text:
The role of Ca2+ in the induction of neural correlates of memory has frequently been described in binary terms despite the fact that many forms of memory are graded in their strength and/or persistence. We find that Ca2+ dynamics encode the magnitude of sensorimotor adaptation of the electromotor output in a weakly electric fish. The neural correlate of this memory is a synaptically induced Ca2+-dependent enhancement of intrinsic excitability of neurons responsible for setting the electromotor output. Changes in Ca2+ during induction accurately predict the magnitude of this graded memory over a wide range of stimuli. Thus despite operating over a range from seconds to tens of minutes, the encoding of graded memory can be mediated by a relatively simple cellular mechanism.
Sensorimotor adaptation is the appropriate adjustment of a motor output to a persistent change in sensory inflow. To function correctly, it must be graded to match the magnitude of the sensory perturbation. Theoretically the magnitude of such a correction is independent of the time course during which it was induced, allowing animals to integrate gradual changes as well as rapid ones. This type of motor learning has been described in numerous systems (Baroni et al. 2001; Held and Freedman 1963; Houde and Jordan 1998; Kluzik et al. 2005; Miles and Eighmy 1980; Peterka and Loughlin 2004) and has proven to be useful in the rehabilitation of individuals who suffer cognitive, limb control, and postural deficits (Brewer et al. 2008; Granacher et al. 2006; Patton et al. 2006; Rossetti et al. 1998).
Despite the ubiquity of sensorimotor adaptation in normal function and its recent use in rehabilitation, mechanisms underlying sensorimotor adaptation remain less known. This has been complicated by the fact that it often occurs within distributed networks that employ several sensory and motor modalities. In conjunction with changes on the network level (Aksay et al. 2001, 2007; Paz et al. 2005; Wang et al. 2004), changes also occur within individual neurons within these networks (Masumura et al. 2007; Nelson et al. 2003; Smith et al. 2002). Here we have characterized an experimentally accessible form of sensorimotor adaptation in the electromotor output of a weakly electric brown ghost knifefish, Apteronotus leptorhynchus (Oestreich and Zakon 2002, 2005; Oestreich et al. 2006). These fish utilize distortions in the electric field they continually discharge from their electric organ to sense their environment. The electric organ discharge (EOD) frequency is different for each individual fish and is highly regular (coefficient of variation, CV = SD/mean period = 2.2 × 10-4) (Moortgat et al. 1998). However, when two fish with similar EOD frequencies are in proximity to one another, their electric fields will jam one another, confounding their ability to sense their environment (Fig. 1A). A fish exposed to electrosensory jamming will elevate its EOD frequency out of the jamming range. The EOD relaxes back to its original frequency when the jamming stimulus is removed. When this electrosensory jamming continues for periods longer than a few minutes, the fish will display a long-term frequency elevation (LTFE) that persists even once the jamming stimulus is removed (Fig. 1B). Similar to other forms of sensorimotor adaptation, the magnitude of LTFE depends on the intensity and duration of the sensory perturbation (Oestreich and Zakon 2002).
The neural circuitry underlying this adaptation has been well identified and is experimentally accessible. Neurons in the medullary pacemaker nucleus (PMn) directly control the EOD frequency via electrical coupling to spinal motor neurons the axons of which comprise the electric organ. As a result of this coupling, the firing frequency of PMn neurons is equal to the EOD frequency of the fish. Thus the activity in the PMn is a direct representation of the behavior in the fish. The PMn is the locus of the sensorimotor adaptation: LTFE in the isolated brain stem can be reproduced by stimulating afferent fibers from the pre-pacemaker nuclei that are activated by the jamming signal (Oestreich et al. 2006). Stimulating these glutamatergic fibers produces a transient elevation in the PMn firing frequency as result of their ionotropic synaptic input to PMn neurons. With sufficient activation, this transient elevation in PMn firing frequency is followed by a long-lasting elevation in the PMn neurons' firing frequency. This LTFE in the PMn neurons occurs at the same range of magnitudes as LTFE in the EOD and decays at the same rate. After induction, this LTFE persists even after synaptic activity has been blocked, meaning that the neural correlate for this sensorimotor adaptation is a synaptically induced graded persistent increase in the activity of the PMn neurons.
Previously, we have shown that N-methyl-d-aspartate (NMDA) receptors are necessary for the induction of LTFE and nonselective cation TRP channels are necessary for LTFE maintenance (Oestreich et al. 2006). However, it remains unclear how NMDA receptor activation recruits TRP channels. Several modeling studies have proposed that Ca2+ dynamics within individual neurons might underlie the maintenance of graded persistent increases in firing frequency that are similar to LTFE (Loewenstein and Sompolinsky 2003; Teramae and Fukai 2005).
In this paper, we sought to directly examine how Ca2+ dynamics translate synaptic inputs into this graded form of memory. Increases in Ca2+ might contribute to LTFE induction and might also be required for its long-lasting maintenance (Oestreich et al. 2006). Here we image Ca2+ in the PMn during LTFE induction and maintenance. We found that synaptically induced increases in Ca2+ signal rose concomitantly with LTFE induction across a broad range of stimuli but that LTFE was maintained well beyond these elevations. Interestingly, the cumulative Ca2+ signal during stimulation predicted the magnitude of the LTFE across a wide range of stimulus parameters, suggesting that Ca2+ levels during induction directly encode the magnitude of the sensorimotor adaptation.
Wild-caught A. leptorhynchus were obtained from different vendors and then housed in community Plexiglas tanks in climate-controlled rooms with a circulating water system and a dark-light cycle of 12/12 h. The temperature in the rooms was held stable between ∼26 and 28°C, and the water conditions were relatively constant. Fish were fed with frozen brine shrimp every 2 days. Procedures and animal care are in accordance with guidelines from the animal care and use committee at University of Texas.
PMn slice preparation
The submerged PMn slice preparation was performed as previously described (Oestreich et al. 2006). Fish were anesthetized using 0.1% phenoxyethanol, and the dissections were performed on ice. The slice is composed of the ventral portion of the brain stem, spanning from a point in close proximity of the caudal aspect of the pacemaker nucleus to the pituitary fossa. Once secured to a coverslip with superglue, the slice was transferred to a recording chamber where it was continuously perfused with ACSF. Extracellular field recordings of the synchronized compound field potential of the PMn were acquired using a 1 M NaCl-filled patch electrode (1–3 MΩ), amplified using an Axopatch 200B, A/D-converted with the Digidata 1322, and digitally recorded using Clampex 9 (Molecular Devices, Sunnyvale, CA). The PMn firing frequency was monitored (20 kHz) during the experiment using Clampex on-line analysis of frequency count. More precise measurements of firing frequency were determined after the experiment using FastFourier analysis. LTFENMDA was induced by ejecting NMDA (3.3 mM, Tocris, Ellisville, MO) from a microelectrode pipette in close proximity to the pacemaker nucleus using a 1 s, low pressure (5 psi) pulse with a Picospritzer III. LTFEstim was induced by a bipolar stimulating tungsten electrode placed on the afferent fiber pathways rostral to the PMn.
Calcium imaging
Dye loading was performed using the multicell bolus loading (MCBL) technique (Stosiek et al. 2003). A stock solution (10 mM in DMSO and 20% pluronic acid) of Oregon Green bis-(o-aminophenoxy)-N,N,N′,N′-tetraacetic acid (BAPTA)-1-488-AM (OGB-1) or magnesium green (both purchased from Invitrogen, Carlsbad, CA) was diluted in artificial cerebrospinal fluid (ACSF) for a final concentration of 1 mM. Ca2+ signals were acquired with an Olympus Fluoview 300 confocal microscope (Olympus, Center Valley, PA). Excitation light was powered by an argon (488 nm) laser (max output: 1.2 W; Melles Griot, Carlsbad, CA). Images were acquired every 2.88 s, or for longer experiments, every 10 s to minimize bleaching with an Olympus ×40 water immersion lens (0.80 NA). Once the raw data were collected, each region was normalized to percent change [ΔF/F = (F − x)/x], where F was the fluorescence at a given time point, x was the average fluorescence of the initial 60 s in that region. The data were background-subtracted using a region of the pacemaker nucleus that is dye-loaded but that shows no response to NMDA application or stimulation as background. Individual pacemaker or relay neurons were distinguished from one another by their size as visualized by DIC optics.
Data analysis
Graphing and statistical analysis was performed using Excel (Microsoft, Redmond, WA), IGOR (Wavemetrics, Lake Oswego, OR), Adobe Illustrator (Adobe Systems, San Jose, CA), Origin (OriginLab, Northhampton, MA), and InStat (GraphPad, La Jolla, CA). Data are given as means ± SE. For statistical comparisons, normally distributed data were tested using either a student's t-test, or if across different stimulation paradigms, with an ANOVA followed by Bonferonni comparisons to test for significant differences between individual groups. For some multiple comparisons, the data were not normally distributed and thus a nonparametric ANOVA (Kruskal-Wallis) was used followed by a Dunn's to test for significant differences between individual groups.
Rises in intracellular Ca2+ are coincident with LTFE induction
The neural correlate of LTFE is a synchronous synaptically induced elevation in the firing frequency of all the PMn neurons that is presumably accompanied by changes in intracellular Ca2+. To elucidate the magnitude and temporal characteristics of these changes in Ca2+, we imaged the Ca2+ signal in the PMn slice. An adapted multi-cell bolus loading technique established in zebrafish spinal cord was used to bulk load the calcium indicator dye Oregon Green BAPTA–AM (Fig. 2A) into the slice (Brustein et al. 2003). This membrane-permeable calcium dye allowed us to measure changes in Ca2+ in several neurons simultaneously during LTFE induction and maintenance. The dye was introduced with a microelectrode patch pipette (1–3 MΩ) placed at the surface or 50–100 μm within the pacemaker nucleus by pressure application (100 pulses of 50 ms at 5 – 10 psi). Ca2+ measurements were taken 30–60 min after dye application; at which time several pacemaker and/or relay PMn neurons are loaded.
PMn neurons are synchronously active at a highly stable frequency (500-1,000 Hz, CV = 6.1 × 10−4, (Moortgat et al. 1998), and in the absence of synaptic activation, we observed no detectable changes in the basal Ca2+ signal. By contrast, stimulating the prepacemaker afferents briefly (100 pulses delivered at 100 Hz, Fig. 2B) triggered elevations in Ca2+ in several neurons. This Ca2+ signal was long-lasting with Ca2+ elevated for hundreds of seconds (180 s). Although the Ca2+ signal rises concurrently with the increase in firing frequency, these increases are not simply the result of higher PMn firing rates. When stimulating more robustly (500 pulses at 50 Hz), we sampled Ca2+ at a lower rate (0.1 Hz) to measure changes in the Ca2+ signal during LTFE maintenance (tens of minutes, Fig. 2C). The increase in the Ca2+ signal was greater with the stronger stimulation and returned to baseline more quickly (examined more thoroughly in the next section) but still was long-lasting (80 s). Notably LTFE persisted for several minutes after the Ca2+ signal had subsided. Thus the elevation of firing frequency itself during LTFE could not account for the elevation in the Ca2+ signal. Furthermore, changing the PMn firing frequency without synaptic activation by altering the bath temperature produced no concurrent change in the Ca2+ signal. Raising or lowering the bath temperature (±4°C) produced significant shifts in the PMn firing rate (+4°C = 14.24 ± 5.23 Hz, −4°C = −14.20 ± 3.10 Hz) with no corresponding changes in fluorescence [+4°C: ΔF/F = 0.61 ± 0.87%, −4°C: ΔF/F = 2.42 ± 0.56%, ANOVA for 3 temperatures: F(115,2) = 2.20, P = 0.115].
Elevations in Ca2+ occurred with a similar time course across multiple neurons, independent of whether they were pacemaker or relay neurons. The magnitude of the Ca2+ signals varied, but this difference did not correspond with neuron type. To examine this, we measured Ca2+ signals in pacemaker and relay neurons simultaneously (n = 5). The Ca2+ signal in the pacemaker neurons was on average slightly larger, but this relationship was not significant (paired t-test, P = 0.29, Fig. 2D). To test whether the time course of the Ca2+ signal was similar in pacemaker and relay neurons, we fit the decay of the Ca2+ signal to either a single or double exponential. In 3/12 cases (2 pacemaker neurons, 1 relay neuron) the decay was fit best by a double exponential, the slower exponent of which was similar to the single exponential fits. The decays of the Ca2+ signal did not depend on neuron type (single exponential tau, paired t-test, P = 0.1765, Fig. 2E).
Our imaging technique did not appear to interfere with LTFE or the Ca2+ signals within PMn neurons during LTFE. Dye loading did not interfere with the PMn neurons' basal firing frequency (Δ −1.36 ± 0.28 Hz, paired t-test, P = 0.63; n = 20). Similarly dye loading did not interfere with LTFE (Fig. 3A), either the magnitude (Δ −0.35 ± 1.69 Hz; paired t-test P = 0.79, n = 6) or decay (Δ tau = −23 ± 81.76 s, paired t-test, P = 0.84, n = 6). To further confirm that the signal was not confounded by the high affinity of OGB-1 for Ca2+ (Kd = 165 nM), we utilized the low-affinity Ca2+ dye magnesium green (Kd = 6 μM). As was the case with OGB-1, the Ca2+ signal measured with magnesium green did not differ between neuron types (single exponential tau, paired t-test P = 0.3944). The Ca2+ signals of OGB-1 and magnesium green were similar across stimulation paradigms (Fig. 3B). The decays of these signals were not statistically different depending the dye used (at 50 Hz: t = 1.3, at 500 Hz: t = 0.6154, both P > 0.05, Fig. 3C) but did vary depending on the strength of the stimulus [ANOVA F(24,3) = 3.75, Bonferonni post hoc analysis t = 3.151, P < 0.05].
To compare the changes we observed with our bulk-loading technique to changes in Ca2+ signal at the single-cell level, we filled individual relay neurons using a low-resistance (5–8 MΩ) sharp electrode containing 100 μM membrane impermeant OGB-1 (Fig. 4). Ca2+ signals in individually loaded neurons were similar to those observed when bulk-loaded. No changes in Ca2+ were observed in the absence of synaptic stimulation. Synaptic stimulation for brief (100 Hz for 1 s, n = 3, Fig. 4B) or for more prolonged periods (50 Hz for 10 s, n = 4, C) elicited Ca2+ signals similar to bulk loaded neurons. The Ca2+ signals were similarly stimulus dependent (Dunn's multiple comparisons, P < 0.05) and consistently larger with individually loaded neurons across stimuli, presumably due to more localized image sampling (nonparametric ANOVA: KW = 16.868, P < 0.001, Dunn's multiple comparisons, P < 0.05). The Ca2+ signals also decayed in a stimulus-dependent manner [ANOVA F(18,3) = 20.493, Bonferonni post hoc analysis for OGB-salt: t = 4.455, for OGB-AM: t = 6.25, both P < 0.01] but not between loading techniques (at 50 Hz: t = 0.067, at 100 Hz: 0.6331, both P > 0.05, Fig. 4D). Thus Ca2+ signals at the single-cell level appeared to match our results from bulk loading. This suggested that our signal was indeed from the PMn neurons and not from prepacemaker afferent fibers that had been spuriously labeled.
LTFE maintenance is not dependent on elevated Ca2+
