Effect of fruit maturity,seed weight and storage time on the viability and germination of the seed of candelilla (<i>Euphorbia antisiphylitica</i> Zucc.)

Candelilla (Euphorbia antisiphylitica Zucc.) is a very important plant resource in the arid lands of Northern Mexico. This is because the wax content coating the stem has unique properties which have been useful for multiple applications in the food industry, electronics, cosmetics, etc. However, the intensive exploitation of this resource has caused a great decrease in the populations of this species making necessary to consider strategies for their conservation and sustainable use. One of the primary needs with regeneration purposes is to know their reproductive processes, particularly the biotic and/or abiotic factors that determine the viability and germination of seeds. The present study evaluated the (1) germination and seed viability in relation to the ripeness degree of the fruit at the time of collection, (2) weight of the seed (low, medium and high), and (3) storage time (1, 3, and 5 months). Fruits from four locations, two in the State of Coahuila (Las Coloradas and Candela) and two in the State of Nuevo Leon (Icamole 1 and Icamole 2), were collected. Three germination assays were carried out corresponding to each month of storage. Seed viability was determined by the tetrazolium test. The average weight of the candelilla seeds was 0.0029 ± 0.0010 g, with extreme average values of 0.0018 ± 0.0006 g at Las Coloradas and 0.0036 ± 0.0010 g in Icamole 2. Those seeds with heavier weight obtained from red fruits and with 1 month of storage showed the highest average percentage of viability (66.87 ± 24.19%). At the same time, seeds with around average weight, obtained from red fruits and five months of storage, showed the highest average germination percentage (50.00 ± 9.42%).     

Correlation between thyroid hormone status and hepatic hyperplasia and hypertrophy caused by the peroxisome proliferator-activated receptor alpha agonist Wy-14,643

BACKGROUND: The metabolic inhibitor rotenone inhibits hepatocellular proliferation and the incidence of liver cancer resulting from exposure to the PPARalpha agonist Wy-14,643, via unknown mechanisms. Since the absence of thyroid hormones diminishes hepatomegaly, an early biomarker for the hepatocarcinogenicity induced by PPARalpha agonists, this study was undertaken to investigate whether rotenone might interference with the ability of Wy-14,643 to alter the animal thyroid status. METHODS: Male B6C3F1 mice were given Wy-14,643 (100 ppm), rotenone (600 ppm) or a mixture of both, in the feed for 7 days. Bromodeoxyuridine (BrDU), marker of cell replication, was delivered through subcutaneously implanted osmotic mini-pumps. At the end of the experiment, sera were collected and corticosterone and thyroid hormone levels were measured by solid-phase radioimmunoassay kits. In addition, liver tissue samples were stained immunohistochemically for BrDU to determine percentages of labeled cells. Further, cell surface area was determined from images generated by a Zeiss Axioplan microscope equipped with a plan Neofluar x40 0.75 na objective. Tracings of individual hepatocyte perimeters were then analyzed and cell-surface areas were calculated using MicroMeasure FL-4000. RESULTS: Wy-14,643 caused a significant increase in liver weights, hepatocyte BrDU labeling index (LI), and hepatocyte surface area. In animals which received both Wy-14,643 and rotenone simultaneously, all of these effects were significantly less pronounced compared with mice that received Wy-14,643 alone. Rotenone alone decreased liver weights, LI and surface area. The Free Thyroid Index (FTI), which provides an accurate reflection of the animal's thyroid status, was 5.0 +/- 0.3 in control mice. In animals exposed to rotenone, these values decreased to 2.0 +/- 0.9, but in animals which received Wy-14,643, levels increased significantly to 7.7 +/- 0.9. FTI values decreased to 3.4 +/- 0.8 in mice receiving both rotenone and Wy-14,643. CONCLUSION: A strong correlation was observed between the animal thyroid status and both, hepatocyte proliferation (r2 = 0.62), and hepatocyte surface area (r2 = 0.83). These results support the hypothesis that the thyroid status of the animal plays a role in PPARalpha-induced hepatocellular proliferation and liver cell enlargement. Both these events are known to contribute to the expression of liver cancer in response to the activation of PPARalpha.     

