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A family who spent $100,000 on beanie babies went bankrupt because of their obsession.
Chris Robinson, a burgeoning film-maker, has now completed a documentary that revealed the extent of their purchases, which began after his father was told that the cuddly toys would be a great investment.
The Robinson home is now filled to the brim with a variety of cuddly toys and beanie bags, which range from soft pandas, giraffes, and bunnies, but its value still remains quite low.
Chris went on to state that the family used to munch down McDonald's takeaway burgers just so they could then get their hands on the Teenie Beanies in Happy Meals.
Chris Robinson explained, "During the time we were actively collecting Beanie Babies, there really weren't many aspects of our lives that weren't affected by it. It became this all-consuming family activity."
He discussed this in depth with Dazed and Confused, stating that the toys were meant to have a huge resale value and be pined for by collectors.
Robinson continued, "My father bought into everything these people were telling him about how valuable they were going to be and how exclusive some of them were, and he passed that along to us and created this excitement in our family about what we were doing."
He added, "For a while there was a real belief that we were going to make money from collecting these, and I'm sure some other families thought the same thing. At least I hope that we weren't the only ones."
However, Robinson also admitted, "We actually never really tried selling them. We just collected them for a few years, finally saw the error in our ways, and then packed them away in hopes that maybe someday they'd rebound and we could get some money back.
Mr Robinson then divulged what they intended to do with the vast sum of money they expected to received. "The plan going in was for them to pay for our college tuition, but it became pretty clear that wasn't going to happen for us," he stated. "Maybe by the time our kids graduate high school they'll have made a comeback. Or we can just burn them for warmth in the event of the Apocalypse."


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Monday, March 30, 2009
Your thoughts on sugar
Regarding the sugar, I am definately a victim of craving more sugary foods once I have a tiny amount, especially with lollies which are my all time favourite, I have to really watch my portions as I can eat a lot more than I need. I tend to stick to having some lollies at the movies as my treat (I don't go to the movies that often for it to be a problem) and that way I can put a small amount of my faves (pineapples and strawberries and cream) into the bag to enjoy. I never buy too many as the lollies at the cinema are too expensive! But it is better than buying a whole bag and eating the lot, which I could very easily do! (1 square of chocolate is enough for me - but lollies is another story
Posted by Susie Burrell at 8:13 AM

subset: commoncrawl


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Spicy Guacamole
I'm all for trying different foods – at least once. Sometimes one bite is all it takes to make me a convert – other times, that first bite will be the only bite of that food that will ever cross my lips. For instance, I've eaten a raw oyster and a frog leg once – I'll never consume either of these foods again. Recounting those moments gives me the shivers. But guacamole – that's a totally different story.
Up until about two years ago avocados and I weren't on good terms. And if I'm being honest, I hated them before even giving them a chance. I think I had a negative textural perception of them. I'm big on textures and if something falls into the categories of slimy, gritty, jiggly or velvety (unless it's ice cream) I immediately deem it as suspect.
A couple of years ago I was wandering around Whole Foods and lost my friend for a minute. When I found him, he was standing in the middle of the produce section oohing and aahing over a tub of $9.00 guacamole. I went over to inspect the situation and was welcomed with a guacamole covered chip shoved near my face. Of course, I got all dramatic, recoiled and claimed I hated the stuff. However, I decided to take a tiny nibble – just to prove to him that I did indeed loathe the velvety green dip. But I was wrong. I actually loved the stuff and immediately tossed the pricy container into our cart. From that day forward, I was a guacamole lover.
This guacamole recipe is one of the best I've found, and certainly my favorite. Try it at your next barbecue, or heck – on a random Tuesday night. Then come back here and tell me what you think.
Spicy Guacamole
3 ripe avocados, halved, seeded and scooped
2 limes, juiced
3/4 teaspoon kosher salt
1/2 teaspoon ground cumin
1 teaspoon Sriracha* (use less or more to taste)
1/2 cup diced red onion (about 1 small onion)
1 jalapeno pepper, seeded and diced (leave some seeds in if you want to crank up the heat)
2 Roma tomatoes, seeded and diced
2 tablespoons chopped cilantro
1 clove garlic, minced
1. In a large bowl, toss the scooped avocados with the lime juice.
2. Mash the avocados using a fork or potato masher.
3. Add the salt, cumin, and Sriracha and mash.
4. Fold in the onion, jalapeno, tomatoes, cilantro, and garlic.
5. Let sit at room temperature for 1 hour and then serve.
- If you don't have Sriracha, you can substitute another hot sauce or use 1/4-1/2 teaspoon of cayenne pepper.
- Play around with the measurements - If you like more garlic, add more, etc.
- If you don't like your guacamole spicy, simply omit the jalapeno and cut back on the Sriracha.
- Recipe adapted from Alton Brown
Copyright © My Baking AddictionAll images and text ©
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|Statistics and Usage
Oat production in the World
Cereals in Finland
The pictures below describe the prodution and consumption of different cereals in Finland.
Cereals in Norway
The numbers below are based on annual numbers for trading of cereals in Norway.
Food and Agriculture Organization of the United Nations (http://faostat3.fao.org/home/index.html)
Finnish Agriculture Statistics (http://www.maataloustilastot.fi/satotilasto)