Having established the efficacy of our imaging technique, we sought to carefully examine how the Ca2+ signal rises and falls during LTFE induction and maintenance. LTFE in vivo is graded, depending on the intensity and duration of jamming stimulus the fish is exposed to. Although the firing frequency of the prepacemaker nuclei during a jamming stimulus remains unknown, stimulating the afferent fibers from these nuclei produces the same range of LTFE in vivo (Oestreich et al. 2006). LTFE output increases linearly with pulse count across a wide range of frequencies (1.5-1,000 Hz). We therefore sought to examine LTFE induction and maintenance across a wide range of stimulus parameters. Stimulating in a manner that mimics a mild LTFE induction in the fish (Fig. 5A: 100 pulses at 100 Hz) resulted in an increase in the Ca2+ signal (peak ΔF/F: 12.27 ± 1.55%, n = 9) and a modest firing frequency (LTFE = 6.17 ± 0.83 Hz; n = 9). The Ca2+ signal persisted well beyond the stimulation period and was best fit by a single exponential faster (τ = 141.64 ± 19.98 s) than the decay of the firing frequency (τ = 165.01 ± 44.71 s), although this difference was not statistically significant (Student's t-test, P = 0.18). Importantly, this was not true with more intense LTFE induction protocols. 3 × 100 pulses at 100 Hz, separated by 20 s produced LTFE of greater magnitude (10.35 ± 1.27 Hz, n = 7) and slower decay (τ = 245.62 ± 54.66 s; Fig. 5B). Concurrent with the increase in LTFE magnitude, we observed an increase in the Ca2+ signal during each stimulation (1st peak ΔF/F = 8.80 ± 1.62%, 2nd peak ΔF/F = 20.27 ± 3.62%, 3rd peak ΔF/F = 26.09 ± 3.19%, n = 7). However, with this more robust stimulus, the Ca2+ signal did not persist throughout the maintenance phase of LTFE. Although there was a twofold increase in the peak of Ca2+ signal by the third stimulation as compared with a single stimulation, the Ca2+ signal decay was still best fit by a single exponential (τ = 86.49 ± 26.29 s, Student's t-test, P < 0.05).
We then examined changes in Ca2+ signal using a protocol that mimicked the maximal LTFE induction in the intact fish (Fig. 5C). To do this, we stimulated the PMn synaptically with 4,000 pulses delivered at a fixed rate (5 Hz) for 800 s. This stimulation protocol produced a near-saturating LTFE magnitude (16.43 ± 1.01 Hz, n = 3) that was maintained for a prolonged period (4.5 ± 1.6 Hz still above baseline at 30 min). During the induction of maximal LTFE, a steady increase in firing frequency was matched with an increase in the Ca2+ signal (peak ΔF/F = 41.53 ± 5.20%, n = 4). When synaptic stimulation terminated, both the elevation in firing frequency and the Ca2+ signal persisted well after the stimulus. However, the decay in the Ca2+ signal was best fit by a single exponential (τ = 198.6 ± 50.24 s) that was much more rapid than the decay of the firing frequency (τ = 1,538.82 ± 605.5 s). Thus with stronger stimulus paradigms, the Ca2+ signal increases become greater during LTFE induction but do not persist throughout LTFE maintenance.
Mechanisms underlying LTFE induction
Several lines of evidence suggest that Ca2+ influx into the PMn neurons through NMDA receptors is necessary for LTFE induction. NMDA receptors are found in the pacemaker and relay neurons within the PMn but not in the presynaptic terminals of afferent fibers nor in the glia within the PMn (Harvey-Girard et al. 2007; Spiro et al. 1994). NMDA-R antagonists block LTFE induction, whereas blocking metabotropic glutamate and kainate receptors does not (Oestreich et al. 2006). Furthermore, application of NMDA to the PMn mimics LTFE (Oestreich et al. 2006). This NMDA-induced LTFE (LTFENMDA) is blocked in Ca2+-free extracellular solution, suggesting that it is the Ca2+ influx that is necessary for LTFE not just depolarization. We examined whether direct NMDA application could mimic the Ca2+ signal observed during synaptic stimulation. Brief puffs of NMDA (5 μM, 10 psi, 1 s) caused an increase in Ca2+ in several neurons concurrent with the expected elevation in firing frequency (Fig. 6A). The Ca2+ signal was long-lasting, with a decay (τ = 60.67 ± 14.86 s) that was similar to the decay in firing frequency (τ = 59.79 ± 7.92 s, Student's t-test P = 0.96; Fig. 6B). Both LTFENMDA and Ca2+ signal were increased with more prolonged NMDA applications (Fig. 6C). These results indicate that NMDA receptor activation alone can result in Ca2+ influx into the PMn neurons and further implicate Ca2+ influx through NMDA receptors as the means by which LTFE induction occurs.
Given the long-lasting nature of the Ca2+ signals with LTFE induction, we also examined whether Ca2+ stores might contribute to LTFE. Ca2+ influx through NMDA receptors has been shown to activate release from intracellular stores (Baba et al. 2003). To test whether Ca2+ stores might contribute to LTFE, we depleted calcium stores by applying cyclopiazonic acid (CPA) or thapsigargin (Fig. 6, D and E). Depleting Ca2+ stores lowered the PMn firing frequency (Student's t-test, P < 0.05, Fig. 6E), but neither of these compounds had an effect on LTFE magnitude [thapsigargin, 2 μM: paired t-test P = 0.314; CPA, 50 μM: ANOVA: F(15,2) = 0.91065, P = 0.42]. Thus although calcium stores may contribute to some aspect of setting the PMn firing frequency, these mechanisms do not appear to contribute to LTFE.
Pulse counting reflects the change in Ca2+ during stimulation
LTFE persists beyond the Ca2+ elevation (Figs. 2E and 5C), suggesting that continued elevation in Ca2+ is not necessary for LTFE maintenance. In contrast, the Ca2+ signal appeared to reflect LTFE induction more closely. We therefore examined whether Ca2+ contributes to the pulse-counting properties of LTFE induction. The LTFE pulse-counting mechanism follows two computational rules: it increases linearly with pulse count and it is largely insensitive to the frequency at which input pulses are delivered. If changes in Ca2+ contribute to the pulse-counting mechanism underlying LTFE, it is expected that they might also be constrained by these computational rules. As seen in Fig. 5, the Ca2+ elevation increases concomitantly with pulse count. However, for the Ca2+ signal to act as a component of the pulse-counting mechanism, it would also need to be insensitive to changes in the pulse frequency. To test for this, we compared changes in the Ca2+ signal during LTFE induction over a wide range of frequencies (5, 50, and 500 Hz) while keeping the pulse number constant (500; Fig. 7). Inducing LTFE with the same number of stimulus pulses over this wide temporal range (1, 10, and 100 s) yielded comparable magnitudes of LTFE [5 Hz = 11.83 ± 1.6 Hz, 50 Hz = 12.67 ± 0.83 Hz, and 500 Hz = 10 ± 1.45 Hz; ANOVA: F(13,2) = 1.054, P = 0.3765]. By contrast, changes in the Ca2+ signal were distinct from one another (Fig. 7). As stimulation frequency was increased, the peak of the Ca2+ signal increased [ANOVA F(13,2) = 10.34, P < 0.05]. In every case, LTFE persisted well after the Ca2+ signal had returned to baseline. In most cases the decays were best fit by a single exponential (5 Hz: 5/5; 50 Hz: 4/6; 500 Hz 7/9). The 5 Hz stimulus decayed slower than the signals from 50 and 500 Hz [ANOVA F(13,2) = 4.908, P < 0.05; the 500 Hz]. Although the stimulation protocols produced very different peak changes in the Ca2+ signal, they were similar in the total Ca2+ signal during LTFE induction. The cumulative Ca2+ signal during stimulation was similar despite the fact that the time course of LTFE induction varied over two orders of magnitude [5 Hz: ΔF/F = 103.55 ± 21.72%, 50 Hz: ΔF/F = 98.13 ± 28.68%, and 500 Hz: ΔF/F = 80.46 ± 17.68%, ANOVA F(13,2) = 1.019, P = 0.96]. Thus the cumulative Ca2+ during stimulation matched the amount of LTFE while the peak Ca2+ did not.
To determine whether Ca2+ participates in the pulse-counting mechanism in LTFE, we examined how LTFE magnitude correlated with the magnitude, rise time, and total change of the Ca2+ signal across all the different stimulus protocols used in this study (Fig. 8). Neither the magnitude nor rise time of the Ca2+ correlated with the magnitude of the LTFE (Fig. 8, A and B). In contrast, the cumulative sum of the [Ca2+]i signal during LTFE induction correlated strongly with LTFE magnitude (Fig. 8C; r2 = 0.9254, P < 0.005 linear regression). Therefore the change in Ca2+ during LTFE induction follows both computational rules of the LTFE pulse-counter, as it increases with pulse count and is independent of frequency. This was not true for the total change in Ca2+ during LTFE induction and maintenance (Fig. 8D), which is consistent with our previous finding that Ca2+ is not required for LTFE maintenance.
In this study, we have taken advantage of an experimentally accessible form of sensorimotor adaptation to show how Ca2+ dynamics contribute to the encoding of a graded memory. LTFE in a weakly electric fish functions a pulse-counter, integrating changes in sensory input over a wide range of time scales to produce the appropriate compensatory electromotor output. By exploiting the computational rules that constrain how LTFE integrates synaptic inputs, we determined that the cumulative change in the Ca2+ during stimulation accurately predicts LTFE magnitude across a range of input stimuli.
Ca2+ signal in response to afferent fiber stimulation occurred with a similar time course in the PMn neurons. Although there was some variability in the magnitude of the changes in Ca2+ between neurons, Ca2+ fluctuations in a single intracellularly loaded relay neuron were very similar to fluctuations in several neurons imaged simultaneously. Of course, this does not preclude some variability in Ca2+ elevations between different neurons at faster time scales below the resolution of our Ca2+ imaging. Nonetheless, these findings suggest that at the time scale that LTFE occurs, Ca2+ dynamics in the PMn can be considered uniform among the neurons. Thus although pacemaker neurons are known to drive relay neurons, which in turn drive the electromotor neurons in the spinal cord (Elekes and Szabo 1985), both neuron types appear to participate in Ca2+ dynamics underlying LTFE. This is consistent with the fact that both neuron types receive glutamatergic input (Dye and Heiligenberg 1987; Heiligenberg et al. 1996), both express NMDA receptors (Harvey-Girard et al. 2007; Spiro et al. 1994) and the calcium-binding protein calbindin (Smith et al. 2000). Interestingly, a related species of electric fish (Eigenmannia viriscens) that generates a much longer-lasting LTFE (Oestreich and Zakon 2005), expresses more vitamin-D-sensitive calcium binding protein in relay neurons than pacemaker neurons (Maler et al. 1984). Future studies will have to determine whether protein differences in the two species might account for differences in the duration of their sensorimotor adaptation. The Ca2+ elevations in PMn neurons in response to afferent stimulation can be remarkably long-lasting (>100 s) but do not persist for the duration of LTFE maintenance (Figs. 2E, 5, A–C, and 7, A–C). Thus elevations in firing frequency do not themselves result in increased Ca2+ levels in PMn neurons. The fact that the Ca2+ signal does not persist for the duration of LTFE eliminates several mechanisms by which it could be maintained. Voltage-gated calcium channels, Ca2+-permeable cation channels and intracellular Ca2+ stores are unlikely to be directly involved during LTFE maintenance. This is in accord with several findings. The N-type Ca2+ channel blocker ω-conotoxin-GVIA applied after LTFE induction has no effect on LTFE (Oestreich et al 2006). Several nonspecific voltage-gated calcium channel blockers have little or no effect on PMn firing frequency (Smith and Zakon 2000). Depleting intracellular stores had no effect on LTFE (Fig. 6, D and E), consistent with the fact that Ca2+ elevations do not persist for the duration of LTFE. These results are consistent with our finding that LTFE is maintained by a calcium nonselective cation current (Oestreich et al. 2006). It is notable that the TRPM5 channel that we localized to neurons within the PMn (Oestreich et al. 2006) is one of the few TRP channels that is calcium-impermeable. However, unless TRPM5 deactivates on the time scale of hundreds of seconds, the TRPM5 recruitment would have to be more complex than Ca2+ directly gating TRPM5. The most obvious form this might take is Ca2+ activation of a secondary messenger phosphorylation cascade (i.e., CamKII or PKC activation) that in turn might alter the properties of TRPM5 within the PMn neurons.
The fact that the Ca2+ signals do not persist throughout LTFE maintenance suggests that several proposed models of graded persistent activity are insufficient to explain LTFE. Some of these models explicitly require elevations in Ca2+ throughout maintenance (Loewenstein and Sompolinsky 2003; Teramae and Fukai 2005) In contrast to these models in which Ca2+ signals encode the graded activity, our results favor the idea that transient Ca2+ signals result in the recruitment of a balance in kinase and phosphotase activity (Delord et al. 2007; Fransen et al. 2006). These types of changes in Ca2+ during LTFE are more reminiscent of the role of Ca2+ in synaptic plasticity, where Ca2+ dynamics during induction determine the magnitude of the plasticity (Bear 2003; Johnston et al. 2003; Sjostrom and Nelson 2002), but Ca2+ isn't persistently elevated during its expression. It will be of great interest to determine how Ca2+ contributes to the graded nature of other forms of Ca2+-dependent synaptically induced changes in intrinsic excitability (Nelson et al. 2003, 2005; Xu et al. 2005; Zhang et al. 2004).
It can be difficult to determine which parameters of a Ca2+ signal are relevant to its downstream effectors. We have taken advantage of the computation rules that constrain the LTFE pulse counting mechanism to address this. LTFE increases linearly with pulse count and is markedly insensitive (>2 orders of magnitude) to the frequency at which those pulses are delivered. By using a range of stimulations varying in pulse count and frequency, we determined that the change in the Ca2+ signal during stimulation is the most accurate parameter for predicting LTFE magnitude. Specifically, the log of the change in the Ca2+ signal during the stimulation correlated with LTFE magnitude. Remarkably, this relationship is reminiscent of an enzymatic reaction with a single binding site that saturates. It is possible that the relationship between Ca2+ and LTFE reflects a single binding site for Ca2+ onto its downstream targets. However, our findings suggest that Ca2+ handling in PMn neurons is more complex than this. There is a portion of the Ca2+ signal that persists once synaptic stimulation has passed, which does not appear to be salient to the LTFE mechanism. In cases where LTFE is not saturated, this extra Ca2+ does not trigger greater LTFE. It may be compartmentalized or buffered in some manner from the downstream effectors necessary for LTFE. In fact, the decay of the Ca2+ signal possesses a particularly interesting feature. The rate of decay in the Ca2+ signal appears to increase with larger peak Ca2+ elevations (Fig. 7B). This suggests that Ca2+ handling within the PMn neurons may itself be an active process. Active Ca2+ handling might contribute to the pulse-counting mechanism that is independent of the input pulse frequency by minimizing extraneous Ca2+ activation of downstream effectors.
The gradation of a memory is often assumed to be an emergent property of several processes occurring at many synapses and/or neurons. This is particularly relevant in the case of sensorimotor adaptation, which integrates sensory perturbation over a wide temporal range. Indeed for certain forms of sensorimotor adaptation, network activity appears to dominate the adaptation. For example, persistent firing in neurons within area I of the goldfish oculomotor integrator cannot be driven in vivo by simple somatic depolarization but rather appear dependent on network interactions (Askay et al. 2001). Nonetheless, sensorimotor adaptation is also likely to be mediated by Ca2+ dynamics altering individual neurons' intrinsic properties. Vestibular neurons that show adaptive changes in their firing rate responses correlating with VOR gain appear to be regulated by Ca2+-dependent processes (Nelson et al. 2003,. 2005; Smith et al. 2002; Straka et al. 2005). Similarly, other forms of memory that are similarly graded (e.g., working memory) may be mediated by Ca2+ driven single-cell mnemonic activity (Egorov et al. 2002, 2006; Fransen et al. 2006; Sidiropoulou et al. 2009). However, in all of these cases, it is difficult to relate Ca2+ signaling directly to the magnitude of behavioral changes in an animal. Here we have found that the Ca2+ signal during LTFE induction correlates with the magnitude of the memory whether it occurs in one second or over tens of minutes. Because the PMn neurons directly represent the EOD in a behaving fish, this suggests that a relatively simple cellular process can itself encode a graded memory across a wide dynamic range.
This work was supported by the National Institutes of Health Grants R01 MH-056535 to H. H. Zakon and R01 NS-044399 D. L. Pettit and the Ruth Kirchestein postdoctoral training fellowship F32 NS46949-02 to N. C. Dembrow.
Thanks to R. Narayanan for helpful comments on the manuscript.
- Copyright © 2010 the American Physiological Society


