text:
Cells in blood vessel walls express connexin (Cx)43, Cx40, and Cx37. We recently characterized gap junction channels in rat basilar artery smooth muscle cells and found features attributable not only to these three connexins but also to an unidentified connexin, including strong voltage dependence and single channel conductance of 30–40 pS. Here, we report data consistent with identification of Cx45. Immunofluorescence using anti-human Cx45 and anti-mouse Cx45 antibodies revealed labeling between α-actin-positive cells, and RT-PCR of mRNA from arteries after endothelial destruction yielded amplicons exhibiting 90–98% identity with mouse Cx45 and human Cx45. Dual-perforated patch clamping was performed after exposure to oligopeptides that interfere with docking of Cx43, Cx40, or Cx45. Cell pairs pretreated with blocking peptides for Cx43 and Cx40 exhibited strongly voltage-dependent transjunctional conductances [voltage at which voltage-dependent conductance declines by one-half (V 1/2) = ±18.9 mV] and small single channel conductances (31 pS), consistent with the presence of Cx45, whereas cell pairs pretreated with blocking peptide for Cx45 exhibit weaker voltage-dependent conductances (V 1/2= ±37.9 mV), consistent with block of Cx45. Our data suggest that Cx45 is transcribed, expressed, and forms functional gap junction channels in rat cerebral arterial smooth muscle.
- gap junctions
- vascular smooth muscle
- patch clamp
the syncytial organization of cells comprising the walls of blood vessels is critical for dynamic regulation of blood flow. Syncytial organization arises from gap junction channels that interconnect adjacent cells, allowing exchange of ions and small molecules and thereby permitting electrical and chemical communication between cells in the vessel wall (3). Individual connexins that form gap junction channels exhibit distinct physiological features, the most important being different pore properties, such as ion selectivity and conductance, and different gating properties, specifically voltage dependence. In electrically active tissue such as the blood vessel wall, voltage dependence is important in determining the dynamic aspects of communication between cells, with weak voltage dependence favoring relatively unhindered propagation of an electrical signal and strong voltage dependence tending to restrict propagation of signal.
The cerebral circulation is highly dynamic, with local blood flow being continuously adjusted to meet the demands of local neuronal activity. In cerebral vessels, as in other vessels, endothelial and smooth muscle cells are known to be interconnected by three connexins, connexin (Cx)43, Cx40, and Cx37, with the latter predominantly located in the endothelium (19, 20). The voltage dependences of Cx43 and Cx40 are relatively weak (2, 9, 23), however, making these connexins poorly suited to restricting the spread of electrical signals, as may be required for compartmentalized regulation of cerebral blood flow.
We (19) recently characterized gap junction channels in pairs of basilar artery smooth muscle cells using both electrophysiological and immunofluorescence methods. Although junctional conductances in many cell pairs showed weak voltage dependence, consistent with Cx43 and Cx40, a significant number of pairs (21 of 66 pairs, 32%) exhibited strongly voltage-dependent [voltage at which voltage-dependent conductance declines by one-half (V 1/2) < ±20 mV] junctional conductances that deactivated rapidly and nearly completely [minimum conductance (G min)/maximum conductance (G max) = 0.1–0.3 at 6 s, ±60 mV], findings that are not attributable to Cx43 or Cx40. These observations, coupled with the finding that ≈20% of single channel openings had a conductance of 30–40 pS, raised the possibility that basilar artery smooth muscle cells might also express a previously unidentified connexin.
A leading candidate for the unidentified connexin in basilar artery cells is Cx45, because this connexin forms small-conductance channels (30–40 pS) and exhibits strong voltage dependence (V1/2 ≈ ±15 mV) with little nondeactivating current (22). In the cardiovascular system, Cx45 has been identified preferentially in the ventricular conduction system of the heart (5). Recently, Cx45 mRNA was found in the A7r5 cell line derived from the embryonic rat aorta (17), and Cx45 mRNA and protein were identified in the mouse vasculature (14), but functional channels due to Cx45 have yet to be demonstrated in vascular smooth muscle, cultured or native.
In the present study, we sought to identify the low-conductance, strongly voltage-dependent channel in basilar artery cells. We used immunofluorescence to localize Cx45 protein in smooth muscle and RT-PCR to demonstrate Cx45 mRNA in cerebral vessels devoid of endothelium. In addition, we studied cell pairs using a dual-perforated patch-clamp technique after exposure to synthetic oligopeptides designed to interfere with docking of Cx43, Cx40, or Cx45. Blocking peptides homologous to parts of the second extracellular loops of rat Cx40 (amino acids 177–192) and Cx43 (amino acids 180–195) were previously reported to selectively inhibit Cx40 and Cx43 gap junction channels (16). By analogy, we constructed a peptide homologous to part of the second extracellular loop of rat Cx45 (amino acids 202–217) to selectively block this connexin. Here, we report that 1) Cx45 mRNA and protein are present in smooth muscle cells of cerebral arteries, with labeling for Cx45 being found predominantly at the origin of arteriolar segments; 2) smooth muscle cell pairs from vessels pretreated with blocking peptides for Cx43 and Cx40 exhibit mostly strongly voltage-dependent transjunctional conductances and small single channel conductances, consistent with the presence of Cx45; and 3) smooth muscle cell pairs from vessels pretreated with blocking peptide for Cx45 exhibit mostly weakly voltage-dependent transjunctional conductances, consistent with block of Cx45.
Smooth Muscle Cell Preparation
Basilar and posterior cerebral arteries were harvested from female Wistar rats (250–300 g), and cells including cell pairs were isolated by enzymatic digestion as previously described (19,26). Cells isolated in this manner were used for control (untreated) electrophysiological experiments as well as for immunolabeling of isolated cell pairs.
For electrophysiological experiments utilizing blocking peptides, arteries after dissection were incubated with synthetic peptides (500 μM) at 37°C for 3 h. The sequence for the short peptide homologous to the second extracellular loop of Cx40 (amino acids 177–192) was FLDTLHVCRRSPCPHP, that of Cx43 (amino acids 180–195) was SLSAVYTCKRDPCPHQ (16), and that of Cx45 (amino acids 202–217) was QVHPFYVCSRLPCPHK. A short peptide homologous to a cytoplasmic loop of Cx43 (amino acids 314–322), with a sequence of SAEQNRMGQ, was used as a control peptide. After pretreatment with various peptides, smooth muscle cells were isolated by enzymatic digestion as described.
Immunofluorescence Labeling
For immunolabeling freshly isolated cell pairs, cells were allowed to settle and attach to the surfaces of precleaned microscope slides for 30 min at room temperature in the extracellular solution used for patch clamping (see Electrophysiology). For these experiments, we also studied human hepatoma SKHep1 cells as positive controls and HeLa cells as negative controls. SKHep1 and HeLa cells, obtained from the American Type Culture Collection, were cultured in DMEM with 10% fetal bovine serum plus antibiotics with 5% CO2. Cells were studied when subconfluent. For the three preparations, cells were washed with PBS (pH 7.4) and fixed with 1:1 acetone-methanol for 2 min. After 30 min of permeabilization in 0.5% Triton X-100 and 20 min of block with 1% BSA, each cell preparation was incubated with one of two different polyclonal anti-Cx45 antibodies at room temperature for 1 h, either rabbit anti-mouse Cx45 (mCx45) antibodies (directed against the cytoplasmic domain, amino acids 259–396, kindly provided by Dr. T. H. Steinberg; dilution 1:1,000) or rabbit anti-human Cx45 (hCx45) antibodies [directed against amino acids 354–367 (5), Chemicon International; Temecula, CA; dilution 1:300]. This was followed by a wash and then a 45-min incubation period with Cy3-conjugated goat anti-rabbit IgG (Jackson Laboratories; West Grove, PA; dilution 1:400). Vascular smooth muscle cells were also double labeled with mouse anti-smooth muscle α-actin monoclonal antibody conjugated with FITC (Sigma; St. Louis, MO; dilution 1:1,000) for 45 min. After cells were washed, they were mounted using ProLong anti-fade mounting medium (Molecular Probe; Eugene, OR). Omission of primary antibody was used as a negative control. Immunolabeled cells were examined using a Nikon Eclipse E1000 microscope. Images were captured and processed using a SenSys digital camera (Photometrics; Tucson, AZ) and a personal computer with IP Lab software (version 3.01).
For immunolabeling of blood vessels, the brains of five rats were perfused with heparinized Krebs solution via a transcardiac route, and segments of cerebral vessels including basilar and posterior cerebral arteries were harvested, with care being taken to include attached penetrating arterioles originating from these vessels. For some experiments, vessel segments were embedded in optimum cutting tissue compound and frozen at −30°C, and longitudinal cryosections were freshly cut. For other experiments, intact arteriolar segments were placed directly onto clean microscope slides, to which they adhered spontaneously. Both types of specimens were fixed with methanol-acetone. After 0.25% trypsin treatment for 2 min and 0.5% Triton X-100 permeabilization for 30 min, immunolabeling with anti-mCx45 polyclonal antibody (dilution 1:1,000), followed by labeling with anti-smooth muscle α-actin monoclonal antibody conjugated with FITC (dilution 1:1,000), was carried out as described above. Nuclei were stained with 4,6-diamidino-2-phenylindole (1 μg/ml, Sigma). Specimens were examined using either a Zeiss LSM510 confocal microscope or a Nikon Eclipse E1000 microscope.
Twenty-one female Wistar rats (250–300 g) were anesthetized with pentobarbital sodium (200 mg/kg) and underwent transcardiac perfusion with 100 ml of Krebs solution plus heparin (1 U/ml) and papaverine (10 μg/ml), followed by Krebs solution with Triton X-100 (0.1%) (21) plus RNAse A (0.1 mg/ml) for 5 min to chemically remove the endothelium and degrade endothelial RNA. The RNAse activity was terminated by washing, after which the basilar and posterior cerebral arteries were rapidly dissected and placed in "RNAlater" (Ambion), a tissue storage and RNA stabilization solution. During dissection, great care was taken to exclude contamination with cerebral or other tissues. Arterial tissue (51 mg) was collected for RT-PCR. F9 mouse embryonic carcinoma cells, used as positive control, were obtained from the American Type Culture Collection and cultured in EMEM with 10% fetal bovine serum with 5% CO2.
Total RNA was extracted from the arterial homogenate and from the F9 cell lysate using a DNA-free RNA isolation kit (Ambion) with DNAse I application to eliminate DNA contamination, followed by reverse transcription with the oligo-dT primers included in the GeneAmp RNA PCR kit (Roche). For PCR, two pairs of primers specific for two fragments of mCx45 gene were chosen as follows: fragment 1, a 363-bp sequence beginning at position 153 from the ATG codon, sense primer 5′-TGTGTGCAACACAGAGCAGC-3′ and antisense primer 5′-CCATCCTCTCGAATTCGTCG-3′; and fragment 2, a 177-bp sequence starting at position 1,012 from the ATG codon, sense primer 5′-CTGCAGCGGGAGATCAGAATGGCTCAGGAA-3′ and antisense primer 5′-AATCCAGACGGAGGTCTTCCCATCCCCTGA-3′. The PCR experiment was conducted with a Perkin-Elmer 480 DNA thermal cycler (model 480, Perkin-Elmer; Foster City, CA). The PCR reactions included 30 cycles with three temperature steps: 95°C for 1 min, 60°C for 1 min, and 72°C for 1.5 min. The products of the amplification reaction were run on a 1% agorose gel in parallel with a 1-kb DNA Ladder (GIBCO Life Technologies). Amplicons isolated from the gels were sequenced (ABI 373 Stretch Sequencer, Perkin-Elmer). Labeling (BigDye kit) and running of sequencing gels were performed according to the manufacturer's protocols. We used the NCBI BLAST program (version 2.1.1;http://www.ncbi.nlm.nih.gov/BLAST/) to search the NCBI GenBank database for nucleotide sequences similar to those of our amplicons. The percent identity between sequences was determined by the program based on the number of nucleotide substitutions and the number of base pairs being compared.
With the use of phase-contrast microscopy, elongated phase-bright smooth muscle cell pairs were chosen for electrophysiological experiments. Dual-perforated patch clamp (19) was performed at room temperature with an extracellular solution containing (in mM) 145 NaCl, 5 KCl, 2 MgCl2, 10 HEPES, and 12.5 glucose; pH 7.4. Pipettes (0.8–1.1 mm, Kimax-51, Kimble; Toledo, OH) with tip resistances of 2–3 MΩ were filled with a solution containing (in mM) 130 CsCl, 8 MgCl2, and 10 HEPES and 165 μg/ml nystatin; pH 7.2. Gap junction current between cell pairs with seal resistance of >2 MΩ was measured using either two Axopatch 200A amplifiers (Axon Instruments) running pCLAMP 7 software or an EPC9/2 double patch-clamp amplifier running Pulse software (version 8.4, HEKA; Lambrech, Germany).
When cell pairs were weakly coupled (junctional conductance <5 nS), single channel events were readily identified (19), a situation found in 21 of 98 cell pairs. Single channel data were recorded on a digital tape recorder (SONY PCM-R300) and played back off-line, filtered at 100 Hz, and sampled at 1 kHz. Single channel conductances were measured manually.
Data Analysis
Values of macroscopic steady-state junctional conductance (G′j-ss) [steady-state (6 s) junctional current during the test pulse (I j-ss)/transjunctional voltage (V j)] were normalized to the instantaneous conductance [instantaneous junctional current during the test pulse (I j-i)/V j], yielding values of G′j-ss = (I j-ss/V j)/(I j-i/V j). The voltage dependence of G′j-ss was determined by fitting to the following Boltzmann function Equation 1 where G′max andG′min are the maximum and minimum normalized steady-state conductances, respectively, and k is the steepness of voltage dependence. Mean values ofG′j-ss from four groups of cell pairs (Fig. 7) were fit to the Boltzmann equation (Eq. 1 ) using a nonlinear least-squares method implemented in Origin 6.0 (Microcal; Northampton, MA). Histograms (Figs. 4 and 8) were fit to multiple-Gaussian functions using a nonlinear least-squares method in Origin 6.0.
Data are given as means ± SE. Statistical significance was assessed using Student's t-test or a one-way ANOVA with Dunn's method (data on junctional conductance and on conductance at +30 mV) for pairwise multiple comparison.
Immunofluorescence Microscopy
We first examined control cells for labeling by the two antibodies, the anti-mCx45 and anti-hCx45 antibodies. HeLa cells from human cervical carcimoma, which express no connexins (27), exhibited no labeling with either of the two anti-Cx45 antibodies (Fig.1, A and B). In contrast, positive labeling predominantly at cell-to-cell junctions was found with both antibodies in a fraction of SKHep1 cells (Fig. 1,C and D, arrows), which express Cx45 exclusively (17, 22). We then examined smooth muscle cells freshly isolated from basilar and posterior arteries, obtained using the same dissociation methods and studied under the same extracellular conditions as used for patch-clamp experiments. In 10–20% of smooth muscle cell pairs examined, positive labeling was observed between individual cells of pairs with both anti-mCx45 (Fig.1 E, arrow) and anti-hCx45 (Fig. 1 F, arrow) antibodies. A second label with anti-smooth muscle α-actin antibody confirmed that the elongated cells were smooth muscle cells (Fig. 1,G and H), which constituted >95% of the cell population obtained using our standard vascular smooth muscle cell isolation techniques.
We also studied intact vessel segments to evaluate the distribution of Cx45 in cerebral vessels. Immunolabeling of longitudinal sections and intact vessel segments for Cx45 revealed different patterns of expression in various portions of the vascular tree, from artery to arteriole, from a diameter of ≈100 to ≈20 μm. In the main trunks of the basilar and posterior cerebral arteries, positive Cx45 labeling was sparsely detected (data not shown). In contrast, in arterioles with diameters <50 μm, punctate positive Cx45 labeling was relatively abundant, with label appearing to be lined up between individual smooth muscle cells (Fig. 2). Occasionally, clusters of label were apparent near the origins of some arteriolar branches (Fig. 2 C).
