Stoichiometric binding of diacylglycerol to the phorbol ester receptor

The major phorbol ester receptor is the Ca++-activated, phospholipid-dependent protein kinase C. Diacylglycerol stimulates protein kinase C in a fashion similar to the phorbol esters. Likewise, it inhibits phorbol ester binding competitively. Both results suggest that diacylglycerol is the/an endogenous phorbol ester analogue. Alternatively, the diacylglycerol might simply be acting to modify the phospholipid environment of the protein. If diacylglycerol were indeed functioning as an analogue, it should interact with the receptor stoichiometrically. This interaction can be quantitated by measuring the perturbation in apparent diacylglycerol binding affinity as a function of the ratio of diacylglycerol to receptor. We report here that 1,2-dioleoylglycerol interacts with the receptor with the predicted stoichiometry.     

Sperm aging in the male after sexual rest: Contribution to chromosome anomalies

The chromosome complement was studied in first-cleavage metaphases of mouse zygotes resulting from sperm aged in the male physiologically, after sexual rest. Females were inseminated by control males mating at 3-day intervals while experimentals mated to males that had had a sexual rest of 14 or more days. A total of 1954 eggs were collected 33–35 h post-HCG from 101 superovulated females mated to 42 controls and 43 experimental males. The fertilization rate was similar in both groups, being 84% and 85%, respectively. G-banded or Q-banded chromosomes were analyzed in 301 (68.3%) controls and 392 (49%) experimental first-cleavage metaphases. The overall rate of chromosome anomalies in controls was 4.45% as compared to 10.94% in experimentals, a highly significant difference. In the experimental group compared to controls, the frequency of trisomy, triploidy, structural rearrangements, and tetraploidy increased from 3.9% to 6.9%, 0% to 1.6%, 0.8% to 2.8%, and 0% to 1.3%, respectively. The genomic source of origin of the abnormalities was determined on the basis of differential condensation of the genomes. In the experimentals, grossly unbalanced sperm (diploids, disomics, double disomics, and those with large fragments) fertilized significantly more oocytes compared to controls. Our results implicate an advantage either in numbers or fertilizing capability for chromosomally abnormal sperm in a physiologically aged population.     

Glycinebetaine content of halophytes: Improved analysis by liquid chromatography and interpretations of results

The glycinebetaine content of plants can be determined by simple isocratic high performance liquid chromatography. The method is applicable to extracts from a wide range of species and, in most cases, is suitably rapid and specific to be preferable to other methods of analysis. The chromatographic system employed permits accurate and sensitive ultraviolet detection, free of most interferences. Because the principle plant carbohydrates elute well before glycine betaine, preparative ion exchange procedures can be simplified. Twenty-seven species, mostly inland halophytes, were screened by these methods and 13 were found to be glycinebetaine accumulators. On a dry weight basis, the glycinebetaine content of Salicornia europaea L. actually declined with exposure to progressively higher levels of NaCl. When expressed as a proportion of plant organic matter, however, patterns were more typical (up to 7.7% at higher salt concentrations).     

Germacranolides from Schkuhria anthemoidea

The isolation of eucannabinolide and three new sesquiterpene lactones from Schkuhria anthemoidea is reported. The structures and stereochemistries of the new compounds were established by chemical and spectroscopic means. The structure of santhemoidin B was confirmed by X-ray crystallography.     

Enrichment and characterization of clonogenic epithelial cells from adult rat liver and initiation of epithelial cell strains

Summary A highly efficient method is described for obtaining prolifertive epithelial cells from adult rat livers for the reproducible establishment of liver epithelial cell strains. When cells were isolated from livers of 10-to 15-wk-old male Fischer 344 rats by a collagenase-perfusion method, collected by centrifugation at 50×g for 5 min, and cultured in Williams' medium E containing fetal bovine serum and dexamethasone, colonies of epithelial cells different in size and morphology from hepatocytes were obtained. Sequential perfusion with collagenase and dispase yielded numerous epithelial cell colonies. When isolated cells were fractionated by differential centrifugation, the great majority of hepatocytes were sedimented at 50 ×g for 1 min, whereas many non-hepatocytic cells remiined in the supernatant and could be sedimented by a second centrifugation at 50×g for 5 min. Culture of the two fractions revealed that almost all the epithelial cell colonies were derived from cells in the non-hepatocytic cell fraction. The epithelial cells were cytochemically negative for γ-glutamyl transpeptidase activity, whereas an increase in the activity was detected in hepatocytes with duration in culture. Ultrastructural characteristics of hepatocytes were not found in the cells of newly established cell strains. These results suggest that adult rat liver epithelial cells propagable in culture were derived from a cell type other than the hepatocyte.     