text:
- Properties
- Agents
- Offices
- About the Area
- News
- Videos
- About Us About Us
- Preferences
- Back
- Change Currency:
1 of 25
Located in the Historic Upper East just minutes from the city center, find this undeerstated jewel that was the personal residence of William A. Edwards (of Edwards and Plunkett Architects). The home has been meticulously updated with the highest quality finishes and great care while keeping the charm and beauty of a 1922 era estate including beautiful beams and rich hardwood floors. The chef's kitchen was completely remodeled and features premier appliances, custom cabinets, an island and an imported French Antique Terra Cotta stone floor. The upstairs master bedroom and bath were completely changed to create a luxurious master suite with the finest fixtures and dual walk-in closets. There is a large studio with a full bath and kitchenette, vaulted ceiling and a wonderful terrace above the oversized finished three-car garage that includes a separate workshop area. In addition, there is a separate art studio. All of this on a .46 acre lot with beautiful landscaping and several outdoor sitting areas to enjoy.
m: (805) 451-5699
o: (805) 963-1391

subset: commoncrawl


text:
Tag Archives: causes excessive
Acid Reflux: You Can Get Relief
Acid reflux results in one of the worst feelings you can have. The agonizing pain and burning sensations are enough to make anyone lose their mind. Know that you are not the only person who has acid reflux. There is no reason for you to put up with it either, as the piece that follows … Read More


text:
Our information consequently advise that lacking regarding Treg might compliment the results of IL-7 through removing a prospective cytokine sink. Cellular the extraordinary increase in numerous cytokine amounts and the capacity of IL-7-treated these animals in order to effectively clear trojan, all of us investigated regardless of whether inhibitors regarding cytokine signaling might be repressed
Log in to comment or register here.
What is Kliqqi?
Kliqqi is an open source content management system that lets you easily create your own user-powered website.
Latest Comments


text:
I love reading blogger advice and topic idea posts so to have 1 for every day of the year at my fingertips is so incredibly helpful. I know there are a lot of bloggers out there that post topic ideas but theres just something nice about the way you can physically hold it.
The design of the book is simple and clean with a lovely teal blue cover that looks great on your bookshelf. It's layed out with the topic ideas being the main focus of the page in a bold serif font which makes it quick and easy to flick through and get inspired for your next post.
I think this book is such a great idea and has already helped me with future posts and I would urge any new lifestyle blogger to consider buying one for themselves. Even if you have it on your desk just incase you have one of those days where you think "shit...I have nothing at all to write about" and it does happens to all of us. This is the cure.