A conserved mechanism of synaptogyrin localization

We have studied the localization of synaptogyrin family members in vivo. Both native and green fluorescent protein (GFP)-tagged Caenorhabditis elegans synaptogyrin (SNG-1) are expressed in neurons and synaptically localized. Deletion and mutational analysis with the use of GFP-tagged SNG-1 has defined a 38 amino acid sequence within the C terminus of SNG-1 and a single arginine in the cytoplasmic loop between transmembrane domain 2 and 3 that are required for SNG-1 localization. These domains may represent components of signals that target synaptogyrin for endocytosis from the plasma membrane and direct synaptogyrin to synaptic vesicles, respectively. In chimeric studies, these regions were sufficient to relocalize cellugyrin, a nonneuronal form of synaptogyrin, from nonsynaptic regions such as the sensory dendrites and the cell body to synaptic vesicles. Furthermore, GFP-tagged rat synaptogyrin is synaptically localized in neurons of C. elegans and in cultured hippocampal neurons. Similarly, the C-terminal domain of rat synaptogyrin is necessary for localization in hippocampal neurons. Our study suggests that the mechanisms for synaptogyrin localization are likely to be conserved from C. elegans to vertebrates.     

rpm-1, a conserved neuronal gene that regulates targeting and synaptogenesis in C. elegans

Little is known of mechanisms regulating presynaptic differentiation. We identified rpm-1 in a screen for mutants with defects in patterning of a presynaptic marker at certain interneuronal synapses. The predicted RPM-1 protein contains zinc binding, RCC1, and other conserved motifs. In rpm-1 mutants, mechanosensory neurons fail to accumulate tagged vesicles, retract synaptic branches, and ectopically extend axons. Some motor neurons branch and overgrow; others show altered synaptic organization. Expression of RPM-1 in the presynaptic mechanosensory neurons is sufficient to rescue phenotypes in these cells. Certain rpm-1 phenotypes are temperature sensitive, revealing that RPM-1 function can be bypassed by maintaining mutants at the permissive temperature at stages commensurate with synapse formation in wild-type animals. These results indicate that RPM-1 functions cell autonomously during synaptogenesis to regulate neuronal morphology.     

Short-term hypoxia/reoxygenation activates the angiogenic pathway in rat caudate putamen

In response to hypoxia, tissues have to implement numerous mechanisms to enhance oxygen delivery, including the activation of angiogenesis. This work investigates the angiogenic response of the hypoxic caudate putamen after several recovery times. Adult Wistar rats were submitted to acute hypoxia and analysed after 0 h, 24 h and 5 days of reoxygenation. Expression of hypoxia-inducible factor-1 alfa (HIF-1α) and angiogenesis-related genes including vascular endothelial growth factor (VEGF), adrenomedullin (ADM) and transforming growth factor-beta 1 (TGF-β1) was determined by both RT-PCR and ELISA. For vessel labelling, lectin location and expression were analysed using histochemical and image processing techniques (fractal dimension). Expression of Hif-1α, Vegf, Adm and Tgf-β1 mRNA rose immediately after hypoxia and this increase persisted in some cases after 5 days post-hypoxia. While VEGF and TGF-β1 protein levels increased parallel to mRNA expression, ADM remained unaltered. The quantification of the striatal vessel network showed a significant augmentation at 24 h of reoxygenation. These results reveal that not only short-term hypoxia, but also the subsequent reoxygenation period, up-regulate the angiogenic pathway in the rat caudate putamen as a neuroprotective mechanism to hypoxia that seeks to maintain a proper blood supply to the hypoxic tissue, thereby minimizing the adverse effects of oxygen deprivation.     

Assessment of lexical semantic judgment abilities in alcohol-dependent subjects: An fMRI study

Neuropsychological studies have shown that alcohol dependence is associated with neurocognitive deficits in tasks requiring memory, perceptual motor skills, abstraction and problem solving, whereas language skills are relatively spared in alcoholics despite structural abnormalities in the language-related brain regions. To investigate the preserved mechanisms of language processing in alcohol-dependents, functional brain imaging was undertaken in healthy controls (n=18) and alcohol-dependents (n=16) while completing a lexical semantic judgment task in a 3 T MR scanner. Behavioural data indicated that alcohol-dependents took more time than controls for performing the task but there was no significant difference in their response accuracy. fMRI data analysis revealed that while performing the task, the alcoholics showed enhanced activations in left supramarginal gyrus, precuneus bilaterally, left angular gyrus, and left middle temporal gyrus as compared to control subjects. The extensive activations observed in alcoholics as compared to controls suggest that alcoholics recruit additional brain areas to meet the behavioural demands for equivalent task performance. The results are consistent with previous fMRI studies suggesting compensatory mechanisms for the execution of task for showing an equivalent performance or decreased neural efficiency of relevant brain networks. However, on direct comparison of the two groups, the results did not survive correction for multiple comparisons; therefore, the present findings need further exploration.     