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|image credit
Ok, so he's not an emotional vampire but he is my fave True Blood vamp. ;-)
Each of us is a work in progress. There are small parts of us that are broken and in need of repair. These parts need mending so they can work better than new. Why? Because if you have, say, a broken heart, you work though this brokenness and mend your heart. Your heart isn't as good as new, it's better than new. The stitches and scars make you a better person. While these parts are still in need of repair, you attract like. What we haven't resolved with ourselves is what we tend to attract. This is where the vampires stroll along. Why? Like attracts like. Here is where a change of perspective comes into play. The emotional vampires annoy you enough that you begin to work on those hurt areas, you begin to heal, and you take care of you. Guess what? The emotional vamps usually disappear at this point because of your self-care. You've desensitized yourself to them. They no longer have a need for you. What made them gravitate towards you has been demagnetized. The question here is, would you have had the confidence and clarity to work on these pieces had the drainer not come along? Perhaps eventually, but their presence brought these vulnerable, hurt places to your attention.
That's not to say that emotional vampires can't hurt you with their dark cloud of doom. If these sore spots are left untreated, the vamps can enlarge that wound. A little salt for your open wound?
When an emotional vampire moves in for the kill –
-look within and explore what may have attracted them your way.
-Ask yourself, "What can this emotional vampire teach me?"
-Kill them with kindness. Nothing stops a difficult person like a loving dose of empathy. Confusion, my dear, confusion.


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Unitarians and Slavery--Rabbi Wise--The Abbé Miel--Free Lances of the Pulpit--Literary Studies--Evolution--Darwin's work--Emerson in Cincinnati--Edward Everett--My Marriage--Robert Collyer --The Woman Movement--Chess--Paul Morphy.
THERE appeared to me no cloud on the horizon when I found myself in Cincinnati with an anti--slavery congregation. Everywhere were signs of increasing anti-slavery sentiment. The Conference of Western Unitarian Churches ( 1858) passed a resolution that the cause of the slave was moral and religious, rightly belonging to our pulpits. But two of the societies were in slave States, that of the Rev. Mr. Heywood of Louisville, and that of the veteran Dr. Elliot of St. Louis, men of New England birth. Notwithstanding the moderation of our resolution, we had the sorrow of seeing Dr. Elliot and his strong delegation file solemnly out, never to return.
This action of the conference, reversing a timid resolution of three years before, was a relief to me. It had always been a burden to preach about slavery, and it was now less necessary to deal much with the subject. The incident was widely discussed in the papers, and the Cincinnati Enquirer (anti-Republican) described me as an ambitious agitator. I said to my people that inhumanity in man or nation must always prove a demon of unrest. A legend on which twenty-three years later I published a volume then first arose before me as a prophecy: "That fable of the Wandering Jew shall be a dread reality to the heart which knowingly drives from its threshold the Christ who falls there in the form of those who now bear the cross of wrong and oppression, and toil up the weary hills of life to their continual crucifixion."
About that time a little recrudescence of prejudice against Jews occurred in connection with an organisation called the Cincinnati Zouave Guard, against which I protested in the papers; and I even attacked Shakespeare on account of the


