Observation of calcium uptake by isolated sarcoplasmic reticulum employing a fluorescent chelate probe

The fluorescent chelate probe technique is employed to observe the accumulation and binding of Ca⁺⁺ to isolated sarcoplasmic reticulum from skeletal and cardiac muscle. Chlorotetracycline serves as a fluorescent chelate probe which chelates to membrane bound Ca⁺⁺ giving rise to an intensely fluorescence adduct. An increase in fluorescence of chlorotetracycline is caused by ATP induced Ca⁺⁺ transport in both skeletal and cardiac muscle microsomes. The fluorescence spectra indicate that Ca⁺⁺ lies on the membrane surface in a relatively polar environment.     

Effect of Nalidixic Acid on Semiconservative Replication and Repair Synthesis After Ultraviolet Irradiation in Escherichia coli

Both semiconservative deoxyribonucleic acid replication and "extensive repair" synthesis, after ultraviolet irradiation, appear to be blocked by nalidixic acid. These findings suggest that the agent(s) responsible for both of these modes of replication, or some necessary common process or structure, is affected by this drug.     

Ultraviolet Light-Induced Division Delay in Synchronized Chinese Hamster Cells

The age-dependent, ultraviolet light (UVL) (254 nm)-induced division delay of surviving and nonsurviving Chinese hamster cells was studied. The response was examined after UVL exposures adjusted to yield approximately the same survival levels at different stages of the cell cycle, 60% or 30% survival. Cells irradiated in the middle of S suffered the longest division delay, and cells exposed in mitosis or in G₁ had about the same smaller delay in division. Cells irradiated in G₂, however, were not delayed at either survival level. It was further established, after exposures that yielded about 30% survivors at various stages of the cycle, that surviving cells had shorter delays than nonsurvivors. This difference was not observed for cells in G₂ at the time of exposure; i.e., neither surviving nor nonsurviving G₂ cells were delayed in division. The examination of mitotic index vs. time revealed that most cells reach mitosis, but all of the increase in the number of cells in the population can be accounted for by the increase of the viable cell fraction. These observations suggest strongly that nonsurviving cells, although present during most of the experiment, are stopped at mitosis and do not divide. Cells in mitosis at the time of irradiation complete their division, and in the same length of time as unirradiated controls. Division and mitotic delays after UVL are relatively much larger than after X-ray doses that reduce survival to about the same level.     

Effect of Prehumidification on Sampling of Selected Airborne Viruses

Studies were undertaken to determine if a prewetting device (humidifier bulb) used in combination with an all glass impinger (AGI-30) would increase the recovery of airborne mengovirus-37A, vesicular stomatitis virus (VSV), and the S-13 coliphage. Suspensions of T3 coliphage with mengovirus-37A, VSV, or S-13 were aerosolized and collected by using the AGI-30-humidifier bulb combination to sample the aerosols before and after shifts in relative humidities (RH). These studies revealed the following. (i) At low RH values there was a 3 to 4 log increase in recovery of airborne T3 phage; (ii) concomitantly, the recovery of mengovirus-37A and VSV decreased; and (iii) only at the mid-range RH values was the recovery of S-13 enhanced. The prehumidification technique significantly increased the recovery of airborne T3 phage but decreased the recovery of the two animal viruses tested.     

Evidence for a dipyrromethane cofactor at the catalytic site of E. coli porphobilinogen deaminase

Porphobilinogen deaminase isolated from Escherichia coli is shown to contain a dipyrromethane cofactor (DPMC) linked covalently to the enzyme. The structure of the cofactor is proposed on the basis of its reaction with Ehrlich's reagent and from its chemical properties. The cofactor is involved in the binding of intermediates during the catalytic reaction but is not incorporated into the product preuroporphyrinogen, E. coli strains containing the cloned porphobilinogen deaminase gene (hemC) when grown on 5-amino[14C]-levulinic acid incorporate 14C radioactivity specifically into the dipyrromethane cofactor of porphobilinogen deaminase.     

Mutagenicity in salmonella of hazardous wastes and urine from rats fed these wastes

15 hazardous industrial waste samples were evaluated for mutagenicity in the Salmonella plate-incorporation assay using strains TA98 and TA100 in the presence and absence of Aroclor 1254-induced rat liver S9. Dichloromethane/methanol extracts of the crude wastes were also evaluated. 7 of the crude wastes were mutagenic, but only 2 of the extracts of these 7 wastes were mutagenic; extracts of 2 additional wastes also were mutagenic. In addition, 10 of the crude wastes were administered by gavage to F-344 rats, and 24-h urine samples were collected. Of the 10 raw urines evaluated, 3 were mutagenic in strain TA98 in the presence of S9 and beta-glucuronidase. The 3 crude wastes that produced these 3 mutagenic urines were, themselves, mutagenic. Adequate volumes of 6 of the 10 raw urines were available for extraction/concentration. These 6 urines were incubated with beta-glucuronidase and eluted through Sep-Pak C18 columns; the methanol eluates of 3 of the urines were mutagenic, and these were the same 3 whose raw urines also were mutagenic. In general, the C18/methanol extraction procedure reduced the cytotoxicity and increased the mutagenic potency of the urines. To our knowledge, this is the first report of the mutagenicity of urine from rodents exposed to hazardous wastes. Based on the present results, the use of only strain TA98 in the presence of S9 might be adequate for general screening of hazardous wastes or waste extracts for genotoxicity. The urinary mutagenesis assay does not appear to be a useful adjunct to the Salmonella assay for screening hazardous wastes. The problems associated with chemically fractionating diverse types of hazardous wastes for bioassay are also discussed.     

Dose-dependent elimination of 5-fluoro-2'-deoxyuridine in the monkey

The kinetics of 5-fluoro-2'-deoxyuridine (FdUrd) and 5-fluorouracil (FUra) disposition after bolus intravenous injection were determined in anesthetized rhesus and cynomolgus monkeys. FdUrd disappearance from plasma was an apparent triexponential process with average half-lives of 0.5, 2, and 8 min; FUra disappearance was biphasic with average half-lives of 2 and 13 min. After FdUrd injection, FUra reached peak plasma concentrations of 15-30% of the initial FdUrd concentrations within 3 min, and then disappeared more slowly than FdUrd. Total FdUrd clearance fell from 105 to 73 to 56 ml/kg/min as the dose increased from 10 to 20 to 40 mg/kg. Metabolic clearance was about 85% of total clearance and fell similarly with increasing dosage. Total and metabolic FUra clearances were about 30% of FdUrd values at an equimolar dose. Renal FdUrd clearance exceeded glomerular filtration rate and was decreased by probenecid, indicating tubular secretion; renal FUra clearance was close to glomerular filtration rate. There was no apparent correlation between dose and renal clearance or volume of distribution. It was concluded that FdUrd, like FUra, is eliminated primarily by a dose-dependent process. The metabolic basis of the dose-dependent kinetics remains to be determined.     

Stress induces an increased hexose uptake in cultured cells

Temperature-sensitive mutants have revealed a region of the herpes simplex virus 1 genome that affects both the uptake of hexose and the synthesis of heat shock proteins. Other inducers of heat-shock proteins, namely heat shock itself and arsenite, likewise induce an increased uptake of hexose. The increased uptake, like that induced by insulin, is insensitive to the presence of actinomycin D or cycloheximide. It is concluded that an increased hexose uptake, reflecting an activation or relocation of existing hexose transport protein, is a general biochemical response of stressed cells.