Slit1a inhibits retinal ganglion cell arborization and synaptogenesis via Robo2-dependent and -independent pathways

Upon arriving at their targets, developing axons cease pathfinding and begin instead to arborize and form synapses. To test whether CNS arborization and synaptogenesis are controlled by Slit-Robo signaling, we followed single retinal ganglion cell (RGC) arbors over time. ast (robo2) mutant and slit1a morphant arbors had more branch tips and greater arbor area and complexity compared to wild-type and concomitantly more presumptive presynaptic sites labeled with YFP-Rab3. Increased arborization in ast was phenocopied by dominant-negative Robo2 expressed in single RGCs and rescued by full-length Robo2, indicating that Robo2 acts cell-autonomously. Time-lapse imaging revealed that ast and slit1a morphant arbors stabilized earlier than wild-type, suggesting a role for Slit-Robo signaling in preventing arbor maturation. Genetic analysis showed that Slit1a acts both through Robo2 and Robo2-independent mechanisms. Unlike previous PNS studies showing that Slits promote branching, our results show that Slits inhibit arborization and synaptogenesis in the CNS.     

Distribution of MT1 Melatonin Receptor Promoter-Driven RFP Expression in the Brains of BAC C3H/HeN Transgenic Mice

The pineal hormone melatonin activates two G-protein coupled receptors (MT₁ and MT₂) to regulate in part biological functions. The MT₁ and MT₂ melatonin receptors are heterogeneously distributed in the mammalian brain including humans. In the mouse, only a few reports have assessed the expression of the MT₁ melatonin receptor expression using 2-iodomelatonin binding, in situ hybridization and/or polymerase chain reaction (PCR). Here, we described a transgenic mouse in which red fluorescence protein (RFP) is expressed under the control of the endogenous MT₁ promoter, by inserting RFP cDNA at the start codon of MTNR1a gene within a bacterial artificial chromosome (BAC) and expressing this construct as a transgene. The expression of RFP in the brain of this mouse was examined either directly under a fluorescent microscope or immunohistochemically using an antibody against RFP (RFP-MT₁). RFP-MT₁ expression was observed in many brain regions including the subcommissural organ, parts of the ependyma lining the lateral and third ventricles, the aqueduct, the hippocampus, the cerebellum, the pars tuberalis, the habenula and the habenula commissure. This RFP-MT₁ transgenic model provides a unique tool for studying the distribution of the MT₁ receptor in the brain of mice, its cell-specific expression and its function in vivo.     

Melanin in <Emphasis Type="Italic">Fonsecaea pedrosoi</Emphasis>: a trap for oxidative radicals

Background  

The pathogenic fungus Fonsecaea pedrosoi constitutively produces the pigment melanin, an important virulence factor in fungi. Melanin is incorporated in the cell wall structure and provides chemical and physical protection for the fungus.     

The Caenorhabditis elegans Kinesin-3 motor UNC-104/KIF1A is degraded upon loss of specific binding to cargo

UNC-104/KIF1A is a Kinesin-3 motor that transports synaptic vesicles from the cell body towards the synapse by binding to PI(4,5)P(2) through its PH domain. The fate of the motor upon reaching the synapse is not known. We found that wild-type UNC-104 is degraded at synaptic regions through the ubiquitin pathway and is not retrogradely transported back to the cell body. As a possible means to regulate the motor, we tested the effect of cargo binding on UNC-104 levels. The unc-104(e1265) allele carries a point mutation (D1497N) in the PI(4,5)P(2) binding pocket of the PH domain, resulting in greatly reduced preferential binding to PI(4,5)P(2)in vitro and presence of very few motors on pre-synaptic vesicles in vivo. unc-104(e1265) animals have poor locomotion irrespective of in vivo PI(4,5)P(2) levels due to reduced anterograde transport. Moreover, they show highly reduced levels of UNC-104 in vivo. To confirm that loss of cargo binding specificity reduces motor levels, we isolated two intragenic suppressors with compensatory mutations within the PH domain. These show partial restoration of in vitro preferential PI(4,5)P(2) binding and presence of more motors on pre-synaptic vesicles in vivo. These animals show improved locomotion dependent on in vivo PI(4,5)P(2) levels, increased anterograde transport, and partial restoration of UNC-104 protein levels in vivo. For further proof, we mutated a conserved residue in one suppressor background. The PH domain in this triple mutant lacked in vitro PI(4,5)P(2) binding specificity, and the animals again showed locomotory defects and reduced motor levels. All allelic variants show increased UNC-104 levels upon blocking the ubiquitin pathway. These data show that inability to bind cargo can target motors for degradation. In view of the observed degradation of the motor in synaptic regions, this further suggests that UNC-104 may get degraded at synapses upon release of cargo.