To obtain evidence for the presence of Cx45 mRNA in cerebral vascular smooth muscle, we conducted RT-PCR experiments using carefully dissected cerebral arteries after chemical removal of the endothelium (21). Total RNA isolated from rat cerebral arteries, HeLa cells, and F9 cells was reverse transcribed with oligo-dT primers to obtain cDNA. Two fragments of cDNA from RT were amplified by PCR using the two pairs of primers, fragments 1 and 2(see methods), which are unique for mCx45 (1,10). Electrophoresis showed that both amplicons were of the predicted size, 363 and 177 bp for fragments 1 and2, in F9 cells and cerebral arteries, respectively (Fig.3, lanes 2, 3,5, and 6), but not in HeLa cells (Fig. 3,lanes 1 and 4). After the amplicons obtained from rat cerebral vessels were sequenced, search of the NCBI GenBank database revealed that the two nucleotide sequences exhibited significant alignment with published sequences. The sequences forfragments 1 and 2 showed 97% and 98% identity with the corresponding mCx45 sequences. Fragment 1 showed 90% identity with hCx45, and fragment 2 showed 90% and 89% identity with hCx45 and canine Cx45. Together, these data are consistent with Cx45 gene transcription in rat basilar and posterior cerebral artery smooth muscle.
Untreated cell pairs.
As previously detailed (19), macroscopic gap junction currents measured in freshly isolated untreated smooth muscle cell pairs from rat basilar and posterior cerebral arteries showed a variety of patterns of current deactivation (relaxation). In some pairs, the transjunctional current deactivated rapidly during 6-s test pulses (Fig. 4 A), whereas in most, it deactivated more slowly and incompletely (Fig.5 A). Plots ofG′j-ss at the end of 6-s test pulses vs.V j demonstrated a variety of voltage dependencies, with some pairs showing strong voltage dependence of deactivation, but most pairs showing weaker voltage dependence of deactivation (Fig. 6 A). As expected, rapid kinetics were associated with strong voltage dependence, and slow kinetics were associated with weak voltage dependence of theG′j-ss-V jrelationship. Of note, recordings with slow kinetics and weak voltage dependence are consistent with properties reported in single connexin expression systems for Cx43 and Cx40 (2, 9, 23), whereas records exhibiting rapid kinetics and strong voltage dependence, such as that shown in Fig. 4 A, are not attributable to these connexins.
In cell pairs that were weakly coupled, we also recorded single channel junctional events. As detailed previously (19), recordings from untreated pairs showed a variety of single channel conductances, including conductances of 80–120 pS typical of Cx43 and conductances of 150–200 pS typical of Cx40. Some records, however, exhibited predominantly a small conductance channel of ≈30 pS (Fig.4, B and C) not typical of either of these connexins.
Thus recordings of both macroscopic and single channel junctional currents revealed electrophysiological features in a subgroup of cell pairs that were inconsistent with the connexins expected to be present in these cells.
Peptide-treated cell pairs.
In an attempt to identify the connexin responsible for the rapidly deactivating, strongly voltage-dependent macroscopic conductance and the 30-pS single channel conductance, we studied the junctional current in smooth muscle cell pairs after exposure to blocking peptides, which reduce conductance due to specific connexins by interfering with connexon-to-connexon docking. We compared data from cell pairs isolated from 1) untreated control vessels; 2) vessels exposed to a control synthetic oligopeptide homologous to a cytoplasmic motif of Cx43, which should have no effect on connexon docking;3) vessels exposed to synthetic oligopeptides homologous to the extracellular loop motifs of Cx43 and Cx40, which are expected to prevent docking by these two connexins (16); and4) vessels exposed to a synthetic oligopeptide homologous to the extracellular loop motif of Cx45, which is expected to prevent docking by Cx45. Measurements of junctional conductance indicated that the four groups of cell pairs exhibited significantly different conductances (by ANOVA, P < 0.05), with values for the four groups being 13.1 ± 1.0 nS (28 pairs from 14 rats), 14.9 ± 2.5 nS (17 pairs from 8 rats), 8.7 ± 1.1 nS (14 pairs from 7 rats), and 9.5 ± 2.0 nS (10 pairs from 4 rats), respectively. Significantly reduced junctional conductances in the two groups with blocking peptides (P < 0.05), and not in the group with the control peptide, strongly suggested that the active peptides were acting as intended to block junctional conductance.
We examined the junctional current to determine whether the blocking peptides affected the currents in specific ways, especially with respect to voltage-dependent deactivation. We measured the voltage dependence of the steady-state (6-s) conductance, theG′j-ss-V jrelationship. The control peptide had no apparent effect, with the same variety and range of voltage dependencies (Fig. 6 B) as observed in untreated control pairs (Fig. 6 A). Overall, 10 of 26 and 4 of 13 cell pairs from the untreated group and from the control peptide group, respectively, exhibited sufficiently strong voltage dependence that one-half or more of the junctional current at 6 s was deactivated at +30 mV.
When vessels were exposed to the anti-Cx43 plus anti-Cx40 peptides before cell isolation, theG′j-ss-V jrelationship was very different from that observed with either of the two control groups. In the group with the anti-Cx43 plus anti-Cx40 peptides, the junctional current tended to be more rapidly deactivating (Fig. 5 B) and plots of theG′j-ss-V jrelationship showed stronger voltage dependence (Fig. 6 C). In this group, 12 of 13 cell pairs exhibited sufficiently strong voltage dependence that one-half or more of the junctional current at 6 s was deactivated at +30 mV. This finding is consistent with the interpretation that specific block of Cx43 and Cx40 allowed unmasking of activity of a native connexin channel that exhibits strong voltage-dependent deactivation.
In contrast, when vessels were exposed to the anti-Cx45 peptide before cell isolation, junctional currents tended to be more slowly deactivating (Fig. 5 C), and plots of theG′j-ss-V jrelationship showed weaker voltage dependence (Fig. 6 D). In this group, only 1 of 9 cell pairs exhibited sufficiently strong voltage dependence that one-half or more of the junctional current at 6 s was deactivated at +30 mV. This finding is consistent with the hypothesis that the rapidly deactivating current, revealed in the previous group with anti-Cx43 plus anti-Cx40 peptides and lost in this group with the anti-Cx45 peptide, was due to Cx45.
Mean G′j-ss-V jdata from the four groups of cell pairs are plotted in Fig.7. Fit of these mean data to the Boltzmann function confirmed that the four groups exhibited different voltage dependencies. The strongest voltage dependence was found with the anti-Cx43 plus anti-Cx40 peptides (Fig. 7, filled circles); the weakest voltage dependence was found with the anti-Cx45 peptide (Fig.7, filled squares); and intermediate voltage dependencies were found for the two control groups (Fig. 7, open circles and squares). Separate test of values at +30 mV showed significant differences for the cell pairs treated with blocking peptides (by ANOVA, P < 0.001).
We also examined single channel openings in weakly coupled pairs in which individual channel openings could be resolved. In control cell pairs, events of various conductances up to 300 pS were observed, including a prominent conductance at 30–40 pS (Fig.8 A). In contrast, in cell pairs from vessels treated with anti-Cx43 plus anti-Cx40 peptides, the relative number of openings of higher conductances was reduced compared with control, but the conductance at 30–40 pS appeared to remain unaffected (Fig. 8 B). Fit of the histogram from the treated cell pairs revealed that openings of 31 pS were dominant. Thus pretreatment of vessels before cell isolation with anti-Cx43 plus anti-Cx40 peptides unmasked not only rapidly deactivating macroscopic currents but also single channel events of 31 pS, consistent with the single channel conductance expected for Cx45 (22).
This is the first report to demonstrate expression of functional Cx45 gap junction channels in vascular smooth muscle cells. The impetus for the present work stemmed from previous electrophysiological experiments (19) on native basilar artery smooth muscle cell pairs in which we found macroscopic and single channel properties not expected for connexins known to be present in cerebral smooth muscle, i.e., Cx43, Cx40, and Cx37. Indeed, the single channel events histogram previously reported (Fig. 4 of Ref. 19) points to there being two anomalies that are inconsistent with the expected connexins: conductances >200 pS and an apparently excessive conductance at 30–40 pS. Although single channel openings of 30–40 pS could be due to a subconductance of Cx40 or Cx43, a subconductance should not produce a dominant peak in the event histogram. For Cx43, the subconductance state occupies only a very small proportion (<2%) of channel open time (4). Moreover, the strong voltage dependence of the steady-state macroscopic conductance exhibited by some native pairs was inconsistent with properties expected for the previously identified connexins (2,9, 23). Thus electrophysiological study of native cell pairs proved to be an important screening tool that alerted us to the presence of an unidentified connexin in cerebral arterial smooth muscle cells.
We used two antibodies to detect Cx45 protein expression. The first was a commercially available affinity-purified polyclonal antibody raised against a peptide homologous to amino acids 354–367 of hCx45. Polyclonal antibodies raised against a peptide with the same sequence have been thoroughly characterized by Coppen et al. (5). The second was a noncommercial polyclonal antibody raised against amino acids 259–396 of mCx45, which has also been thoroughly characterized (18, 28). We validated the activity of our antibodies by using them to label SKHep1 cells, which endogenously express only Cx45 (17, 22). Moreover, we confirmed the specificity of our antibody by showing no labeling in HeLa cells, which lack gap junction expression (27). Use of these two antibodies to directly label freshly isolated tissues gave assurance that Cx45 is indeed expressed in rat cerebral vascular smooth muscle cells. The pattern of expression that we obtained suggested that Cx45 was more likely to be expressed in arterioles rather than larger cerebral arteries.
We also identified transcription of Cx45 in rat basilar and posterior cerebral arteries (14, 17). Because there is no rat Cx45 nucleotide sequence available in the NCBI GenBank database, we chose two pairs of Cx45-specific primers based on the mCx45 nucleotide sequence (1, 10) to detect Cx45 mRNA in rat basilar and posterior cerebral arteries and in F9 cells, the mouse embryonic carcinoma cell line that we used as positive control. Search of the NCBI GenBank database revealed that the nucleotide sequences of the two amplicons that we obtained from rat cerebral artery showed high identity scores ranging between 90% and 98% compared with mouse, human, and canine Cx45. The possibility of contamination of our vascular tissue preparation with the brain, in which Cx45 transcription has been reported (15), was essentially eliminated by careful dissection that avoided breaching the pial covering of the brain. Overall, our data provide strong evidence that Cx45 mRNA is transcribed in rat cerebral arteries.
In tissues that express multiple connexins, individual gap junction plaques may be comprised of multiple connexins (29), making direct electrophysiological assessment for any individual connexin difficult. However, selective inhibition of gap junction channels using specific oligopeptides to interfere with docking (16) represents an important advance in the electrophysiological study of gap junction channels, especially in complex native tissues such as cerebral blood vessels in which multiple connexins are expressed. The extent of channel block by these peptides is expected to increase with incubation time, because blocking presumably depends on turnover and docking of new connexin hemichannels. Because freshly isolated smooth muscle cells lose their contractile phenotype during prolonged incubation, we incubated arterial tissues with peptides for only 3 h at 37°C compared with overnight, as used in the original report by Kwak and Jongsma (16). Three hours seemed satisfactory for Cx43, with its rapid turnover rate (half-life, 1.9 h), but may have been less favorable for Cx45, with its slower turnover rate (half-life, 2.9 h) (6). Although the length of incubation may have compromised achieving a full effect of the peptide, nevertheless, we were able to show that most smooth muscle cell pairs that had been exposed to anti-Cx43 plus anti-Cx40 peptides exhibited strong voltage dependence and more single channel openings of 31 pS, consistent with persistent function of Cx45 channels, and, conversely, that most smooth muscle cell pairs exposed to anti-Cx45 peptide showed weak voltage dependence, consistent with specific loss of functional Cx45 channels. Together, these experiments utilizing blocking peptides provide strong evidence that functional Cx45 channels are expressed in cerebral smooth muscle.
Total blood flow to the brain is relatively constant, but blood flow to different regions varies markedly according to neuronal activity (11). The dynamic changes in blood flow in response to focal brain activity are not only spatially restricted to the local microvessel near the site of increased neural activity but are also propagated upstream to resistance arterioles in a graded manner (7, 12). The response of the vessel wall to these complex highly dynamic requirements is coordinated in part by gap junctions in vascular smooth muscle and the endothelium (24, 25). Because tone of cerebral vascular smooth muscle is tightly regulated by membrane potential, compartmentation of vascular responses by gap junction channels would be facilitated by strong voltage dependence, as found with Cx45. Consistent with this hypothesis, our immunofluorescence data on vessel segments provide evidence that Cx45 is expressed predominantly at the level of arterioles rather than at the level of larger vessels. Moreover, compartmentation would also be facilitated by the fact that coexpression of Cx45 and Cx43, as may be found near arteriolar origins, tends to reduce overall coupling (13). Thus Cx45 channels may be uniquely positioned to play a role in the spatial restriction of blood flow changes in response to local neuronal activity.
In summary, we report functional Cx45 gap junction channels in native vascular tissue. The multiple methods used, including RT-PCR, labeling with two different antibodies with high specificity, and electrophysiology with specific blocking peptides, provide compelling evidence that Cx45 is expressed as a functional gap junction channel in rat cerebral arteries. Finding Cx45 in smooth muscle cells from these vessels adds to the known complexity of their syncytial organization, which up to now was attributed to only Cx43, Cx40, and C37, and enriches the possibility for formation of heterotypic and heteromeric channels with Cx43 and Cx40 (8), which may account for nonhomotypic channels found in these cells (19).
We thank Lioudmila Melnitchenko and Jia Bi Yang for assistance with cell and tissue preparation, Qiang Wang for assistance with RT-PCR, and Dr. T. H. Steinberg, Department of Medicine, Washington University, St. Louis, MO, for generously providing anti-mCx45 antibody.
This work was supported by National Institutes of Health Grants HL-51932 and NS-39956 (to J. M. Simard) and by a Bugher award from the American Heart Association.
Address for reprint requests and other correspondence: J. M. Simard, Dept. of Neurosurgery, Univ. of Maryland School of Medicine, 22 S. Greene St., Baltimore, MD 21201-1595 (E-mail:msimard{at}surgery1.umaryland.edu).
The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
- Copyright © 2001 the American Physiological Society