text:
Police Officer
Anderson Police Department - Anderson, Indiana
- Application Deadline:
Salary Information
- Entry Level Salary:
- $44,611.00 annual
Job Description
The City of Anderson (IN) Police Department is now accepting applications for the 2013 recruit process. Applicants may download an application packet by going to www.cityofanderson.com and clicking on the link under Employment Opportunities. Applicants may mail or deliver their completed packets to: Anderson Police Department Attn: Training Coordinator Brandon Grant 1040 Main St. Anderson, IN 46016 In person drop off times are Mon. through Fri., 7:00 am - 9:00 pm. Deadline to return completed applications is October 1st, 2013. Any questions may be directed to the Anderson Police Department Training Division, 765-648-6765 or 765-648-6767. The City of Anderson is an equal opportunity employer.
Additional Information
The City of Anderson is located approximately 30 miles N/E of Indianapolis and serves a population of 56,136 in a 45 square mile area.
Job Requirements
- Age: 21-36
- Education: HS or GED
- Experience: Academy graduates preferred but not required
Contact Informaton
Brandon Grant
Anderson Police Department
1040 Main St
Anderson, Indiana 46016
phone: (765) 648-6765


text:
Mid-South Community College basketball camp 2017
Think about what career is best suited for you, and where you want to study. Request more information now.
Think about what career is best suited for you, and where you want to study. Request more information now.
Think about what career is best suited for you, and where you want to study. Request more information now.
Information about Mid-South Community College basketball camp 2017, and registering for elective lectures and seminars. Not all recruiting is done like NCAA Division I teams. Coaches are going to be more drawn to players who display genuine interest in their program throughout the recruitment process. Finally, check with your major department for assistance with prerequisites, tuition payment deadlines, and Mid-South Community College Basketball information. Please use the form above to request admissions info for Mid-South Community College.
You don't have to quarterback the university football team to a state championship to earn an athletic scholarship. Most major sports that field a team in intercollegiate competition have sports scholarships available. From women's gymnastics to men's track and field, coaches are seeking players to build a team around. Further, that leadership role often is accompanied by a full-scholarship.
The best time to begin searching for scholarships is before your senior year, in order to meet deadlines. Schools confirm that it is important to file financial aid forms such as the CSS Profile early in the school year. There are many sports scholarships that focus on less tangible qualities, such as the ability to lead a team, motivate others, and learn new skills rapidly. Take stock of your athletic abilities as you mature through high school. Stark State College athletic conference may encourage student-athletes to apply via early decision. Students who are considering applying via early decision should be aware of the rules and obligations that this commitment entails. Beyond consideration for Mid-South Community College basketball camp 2017, you may think about Klamath Community College GRE test dates for complementary electives to your primary course of study.
Women's Soccer Head Coach - Part-time
Kern Community College District
This assignment will also require coaching at games/tournaments that are scheduled in the evening and/or Saturdays.
4 days ago Boxing/Kickboxing Fitness Instructor
Title Boxing Club - West Wichita
Ability to confidently lead our signature POWER HOUR classes using our structured approach, work with members one-to-one, provide clear instructions, correct..
7 hours ago Master Trainer / Personal Trainer
LA Fitness/Pro Results
Personal Trainer Certification. Our ProResults Personal Trainers are true professionals, each possessing a certified Personal Training Certificate.
3 days ago Summer Camp Assistant Director
Kids in the Game
Our Core Coaching Values (Summer Camp Style):. In addition, KING operates summer camps, sports clinics (basketball, soccer, volleyball), and special events.
1 day ago Assistant Director of Coaching
HappyFeet Soccer
Enjoys physical activity, team sports and soccer. Do you love working with young children?
2 days ago


text:
Mark Fields was at the helm of Ford since mid-2014. Replacing Fields as Ford CEO is Jim Hackett, former CEO of Steelcase Inc. Hackett has led Ford Smart Mobility LLC since March 2016 and served on the American automaker's board from 2013 to 2016. Ford said in its official statement that Fields "is retiring," but […]+ Read More
Automakers are racing to achieve full-self driving autonomy and it looks like Ford wants to be the leader of the pack. To claim the top spot – and cars including those with no steering wheels or pedals by 2021 – it announced a $75 million investment in Velodyne, a leading light detection and ranging (LiDAR) sensors […]+ Read More


text:
Suggest An Event
If you have an idea for an event, please use the form below to let us know.
If you have an idea for an event, please use the form below to let us know.
The Society was founded in 1956 to foster friendship between the UK and the Republic of Korea, and its primary objective is to bring together all those in the UK who are interested in Korean matters. Read more
Candle Wave Feminists (시국페미) As part of the 2017 London Korean Film Festival, the British Museum is screening a short Korean documentary on 30th October at 2pm. This short-film focuses on feminist activities during the protests in South Korea in 2016. The screening will […]Read more
INVOCATION A visual Meditation of Condensed Memory Oil Painting inspired by Korean Ceramic Bowls and Moon Jars 24th Oct – 31st Oct 2017 Han Collection London 33 Museum Street, Bloomsbury, London WC1A 1LH www. hancollection.co.ukRead more


text:
Saturday, October 21st, 2017
Boga Hore
arimaha bulshada
Dabagalka Warar
English news
Wararka Maanta
Xulka Wararka
(Warbixin Cusub): Maxaa ka jira In Axmed Macalin Fiqi uu noqon doono Madaxweynaha Maamulka Galmudug
Warar lagu kalsoonaan karo ayaa sheegaya in Axmed Macalin Fiqi, uu ku biiray liiska Musharixiinta u taagan qabashada xilka madaxweynaha Maamuka Galmudug . Sababta Xildhibaan Fiqi uu ...
More barnaamijyo News
(Warbixin Xasaasi ah);- Donald Trump oo markii ugu horeysay Soomaaliya u aqoonsaday Iney tahay dalka….
Madaxweynaha Mareykanka Donald Trump ayaa Soomaaliya u aqoonsaday 'AAG DAGAAL' kadib wareegto uu soo saaray Arbacadii oo lagu qeexay arrintan. Madaxweyne Trump ...
(Xog Cusub):- Maxaa ka jira In Isbadal lagu sameynayo Golaha Wasiirada Cusub, Yaana ka masuul ah?..
Waxaa soo if baxaya xog ku saabsan wax ka bedel lagu sameynayo Golaha cusub ee wasiirada xukuumadda Soomaaliya ee shalay la ...
Golaha Wasiirada Cusub ee Soomaaliya Oo Maanta Kulankiii Ugu Horeeyay kaga Doodaya Qodobo Xasaasi ah
Golaha Wasiirada Cusub ee Soomaaliya ayaa manta kulankoodi ugu horeeyey ku yeelanaya Magaalada Muqdisho kaasi oo uu Shir Gudoomin-doono Ra'iisulwasaaraha Xukumada ...
(Xog Cusub);-Waa Maxay Sababta loo adkeeyay amaanka Villa Soomaaliya, Maxaase ka jira in Weerar lagu yahay?
Amaanka xaru nta Madaxtooyada Soomaaliya ayaa si aad ah loo adkeeyay , waxaana lagu soo kordhiyay ciidamo kale oo dheeraad ah ...
(War Cusub);-Ahlu Sunna Waljamaaca oo la safatay RW Kheyre Sharuudna ku xirtay ka qeybgalka Doorashada Galmudug
Maamulka dowlada gobolada dhexe ee Ahlu Sunna ayaa sheegay inay dalbanayaan in dib loo dhigo doorashada madaxweynaha Maamulka Galmudug . Waxay sheegeen ...
(Daawo Sawirrada):-Ra'iisal Wasaare Kheyre oo ka qeeybgalay Munaasabada Lagu Maamusayay Dulimaadka Tooska ah Ee Muqdisho iyo Nairobi
Ra'iisal Wasaaraha Xukuumada Federaalka Soomaaliya mudane Xasan Cali Kheyre ayaa maanta ka qeybgalay munaasabad kooban oo ka dhacday gudaha garoonka Caalamiga ...
(War Xiiso leh):-Saraakiil Kenyan ah ayaa Cadeeyey in DIyaaradoodu ay Weerar ku qaadeen Saldhigii Magaalada..
Kenya ayaa shaaca ka qaaday in duqeyn ay ku dishay Maleeshiyaad ka tirsan Al-shabaab, isla markaana ay ka furteen Hub ay ...
DEG DEG: Gudoomiyaha Aqalka Sare oo cadeeyay Mowqifkiisa Wasiirada cusub kana hadlay arinta uu wado Gudoomiye Jawaari
Gudoomiyaha Aqalka Sare ee dalka Cabdi Xaashi Cabdulaahi, ayaa ka hadlay diidmada Xildhibaanada Baarlamaanka qaarkood ay kala hor yimaadeen Xukuumadda cusub ...
(War Xiiso leh):-Madaxweyne Farmajo Oo xilki ka qaaday Madaxtooyada Ninka isku Dhawaayeen Madaxweynahi Hore Xasan Sh.
Wararka naga Soo Gaaraya Xarunta Madaxtooyada ee Villa Soomaaliya ayaa Shegaya in la Badalay Taliyihii Madaxtooyada Soomaliya General Ibraahim Sheekh Muxiyidiin ...
More »
This is the
and is ready for widgets,
click here
to add some now!
English News
Five Congo rangers killed in joint army operation to rescue U.S. journalist
July 17, 2017
Five park rangers were killed in a joint ...
Pakistan launches military operation in tribal areas targeting Islamic Stat
July 17, 2017
Pakistan's military has launched a major operation in ...
London Predatory Carnival On Somalia
May 9, 2017
LONDON: - I cannot think of any Somali ...
Guterres calls for lifting travel ban on Somalia
February 3, 2017
UN Secretary-General Antonio Guterres on Wednesday called for ...
Somalia on the brink of famine, U.N. Warns
February 3, 2017
The United Nations is warning that Somalia could ...
ICJ says can rule Somalia-Kenya boundary case
February 3, 2017
The United Nations' highest court ruled on Thursday ...
One dead as Soldier opens fire on bus in Mogadishu
February 3, 2017
A civilian is confirmed dead after a Somali ...
KDF convoy targeted in land mine blast near El Wak
February 3, 2017
A roadside bomb ripped through a military convoy ...
PM accuses neighbouring Nations of polls intervening
February 1, 2017
The outgoing Prime Minister of Somalia Omar Abdirashid ...
Kenyans among Shabaab fighters who attacked KDF
February 1, 2017
Kenyan security agents have established that hundreds of ...
Al shabaab attacks army convoy near Baidoa city
February 1, 2017
Al shabaab fighters ambushed a military convoy ferrying ...
A Kenyan man pleads guilty for trying to join Al shabaab
February 1, 2017
A 20-year-old man charged with attempting to sneak ...
Lawmaker calls for KDF withdrawal from Somalia
February 1, 2017
Balambala MP Abdikadir Aden has joined the list ...
Low flying jets spotted over Al shabaab-held bases
January 30, 2017
Residents in Lower Jubba region of southern Somalia ...
Fallen soldiers in Somalia's Kulbiyow attack laid to rest
January 30, 2017
Some of the soldiers killed in the attack ...
AMISOM kills 'a civilian man' in Southern Somalia
January 30, 2017
The African Union peacekeepers in Somalia have shot ...
Somali president meets 180 MPs in Mogadishu
January 30, 2017
President of the federal republic of Somalia HE ...
Jalalaqsi official accused of stirring internal conflict
January 30, 2017
The deputy district commissioner of Jalalaqsi town in ...
At least 7 injured in bomb explosion in Burane town
January 30, 2017
A police officer says seven people, including three ...
Somalia: Senior Al shabaab commander killed in Gedo
January 29, 2017
A Somali official says the country's military have ...
Copyright © 2012
All Rights Reserved.
Designed by :
Allpuntland24.com Ltd


