Choline acetyltransferase activity of spinal cord cell cultures increased by co-culture with muscle and by muscle-conditioned medium

Activity of the enzyme choline acetyltransferase (CAT), which mediates the synthesis of the neurotransmitter, acetylcholine, was increased up to 20- fold in spinal cord (SC) cells grown in culture with muscle cells for 2 wk. This increase was directly related to the duration of co-culture as well as to the cell density of both the SC and muscle involved and was not affected by the presence of the acetylcholine receptor blocking agent, α-bungarotoxin. Glutamic acid decarboxylase (GAD) activity was often markedly decreased in SC-muscle cultures while the activities of acetylcholinesterase and several other enzymes were little changed. Increased CAT activity was also observed when SC cultures were maintained in medium which had been conditioned by muscle cells or by undifferentiated cells from embryonic muscle. Muscle-conditioned medium (CM) did not affect the activities of SC cell GAD or acetylcholinesterase. Dilution or concentration of the CM directly affected its ability to increase SC CAT activity , as did the duration and timing of exposure of the SC cells to the CM. The medium could be conditioned by muscle cells in the presence or absence of serum, and remained effective after dialysis or heating to 58 degrees C. Membrane filtration data were consistent with the conclusion that the active material(s) in CM had a molecular weight in excess of 50,000 daltons. We conclude that large molecular weight material that is released by muscle cells is capable of producing a specific increase in CAT activity of SC cells.     

A stem–loop structure directs oskar mRNA to microtubule minus ends

mRNA transport coupled with translational control underlies the intracellular localization of many proteins in eukaryotic cells. This is exemplified in Drosophila, where oskar mRNA transport and translation at the posterior pole of the oocyte direct posterior patterning of the embryo. oskar localization is a multistep process. Within the oocyte, a spliced oskar localization element (SOLE) targets oskar mRNA for plus end-directed transport by kinesin-1 to the posterior pole. However, the signals mediating the initial minus end-directed, dynein-dependent transport of the mRNA from nurse cells into the oocyte have remained unknown. Here, we show that a 67-nt stem–loop in the oskar 3′ UTR promotes oskar mRNA delivery to the developing oocyte and that it shares functional features with the fs(1)K10 oocyte localization signal. Thus, two independent cis-acting signals, the oocyte entry signal (OES) and the SOLE, mediate sequential dynein- and kinesin-dependent phases of oskar mRNA transport during oogenesis. The OES also promotes apical localization of injected RNAs in blastoderm stage embryos, another dynein-mediated process. Similarly, when ectopically expressed in polarized cells of the follicular epithelium or salivary glands, reporter RNAs bearing the oskar OES are apically enriched, demonstrating that this element promotes mRNA localization independently of cell type. Our work sheds new light on how oskar mRNA is trafficked during oogenesis and the RNA features that mediate minus end-directed transport.     

Small molecule inhibitors reveal an indispensable scaffolding role of RIPK2 in NOD2 signaling

RIPK2 mediates inflammatory signaling by the bacteria‐sensing receptors NOD1 and NOD2. Kinase inhibitors targeting RIPK2 are a proposed strategy to ameliorate NOD‐mediated pathologies. Here, we reveal that RIPK2 kinase activity is dispensable for NOD2 inflammatory signaling and show that RIPK2 inhibitors function instead by antagonizing XIAP‐binding and XIAP‐mediated ubiquitination of RIPK2. We map the XIAP binding site on RIPK2 to the loop between β2 and β3 of the N‐lobe of the kinase, which is in close proximity to the ATP‐binding pocket. Through characterization of a new series of ATP pocket‐binding RIPK2 inhibitors, we identify the molecular features that determine their inhibition of both the RIPK2‐XIAP interaction, and of cellular and in vivoNOD2 signaling. Our study exemplifies how targeting of the ATP‐binding pocket in RIPK2 can be exploited to interfere with the RIPK2‐XIAP interaction for modulation of NOD signaling.     