The nutritional ecology of the ctenophore Bolinopsis vitrea: comparisons with Mnemiopsis mccradyi from the same region

Bolinopsis vitrea is a warm water lobate ctenophore which doesnot overlap in its distribution with Mnemiopsis mccradyi incontiguous waters. We examined its feeding ecology on a seriesof cruises. B. virrea ingested increasingly more prey at higherfood concentrations (2–100 prey l^–1) but feedingeffort (clearance rate) decreased with increasing food availability.On a dry weight basis, smaller tentaculate Bolinopsis ingestedseveral times more than larger lobates, but based on carbonweight, specific ingestion was fairly uniform over the entiresize range investigated (6–60 mm total length). Bolinopsiscollected during the daytime in the Bahamas rarely had morethan three prey items in their guts. These results and laboratorymeasurements of digestion times (av. = 1.9 h) allowed computationof daily rations, which could not account for the metabolicrequirement as measured on the same cruises. Results of feedingexperiments, however, implied that prey densities in excessof 11^–1 were sufficient to sustain a growing populationof Bolinopsis. Prey concentrations about an order of magnitudehigher were required for M. mccradyi based on similar experiments.These results were in general agreement with observed densitiesand distributions of ctenophores and their zooplankton preyin the Bahamas and coastal South Florida.     

Ethanol effects on voltage-dependent membrane conductances: comparative sensitivity of channel populations inAplysia neurons

The study of ethanol (EtOH) action is interesting because of its clinical relevance and for the insights it provides into structure-function relationships of excitable membranes. This paper describes the concentration dependencies of various parameters of four currents in Aplysia cells. ICa is the most sensitive of the currents studied. There was a significant reduction of ICa at concentrations of 50 mM EtOH. At low concentrations, the reduction of amplitude was the primary effect of ethanol, with the kinetics and voltage dependency of activation not affected. INa and IA were also affected, but at EtOH levels higher than those which altered ICa. The primary effect of EtOH on INa was a reduction in its amplitude, although the time to peak current flow was increased by EtOH. The effects of EtOH on IA were cell specific and, for the purposes of this paper, we examined the giant metacerebral cell (MCC). In MCC, the primary effect of EtOH on IA was an increase in the time course of inactivation. The time to peak IA was also increased by high concentrations of EtOH, but its amplitude was unaffected even at high concentrations. The delayed rectifier current, IK, was the most EtOH resistant of the currents examined. High EtOH concentrations augmented the amplitude of IK, although even at 600 mM concentrations, the percentage change was only 30%. Our results indicate that the calcium channel is very susceptible to the influence of ethanol and is a serious candidate to be the primary target of EtOH action in the nervous system. The differential sensitivity of voltage-dependent currents and individual components of a given current suggests further experiments to probe the relationship between membrane structure and channel function in excitable membranes.     

Androgen-induced alterations in vocalizations of femaleXenopus laevis: modifiability and constraints

Summary We examined effects of exogenous androgen (testosterone and dihydrotestosterone) on vocalizations of ovariectomized, adult female South African clawed frogs,Xenopus laevis. When paired with sexually active males, all ovariectomized females exhibited ticking, the unreceptive or release call. Ticking consists of low amplitude, regularly spaced clicks with a mean interclick interval of 154 ms. When androgen-treated and paired with sexually active males, these ovariectomized females also exhibited an aberrant call (atypical ticking) in which click multiples replaced the single clicks of ticking. Mean ICI's for atypical ticking were 37 ms for click doublets and 22 ms for click quadruplets. Androgen treatment decreased the total time spent vocalizing (typical and atypical ticking) by ovariectomized females.All androgen-treated females were then tested repeatedly with sexually receptive females in an attempt to elicit the male-typical vocalization, mate calling. Six of 17 females did not vocalize at all, even when gonadotropin injected. Eight females gave rapid (mean ICI, 36 ms) trains of clicks in an irregular temporal pattern (tick-like calls). Three females gave brief trills with alternating fast and slow components. Comparison of mate calllike vocalizations of androgen-treated females to mate calling of males reveals that calls in females are considerably shorter in duration (female: 0.32 min versus male: 45 min) and slower in tempo (ICI's; fast trill, female: 21 ms, male: 14 ms; slow trill, female: 36 ms, male: 28 ms). Incomplete masculinization of the vocal pattern of females by androgen treatment in adulthood may be due to developmental constraints on the modifiability of the neurons and muscles responsible for calling.Abbreviations C cholesterol - DHT dihydrotestosterone - HCG human chorionic gonadotropin - IBI interburst interval - ICI interclick interval - ovx ovariectomized - T testosterone     

Spontaneous binding ofYersinia enterocolitica to murine leukocytes

Twelve strains ofYersinia enterocolitica were examined for their ability to bind spontaneously to murine leukocytes. Each of eight HeLa cell invasive strains exhibited nonselective binding to peritoneal leukocytes, lymph node leukocytes, and thymocytes, whereas four noninvasive strains lacked binding properties. Like the HeLa cell invasion, the binding ofY. enterocolitica to leukocytes was much less efficient for bacteria grown at 37°C than for bacteria grown at 22°C. The binding properties were not influenced by the virulence plasmid that codes for Vwa⁺ phenotype. This leukocyte binding test is proposed as a simple assay for invasive properties ofY. enterocolitica.