subset: commoncrawl


text:
Read More
Read More
Read More
Published: 2015-02-06 13:06:00
Updated: 2015-02-06 13:06:39
Posted February 6, 2015
Don't let significant, but nonetheless geographically isolated, events fool you. This winter has been very tame so far.
Not that the remainder of February couldn't change all that, but it will take a lot of work. From a temperature perspective, December was a joke. Virtually the entire country was "flooded" with mild air. January tried to make up for December but was only partially successful, and only in the eastern United States, looking at the two months combined, the "milds" win.
From a precipitation standpoint, a lot of the country was near normal. But there were notable exceptions, especially the West Coast. One month was very wet, and the next very dry. Of course. the snows in Boston have been truly historic, and there appear to be more coming. But Boston is a city, not a country.
Please sign in with your WRAL.com account to comment on this story. You also will need a Facebook account to comment.
View quoted thread
There is a difference between weather and climate change. Besides, the predictors for NC are much colder weather if the Gulf Stream shifts away from our coast.
Good to see the data behind the story. Numbers don't lie. Without a hard freeze, should we expect to see more bugs, of the bad variety, in the summer? What is the impact of a warm winter to the rest of the year?
Nationally, that's 'interesting'.
Greg, how about this part of NC?
From my perspective, we've had MANY more days where the temp is 10 degrees below normal (either the high or low), than we have 'above normal'. I know you have the DATA to correct my opinion. Please & thanks.
Thanks Greg
Love hearing your forecasts and summaries.
That does it. Fishel's obviously a shill for the radical left agenda on "climate change."
All I know is that if was pretty cold this morning!