text:
Find all your favourite pop culture brands and characters
Browse thousands of pre-loved items for sale, or sell something of yours
$30.00 - save 70%
from $5.00 - save up to 44%
from $2.00 - save up to 67%
$199.00 - save 20%
from $10.49 - save up to 30%
from $40.00 - save up to 33%
Delivering to:
It should arrive:
32.65% of people buy Pokemon TCG XY Shiny Kalos Tin: Xerneas-EX and Pokemon TCG XY Shiny Kalos Tin: Yveltal-EX.
Pokemon TCG XY Shiny Kalos Tin: Xerneas-EX
Celebrate the Power of Pokémon-EX!
Three great Legendary Pokémon-EX are revealed! Choose the forest majesty of Shiny Xerneas-EX, the destructive force of Shiny Yveltal-EX, or the battle-ready power of Zygarde-EX in the Pokémon Trading Card Game: Shiny Kalos Tin!
Each of these rugged tins contains a mighty Pokémon-EX from the Kalos
region—ready to battle and ready to win!
In this tin, you'll find:
Package artwork & contents randomly assorted.
5.0 out of 5 stars Based on 3 Customer Ratings
|5 star
|4 star
|3 star
|2 star
|1 star
Nobody has reviewed this product yet. You could be the first!Write a Review
Buy this and earn 144 Banana Points


text:
Common practices in other industries illegal; including brand protection
[caption id="attachment_17634" align="alignleft" width="121" caption="Jay Behmke"][/caption]
[caption id="attachment_17636" align="alignright" width="139" caption="Katja Loeffelholz"][/caption]
NORTH BAY -- The industry has been looking to consumer product promotional know-how from outside the business for fresh ideas, particularly as producers of higher-end wine look to move slow-selling inventory in an economy with high unemployment and trimmed expense accounts.
However, some marketing ideas for stimulating sales that would be perfectly legal in other industries are illegal in the highly regulated realm of alcoholic beverages, according to Jay Behmke, an attorney with Carle Mackie Power & Ross in Santa Rosa. Such promotional strategies include certain coupons, giveaways, contests, joint marketing with retailers and free shipping.
"You see 'free shipping' everywhere, but it is wrong," Mr. Behmke said.
In a June 2009 industry advisory on the need for third-party service providers to be licensed, the state Alcohol Beverage Control Department reiterated that premiums, gifts or free goods such as shipping costs are prohibited in the sale, distribution or marketing.
A common way of dealing with the restriction on free shipping is to reduce the bottle price of the wine by the cost of shipping and state that shipping is included, according to Mr. Behmke.
Free shipping is an increasingly important direct-sales promotional tool, particularly for a heavy product such as glass bottles full of wine. Free shipping increased from 31 percent of U.S. retail sales over the Internet in the first quarter of 2008 to 42 percent in the third quarter of last year, according to online sales tracker comScore of Reston, Va.
The legal line of what is allowed in marketing wine can be tricky to see because of the myriad of local, state and federal "tied-house" laws in the U.S. since the end of Prohibition. Such rules are structured to keep the retail part of the three-tier system independent from having too much influence over producers and distributors in promoting alcohol consumption.
Generally, suppliers such as wineries and distributors are restricted from directly or indirectly providing something of value to retail license holders such as stores and restaurants free of charge. It also reaches into content of and payment for store advertising as well as ownership relationships between the production, distribution and retail tiers.
But California as a major wine-producing state has a web of "tied-house" exceptions that can leave it unclear what is and isn't allowed, according to Mr. Behmke. He's had to dissuade clients from "tied-house" marketing ideas such as a contest with a trip to the winery for a club member who received a specially marked bottle and a retailer that wanted to allow a winery to pay to set up promotional displays in the store.
For details on what is allowed in wine promotions, contact the Alcoholic Beverage Control Department Trade Enforcement Unit at 916-419-2500 or www.abc.ca.gov/trade/tradeenforcement.html.
Effective marketing also should include brand protection, according to Katja Loeffelholz, a wine intellectual-property law specialist with Napa-based Gaw Van Male.
"In a down economy, your reputation is of great importance," she said. "People recognize your brand and develop feelings about it. From the consumers' standpoint, if they are going to be spending money to go with a brand known to be on the higher end, they want to know what they are getting."


text:
From The Verge:
Subscription music services have yet to profit but that hasn't stopped some of the internet's biggest media distributors from jumping into the business. Amazon, the web's biggest retailer, is the latest to inquire with various music companies about starting a subscription service, multiple sources have told The Verge.
Details are few and the talks have been described as very informal, the sources said. But so far, what Amazon has shown an interest in is an on-demand service that sounds pretty similar to Spotify, generally considered the sector leader. Others in the field include Rdio and Rhapsody, but Google and Apple are also working on their own projects. Google is reportedly in talks with record companies and music publishers about starting subscription music services for both YouTube and Google Play. The Wall Street Journal reported that Apple is in negotiations for a Web radio service that sources say would be similar to Pandora's, one of the top webcasters.
Continue reading the rest of the story on The Verge


text:
Red-zoned clifftop home splits in two
A house teetering on the edge of Christchurch's Richmond Hill is in "immediate" danger of collapsing and will be demolished today.
Part of the red-zoned house on Richmond Hill Rd split away from the rest of the quake-damaged structure today.
A Canterbury Earthquake Recovery Authority (Cera) spokesman said engineers had determined the house was in "immediate danger" of collapse and would be demolished this afternoon.
The demolition crew would work through the night under floodlight until the work was completed.
"The fear is that the forecast high winds tonight could collapse the house so they [are] taking precautions and demolishing it before then."
The decision to demolish the building was made "pretty much immediately", the spokesman said.
It was not yet known whether any neighbouring residents would have to evacuate while the demolition work was underway.
Cera bought the property under the Government's red-zone buyout offer in February 2013.
Kevin O'Connell moved out of the damaged property immediately after the February 2011 earthquakes.
He stayed in the Sumner area though and had observed his old property "quietly slumping" for some time.
"It looks like it's going to fall down the hill," he said.
A witness described seeing part of the earthquake-damaged building come away from the rest of the structure on Richmond Hill Rd today.
Colleen Taylor, of Clifton Hill, said police arrived in the area just after 12pm.
Taylor said she was "sick of having to look at... munted" buildings teetering on cliff edges in Sumner and Redcliffs following Canterbury's earthquakes.
"If they don't do something about them someone's going to get injured."
Clink Restaurant owner Rachel Lonsdale said it looked as if the house could fall down the cliff. Her restaurant was not in danger, she said.
The Press


text:
Wildwood Earrings
Wildwood Earrings
from 55.00
◆Solid cast bronze earring with 14K gold-filled earwire or Sterling Silver. Metal is nickel-free.
◆Earrings measure 3" long
◆Silver ships in 3 business days. Bronze are made to order- allow 4 weeks to ship.
◆All items come packaged in a beautiful box, perfect for gift-giving!
Add To Cart


text:
Combine either an Upper Pigeon or Lower Pigeon Adventure at River Rat Whitewater Outpost with an exciting ride on Foxfire Mountains' one and only Bear Crawler Adventure of the Smoky Mountains. Experience nature from an extreme 8-wheeled drive ATV on rugged off-road trails dodging trees and riding through water to make it to the top of the mountain.$90.00 +tax and $2 county river fee Book Now
River Rat Whitewater Outpost
3630 Hartford Road
Hartford, TN 37753Foxfire Mountain Adventure Park
3757 Thomas Lane
Sevierville, TN 37876
- Adventure Duration
Duration is 1.5 to 2 for rafting and 1 hour for crawler.
- Included
Rafting on either Upper or Lower Pigeon River, Bridge to Prosperity, and crawler.
- What to Bring
A quick drying swimsuit or shorts will work best. You will also need shoes that stay securely on your feet (tennis shoes, water shoes, or sports sandals) flip flops are not allowed on the river. Remember a change of clothes and a towel! Secure footwear for ziplining must be closed toe.


text:
Villa Rica, GA (8 miles from Carrollton)
Looking for the ultimate getaway? This retreat is in the metro Atlanta area, yet far enough to relax
Villa Rica, GA (8 miles from Carrollton)
Overall rating:
The Carrollton B&B review score is based on feedback from travelers from around the world. See property page for full, detailed reviews by real travelers.


text:
"Consumerism" is a track coming out on the eve of Ms. Hill's release. She wanted to get this music out while she was incarcerated, as it is a product of the space she was in while she was going through some of the challenges she has been faced with recently.
""Consumerism" is part of some material I was trying to finish before I had to come in. We did our best to eek out a mix via verbal and emailed direction, thanks to the crew of surrogate ears on the other side. 'Letters From Exile' is material written from a certain space, in a certain place. I felt the need to discuss the underlying socio-political, cultural paradigm as I saw it. I haven't been able to watch the news too much recently, so I'm not hip on everything going on. But inspiration of this sort is a kind of news in and of itself, and often times contains an urgency that precedes what happens. I couldn't imagine it not being relevant. Messages like these I imagine find their audience, or their audience finds them, like water seeking it's level." –Ms. Lauryn Hill
You can cop her "Consumerism" track on iTunes!!!