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Bernanke Warns Congress About Impeding Recovery
Federal Reserve chairman Ben Bernanke went before lawmakers Wednesday to give his report on the state of the U.S. economic recovery, saying while there were signs of improvement, the unemployment rate, in particular, remained "far from satisfactory."
This transcript has been automatically generated and may not be 100% accurate.
... I ... he later market are improving gradually ... the unemployment rate to seven point six percent in June about a half percentage point lower ... than a month before the federal market Committee initiated its current asset purchase program ... in September ... nonfarm payroll employment increased by an average of about two hundred thousand jobs per month ... so far this year ... despite the scheme the job situation is far from satisfactory ... the unemployment rate remains well above its longer run normal level ... and rates of under employment and long-term unemployment are still much too high ... meanwhile consumer price inflation has been running below the Committee's longer run objective of two percent ... the price index for personal consumption expenditures rose only one percent ... over the year ending in May ... the softness reflects in part some factors that are likely to be transitory ... moreover measures of longer term inflation expectations have generally remained stable ... that should help move inflation back up toward two percent ... however the committee is certainly aware that very low inflation poses risk to economic performance ... for example by raising the real cost of capital investment ... it increases the risk with outright inflation ... consequently Hublot monitor this situation closely as well ... we will act as needed to ensure that inflation moves back quarter of two percent objective ... over time ... at the June FOMC meeting my colleagues and I projected economic growth would pick up the coming quarters ... resulting in gradual progress towards a level of unemployment and inflation ... consistent with the Fed Reserve statutory mandate ... to foster maximum employment and price stability ... specifically most for dissident saw real GDP growth beginning to step up during the second half of this year ... eventually reaching the pace between two point nine three point six percent in twenty fifteen ... it rejected the unemployment rate declined between five twenty six point two percent of the final quarter of twenty fifteen ... Andy saw inflation gradually increasing toward the Committee's two percent objective ... economic growth predicted by most many disciplines partly reflects your view ... the federal fiscal policy will exert somewhat less track over time ... the effects of the tax increases and spending sequestration diminish ... the committee also believes the risk to the economy have diminished since the fall ... reflecting some easing of financial stresses in Europe ... the gains in housing and labor market I mentioned earlier ... and their budgetary position to state and local governments ... and stronger household and business balance sheets ... that said the risk remained tight federal fiscal policy will restrain economic growth over the next few quarters ... but more than we currently expect ... or that the debate concerning other fiscal policy the key issues such as the status of the debt


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Little Awkward Studio
Ep 10. Random End of Year Thoughts & Holiday CheerDecember 22, 2017 Amy Frances and Megan Elizabeth
In our 10th Episode and the very last episode of 2017, we talk about nothing and everything! How to deal with "slow seasons" in your business and a whole lot of holiday randomness. Amy sings, and deals with her new identity as a scrooge. Join us!