Effect of long-term exposure to constant dim light on the circadian system of rats

The newly discovered multi-oscillatory nature of the mammalian circadian clock system and the cloning of the genes involved in the molecular mechanism that generates circadian rhythmicity have opened new approaches for understanding how mammals are temporally organized and how the mammalian circadian system reacts to the lack of normal synchronization cues. In the present study we investigated the effects of long-term exposure to constant red dim light on the pattern of the expression of Period 1 in the suprachiasmatic nuclei of the hypothalamus and of Arylalkylamine N-acetyltransferase(Aa-nat) in the retina and pineal gland. Our data demonstrate that Period 1 mRNA expression in the suprachiasmatic nuclei of the hypothalamus was not affected by exposure to constant red dim light for 60 days, whereas Aa-nat mRNA expression in the retina and in the pineal gland was significantly affected, since in some animals (20-30%) Aa-nat mRNA levels were found to be higher during the subjective day. A circadian rhythm of serum melatonin and locomotor activity was present in all the animals tested. In 4 animals serum melatonin levels were high during the subjective day. Our data suggest that long-term exposure to constant red dim light may induce desynchronization between the circadian rhythm of locomotor activity and serum melatonin levels.     

Long-term change in calcareous grassland vegetation and drivers over three time periods between 1970 and 2016

Plant Ecology - Analysis of long-term vegetation change is limited. Furthermore most studies evaluating change only examine two snapshots in time, which makes it difficult to define rates of change...     

Pten deletion in Dmp1-expressing cells does not rescue the osteopenic effects of Wnt/β-catenin suppression

Skeletal homeostasis is sensitive to perturbations in Wnt signaling. Beyond its role in the bone, Wnt is a major target for pharmaceutical inhibition in a wide range of diseases, most notably cancers. Numerous clinical trials for Wnt-based candidates are currently underway, and Wnt inhibitors will likely soon be approved for clinical use. Given the bone-suppressive effects accompanying Wnt inhibition, there is a need to expose alternate pathways/molecules that can be targeted to counter the deleterious effects of Wnt inhibition on bone properties. Activation of the Pi3k/Akt pathway via Pten deletion is one possible osteoanabolic pathway to exploit. We investigated whether the osteopenic effects of β-catenin deletion from bone cells could be rescued by Pten deletion in the same cells. Mice carrying floxed alleles for Pten and β-catenin were bred to Dmp1-Cre mice to delete Pten alone, β-catenin alone, or both genes from the late-stage osteoblast/osteocyte population. The mice were assessed for bone mass, density, strength, and formation parameters to evaluate the potential rescue effect of Pten deletion in Wnt-impaired mice. Pten deletion resulted in high bone mass and β-catenin deletion resulted in low bone mass. Compound mutants had bone properties similar to β-catenin mutant mice, or surprisingly in some assays, were further compromised beyond β-catenin mutants. Pten inhibition, or one of its downstream nodes, is unlikely to protect against the bone-wasting effects of Wnt/βcat inhibition. Other avenues for preserving bone mass in the presence of Wnt inhibition should be explored to alleviate the skeletal side effects of Wnt inhibitor-based therapies.     

Consequences of limited seed dispersal within simulated annual populations

Summary Theoretical analyses of the consequences of dispersal have been almost entirely limited to studies of colonization. A simulation model is constructed and analyzed to study the effects of dispersal within local plant populations. Rules for the growth and reproduction of individuals are deterministic and related only to competition with other plants. Rules for the dispersal of individual seeds are stochastic; the generalized seed shadow is negatively exponential. The analysis has several interesting results. 1) In small, dense populations of annuals, mortality and growth behavior are significantly affected by small-scale changes in dispersal. 2) Stable population size, at any life stage, is not environmentally fixed (e.g. by the number of safe sites) but is also a function of reproductive capacity and dispersal. 3) There is a minimum productive capability which must be exceeded for a population to be able to respond to increased dispersal. The responsive range of the parameters may be related to the grain of safe site distribution. 4) When the total number of seeds per generation is large, high dispersal decreases the number of germlings that are able to establish. 5) For plants that reach the growing stage, however, expected reproduction can exceed that of genotypes with greater reproductive capacity but less dispersal. 6) An uneven distribution of seeds is essential to a spatial patterning of mortality which can deter competitive extinction.Present address until August 1977: c/o United States Embassy, B.P. 817, Yaounde, Cameroun, West Africa     

