentrifugation a complex of actophorin and actin sediments more rapidly than either actin monomers or actophorin. Although actophorin inhibits elongation at both ends of actin filaments, it accelerates the late stage of spontaneous polymerization like mechanical shearing and theoretical predictions of polymer fragmentation. Low concentrations of actophorin decrease the length and the low shear viscosity of actin filaments. High concentrations cause preformed filaments to shorten rapidly. Ca2+ is not required for any of these effects. Muscle and amoeba actin are equally sensitive to actophorin.