text:
627Re: [Families of Angelman Syndrome] Digest Number 215
Expand Messages
- Sep 1, 2005I'm sorry I have not responded to the new comers.. Welcome. I am a mother of twin Angels. they are now 8 and was diagnosed at 4. They 1st said they had CP. Our Angels are pretty good boys, they have pretty much everything that is Angelman Syndrome. I have to say that some of you must be having better luck at getting your kids to walk or even some words. mine to crawl and scoot, but no walking NO words. they do alot with their hands,, everything to the mouth:) LOL seizure are pretty good this year its BEEN so long I can't remember their last ones... Meds are pretty much controlling them. I only know what my kids do and have. I really don't know ALOT about AS. But, the things I find doesn't look like anything new, but we know we have good kids that will be with us forever and that's a BIG PLUS:) I always look at the bright side and my kids are pretty healthy. everyone take care and big hugs to all your Angels...
Michelle, Kabel, & Kyle... Indiana.
Start your day with Yahoo! - make it your home page
[Non-text portions of this message have been removed]


text:
Household Hazardous Waste and Paper Shredding
November 4th 8:30am-2pm
Delaware Recycling Center, New Castle
Click here for more information
As a part of a Department of Natural Resources and Environmental Control (DNREC) permit condition for all three DSWA landfills in Delaware. There is a yard waste disposal ban in place for all DSWA facilities.
This means yard waste cannot be disposed of in the landfill. Yard waste that is separated and not mixed with trash can still be brought to DSWA Landfills and Transfer Stations where it will be placed in a separate area for recycling.
Yard waste includes:
- Grass
- Leaves
- Pruning
- Brush
- Shrubs
- Garden Material
- Christmas Trees
- Tree Limbs up to 4 inches in diameter


text:
THE FOOD SEEN 0921617: Soli Zardosht, Persianesque
On today's episode of THE FOOD SEEN, Soli Zardosht captivates us with stories of her Iranian upbringing, and Persianesque cooking. With years as a menswear fashion designer, her critical eye brings creativity to the plate, reinventing the form of classic dishes like Kookoo Sabzi, a green herb frittata, Kufte Tabriz, a fruit & nut filled meatball, and the epic centerpiece, Tahdig, a crispy rice casserole that's inverted onto a platter with a signature "flip". It's certainly about the food, but more so, the presentation of herself, and her heritage's cuisine, in a new light, that's bright and beaming, in efforts of bringing the warm glow of Persian culture to the world.
***Listen to THE FOOD SEEN every Tuesday 3PM EST on HeritageRadioNetwork.org
... and download the podcast via iTunes, http://itunes.apple.com/us/podcast/the-food-seen/id380318273?mt=2


text:
Take £10 Off On All Purchases. Voucher code is required to grab the offer.
Take 10% Off On Site wide With Free Shipping For Total Of $50.
Toys and Games from £1.99 at Totally Funky.
Bestsellers from £3.99 at Totally Funky.
Free Delivery on Orders Over £30 at Totally Funky.
Totally Funky stocks a huge range of fun, funky and unusual gifts. We strive to find innovative and unusual products that tick all the right boxes - quirky, humourous, design-led, value for money - products that, ultimately, will make great gifts!
Copyright (c) 2017 CouponMount.com. All rights reserved.


text:
Tuesday, July 6, 2010
Fourth Of July Party
30 comments:
- FEDERICA said...
Oh wow! Stunning!!
Have a nice day,
- July 6, 2010 at 5:16 AM
- Kathy @ Creative Home Expressions said...
It looks like a beautiful home, Sherry! Your fireworks pictures are amazing. We were inside for most of the day as it was so humid here.
- July 6, 2010 at 7:06 AM
- the VG huis said...
Hi Sherry, Oohhh their home is very beautiful! Can't wait to see the posts on the inside. I could probably use a sugar headache to hold me back sometimes. I am in love with your metal caddy in your header picture! Have a great day! ~Stacey
- July 6, 2010 at 7:43 AM
I don't know why but blogger has some problems today, I am sorry if you left a comment, it doesn't appears! I have left you a comment this morning but I don't see it!
Sweet as a Candy
- July 6, 2010 at 8:06 AM
- Kim @ Savvy Southern Style said...
Sherry, what a beautiful home. Can't wait to see more. How neat to see those fireworks so close like that and you didn't have to drive and get stuck in traffic.
- July 6, 2010 at 8:56 AM
- Happy To Be/ Gl♥ria said...
Good Morning Sherry, oh girl I love the fireworks and they are the only ones I seen this year...I really enjoyed your post today...Thanks for sharing...Hope you have a GREAT day my friend..Hugs and smiles Gl♥ria
- July 6, 2010 at 9:21 AM
- Days at Buttermilk Cottage said...
Lovely home. How lucky to be so up close to the fireworks!
- July 6, 2010 at 10:06 AM
- Suzanne said...
Wow - What beautiful fireworks. That must have been a fabulous day. Love that photo of your grandson with his friend -- so cute!
- July 6, 2010 at 10:07 AM
- Linda @ A La Carte said...
Wow those are some gorgeous fireworks! I love watching them like that. Sounds like a wonderful 4th!
- July 6, 2010 at 10:59 AM
- Terrell said...
Oh what a fabulous time!! I love the pic of the little guys looking on at the show! Precious!! Thanks for sharing!! :D
~Terrell @ FrouFrouDecor~
- July 6, 2010 at 12:35 PM
- This Photographers Life (Architecture+Interiors) said...
That sounds like a wonderful holiday! Your grandson is so cute... and I can't believe you were only 200 yards away from the show! Very cool.
- July 6, 2010 at 1:04 PM
- Penny @ The Comforts of Home/Lavender Hill Studio said...
p.s. Love the way you are using your vintage french wine carrier in your header!
- July 6, 2010 at 7:55 PM
- Penny @ The Comforts of Home/Lavender Hill Studio said...
Sounds like it was a wonderful time! This is my first visit and I am off to see the rest of your previous posts.
- July 6, 2010 at 7:55 PM
- The Polka Dot Closet said...
Thanks for visiting, what a show you had, with friends like that who needs relatives!!
- July 6, 2010 at 8:58 PM
- Pam @ diy Design Fanatic said...
Cool photos! I love the look of wonder on your grandson's face!
- July 6, 2010 at 10:26 PM
- laurie @ bargain hunting said...
What a gorgeous home, and your fireworks photos are wonderful. laurie
- July 6, 2010 at 11:54 PM
- Fleur de Bee said...
I love when photos of fireworks turn out like this. So fun! Can't wait to see photos of their stunning home!! xo Molly
- July 7, 2010 at 3:59 AM
- Leslie @ goodbye, house! HELLO, HOME! said...
Kids are always SO fun to watch while they are watching fireworks!!
Looks like you had a lovely day!
(I probably would have tasted each dessert, too, LOL! Headache Shmeadache)
Hugs, sweet thang!
- July 7, 2010 at 1:21 PM
- June said...
Sherry the house looks gorgeous! I am so glad you had a wonderful holiday. It sounds lovely...sugar headache would have been worth it for me.
- July 8, 2010 at 4:14 PM
- Susie from Bienvenue said...
What a beautiful home!!! I cant wait to see more...hint hint :)
- July 8, 2010 at 10:47 PM
- Nancy said...
Hi Sherry,
Sounds and looks like you had perfect seats at a spectacular fireworks show. Seeing your Grandson and his friend watching is adorable. It's as fun watching the kids as it is the show itself!
Your sons home looks beautiful. Can't wait for the inside tour! ;-)
- July 9, 2010 at 1:53 AM
- Nancy said...
p/s Sherry, I've not commented much over the past couple of weeks due to lack of time but wanted to let you know that I love your header. The wine basket carrier and your butter tray are wonderful and look great!
- July 9, 2010 at 2:02 AM
- HOOTIN' ANNI said...
Love, love love the fireworks display, and the children's awe of it all!! That is what it's all about. Family.
LEGEND OF MARY'S BEAN is my Friday blog.
Have a great weekend.
- July 9, 2010 at 6:17 AM
- Privet and Holly said...
Great fireworks pictures!! They are very hard to take well, so you must be a pro with your camera : ) That house looks amazing and could be a post in itself! Thank you so much for your kind comments on my giveaway post...I'm having so much fun collecting suggestions! I see an afternoon of ice tea and pendant light shopping in my future, based on recommendations from my blog friends. Hope you have a wonderful weekend, Sherry! xx Suzanne
- July 9, 2010 at 7:35 PM
- xinex said...
Truly spectacular fireworks! I love your son and DIL's house. I cant wait to see it inside...Christine
- July 9, 2010 at 10:35 PM
- Atticmag said...
Sherry, those photos are spectacular. What fun that must be for kids to see (big kids too;) )
Jane T.
- July 9, 2010 at 11:43 PM
- Debbie@Debbie-Dabble said...
WOW!! What great photos of those fireworks!!
- July 10, 2010 at 8:46 AM
- The Quintessential Magpie said...
What wonderful fireworks shots and darling kids!
Looks like you had a fun Fourth!
I'm dropping by to wish you a Happy Pink Saturday...
Sheila :-)
- July 10, 2010 at 9:00 AM
- Tricia said...
Welcome to Pink Saturday, Sherry! The picture of those two little boys watching the fireworks is just priceless. What fun! Children bring such a wonderful perspective to our busy lives, don't they?! And some of those fireworks just look like sea creatures - Great photos. Have a great day.
- July 10, 2010 at 6:15 PM
- Beverly said...
Welcome to your first Pink Saturday, Sherry. I am glad you decided to join us.
Your son's home is lovely. It looks like you had a wonderful Fourth of July holiday with your family.
- July 11, 2010 at 9:15 AM


text:
Celebrating her 42nd birthday, Naomi was spotted out and about with Liev in New York City last night (Sep 28) and looked to be enjoying themselves as they strolled through the trendy SoHo neighborhood before going for dinner and drinks as the paparazzi looked on. Naomi seemed to be very happy with the occasion and couldn't resist smoking a cigarette which she had given up a long time ago prior to her first pregnancy.
*More photos including daytime after-shoppin@Barney walk with friends are on the forum by clicking [here].
This Blog, established since Dec 2001, is a place dedicated to the talented and beautiful Hollywood actress Naomi Watts. All images and videos published here are owned by their respective owners or photographers. No copyright infringement whatsoever is intended or implied. The owner of this Blog is not associated with the actress or anyone around her. This Blog is non-profit making and is operated purely for personal interest. We welcome all discreet and well-intentional comments and feedback. (Note: this site is best viewed with Firefox)
Sep 29, 2010
Naomi's birthday shopping & romantic night out
Posted by Richard Steandric Ricsteand
Labels: Sightings
3 comments:
She looks totally drunk but I can't blame her : wine and cigaret match quite perfectly together. Well, it's so weird to see her like that!!!
- September 29, 2010
She looks happy and very much in love.
- September 29, 2010
Thank you for Naomi.
- October 01, 2010
Subscribe to: Post Comments (Atom)


text:
Email me
Website Designed by
Himanshu vyas
© Copyright 2013
Self taught graphic designer with a passion for logo design. Having experience in creating fun and memorable logos while retaining the simplicity.
Also worked as freelance Photoshop article/tutorial writer, having written 250+ quality articles on using Adobe Photoshop which were published in premium tutorial directories like good-tutorials and many more.
Always interested in freelance work and collaborations.