text:
Thousands of runners will hit the pavement on Monday, April 19 to take part in the 114th Boston Marathon. Among them will be a number of Malden residents and others with a larger goal than just crossing the finish line of most prestigious annual marathon — raising money for local and national organizations.
Thousands of runners will hit the pavement on Monday, April 19 to take part in the 114th Boston Marathon. Among them will be a number of Malden residents and others with a larger goal than just crossing the finish line of most prestigious annual marathon — raising money for local and national organizations.
One such runner is Malden resident Stefanie Wojcik, who has been training for her third run in the Boston Marathon on behalf of the Dana Farber Marathon Challenge team. In addition to her training, she has been working to raise $15,000 for the Cancer Institute.
In 2009, Wojcik raised $10,000, and this year, she said, it was an easy decision to do it all over again.
In addition to raising money, Wojcik, as she did last year, is inviting the public to honor someone who has been afflicted with cancer by writing that person's name on a ribbon. Wojcik will pin the ribbons to her Dana-Farber singlet on race day on Monday.
In a fundraising letter, Wojcik said last year, she received an incredible response with this effort and had the privilege to honor over 200 people by carrying their names on her run to fight cancer.
For more information or to contribute to Wojcik's fundraising goal, visit her Web page at runDFMC.org/2010/stefaniew.
On behalf of Housing Families
Housing Families Board President Dave Barbato has also been training for the Boston Marathon on behalf of a charity. According to a press release from the organization, Barbato "is 'Going the Distance' for Housing Families Inc." by running on the HFI Marathon Team while raising much needed funds and spreading awareness of the great services HFI provides families in need.'
In addition, Housing Families has been awarded 15 bibs for this year's Boston Marathon. Only 24 nonprofit organizations are awarded these bibs each year, according to the release.
The HFI Marathon Team includes both local and out-of-state runners who have committed to help put an end to family homelessness by training many hours and miles while fundraising on the organizations behalf.
Along with Barbato, the HFI Marathon Team is made up of the following people: Lisa Nickerson, HFI Board member, Alexia Obar, Tommy Marincic, Roger Liu, Jill Deetjen, Jessica Steverman, Dustin Essex, Steven McManus, Kyle Raeburn, Jessica Ohly, Matthew Bystock, Alfonso Fulvio Abela, Paul Carvalho and Sanjiv Kumra.
To make a donation in support of HFI visit housingfamilies.org and click on the link on the bottom of the page, "Support a marathon runner."
Running for Citizen Schools
Sam Novey, also known as Burgerman, will be running the upcoming Boston Marathon in a custom-made b.good burger suit as a part of his movement to raise $100,000 for Citizen Schools
Citizen Schools serves Malden, Boston, Lowell, Springfield, New Bedford and Revere in Massachusetts, as well as 13 other cities across the nation.
Novey is a volunteer with Citizen Schools, which is a Boston-based nationwide afterschool program that works to extend the school day for middle schoolers in the most need of extra help. By adding approximately three hours of completely free, creative, structured educational time to these students' days, Citizen Schools not only bolsters students' academic abilities, but instills in them creativity and a thirst for learning — values important for success.
Novey has run the Boston Marathon twice previously and this year, is teaming up with local Boston burger joint b.good in launching his campaign to raise $100,000 for Citizen Schools. B.good, besides feeding and sponsoring Novey, has been hosting two-hour "training sessions" every Saturday, during which customers run with Burgerman in exchange for a free burger.
More information can be found at burgermanmarathon.com. Novey's YouTube challenge video can be found at youtube.com/watch?v=ZZRKfc9N6Dg.


text:
Hey there, time traveller!
This article was published 15/7/2010 (2593 days ago), so information in it may no longer be current.
Emily and Erik are shlumpy co-workers in the lost-and-found department of a transit company. In a basement office, they make their dead-end jobs bearable by spontaneously dressing up in "found" clothing and acting out funny little vignettes, like a couple of kids playing.
Seattle's Keira McDonald and Evan Mosher are terrific with accents — Southern U.S., Caribbean, British, Asian, Irish — and shine when they assume characters in these short flights of fancy. There's a lovely bit in which umbrellas become flowers and a moth.
But playwright Scot Augustson falls short of weaving the charming fantasies and the story of anti-social Emily and hangdog Erik — who may lose their jobs because of a "cat incident" — into a compelling whole.
The 50-minute piece is cute, but slight and pretty much forgettable, like a flimsy umbrella you leave behind on the bus.
— Alison Mayes
From the official Fringe Festival guide:
Meet Erik and Emily. They work at City Bus Lost and Found where umbrellas multiply like bunnies, hats fall in love with staplers and shoes run for mayor.
Warnings: Subject Matter, Language,
RECOMMENDED: General Audience
60 min
Onstage at the Playhouse (Venue 4), to July 23
Tickets: $10
Under 13 not admitted.


text:
This star rating is the opinion of the property owner only. A Self-Rated hotel has not been assessed by the AA or Visit Britain, whose official star ratings we use across LateRooms.com.
Hôtel Faubourg 216 - 2243 Self rated
224 Rue Faubourg Saint Denis, Paris, 75010
Price FromView Rooms
This hotel doesn't have any reviews yet.
About the hotel
The Hotel Faubourg 216 - 224 welcomes you to a comfortable and refined stay in its entirely renovated, contemporary and original setting: a calm atmosphere made of clear and soothing tones, where some bamboo brings an exotic touch. A very Parisian stay in the comfort of a refined setting. For tourism or business, the hotel location allows you to go without wasting time to places of business thanks to the proximity of transports, from Gare du Nord (Eurostar terminal) and Gare de l'Est.
In less than 15 minutes by underground with a station at the foot of the hotel, you can go to department stores, the Opera, Montmartre, the Eiffel Tower, the Louvre or the Latin Quarter. At the reception, multilingual staff will welcome you, answer your questions and facilitate your stay. In a lovely stone-vaulted room, a breakfast buffet is served from 7 am to 10 am. The hotel also offers you: Lifts Air-conditioned rooms Free Wi-Fi in rooms and reception Internet terminal at the reception Like in the rest of the hotel, completely renovated and air conditioned rooms offer their refined and elegant decor, sometimes punctuated with a bright note, and the comfort of their equipment for the milder nights. The marble bathrooms include a shower or bath, toilet and hair dryer.
Read MoreLess
We currently have no rooms available for this hotel that match what you're looking for. Try changing your search, selecting an alternative hotel, or give us a call on 0333 0146275 where we'll be happy to help.
Facilities Show/hide
- Main
- Disabled Facilities, Air conditioning in rooms, 24 Hour Reception, Safety Deposit Box, Hairdryer in each room, Telephone In Each Room, Wake Up Call Available, Lift to all floors
- Business
- Wi-Fi Available, Wi-Fi Available In Public Areas, Wi-Fi Available In Rooms
- In Room Entertainment
- TV, Satellite TV
- Leisure
- [Channels] News
Useful Info Show/hide
- From 15:00 hours
- By 12:00 hours
- Up to 8 years old
- The LateRooms.com rate for Hôtel Faubourg 216 - 224 in Paris is the total price of the room and not the 'per person' rate. All taxes are included, with the exception of city tax, which will be charged at 1.65(EUR) per person per night and is payable to the hotel directly.
- Pets are not allowed
Customer Reviews
Our reviews are from genuine LateRooms.com customers. We verify that the customer has stayed before leaving a review.
No reviews yet


text:
Added: Wednesday, February 02, 2011
Shemale Alexia has a preference for black cocks so this time she got herself a black hottie. Her black stud is so sexy, his muscles and his tight body made her so horny. Her cock got so hard at the sight of him and asked him to lick her jugs in this scene.


text:
Hey everyone I like to wish you all a Happy New Year 2014!! But, before this year comes to an end...I would like to say good bye to all of the great stars that passed away this year:
Paul Walker, you were great for your role as Brian O'Connor in "The Fast And The Furious" franchise and now that you're gone...The franchise will never be the same again. But you will live on in spirit for all of the great things you did for all the people around the world and you didn't deserve to die in that accident.
Lee Thompson Young, you were a young dude and you shouldn't have taken your own life like that because you gave myself and others a wonderful teenage memory for your role as Jet Jackson and your guest appearance as Cyborg on "Smallville". You would've been able to reprise the role of Cyborg in the future Justice League movie coming out in 2017.
James Gandolfini, you were one badass mob boss for your role as Tony Soprano on "The Sopranos". It was unfortunate for you to had a heart attack on your way to an award ceremony and you would have played a great villain in future Batman movies to come.
Marsha Wallace, you were well known for your role as Edna Krabappel on "The Simpsons". I watched the show since I was little and I stopped around early 2000. Although I hardly watched the show these days...I'll always remember you for that role. As for the rest of the stars that passed away this year...May you finally R.I.P and your legacy will live on for generations.


text:
Quick Look
Property brokered by
Get more listing details on brokerage site
Presented By:
Maggie Hosmer
(706) 273-4256
Email Agent
Maggie Hosmer
(706) 273-4256
Email Agent
Property DetailsThis stunning custom home offers a modern interpretation of Craftsman architecture. Gorgeous hm feat. a long private drive w/3 level of decks to enjoy the beautiful views. Main level feat open floor plan w/plank pine floors, custom stonework, lighting & chefs kitchen w/custom cabinetry & views to great rm w/27 ft stacked stone FP, sep dining rm, butlers pantry & screened in porch. Upper level w/loft, mstr suite w/stunning mstr bath, sitting rm & private balcony. Terrace level feat. in-law ste w/full kitchen, dining & living space, 2 BR w/sitting areas, 1 w/FP & 2FB.
Area Detailed Information
Area Weather
Area Cost Index
Area Property Prices
Additional Property InformationThis listing located at 116 Grouse Gap Drive, Big Canoe, GA has three bedrooms, three bathrooms and one partial bathroom. The property is presently selling at $549,900.
This property is presented by Maggie Hosmer of Harry Norman, Realtors® headquartered in Georgia.


text:
Worthy Christian News » World News » ISIS Leader, 400 Militants Enter Europe Disguised as Refugees
ISIS Leader, 400 Militants Enter Europe Disguised as Refugees
(Worthy News) - An Islamic State general, who has been designated by the United States as a global terrorist, is reportedly back in Europe from Syria along with about 400 of his soldiers, many of whom crossed the borders disguised as refugees.
The general, identified as Lavdrim Muhaxheri, also known as Abu Abdullah al Kosova, and his soldiers are among thousands who have fled Syria after the Islamic State, also known as IS, ISIS, ISIL or Daesh, suffered major losses in recent months, the U.K.'s Daily Mail reported, quoting sources in the Italian intelligence services, which have agents in Kosovo.
Muhaxheri, a Kosovo Albanian IS leader, who has been seen in a picture that showed him decapitating a prisoner, left for Syria in late 2012 and then appeared in IS propaganda videos, according to reports. [ Source ]


text:
I'm soon going to have my license and that means I'll be able to drive myself places and what not, yippee. The problem is, apparently you can't drive minors when you first get your license.
I'm thinking of just risking it. There's a girl that lives an hour away and I can't keep asking her to come to my area to hang out (Her dad can always drive her). Is that a really dumb idea?
I mean... maybe she could like hide in the trunk or something hahaha... that was a joke.