Effects of solar radiation on the population dynamics of juvenile flatfish in the shallows of the Wadden Sea

During the first days after their arrival in the Wadden Sea, newly settled plaice Pteuronectes platessa L., flounder, Platichthys flesus L., and sole, Solea solea L., stay at low tide in the residual waters on the tidal flats. Here they can be exposed to high levels of solar radiation and extreme temperature conditions. As a result of the extreme temperatures, all settlers eventually leave the shallows and start to exhibit tidally-phased feeding migrations from the tidal channels on to the submerged fiats, In contrast to warm summers, for example 1982, no formation of a hyaline zone could be detected in the otoliths of surviving O-group plaice in 1990, 4 weeks after such a mass emigration, which could be directly observed on particular days in both hot and cold summers. Analysis of UV-B radiation data indicated that in 1990 cloud cover and wind conditions during the settling period protected the post-larvae from high temperature and radiation stress which is necessary to induce hyaline zones in the otoliths. Nevertheless, histological examination of the dorsal skin revealed cellular changes characteristic of sunburn damage in part of-the O-group plaice population in 1990.
Mortality of O-group plaice during and after such a mass emigration, which was directly observed in the drainage gulleys in 1982, was estimated at approximately 14–27%. This rate is assumed to be close to the maximum mortality which can be expected from abiotic factors for O-group plaice in the Wadden Sea.     

Processing of ampullary input in the brain: comparison of sensitivity and evoked responses among elasmobranch and siluriform fishes

1. Averaged evoked potentials (AEP) in response to pulses and sine waves of electric current, either as a homogeneous field or a large dipole field around the fish, have been recorded from the brain in 5 genera of elasmobranchs and 10 genera of siluriform fishes. 2. In the elasmobranchs, AEP's and spike-hash bursts are described from medulla, cerebellum, mesencephalon and telencephalon. A complex sequence of early and late waves is found in each level. Two sets of factors influence the latency and amplitude of the component waves. One is stimulus parameters including intensity, stimulus orientation, sign of the change of current, position of dipole, serial number in the first few cycles of a train as well as interval or frequency of a regular series. Separate component peaks of the complex AEP appear to have different dependencies upon these factors. To sinusoidal stimuli the best response by a certain criterion is at 20--30 Hz in a shark (Carcharhinus), 10--15 Hz in a ray (Potamotrygon); these are much higher than the values based on behavior from previous authors. The lowest threshold, with moderate averaging, was 0.015 muV/cm (= 0.8 nA/cm2) in marine species, less than 50 muV/cm (= 0.7 nA/cm2) in the freshwater ray. 3. The other set of factors determining AEP is brain parameters, including the recording locus, the type of electrode and electrode advance and the state of the brain (anesthesia, etc.). There is evidence of topographic segregation of some stimulus parameters, but mapping was not undertaken. Responsive regions are described in the several brain levels. 4. The best loci for electric AEP are distinct from those for acoustic and photic AEPs. No interaction between these was found. 5. All of the siluriforms tested show electroreception by the criterion of high sensitivity AEP. No obvious specialization was noted among the diverse species, but most were represented by a single experiment. Taken together with the previously known species, the sample of nine families of this large order suggests that this is a general characteristic of the order. 6. The best responses were found in the region of the torus semicircularis and in the lateral lobe. The lowest threshold by the criterion used was less than 0.15 mV/cm (= 2 nA/cm2), but some were nearly 2 mV/cm. Compared to much lower values in Ictalurus, this suggests species differences although part of the difference may be due to failure to record from the optimal locus. The AEP may be useful as a method of suveying for other electroreceptive groups. It is also potentially useful to reveal discriminable parameters of importance, such as dipole position, orientation, polarity and frequency.