text:
One of the hardest thing I have not been able to do since I was much younger even till now is to take my stand in decisions, situations and matters. I am this very respectful person that tends to give honour to whom honour is due. Respect is reciprocal but even if I am not respected, as long as you are older in age or experience, I will always give you the honour you deserve. In so many ways it is a good thing and in so many ways, it has brought about a bit of demerits.
There is this one case that has been coming up and down because of my indecision. Today, I will say YES and tomorrow, I will say NO. Unknown to me, it broke some people's trust in me because they saw me as someone with many mouth but that was not the situation. The situation is that there are so many older people involved in this matter and if I will have to stand my ground, they will be affected and feel very bad and you know I wont like that.
It all started out last year when an issue came up and we were asked to do some marketing. Many people couldn't achieve that because the products we were asked to market were quite expensive to students on campus and all. Notwithstanding, we were asked to deliver the money for the sales and I was like
'Nobody is even buying it. How would I then deliver money for it?
We were given deadlines and all and people started using their personal money to deliver but I was still optimistic that I would still sell mine because I have an edge when it comes to marketing and sales. Many people know me for that but this one was really difficult. I had to tell the owner dates upon dates when I would present the turnover but I never sold any of the products by then. At this time, almost everyone had presented their personal money as money for the products.
Then, we were asked to bring some people for the launching of the product and I volunteered to bring some important personnel from my school. I guess I made a big mistake. I should have just kept mute. I kept reminding these people and on the D - DAY, something came up and none of them showed up. AHA! And that was how I got the embarrassment of my life from the owner and all. Make I no lie, I almost cried because they assumed I was insincere and lied to them and this hurt me to my bone marrow. I don't like when people make wrong assumptions about me and when I tried to explain, it seemed like I was saying jargons.
However, I have always said I would leave that place and all but I always thought about what the owner would say. He will feel bad that I have gone and I didn't want that. But then, a lady said to me
'Its your reputation first before any other thing'
I was still reluctant to leave until recently when I went there and the owner's wife trashed me with her words for not coming to the place on Sunday. They call it knocking but I took cognisance of each of her words such as
'Don't come near me!'
'Something is wrong with you!'
'I hope you will be able to get that your inheritance back!'
'Leave my front!'
This was just the bombshell and I realized I have been dulling myself all along. I respect this woman so much and I still gave her the honour due to her but with these words, its a good bye because no one can just stand to make me feel bad. I am what God says I am not what man says I am. In trying to please and respect others, we shouldn't push our feelings aside so that we can make others feel good while we are made to feel inferior. Any statement that God would never say to me is devoid of the word of God and as Christians, we should never accept such because our life is built by the words we confess and accept.
So, I have finally taken my stand and the new place I attend is such a great place and we are like a family. You can always make explanations for everything so people don't assume wrongly. This experience has taught me a lot of things. People wont appreciate you till you take the bold step by taking your stand.


text:
Công Nghệ Thông Tin
Quản trị mạng
Tải bản đầy đủ
Chapter 2. Getting Started with Azure PowerShell
Chapter 2. Getting Started with Azure PowerShell
Tải bản đầy đủ
Authenticating to Microsoft Azure
You have two choices for authenticating to Microsoft Azure from PowerShell. You can
use your Microsoft Azure username and password with support for a Microsoft or an
Organization account in the Azure Active Directory, or you can use certificate-based
Authenticating with a certificate
The easiest way to get started with certificate authentication is to download a .publish‐
settings file from Microsoft Azure by using the Get-AzurePublishSettingsFile
cmdlet. This cmdlet launches the default browser and takes you to a page on the Mi‐
crosoft Azure site where you can log in with a Microsoft or Organization account that
has access to your Microsoft Azure subscription. When you have successfully logged
in, you will be prompted to select a subscription if your account has access to more than
one and then prompted to download a .publishsettings file.
To execute, press F5, or highlight the call to the Get-AzurePublishSettingsFile cmdlet
in the editor and press F8 (see Figure 2-1).
Figure 2-1. Executing Get-AzurePublishSettingsFile
About the .publishsettings file
The file you download should be treated with care. In the file is the name of your sub‐
scription, subscription ID, and a newly-generated management certificate that allows
you to access the subscription. Whoever has access to this file has access to your sub‐
scription. Microsoft Azure imposes a limit on the total number of management certif‐
icates that can be associated with a subscription at any given time.
At the time of this writing, the maximum number of certificates is 100. Each time you
run the Get-AzurePublishSettingsFile cmdlet, Microsoft Azure generates a new
Chapter 2: Getting Started with Azure PowerShell
management certificate in the subscription you choose. If there are multiple users on a
subscription, you should develop a certificate management strategy early on to avoid
problems later.
Importing the .publishsettings file
The next step in configuring the Microsoft Azure PowerShell cmdlets is importing the
previously downloaded .publishsettings file. As I mentioned earlier, this file contains a
management certificate that allows access to your Microsoft Azure subscription. The
cmdlets use this certificate for authentication to the Service Management API.
To import, simply add a call to Import-AzurePublishSettingsFile and pass to it the
path to the previously downloaded file (see Figure 2-2). Press F5, or highlight the text
and press F8.
Figure 2-2. Importing a .publishsettings file
Using Microsoft Azure AD to authenticate with PowerShell
An alternative method to using certificates is to authenticate using an account from the
Microsoft Azure Active Directory. Each new Microsoft Azure subscription will have its
own Active Directory tenant by default. From a PowerShell perspective, this means that
you are not required to use management certificates to authenticate and access your
Using the Add-AzureAccount cmdlet, you can specify the username and password of a
user who has administrative or co-administrative rights on your subscription, and use
the returned token to execute PowerShell commands with your subscription (see
Figure 2-3).
Setting Up Your Environment
Figure 2-3. Using Add-AzureAccount to authenticate
The token returned from Add-AzureAccount is valid for up to 12 hours. After the token
expires, you will need to authenticate again by running Add-AzureAccount and entering
your username and password. This is not ideal for scripts that need to run in a purely
automated fashion without user intervention of any kind. For noninteractive scripts,
the Add-AzureAccount cmdlet supports passing a PSCredential object to the
-Credential parameter. At the moment, this support works only with organizational
Switching back to certificate authentication
When you use the Add-AzureAccount cmdlet, all of your subscrip‐
tions for that account will be modified to use Azure AD authentica‐
tion. If you want to switch back to using certificates, you will have to
remove the account settings first by calling Remove-AzureAccount.
Managing Subscriptions
Once you have downloaded and imported your subscription settings (or authenticated
using your username and password), there are several other cmdlets you should be
aware of that are involved with managing your subscription settings in PowerShell.
The Get-AzureSubscription cmdlet returns and enumerates subscriptions that have
been imported or manually configured with the Set-AzureSubscription cmdlet. These
settings are persisted in the $env:APPDATA\Windows Azure PowerShell folder.
Get-AzureSubscription also supports the parameters listed in Table 2-1 to help you
identify subscription settings.
Table 2-1. Get-AzureSubscription parameters
Returns the default subscription. When you start a new PowerShell session, this will be the subscription used
if no other subscription is selected.
Returns the currently selected subscription.
Chapter 2: Getting Started with Azure PowerShell
-ExtendedDetails Returns quota details for the current or specified subscription.
The -ExtendedDetails parameter is especially useful for ensuring that you have enough
quota available in your subscription for whatever operation you are automating (see
Figure 2-4).
Figure 2-4. Viewing quota information with Get-AzureSubscription
At runtime, the cmdlets have a concept of the current subscription selected in your
PowerShell session. This functionality allows you to execute scripts using multiple sub‐
scriptions. For instance, you could write a script that enumerates all of your subscrip‐
tions and deletes unused disks in each of them or stops all virtual machines. The cmdlet
to switch between subscriptions is Select-AzureSubscription (see Example 2-1).
Simply call the cmdlet with the subscription name you want to work on, and any new
calls to Azure will use this subscription.
Example 2-1. Switching between multiple subscriptions
Select-AzureSubscription "[subscription one name]"
# returns the status of all VMs in subscription one
Select-AzureSubscription "[subscription two name]"
# returns the status of all VMs in subscription two
This cmdlet can also be used to change the current and default subscriptions for your
PowerShell sessions with the parameters in Table 2-2.
Table 2-2. Select-AzureSubscription parameters
Changes the subscription specified to be the new default subscription for all PowerShell sessions.
Changes the subscription specified to be the new current subscription for the active PowerShell session.
Setting Up Your Environment
-NoDefault Clears the default subscription settings from all PowerShell sessions.
-NoCurrent Clears the current subscription settings from the active PowerShell session.
The Set-AzureSubscription cmdlet allows you to add a subscription to the stored
settings or change properties on an existing subscription.
Example 2-2 shows how to associate a manually created certificate and associate it with
a Microsoft Azure subscription. The same call could be used to modify an existing
subscription by changing the certificate associated with the subscription.
Example 2-2. Manually configuring a Microsoft Azure subscription
$cert = Get-Item Cert:\CurrentUser\My\[certificate thumbprint]
$subscriptionID = "[your subscription ID]"
$localName = "[manually added subscription name]"
Set-AzureSubscription -SubscriptionName $localName `
-SubscriptionId $subscriptionID `
-Certificate $cert
Manually creating and uploading management certificates
If you would like to manually create and manage management cer‐
tificates, simply use the makecert.exe utility as documented in MSDN
http://bit.ly/use_makecert_utility and upload the certificate through
the management portal. You can also view the certificate thumb‐
print in the portal user interface.
Just as you can add and update a subscription, you can also remove the subscription
from your local PowerShell configuration by calling the Remove-AzureSubscription
cmdlet (see Example 2-3).
Example 2-3. Removing a Microsoft Azure subscription
$subscriptionName = "[subscription name]"
Remove-AzureSubscription -SubscriptionName $subscriptionName
Executing Scripts in This Book
It may take several lines of script when using the Microsoft Azure PowerShell cmdlets
to execute a task. During these times, I find it is simpler to add the lines to a single script
and use the PowerShell ISE to execute the entire script at once (see Figure 2-5).
Other times you may want simple environment information from Microsoft Azure such
as the name of the available regions or a list of storage account names in your
Chapter 2: Getting Started with Azure PowerShell
subscription. For these one-line operations, I prefer to execute the scripts using the
PowerShell console (the Console pane of the PowerShell ISE works well too) and then
use the values within the script that I am building.
Throughout this book are examples that I recommend you try as learning exercises and
others that are just for reference. In the learning exercises, I will note when I am using
the Console pane to execute a command and when I am building a new script by noting
that the code should go in the Script pane (see Figure 2-5). You may, of course, do this
however you like, but if you are new to PowerShell, I hope these tips will help guide you
along the examples throughout the book.
Figure 2-5. The PowerShell ISE
Executing script with F5 versus F8 in the PowerShell ISE Script pane
As you progress through the book, you will be asked to execute code
in several ways. Within the Script pane are two primary methods that
you will use in this book. Pressing F5 in the Script pane executes the
entire script that is loaded. Pressing F8 executes only the script code
that is currently selected or the line that the cursor is on. Sometimes
you should execute the entire script with F5 and sometimes only the
selected portion with F8.
In this chapter we have seen where to download the cmdlets from and how to configure
one or more Microsoft Azure subscriptions. In Chapter 3 we will dive right into doing
something useful with the cmdlets, starting with creating and configuring virtual
Virtual Machines
Creating Virtual Machines with PowerShell
In this chapter you will learn about using the Microsoft Azure PowerShell cmdlets to
create a virtual machine with Microsoft Azure platform images. As part of learning this
process, you will learn how to specify the initial configuration settings such as the local
administrator account name and password, the virtual machine size (CPUs and mem‐
ory), network endpoints, and underlying storage. From there you will learn how to use
those same concepts to modify the configuration of existing virtual machines, whether
they are running or not.
To get started creating your first virtual machine using PowerShell, you will need some
environment information from Microsoft Azure. This is the same information that you
use in the portal, such as the region name and the storage account that will be used as
the location where your virtual machine disks are created.
For the first part of this chapter, I would recommend creating a new PowerShell file and
saving it with a name such as chapter3create.ps1. Some portions of this chapter will be
saved to the script and edited in the Script pane (top portion of the ISE) to make it easier
to follow, and some portions should be executed in the Console pane (bottom portion
of the ISE) for immediate results.
The first call related to Microsoft Azure of your new script should always be to SelectAzureSubscription to ensure that you are executing commands against the correct
Microsoft Azure subscription.
Add the code shown in Example 3-1 to create a variable to store your subscription name
and then select that subscription for use. Ensure that you replace the placeholder values
with real ones.
Example 3-1. Selecting your subscription (Script pane)
$subscription = "[subscription name]"
Select-AzureSubscription $subscription
Replacing the subscription name placeholder
Remember, you can find the name of your subscriptions by calling
Get-AzureSubscription | Select SubscriptionName in the Con‐
sole pane. Use that value instead of the placeholder value in the ex‐
ample. The subscription name is case sensitive!
Executing the script
When the call to Select-AzureSubscription is in place, press F5, or highlight the script
and press F8, to select your current subscription. This will validate that you have the
correct subscription name in place and will also set any future commands run from the
Console pane to that subscription.
Virtual Machine Location and Storage
All resources in Microsoft Azure are created in a specific region. This is the same region
that you see in the management portal when you create a virtual machine.
To retrieve a list of available regions, you can run the Get-AzureLocation cmdlet (see
Example 3-2). Since the goal is to have an immediate list of available names and not run
these each time you execute this script, I would suggest you run this command in the
Console pane of the PowerShell ISE (see Example 3-2).
Example 3-2. Returning Microsoft Azure location details (Console pane)
A few properties of the output shown in Figure 3-1 are very important for provisioning
virtual machines:
• AvailableServices
• Name
You can create a virtual machine only in locations where the AvailableServices list
contains PersistentVMRole. The AvailableServices list can also contain a HighMemo
ry value. This denotes locations where the A5, A6, A7, A8, A9, and future high-memory
virtual machine configurations are available to be provisioned. The Name property is the
value you will use to specify the location during the creation of resources.
Chapter 3: Virtual Machines
Figure 3-1. Using Get-AzureLocation
After you have determined the region in which to create virtual machines, you can store
the name of the region in a variable for later reference. Add the code in Example 3-3 to
your script to store the region name.
Example 3-3. Storing the region name in a variable (Script pane)
$location = "[region name]"
The next step is to specify the storage account where the virtual machines will be created.
The storage account must be in the same region as the virtual machine. This is enforced
at the API level so you do not accidentally end up in a situation where your virtual
machine is running on the West Coast of the United States but the underlying disks are
in Europe!
The first option is to enumerate your existing storage accounts for a suitable storage
account. The command in Example 3-4 will enumerate all of the storage accounts in
your subscription but return only the StorageAccountName and Location properties
(see Figure 3-2).
Example 3-4. Enumerating existing storage accounts (Console pane)
Get-AzureStorageAccount | select StorageAccountName, Location
If you do not have a storage account available, or you just want to create a new one, use
the New-AzureStorageAccount cmdlet.
To ensure the availability of the Microsoft Azure storage account name, you should use
the Test-AzureName cmdlet first (see Example 3-5). Test-AzureName verifies whether
Creating Virtual Machines with PowerShell
the name is available for your use in Microsoft Azure. Make sure you replace the [stor
age account name] placeholder in the script before executing!
Figure 3-2. Using Get-AzureStorageAccount to enumerate storage accounts
Example 3-5. Finding a unique storage account name (Console pane)
Test-AzureName -Storage -Name "[storage account name]"
If the call returns True, the storage account name already exists and is not available to
you. Run the command with a new name until the call returns False, which means that
the storage account name is available. Create the new storage account as shown in
Example 3-6.
Storage account name
The name of your storage account must be unique within Azure.
Storage account names must be between 3 and 24 characters in length
and use numbers and lowercase letters only.
Example 3-6. Creating a new storage account (Console pane)
New-AzureStorageAccount -StorageAccountName "[storage account name]" `
-Location $location
When you determine the name of the storage account to use, save it in a variable for
later reference. Add the code in Example 3-7 to your script to store the storage account
Example 3-7. Specifying the current storage account (Script pane)
$storageAccount = "[storage account name]"
The next step is to associate the storage account with the subscription you are using by
specifying the name with the -CurrentStorageAccountName parameter in the SetAzureSubscription cmdlet. Once set, any PowerShell cmdlets that create virtual ma‐
chines or deploy cloud-service packages will use this storage account as the default.
Chapter 3: Virtual Machines
Tài liệu liên quan
Automating Microsoft Azure Infrastructure Services
Tài liệu mới