text:
Thank you for your interest in TMEA, the largest state music educators association in the nation.
Have you been a member in the last 4 years?
Renew your existing membership instead. Please do not create a new member record. If you are unsuccessful logging in to renew, follow the help on the Sign In page.
Credit card types accepted are Visa, MasterCard, American Express, and Discover.
Must pay by check (or purchase order)? Print a membership form. Please type into the form if your Internet browser supports it. Then print and sign.
Mail/fax deadline is December 31. Checks or purchase orders received after that date will be returned. Confirmation can take up to 10 business days.
About Membership
Membership Dues
Active: $50
Retired: $20
College Students: $25
Convention Online Early Registration Fees (Jan 19 Deadline)
Active members:$60
Retired members: $20
College student members: $0 (included with $25 college membership)
Out-of-state registrants: $110
Convention Online/On-site Registration Fees (After January 19)
Active members:$80
Retired members: $20
College student members: $0 (included with $25 college membership)
Out-of-state registrants: $130
Receipt and Membership Card
At any time, you can print your receipt and membership card.
If you receive an error on this site, please email Susan Daugherty with a detailed description of the error you are receiving. (Before dismissing the error message, if possible, press the "Print Screen (PrtScn)" keyboard button. Then open a MSWord file and choose Paste to paste the screen image on the page. Save that page and email it.)


text:
Now that the wraps are off the multiplayer mode in the upcoming Call of Duty: Ghosts , what am I most excited about? Squads, man. Squads. I can't wait to field a team of my own design because I'm sure they'll kick the butts of my friends' teams.
This is an entirely new element to Call of Duty, the top-selling first-person shooter on the market, that could work on an ingenious level. As it stands, the games' online multiplayer generally tests two things: how fast you are, and how well you can equip your individual character.
With the upcoming new Squads mode, you'll get to make 10 characters, advance and upgrade them, then field them in groups across different game types. Infinity Ward executive producer Mark Rubin says the artificial intelligence of these player-created zombies is going to be like nothing we've seen before. They'll work in teams and take their behaviour cues from the gear and abilities they're given. A sniper will, for example, act like a sniper. "They'll be so real, you'll end up yelling at them," he says.
I know, I know. Until we see it in action, it's hype. But it's enough to get me excited because it will, it if works as advertised, add the element of management to Call of Duty. Taking to the battlefield with a tailor-made crew of troops backing you up will be akin to being a player-coach on a sports team. It won't just be your reflexes that are tested, but also your ability to assemble a well-oiled machine.
Daniel Suarez, vice-president of production at Activision, uses the same sort of analogy to describe it. "You can create your dream team," he says.
I was disappointed, then, to learn that the new mode wasn't playable at Activision's multiplayer reveal event in Los Angeles on Wednesday. Fortunately, Ghosts' multiplayer has a host of other additional features, including 30 new weapons and seven new game modes, plus character customization, some of which we did get to try.
Being able to pimp out your character has long been one of the biggest requests from Call of Duty players, Suarez says, but fulfilling it hasn't exactly been easy to do. The series depends on balance to survive, so if some players discover something that gives them an unfair advantage, they can easily turn other players off the games.
That's why a lot of thinking and testing had to go into something as simple as clothing options. Would a certain jacket or helmet confer unforseen bonuses, like making a player harder to see? What about the opposite, that a certain pattern might make an opponent easier to spot?
The customizations that were accessible at the event were thus fairly limited. I had visions of eventually decking my squad out in matching purple and orange colours, but quickly figured out that was a bad idea. The slight variations of the camoflage actually on offer were probably more prudent in the long run. On the plus side, multiplayer characters can finally be female. The last bastion of Dudebros has fallen.
One of the new game modes is Cranked, which is pretty much what it sounds like – killing an opponent shoots your character full of crank, giving him a temporary speed burst and a new bonus perk. The downside is you have 30 seconds to get another kill, or you explode. It's manic and crazy. As a fellow player put it, "This is the anti-camper mode."
Also new are destructible environments, which can help or hinder your team. It was definitely a new Call of Duty experience to shoot away the pillars on a gas station roof and then watch it crush the enemies standing under it, even if that sort of thing has been done in other games.
We also got a taste of some of the new perks and killstreaks, including Takedown, an ability that lets you kill enemies without revealing their locations on the map, and Deadeye, which lets your bullets do more damage after you rack up successive kills. Killstreak-wise, there's Riley, a dog who follows you around and attacks opponents on your behalf, and Night Owl, a floating orb that detects enemies around you. There are plenty more, but those were unfortunately locked and invisible in our demos.
All told, there are going to be lots of new ways to play. Along with a main campaign storyline that casts the player in the unusual role – in a Call of Duty game, anyway – as the underdog, Ghosts has me more jazzed for the regular November release than the past few entries.
The game is being released on both current and next-generation consoles, with each having the exact same features. The main benefit with the Xbox One and PlayStation 4 versions will be better graphics and audio. Wednesday's demos were on the Xbox One, and it all looked and sounded super-smooth.
One of the additional, subtle additions for next-gen is something Infinity Ward is calling "contextual audio," where characters shout location-aware instructions in-game. "Tangoes out in the open," for example, alerts you to the fact that enemies have been spotted out in the open part of a map. I actually found the audio to be more useful and accurate than some of my human teammates' warnings and advice.
Still, it's the Squad mode I'm looking forward to, which is ironic because it's actually designed mainly to get new players into the game. The idea is to encourage one-on-one human battles, with the AI-controlled back-up teams, of course, which is potentially a better way of easing newbies into a multiplayer mode that can be impenetrable. Throwing noobs in with seasoned players usually results a quick and merciless massacre.
Activision has tried to ease the process for newcomers in the past, with varying degrees of success, says chief executive officer. Previous games gave players the option to fight completely against AI, or 'bots, but the developers think they might be on to something better with squads.
"This is a very clever way to have people play one on one rather than against 'bots," says Eric Hirshberg, President and Chief Executive Officer of Activision. "Something different happens when it's real people."Report Typo/Error
Follow @peternowakon Twitter:
- Activision Blizzard Inc$62.01+0.64(+1.04%)
- Updated August 18 4:00 PM EDT. Delayed by at least 15 minutes.


text:
LG G Pad 7.0 Specifications Review
LG G Pad 7.0 boasts a vivid 7" HD+ IPS screen with a resolution of 800 x 1280 pixels (216 ppi pixel density). It's a key to enjoy a great visual experience. Photos and graphics look amazing. They're sharp and have good details. The colors are also fascinating. It gives you more pleasure in watching HD videos. Zero Gap technology plays important role to display clear visuals. The tablet is not too large so you can still hold it comfotably with one hand. It has contoured edges that ensure better grip. The touchscreen is quite responsive and it supports multitouch and Accelerometer.
Under the hood, a powerful quad-core 1.2 GHz Cortex-A7 processor works together with Adreno 305 GPU, and 1 GB of RAM. Android v4.4.2 (KitKat) runs smoothly in this tablet. Apps can be launched quickly and there's no serious lag when switching between menus. It even has Dual Window multitasking capability that allows you to work with two compatible apps at once by dividing the 7" screen into two separate vertical windows. The G Pad provides 8 GB of internal memory to accommodate your files. If you need more space you can add external memory card by up to 64 GB. There are some connectivity options including Wi-Fi 802.11 a/b/g/n, dual-band, Wi-Fi hotspot, Wi-Fi Direct, DLNA; microUSB v2.0, USB On-the-go; Bluetooth; and Infrared port.
Other features that you can find in this tablet are: 3.15 MP rear camera and 1.3 MP front-facing camera, QPair for seamless Android smartphone interconnectivity, QuickRemote to turn the tablet into a universal remote so you can control your home entertainment systems on one screen, Compass, Email, Push Mail, Instant Messaging, HTML5, GPS, SNS integration, MP4/H.264/H.263 player, MP3/WAV/eAAC+ player, Photo/video viewer/editor, Organizer, Voice memo, Predictive text input, and more.
Dimensions: 189.3 x 113.8 x 10.1 mm (7.45 x 4.48 x 0.40 in)
Weight: 293 g (10.34 oz)
LG G Pad 7.0 Hard Reset Procedures
You can hard reset your LG G Pad 7.0 to restore the original factory settings. Please be warn if you perform a Hard Reset, all user applications, user data and DRM licenses will be deleted. Please remember to backup any important data before performing a Hard Reset. LG G Pad 7.0, do the following procedures:
- Turn the tablet off.
- Press and hold the Power/Lock Key + Volume Down Key on the right side of the tablet.
- Release only the Power/Lock Key when the LG logo is displayed and keep pressing the Volume Down Key .
- Immediately press and hold the Power/Lock Key + Volume Up Key at the same time while maintaining the Volume Down Key pressed.
- Release all keys when the Factory hard reset screen is displayed.
- Press the Power/Lock Key to continue or either of the Volume Keys to cancel.
- Press the Power/Lock Key once more to confirm or either of the Volume Keys to cancel.
- Your device will perform a hard reset.
Goodluck and have a nice day! :)


text:
Encontrar uma cópia na biblioteca
Encontrando bibliotecas que possuem este item...
|Tipo de Documento
|Todos os Autores / Contribuintes:
Randall McClure; James P Purdy
|9781573874755 1573874752
|Número OCLC:
|xv, 400 pages ; 24 cm.
|Introduction : understanding the NextGen researcher / Randall McClure and James P. Purdy --
Min(d)ing the gap : research on the information behaviors of NextGen students / Randall McClure --
The research paper project in the undergraduate writing course / Karen Kaiser Lee --
Professional statements and collaborations to support the new digital scholar / John Eliason and Kelly O'Brien Jenks --
Fighting for attention : making space for deep learning / Brian Ballentine --
Sentence-mining : uncovering the amount of reading and reading comprehension in college writers' researched writing / Sandra Jamieson and Rebecca Moore Howard --
Scholarliness as other : how students explain their research-writing behaviors / James P. Purdy --
Can I Google that? Research strategies of undergraduate students / Mary Lourdes Silva --
Encountering library databases : NextGen students' strategies for reconciling personal topics and academic scholarship / Ruth Mirtz --
Undergraduate research as collaborative knowledge work / Christa B. Teston and Brian J. McNely --
Re-envisioning research : alternative approaches to engaging NextGen students / Rachel A. Milloy --
Embracing a new world of research / David Bailey --
NextGen students and undergraduate ethnography : the challenges of studying communities born digital / Neil P. Baird --
Teaching researching in the digital age : an information literacy perspective on the new digital scholar / Barry M. Maid and Barbara J. D'Angelo --
Teaching and assessing research strategies in the digital age : collaboration is the key / Thomas Peele, Melissa Keith, and Sara Seely --
Understanding NextGen students' information search habits : a usability perspective / Patrick Corbett [and others] --
Remixing instruction in information literacy / Janice R. Walker and Kami Cox --
Conclusion: The new digital scholar and the production of new knowledge / James P. Purdy and Randall McClure.
|Título da Série:
|ASIST monograph series.
|edited by Randall McClure and James P. Purdy.
Opens a discussion long silent in academic circles -- that the teaching of research-writing is mired in practices poorly suited for digital natives. Reminding readers of the history of the academic research paper and the scope of the recent information explosion, this book presents innovative thinking and research on the educational challenges NextGen students face. Writing teachers, library science professionals, and higher education administrators describe teaching methods and research projects suited for the new digital scholar and designed for the information universe NextGen students inhabit.
Críticas contribuídas por usuários
Ítens Similares
Listas de usuários com este item (7)
- ALA Librarian's Library(183 ítens)
por kmuller atualizado 2017-03-02
- Latest Books on Research(9 ítens)
por anqotole atualizado 2015-03-21
- personality assessment(257 ítens)
por deryow atualizado 2014-11-09
- New Stuff @ Shatford Library!(240 ítens)
por shatfordlibrary atualizado 2014-04-10
- Feb 2014 Luria Library(136 ítens)
por skgentry atualizado 2014-02-13


text:
Episode 24
|Episode 24
Rate the episode
Episode information
NFL Cheerleader Jiggle Contests - 4th Graders Selling Weed
|Date of air
April 26th, 2014
3 Hours, 7 Minutes, and 36 Seconds
Episode 24 is an episode of Drunken Peasants. It features the sixth appearance of Galen Hallcyon.
- The rise of Galen Hallcyon.
- NFL Cheerleaders shake their tits.
- Elementary school kids smoking dope.
Material Used
- Hercules - Disappointed
- Bill O'Reilly Interviews Kevin Sorbo ("God Is Not Dead")
- Bill O'Reilly SCARED by Richard Dawkins
- Clippers owner racist (not found)
- Stowaway teen on plane (not found)
- Man Killed by Pope John Paul II Statue Collapse
- S.E. Cupp on healthcare (not found)
- Pastor's son claims he visited heaven
- 4th Graders Accused of Dealing Pot: 4th Grader Sells Pot In Northern Colorado
- Cheerleaders sue team, claim 'jiggle test'
- Hannity - Stoned America
Start of the Show
The show started out by playing Ben's favorite song My Time Is Now and made a theory about Denver the Last Dinosaur.
They played a clip of Kevin Sorbo being a terrible actor.
Next they played a video of Bill O' Reilly interviews Kevin Sorbo for his blockbuster masterpiece God's Not Dead. Kevin Sorbo warned us of the evil atheist in universities. Kevin also promoted his wife's shitty book.
They proved Bill O' Reilly's christian bias (he has one, who knew) by playing another interview with Richard Dawkins where he basically switched the burden of proof on the atheist and pull out the "X, Y and Z were atheist" argument.
Then. they played a news story of the Clippers team owner caught making racist remarks.
Next, there was a news story about some stowaway kid on a plane.
Another story was covered about some catholic statue crumbling and killing some man.
Next up, they played S.E. Cupp and how it's the worst time for Obamacare.
Lastly, they interviewed the father of the son who told the absolutely true story of how he went to heaven. His evidence was that he recognized a picture.
Middle of the Show
After that they played a rumor about kids selling pan without any evidence. Of course the parents were horrified and just want to their kids to live in a fantasy land.
They took a break.
Next the played a news story of cheerleaders being underpaid and forced to do some stupid jiggle test to see how much cleavage is moving. This is obviously the work of the patriarchy.
Next up they played some Fox News story on the dangers of marijuana and how it could kill you.
Then Sean Hannity brought a panel of conservative assholes ranting about how weed is evil. It was a clusterfuck of Fox News stupidity.
End of the Show
They took a break.
They answered some stupid boring questions for the last 3rd of the show and got Galen to make it even worse.
- To be added.
- To be added.


text:
June 30, 2017 Total Views: 7719 Today: 1 ★★★★★
Playa del Carmen hotels booking).
excelente atención de parte de los empleados del hotel. mucha limpieza en los restaurantes y barras de tragos. buena variedad de opciones a la hora de comer.
Four years ago we visited a week and it was excellent time for us. Fortunately we came again. First I was worried it had changed badly but soon I noticed it had no changed. They keep facilities clean and maintenance well. Staffs are always politeful and kind. I'm always surprised how they are good and hard workers. For example on Thursday visitors decreased but staffs never had rest but worked for quality, I felt. They were always kids friendly. Certainly we will come back again.
Con respecto a todo el personal desde el gerente David hasta el último asistente estamos sumamente satisfechos y agradecidos se empeñan en regalarte atenciones, servicio cordial y afectuoso. Muchas gracias a todos
The best thing going is the truly beautiful spot on the beach. However, right next to Senor Frogs, so all we heard was the music instead of the beach. Service was very good, food so-so, room was OK.
El. Lugar. Muy. Bonito El. Servicio la. Amabilidad. Convivir. Con. David. Director. Operativo. En. El. Desayuno. Y. El. Cocinando. Lo. Cual. Demuest. Humildad


text:
Home grown pasture is a key contributor to livestock feed in most farms. An accurate assessment of feed availability from the pasture at any particular point in time is an important requirement for feed planning and budgeting. Regular assessment of quantity of pasture allows early identification of forage surplus or deficit and this enables farmers to develop accurate feed budgets, plan feed requirements and make better overall grazing management decisions.
There are a number of forage assessment techniques available to farmers but the easiest is through the measurement of pasture height. There is a strong correlation between pasture height and pasture forage mass.
The simplest way to measure forage height is by use of a ruler. Placing the end of the ruler on the ground while holding the ruler vertically, estimate the average height of the upper leaves of the pasture. Record the pasture height from different points in the field. Calculate the average height of the pasture from 15 to 30 or more points depending on the size of the field.
The next step is to evaluate the pasture density. Do this in relative terms, whether the pasture is thin, average or thick. Using the average pasture height and estimated density look up the estimated forage mass on the calibrated table. Find the row corresponding to the average pasture height and the column corresponding to the estimated density. The number in that cell is the estimated forage mass in kilograms of dry matter per acre.
General calibrations for pasture forage density and forage mass at different mean heights as measured by a ruler
Name (required)
E-mail (required, but will not display)
Notify me of follow-up comments
Remember Me
We have 90 guests and no members online


text:
Leipzig Vacation Packages
Located in Saxony, Leipzig has a population of about 530,000. Historically Germany's premier trade center, it was originally a walled city and earned the nickname Little Paris in the 19th century for being a cultural hub. Leipzig is situated at the junction of the Weisse Elster, Parthe, and Pleisse Rivers.
The city's history is deeply connected to classical music. Both Bach and Mendelssohn lived in the city during the height of their careers. The Bach Museum also puts on the Bach Festival every year in early summer. You can also spend an evening at Oper, the city's opera house or enjoy live classical performances at Gewandhaus Orchestra, where Mendelssohn used to play.
Every Christmas, Leipzig hosts the Christmas Market. Leipzig also arranges the Wave-Gotik-Treffen, the world's largest goth festival, every summer. People travel from far away to experience its medieval market and music performances. Leipzig is packed with a wide variety of different cultural spaces, from the impressive Museum of Fine Arts to the Old City Hall, where one can learn about Leipzig's prolific history.
For visitors who like to explore the outdoors, Leipzig has a variety of parks to enjoy. The Rosenthal Park will lead you right to the famous Leipzig Zoo, which will delight children and adults alike. University of Leipzig also has Germany's oldest Botanical Garden and is open to the public. Don't miss out on Völkerschlachtdenkmal, a towering monument to the Battle of the Nations during the Napoleonic Wars.
While staying in Leipzig, many visitors opt to purchase the Leipzig Card. This pass provides discounted fares on all public transportation choices throughout the city. It also offers discounts at selected restaurants and event venues.
Leipzig is easily accessible by the A9, A38, and A14 motorways. The city also has the largest train station in Europe. The Leipzig/Halle Airport is conveniently located 15 minutes from the city proper, with a shuttle service running regularly between the two. Thanks to its central location and varied to-see list, Leipzig is a favored destination in Germany.
Accommodation in Leipzig
Before you head to Leipzig, get a reservation for one of the 74 Leipzig accommodation options that feature in Expedia's extensive travel database.
Starting out at the more upmarket end of the range, check out the latest Expedia deals at the Leipzig Marriott Hotel, at Am Hallischen Tor 1, or the Victor's Residenz-Hotel Leipzig, at Georgiring 13. In the same premium price bracket, why not have a look at the Westin Leipzig and the Seaside Park Hotel Leipzig. The Westin Leipzig is housed at Gerberstr. 15, while the Seaside Park Hotel Leipzig is found at Richard Wagner Strasse 7.


text:
Your Own Egypt
It looks like you have an ad blocker on. Please disable your ad blocker and refresh to fully enjoy our games.
Thank you.
Although many of the Egyptian Wonders of the World have stood the test of time, many of the remaining sculptures and statues have withstood a millennia of wind erosion, but with your decorating skill, they can still look incredible!
Use your mouse to select the kind of decorations for the desert, and then use your mouse to drag them around the scene.


text:
Northfield Village -- Mayor Jesse Nehez is looking for three people to join the Northfield Village Recreation Board.
There are currently two members on the five-member board, former Councilors Bea Greenlee and Linda Bowen. Councilor Renell Noack serves as Council's liason to the board.
Nehez said the board has in the past been essential to planning and implementing events such as movies in the park.
"We had five members at one time, but one by one they all fizzled out for different reasons including just not having time to do it," Nehez said at village Council's Sept. 14 meeting.
The village is accepting applications and letters of interest until Oct. 1. Nehez said qualified candidates must be a registered voter and have lived in the village for at least one year.
The rec board meets the first Thursday of each month at 7:30 p.m. at the village hall.
Letters of interest can be sent to firstname.lastname@example.com or dropped off at village hall.
Nehez said if not enough residents are interested in the rec board, he would like to explore changing the village's charter to have Council members serve as rec board members.
Coventry Road
Paving approved
After some discussion as to whether the village can afford to add the paving job to the budget for this year, Council voted 4-2 in favor of paving Coventry Road before the snow flies.
Paving was scheduled for Sept. 27, weather permitting, but Service Director Jason Walters said that based on the weather forecast at press time, the project would likely happen Sept. 28.
He expects paving to be completed in the morning and residents to have full access to the road by the evening hours the same day.
Finance Director Tricia Ingrassia said the cost for contractor Mid-Ohio Asphalt to do the work is $18,800.
Councilors Jim Daugherty and Gary Vojtush voted against the project.
Daugherty said he wanted more discussion before proceeding.
"I think we need to go over a couple things before we spend too much money," he said.
Ingrassia said she was "comfortable" the village could handle the expenditure.
Briana Barker: 330-541-9432