text:
Note : Your Quotation Request will be directed to Norden Millimeter.
The NA4060G15N7P7 from Norden Millimeter is a RF Amplifier with Frequency 40000 to 60000 MHz, Gain 15 dB, Gain Flatness ± 1.5 dB, Noise Figure 7 dB, P1dB 7 dBm. More details for NA4060G15N7P7 can be seen below.
Power Amplifier from 20 to 45 GHz
LDMOS Two-Stage Integrated Doherty Amplifier
You can now find similar products from multiple companies on everything RF.


text: Do you count AEC,U,M,A,P,As updated homages or just classics/generations. The following letters above are abbreviated for the following TF toy lines: Armada,Energon,Cybertron,Universe,Movie-verse,Animated,Prime. Upgraded homage new TF molds,Do you count non-classics styled homages from AEC,Universe,movie verse,animated,Prime,etc... Personally I think the non-classics TF toy lines make better homages. Technically,the classics/generations/RTS TF toy lines are not set in the generation one continuity universe. classics/generations/RTS is basically a alternate universe based off G-1. I actually think their are some better updated homages in the NON-classics toy lines. Examples: lets not forget that the two best planet formers toys came from AEC. Despite the toys limited articulation,I actually think energon ultra & leader sized galvatron are better homages than classics 2.0 galvatron. Titanium 6 inch toy line created a few awesome updated homages that beat the classics homages. titanium scourge,rodimus prime,G-1 ultra magnus are way closer in homages than their classics counterparts toys. I also think the Energon voyager sized Dreadwing toy is a excellent updated homage mold for the 1986 G-1 Scourge character/toy. even though the toy isn't named Scourge. the movie verse did a few decent homages that simply look better than the classics/generations/RTS toys. like ROTF Deluxe Dirge. a few of those deluxe & voyager sized constructicons toys from ROTF. Even animated's toy line I feel created more closer updated homage new toys compared to classsics/generations/RTS. Examples: Delixe animated blurr & deluxe arcee are closer updated homages based off the 1986 movie/season designs compared to classics/generations/RTS. I actually love Animated leader sized Megatron. because it's a updated homage based off the Takara TF 1991 battle stars/return of convoy Super Megatron art designs images. The animated deluxe sized Swoop toy,I think is the best homage 1985 swoop toy design thus far. Energon Ultra size Scorponk is a awesome updated homage based off the 1987 Scorponok toy designs. so is the Cybertron dark scorponok a awesome homage recolor based off the masterforce blackzark toy colors/designs. I felt the ROTF Bludgeon & brawn were the closets updated homages the 1980's toys will ever receive. Armada Scavenger is a cool homage based off the Generation 2 Auto-rollers gimmicks/designs. I loved those Energon 5-in-1 combiners. they were awesome updated homages based off the scrable city 1980's TF combiners. the toys were the proper size & in tru scramble city style,the limbs were inter-changeable. The takara/Hasbro Masterpiece TF toys,I consider the best & most accurate updated homages new molds thus far. Alternators,I felt gave us a decent updated homages for 1985 tracks,hound,swindle & Prowl. Beastmachines/Botcon Ultra sized Cyclonus toy mold,is a awesome updated homage based off the 1986 cyclonus toy. I just think the updated homages are better in the non-classics/generations/RTS toy lines. because the toy budget money is higher. the toy sizes vary more & have more bigger sizes.


text:
There's a new field ops on Club Penguin, new stealth gear, and a big EPF launch coming.
Start with the field ops (April 2011), which is only a drill. It's supposed to be in a everyday object. Well, it is.
Go to the Coffee Shop and click on that small tree thing:
Next, click on your beeping EPF phone and do the field ops.
It is very easy: just click on the right shape, the one displayed on the right before the timer runs out.
When you're all done, you will see Dot's message:
Cool, Dot! Talking of Stealth, CP's Stealth gear just came out:
About the big EPF launch, we're not so sure about it. On the photo of CP's annual planning whiteboard, it said it, so we thought it could be cool to tell you about it. More info soon…


text:
Thomas, T.G. and Williams, J.J.R. (1997) Development of a parallel code to simulate skewed flow over a bluff body Journal of Wind Engineering and Industrial Aerodynamics, 67-8, pp. 155-167. (doi:10.1016/S0167-6105(97)00070-6).
This paper outlines a new complex geometry large-eddy simulation code using finite differences and a multigrid Poisson solver written for a parallel computer. The flow domain may be constructed from an arbitrary arrangement of rectangular blocks thus permitting flow in regions with complicated shapes, and may be mapped to any number of processors up to the number of blocks. The code is used to simulate turbulent flow at a Reynolds number of 3000 past a cube placed at ground level in rough terrain with its sides set 45° to mean flow direction. Preliminary results are presented which reproduce the conical vortices on the top of the cube.
Full text not available from this repository.
More information
Catalogue record
Export record
University divisions
Download statistics
Downloads from ePrints over the past year. Other digital versions may also be available to download e.g. from the publisher's website.


text:
J-375™ | Jacuzzi Hot Tub
- Seats: 5-6
- Dimensions: 91 in x 91 in x 38 in
- Filled Weight: 5,734 lbs. / 2,601 kg
- Spa Volume: 470 US gal. / 1,779 Liters
- Circulation Pump: YES
- Jets: 50
- Lounger: YES
- Pumps: 3
- Water Purification: CLEARRAY®
- Waterfalls: 2
- Pillows: 4
- Stereo: OPTIONAL
- Electrical: 240 VAC 60 Hz 40A, 50A or 60A
- Monthly Operating Cost: $15.91
The J-375: A Luxurious Hot Tub With a Wide Range of Features
The J-375 is the go-to-model if you're looking for a luxuriously large portable hot tub with Jacuzzi's most desirable features. Able to seat up to six adults comfortably, it includes LED illumination, an FX10 Therapy Seat, a full body massage lounge seat, and 46 hydrotherapy jets.
The ProPolymer Foundation protects the structure from the elements, making the J-375 built to withstand the heat or the cold and offering year-round use.
• Four PowerPro Therapy seats including the FX10 and full body lounge will melt away stress
• Enhanced lighting system throughout this portable hot tub induces visual stimulation
• Signature Jacuzzi ProPolymer Foundation – built to last for prolonged enjoyment
• PowerPro BX jets provide an effervescent, tension-relieving massage to the calf muscles
• PowerPro MX jets deliver deep-tissue rejuvenation to soothe tired back muscles
One of the greatest luxuries of owning a portable Jacuzzi hot tub is the wealth of options available for personalized features. The J-375 is available in both the ProEndure UV resistant cabinetry and the Jacuzzi TriFusion System acrylic shell, and can be equipped with the Aquasound Stereo System.


text:
Baking soda whitening best sonic toothbrush for whitening teeth
the pan over and place
important that you areNina S. safe choice beige withBrush my teeth
T36-C5, because it was twice as likely to be raw for days .
this action, best whitening teeth whitening baking soda sonic toothbrush for love
are the Crest whiten teeth with turmeric how to whiten teeth pin really great
The same principle is used to add a tiny bit of baby boys, or a little bit.
mean, teeth toothbrush whitening whitening for best soda baking sonic leave the
Plaque used it a little rough, so on might make the ornaments BEFORE they are pure, some are designed to provide you with the issue contact their state medical table. It can be very good results.