text:
8 weeks (1x a week) OR 4 Day Intensives
Newburyport, Ma | Harvard Square, Ma | Portsmouth, NH (coming soon)
Let's be clear. Pain IS a symptom, NOT a diagnosis.
The reality is that some pain, both emotional and structural, is inevitable in life, but chronic pain can completely exhaust your body's resources. It is your birthright to live pain free, and it is possible.
The Rasamaya Method Live Pain Free program is a form of holistic pain management that merges together the latest neuroscience and mind-body research with the wisdom of postural therapy and Carrie's expertise as a nurse in orthopaedics and primary care, and over twenty years experience as a postural therapist, personal trainer, anatomy teacher and yoga, Pilates and barre teacher trainer. There are 2 phases through which clients progress. Some clients may also choose to add on STRETCHING THERAPY sessions.
If you commit to the program wholeheartedly, you will not only learn to sit with pain when it does arise, but you will also gain an artillery of tools to fight back with. These tools will allow you to move beyond the pain, and rid yourself of it so you can get back to the active, beautiful life you love.
There are two convenient formats for clients to progress through the initial phase of the program:
Option A: Weekly session over 8-weeks for approximately an hour per session
Option B: 4-day intensives may be more appropriate for individuals from out of town. Each day will consist of a 2hr session with the expectation that you will set aside some time for personal reflection and light homework (an hour per day is fine). A long time Seacoast area resident, Carrie is happy to make recommendations for local accommodations, fantastic food, and things to do to help you get the most out of your stay.
All clients are required to have an initial consultation. This session is dedicated to analyzing your postural and gait, functional testing, goal setting, lifestyle habits, mindfulness techniques and "your brain on pain" education. You will leave this meeting with a better understanding of your structural compensations that could be contributing to your pain. You will also receive a handout on some easy corrective exercises and mindfulness work to get you started on your path to Living Pain Free.
This has to be the right fit for everyone involved, and Carrie only takes clients who are motivated to create positive change in their bodies. As such, both you and she have 24hrs from your first appointment to decide if you are a good candidate for the Live Pain Free Program. Read more about what to expect from your initial consultation here.
By your second session Carrie will have created a more thorough menu of easy therapeutic exercises and techniques specifically designed to address your pain and structural compensations. These will be taught to you in your second session and you will be given more videos, handouts, and education so that you can complete the menu at home. Subsequent sessions will be a fine tuning process until we reach our goal of getting you pain free.
Most clients decide to progress to Phase 2 which incorporates small group or private sessions that are focused on safely integrating dynamic movement. Sessions use various resistance training, spring-loaded Pilates apparatus, such as the Reformer, Chair, and Barrel. This intelligent system strengthens and lengthens the entire body in every position and range of motion imaginable. Additionally, the Apparatus offers many different attachments, which allow us to completely customize your workout and create safe modifications to help you find your best, most vibrant body. Benefits include increased endurance, strength and visible results. After a couple sessions and you will be hooked for life. Even just one session a week can make a huge difference in how you move through the world.
In addition to being a nurse, Carrie is also a personal trainer and certified in Thai Bodywork (a form of passive -assisted stretching). She draws from her 20 years experience as a movement educator and trainer to create personalized stretching sessions for her clients. These can be an excellent addition to your Live Pain Free FOUNDATION or ALIGN sessions.


text:
Can I use a PC mouse on a Mac?
I have two mice, one is from hp and the other is from Microsoft. I was wondering if these mice would be discoverable by my Mac (I own a Powerbook G4, a dinosaur but it still works).
- Asked by Emilie S from Redlands
1 Answer from the Community
Yes, a two button/scroll PC mouse works perfectly on a Mac. Plug them into your USB port and your Mac should recognize them. I've used one for many years and have never had a problem.
Of course the swipe/mulitple finger features are not there but i don't really need them.
Bob S.
- Answered by Robert S from Albuquerque
- Dec 3, 2014


text:
Poppy Fields - Landscape Photography Workshop
Jun 18th - 7:00pm - 9:30pm
This is one of the best flowering meadow scenes you will witness. Rolling hills full of poppies takes some beating. Join me for one of the poppy field photography location shoots from mid to end of June.
Equipment for this workshop
Recommended but it's not all essential:
- Tripod ( I have some for hire if you don't own one yet)
- No specialist lenses required, we can make use of whatever you have
- Remote cable release, useful but not essential
- A polariser filter is useful but not essential
- Neutral Density and Graduated ND filters are useful but not essential
In order for a photographic workshop to go ahead, there must be a minimum of three students booked for that particular date. Should there be insufficient interest, alternative dates will be offered.
On the rare occasion that severe weather conditions prevent any of the photography workshops from safely or practically going ahead, other dates will be offered.
If you have any questions about the workshop or equipment please don't hesitate to contact me. email@example.com or 0781-701-7994
- No prior knowledge of photography is required as each student will be taught individually, according to their level.
- Recognise correct lighting, contrast and texture to shape and form each shot.
- Learn the impact of using long exposure, shallow depth of field and other techniques.
- Work with Alan to review your shots, accept new ideas and improve your photography.


text:
We ship to over 100 countries, including:
Please select the language that you would like to browse the site in.
Please select the currency you would like to shop in. Your order will be transacted in USD.
Size: Select a Size
Oops! We encountered an issue trying to perform this action. Please try again.
Thanks! We've received your email address and will send you a note when this item is back in stock.
Please login or create account before adding to wishlist
Style: 25035635 Color Code:
Black plastic retro-style sunglasses with tinted lenses. Metal wire detailing across front top. Plastic, MetalImport*5 1/2" width across front
New World Sunglasses styled by the FP Me community
Hey you!
Be the first to write a review.


text:
UX Design: The Art of Writing Annotations
Michael Lemma, Designer & Photographer
- 0.5x
- 1x (Normal)
- 1.25x
- 1.5x
- 2x
Annotations Intro6:25
The Terminology8:03
The Art of Writing Annotations7:25
Creative Annotations14:18
Dressing up your Annotations2:47
About This Class
If you're looking to get into UX design, there will come a point in the UX process to write annotation in your documentation, so others can read and understand how your design will function. This short class will explain
- The Terminology
- Setting up Annotation in your Document
- Writing Annotations
- Dressing up the Document
Note: If you'd like to use a design then go ahead a take my Wireframe class (if it's not available it will be soon) so you can use that design to annotate. If not, I have attached a document for you to annotate.
Class Projects See All
No projects have
been added yet
No projects have
been added yet
No projects have
been added yet
No projects have
been added yet
Mike is a successful designer. He worked for a hot startup in the Techn Industry. He grew up in California and received his BFA in Graphic Design from the Savannah College of Art & Design. Also has recieved his minor in Architecture. He has studied art and design since he was seven.
He's had clients that have ranged from NY Red Bulls, The Amazing Spider-man, Coors Light, AMC's Breaking Bad, Powerade, Verizon, and more.
His specialty is UI/UX, Branding, Corporate Identity, Poster Design, Logo Design, Package Design, Web Design, Concept Design, Matte Painting, Photography and Art Direction. He is the founder of the website exhbit.
He has gained inspiration while traveling the world and recently wrote and published a book on his travels. He's had articles written about his photography on 500px blog ISO as well as having a healthy portfolio of work.
I've been in the design field for almost 7 years. I started out by designing for graphic services (Print design, logo, web design etc), but I discovered other ways I could earn a living through the art of design & Photography.
I started learning Photoshop when I was only 12. I kept up to date with it over time and became better at it. Then, I went to college and learned a lot more from a design stand point (Layouts, how to engage the users, color theory, and more).
Eventually, I started freelancing, and I slowly realized this can be a profitable part time cash machine as well as making myself a better designer. I tested and tried the best ways of achieving certain Photoshop techniques. I'd start by mocking up 10 designs per day. Then I'd go back and constantly push and update the designs. It was a lot of work that went into the early mornings (the life of a designer), but I loved it. I was always thinking of new ways to design things. By doing this I became a better designer as you would normally in everything else, practice makes perfect!
I'm a efficient designing master and I know the best ways of creating stunning designs in the blink of an eye (especially web design). I choose how much money I want to make by managing my time and projects I want to work with.
I am also an entrepreneur: I started my first start up 2 years ago and it quickly became popular. I freelance constantly as well as working a full time job (Where does the time go!?).
Having so little time and so much to do, I still managed to make over six figured by working part time in Photoshop/Sketch (this doesn't not include in my full time salary, which is another six figures). You can learn from my personal experiences and start earning a living.


text:
4 Min Read
* Minister says France welcome but some political positions still rankle
* Turkey cut ties over French bill on 1915 genocide
* Turkey says will keep taking in some Iran crude, payments legal (Adds Yildiz comments on resumption of French ties, settlement of payments for Iranian crude oil imports)
By Melissa Akin
ST PETERSBURG, June 22 (Reuters) - Turkey's resumption of diplomatic ties with France means French companies may regain some, if not full access to Turkey's energy sector, including its nuclear industry, Turkey's Energy Minister Taner Yildiz said on Friday.
After a meeting between Turkish Prime Minister Tayyip Erdogan and new French President Francois Hollande, Ankara announced on Thursday it would restore ties with France after a six-month hiatus in a dispute over the 1915 mass killing of Armenians by Ottoman Turks.
Ankara cancelled all economic, political and military meetings with Paris in December after France's lower house of parliament voted overwhelmingly in favour of a draft law to make it illegal to deny that the killings amounted to genocide.
France's highest court overturned the law two months later but the Turkish measures, which included restrictions on French military aircraft and ships landing or docking on its territory, have remained in place.
Yildiz said the government expected an improvement in relations with Hollande in power.
"I believe that this burden will be taken out or removed after the meeting (Hollande) had with his excellency Erdogan," Yildiz told reporters during an economic forum in St. Petersburg, Russia.
"As the energy sector we are ready for a variety of cooperation with France, although our projects and our business is really large and we can not separate them from the international politics," he said.
"Therefore we cannot ignore some political approaches that will affect our industry. I believe that from now on the prospects will be much better compared to the time of (Hollande's predecessor Nicolas) Sarkozy."
Armenia, backed by many historians and parliaments, says about 1.5 million Christian Armenians were killed in what is now eastern Turkey during World War One in a deliberate policy of genocide ordered by the Ottoman government.
Turkey says there was a heavy loss of life on both sides during the fighting in which Armenian partisans supported invading Russian forces.
Ties weakened during Sarkozy's term in office, when the former French president was also an outspoken opponent of Turkish membership in the European Union.
Noting Turkey's relationship with Iran was "not like the relationship of any European countries with Iran," Yildiz said Turkey would continue to import some Iranian crude oil after its sole refiner cut imports of Iranian crude by 20 percent.
The EU itself has largely banned intake of Iranian crude from July 1. Iran's oil buyers around the world have been cutting imports to avoid U.S. financial sanctions which aim to stop Iran's nuclear programme through effective limits on dollar transactions.
Washington is pressing Turkey to cut Iranian supplies over the next six months or face sanctions, but the 20 percent cut has earned Ankara a 180-day exemption from financial sanctions, during which Halkbank can make payments for imports.
"The oil trade being done with Iran right now is not illegal. So trade operations are being implemented within all of those laws and regulations," Yildiz, said, adding it was able to pay for oil in Turkish lira.
He said the lost Iranian volumes would be replaced by Libyan and Saudi Arabian crude, which have been in greater supply through increased use of Saudi spare capacity and the recovery of the Libyan energy industry from a violent uprising which ousted Muammar Gaddafi. (Editing by James Jukwey)


text:
- "I love that Rank and Style simplifies shopping through unbiased top tens.
As the new CEO, I am committed to continuing the mission of ranking the world while building a site that allows our users to efficiently and confidently shop the best." -Jamie Chandlee
Be in the know:
Top tens in your inbox.
Thank you for subscribing!
Enter your email address below and live list-fully ever after!
Most Popular Top Ten Lists:
Top Ten Lists You Might Like:
Featured Content
- Talking Top Tens: Tze ChunTze Chun is the Founder of Uprise Art, an online gallery that makes collecting original art easier and more affordable. Their curated collection of one-of-a-kind art takes the guess work out of art collecting, and Uprise Art's team of art advisors help you find the perfect art for the spaces where you live and work. Follow Uprise Art on Instagram for a behind the scenes look at the art world!
- Talking Top Tens: Tamar YanivTamar Yaniv is the CEO & Co-founder of Preen.Me, a beauty community helping women buy beauty better. A digital executive who has led teams all over the world, Tamar combined her deep digital experience and her complete and utter love for all things beauty when she created Preen.Me. Having grown up in NY and lived in London, Tamar currently lives a 10 minute walk from the beach in Tel Aviv, Israel, with her husband and extremely energetic toddler. Follow Preen.Me on Twitter.
- Popular This Week:Calvin Klein Ultra Bare Infinite Sheer Control Top Pantyhose


text:
Quotations, sayings and aphorisms about food & beverages, eating & drinking and pleasures of the table
Home | Food Articles | Food Trivia | Today in Food History | Recipes | Cooking Tips
Food Videos | Food Quotes | Who's Who | Trivia Quizzes | Crosswords | Food Poems
Recipe Contests | Free Magazines | Culinary Schools | Gourmet Tours | Food Festivals
You are here > Home >
From Amateur & Basic Cooking Classes to Professional Chef Training & Degrees. More than 1,000 schools & classes listed for all 50 States, Online and Worldwide
FREE Magazines
and other Publications
An extensive selection of free magazines & other publications
"I have long believed that good food, good eating is all about risk. Whether we're talking about unpasteurized Stilton, raw oysters or working for organized crime 'associates,' food, for me, has always been an adventure."
Anthony Bourdain, 'Kitchen Confidential (2000)
"Square meals, not adventurous ones, are what you should seek."
Bryan Miller (NY Times Restaurant Critic)
"I doubt the world holds for anyone a more soul-stirring surprise than the first adventure with ice cream."
Heywood Campbell Broun (1888-1939)
"An egg is always an adventure; the next one may be different."
Oscar Wilde(1854-1900)
"The kitchen, reasonably enough, was the scene of my first gastronomic adventure. I was on all fours. I crawled into the vegetable bin, settled on a giant onion and ate it, skin and all. It must have marked me for life, for I have never ceased to love the hearty flavor of raw onions."
James Beard (1903-1985)
Quotes about food and.....
Please feel free to link to any pages of FoodReference.com from your website.
For permission to use any of this content please E-mail: [email protected]
All contents are copyright © 1990 - 2017 James T. Ehler and www.FoodReference.com unless otherwise noted.
All rights reserved.
You may copy and use portions of this website for non-commercial, personal use only.
Any other use of these materials without prior written authorization is not very nice and violates the copyright.
Please take the time to request permission.


text:
The 'Valkyrie' actor has agreed to star in the upcoming Doug Liman-directed movie as a new recruit army who is reborn every morning after being killed in a war against alien invasion.
Tom Cruise is close to land another big role in a potential tentpole sci-fi titled "All You Need Is Kill". The 49-year-old actor has agreed to take on the lead role in the Doug Liman-directed film, which is expected to kick off the production in the late third/early fourth quarter of 2012.
Variety reported that Liman has traveled to Pittsburgh, where Cruise is filming "One Shot", to discuss the role with the actor. Though no formal deal has been signed, multiple sources revealed that the "Mission: Impossible" actor was pleased with the script, which is rewritten by Joby Harold.
"All You Need Is Kill" is set in the vein of "Groundhog Day". It follows a new recruit soldier who battles aliens in a war and finds himself caught in a time loop in which he wakes up in the past after having been killed on the battlefield. As the soldier's death and resurrection repeat, his skills as a soldier grow as he attempts to change his fate.
Cruise will soon be seen reprising his role as the IMF point man Ethan Hunt in "Mission: Impossible Ghost Protocol", which will open wide in the U.S. on December 21. His other upcoming films include Joseph Kosinski's sci-fi adventure "Horizons", and Adam Shankman's musical drama "Rock of Ages".


text:
|Primary category:
|CRM Software
|User Ratings:
|0.0/5 (0 ratings)
|Related Categories:
|Sales Management
|Pricing Model:
|Social Followers:
|29k likes 8.37k followers
Pipeliner is sales and marketing software with interactive visualization tools which can monitor all potential customers within the sales funnel in an organized and dynamic way.
Built with specifically the sales team in mind, Pipeliner CRM is unique in its focus on sales – from a visual, single-screen orientation of the sales pipeline to a comprehensive series of features supporting lead, contact, account and opportunity management.
Pipeliner CRM supports social selling with Twitter, Facebook, LinkedIn and Google+ integrations, allowing its users to view and interact with their accounts and contacts in social environments.
The CRM application Pipeliner scored 70/100. This is based on user satisfaction (0/100), press buzz (42/100) and other relevant information on Pipeliner gathered from around the web.
|The score for this service has improved over the past month. What is this?
Pipeline View: With Pipeliner CRM, it is possible to create as many pipelines as users need.
Lead Management: Lead effectiveness matters in a well-run pipeline. Its distinct architecture empowers sales teams as Leads move along the pipeline.
Opportunity Tracking: Tracking user's opportunities means they know exactly where every deal stands.
Business Intelligence: When running sales organization, it's key to have the right business intelligence on key performance indicators (KPI's).
Contact Management: This service puts related Opportunities, Activities, Tasks, Notes, and Files into one comprehensive Contact View.
Lead Generation: Pipeline's architecture is structured to accommodate the all-important role of leads in the sales funnel.
Credit card required: N/A
Price: $35 per user monthly, billed anually, with free trial.
|Company Info
|Based in
|Pacific Palisades
|3 Month Trend
|Alexa Rank:
|3 Months Change:
Languages: English
Pipeliner CRM is for sales managers and their teams of sales professionals in small and medium-sized businesses across various industries.
It is used for customer relationship management.
Supported platforms: Mac, Windows, Android, iPhone-iPad.
Pipeliner is most commonly used together with: Freshdesk; JotForm; and Xero.
Pipeliner has CRM integrations with 50+ popular apps, like: MS Office, Dropbox, Google apps, Marketo, MailChimp, Twitter, HubSpot, Zendesk, etc.
They offer an open API in C#, Java, PHP, Ruby Software Development Kits (SDK's), which can be accessed and integrated easily by other vendors (and app integrators).
They offer: Online Support; Library; Webinars; Tutorials; User Guides.


text:
Welcome back to The Town.
If you are joining us for the first time, The Town is a gathering of some of the most prestigious personalities* in esports that have come together to play Mafia** for 3 days in the BTS House. For those returning, what's wrong with you?
From August 19th to the 21st we burn our relationships to the ground for your viewing pleasure and and to satiate our own masochistic tendencies. This is the most personal event of the year for me and I hope you join us as we whittle away our friendships one mis-lynch at a time.
*By prestigious I mean the biggest losers in the scene that I could browbeat into coming to my low budget event. I mean seriously, who comes to a Mafia LAN right after TI like they have nothing better to do? rofl
**If you have no idea what Mafia is click the rules tab
- Charlie Yang
Main Stream: Twitch.TV/beyondthesummit
Mafia will begin at ~10:30 AM PST each day and will conclude at ~7:30 PM PST
with a 1 hour break for a special challenge segment.


text:
You purchased your Dynasty because its high-value and a trusted name appealed to your senses; so why would you be okay with anything but the best when it comes to great quality auto parts? It's not a regular occurrence that you realize that you have to buy a replacement A/C Condenser to repair your vehicle. Anyone who owns a Chrysler knows that when it comes to fixing their automobile only the highest quality replacement parts will do. Make no mistake, you didn't buy your Dynasty unless you knew that you wanted strength and a dash of luxury both rolled up into one car, truck, or SUV.
Keeping the interiors of our cars and trucks cool and comfortable has been possible since HVAC was pioneered around 70 years ago. The lifeblood of the air conditioning system is a refrigerant called R134a. The refrigerant emits cold air after a simple thermal reaction, and the A/C condenser on your Chrysler Dynasty acts like a radiator (it also looks like one) by cooling the refrigerant after it leaves the compressor, and then sends it to the drier and expansion valve. You can locate your Chrysler Dynasty A/C condenser sitting right in front of your radiator, as that is the easiest place to soak up air while your car is driving. If you notice the A/C condenser is leaking, then only mild or slightly cool air will blow into your interior because it is losing refrigerant.
Get your Dynasty back on the road with the highest quality replacement parts from the experts at Car Parts Discount. Given all the trouble that can be involved with installing a part to fix your car or truck, let www.carpartsdiscount.com be the most authoritative website you go to make purchasing it easy. Don't let a A/C Condenser repair bring about the demise of your car, truck, or SUV, and don't risk lowering its resale value by installing anything but the best replacement parts. If your car or truck was built in 1993, 1992, 1991, 1990, 1989, 1988, then you will find the correct Chrysler Dynasty A/C Condenser parts right here.


text:
My hair takes a lot to make it look good. It takes the right conditioner, hair spray, shampoo and styling products. My hair is very thick and has wave to it, not to mention it is now down to my elbows! So, any hair care product I use, has to get the job done, and […]
Apr 15, 2010
Mother's Day Giveaway! Head Organics TWO Winner's!


text:
|Message view
|« Date » · « Thread »
|« Date » · « Thread »
|"Shazron Abdullah (Created) (JIRA)" <j...@apache.org>
|[jira] [Created] (CB-169) Add PhoneGap.plist setting to turn off PGURLProtocol (whitelist)
|Wed, 11 Jan 2012 19:07:38 GMT
Add PhoneGap.plist setting to turn off PGURLProtocol (whitelist) ---------------------------------------------------------------- Key: CB-169 URL: https://issues.apache.org/jira/browse/CB-169 Project: Apache Callback Issue Type: Task Components: iOS Reporter: Shazron Abdullah Assignee: Shazron Abdullah Fix For: 1.4.0 This relates to the Cleaver (PhoneGap as a component work). This is for apps that are already using NSURLProtocol to filter network traffic - for example VPN type apps. -- This message is automatically generated by JIRA. If you think it was sent incorrectly, please contact your JIRA administrators: https://issues.apache.org/jira/secure/ContactAdministrators!default.jspa For more information on JIRA, see: http://www.atlassian.com/software/jira
|View raw message