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Membrane receptors and ion channels respond to various stimuli and relay that information across the plasma membrane by triggering specific and timed processes. These include activation of second messengers, allowing ion permeation, and changing cellular excitability, to name a few. Gaining control over equivalent processes is essential to understand neuronal physiology and pathophysiology. Recently, new optical techniques have emerged proffering new remote means to control various functions of defined neuronal populations by light, dubbed optogenetics. Still, optogenetic tools do not typically address the activity of receptors and channels native to neurons (or of neuronal origin), nor gain access to their signaling mechanisms. A related method—synthetic optogenetics—bridges this gap by endowing light sensitivity to endogenous neuronal receptors and channels by the appending of synthetic, light‐receptive molecules, or photoswitches. This provides the means to photoregulate neuronal receptors and channels and tap into their native signaling mechanisms in select regions of the neurons, such as the synapse. This review discusses the development of synthetic optogenetics as a means to study neuronal receptors and channels remotely, in their natural environment, with unprecedented spatial and temporal precision, and provides an overview of tool design, mode of action, potential clinical applications and insights and achievements gained.
The need to better understand the function and roles of neuronal membrane proteins, synaptic receptors and ion channels in particular, has become one of the more pressing goals in neuroscience for the reason that their function and dysfunction underlay some of the most prevalent and devastating neurodegenerative diseases, neuropsychiatric disorders, and synaptopathies [1], [2], [3], [4], [5], [6]. The better we understand their mechanisms of actions, in health and disease, the better we may design novel drugs and treatments.
The study of neuronal receptors and channels has been extensively addressed over the years by conventional pharmacology, electrophysiology, structural studies, molecular and genetic manipulations, etc. [7], [8], [9], [10], [11], [12], [13]. Despite the countless insights gained, these and other contemporary techniques display inherent drawbacks, such as specificity and off‐targets, slow kinetics, poor spatial confinement, lack of or slow reversibility, etc. These appreciably limit our understanding of the processes under study [14], [15], [16], [17]. Consequently, new methods are still vigorously pursued.
One novel approach developed for the remote study of neurons is optogenetics [18], [19], [20]. Optogenetics is a momentous and rapidly evolving field that leverages naturally occurring light‐sensitive proteins (hereinafter referred to as photoreceptors, see [21]) and repurposes them into genetically encoded light‐gated cellular actuators. Thereby, optogenetic tools allow the user to control cellular activity by light. The type of modulation depends on the photoreceptor's biological function. For instance, the widespread optogenetic tool ChR2 is a light‐gated cation channel [22]. When expressed at the membrane of neurons, light illumination causes rapid channel opening resulting in strong membrane depolarization and AP firing [23], [24], [25], [26], [27]. This methodology has drastically transformed the way we approach and study biological systems, neuroscience in particular, owing to the method's non‐invasive and reversible nature and, owing to the inherent properties of light, fantastic spatiotemporal resolution. Notably, with cell type‐specific promoters [28], the expression of optogenetic tools can be confined to select cells and neuronal circuits to be exclusively interrogated by light. Optogenetic tools are extensively employed today to control neuronal circuits and behavior by regulating membrane excitability, though they can also control a wide array of additional cellular mechanisms ranging from protein trafficking, to inducing genomic engineering, to humbly name but a few of the achievements and applications described to date [19], [22], [23], [29], [30], [31], [32], [33], [34], [35], [36], [37], [38].
However, optogenetic tools are not without fault. These tools may suffer from slow kinetics of activation or deactivation, small channel conductance, poor expression in mammalian cells, undesired secondary effects (e.g., acidifications), unwarranted basal activity prior to light illumination, unexpected inverse effects (i.e., depolarization instead of hyperpolarization), and more [19], [39], [40], [41], [42], [43], [44]. With regard to neuronal synapses, optogenetic tools are limited in their ability to control endogenous signaling mechanisms. For instance, though several tools have been specifically expressed at the synapse (e.g., [39], [45], [46], [47], [48]), very few were able to provide direct control over native components or endogenous signaling.
This gap can be bridged by an analogous method—synthetic optogenetics—that similarly employs light to regulate the function of proteins but, instead of the use of photoreceptors, relies on the appending of synthetic chromophores to "blind" proteins (i.e., proteins that do not naturally respond to light), denoted photoswitches.
There are many names by which this approach has been described (chemical‐optogenetics, photopharmacology, or optogenetic pharmacology [49], [50], [51]). However, as new tools are appearing in the literature, most names no longer cover all cases. For example, although the method initially employed pharmacophores in the design of photoswitches, several recent advances show the possibility to design pharmacophore‐less photoswitches (e.g., photoswitchable tweezers, see below). Therefore, names embracing the term "pharmaco" are less relevant today, warranting the re‐naming of the approach. The name of synthetic optogenetics deems the most appropriate for several reasons: The method requires the usage of synthetic photoswitches, several key tools are denoted synthetic (e.g., SPARK, see below) and, importantly, as the field of optical control of cellular activity is widely recognized today by the name of optogenetics, the name "synthetic optogenetics" allows it to remain under this umbrella term, whilst emphasizing the distinction of the approach from others. In addition, we hypothesize that this name should also cover future developments in this area too (see discussion).
Here, we first review the historical development of the technique and proceed to provide an overview of modern developments of light‐gated channels and receptors, through examples of their usage in the field of neuroscience. Lastly, we discuss several limitations, how these could be overcome and end with future prospects.
Photosensitizing proteins by synthetic optogenetics
The approach of synthetic optogenetics combines chemical and genetic methods to render potentially any protein sensitive to light. In contrast to optogenetic tools incorporating naturally occurring chromophores, this method employs synthetic, light‐sensitive photoswitches [52]. Akin to chromophores found in nature (e.g., retinal [53], [54]), photoswitches too undergo chemical reactions following the absorption of specific wavelengths. One highly relevant reaction to the design of biological tools is the trans‐to‐cis isomerization of azobenzenes, resulting in a large geometrical change and significant reduction in the length of the molecule (Fig 1A) [55]. Azobenzenes are mostly (> 95%) found in –trans in the dark or following green‐light illumination (e.g., ~500 nm), but isomerize to the shorter bent –cis form (~80%) following near‐UV light absorption (e.g., 350 nm; Fig 1B) [56]. Therefore, isomerization can be utilized to alter/modulate a protein's function. For instance, if the azobenzene‐core is decorated with a specific receptor's ligand, isomerization from trans‐to‐cis could be used to expose the ligand making it accessible to the receptor, indirectly photoregulating it (Fig 1C). Other chemical headgroups can be added onto the azobenzene photoswitch, such as blockers, antagonists, allosteric modulators, conjugating moieties, lipophilic chains (see Fig 1, toolbox). These will typically determine the mechanism of action of the photoswitch. Today, the variety of the modes of action by which photoswitches can regulate protein function is extensive, involving varying the distance between a drug and its binding site, changing local polarity, inducing lateral pressure in the membrane, applying forceps‐like motion to gate channels, piecing a fragmented drug, etc. (Fig 1C–J). Notably, the number of mechanisms employed by photoswitches exceeds the list enumerated here and, in fact, is limited only by the creativity of the designers.
The historical development of synthetic optogenetics
Synthetic optogenetics was pioneered by the Erlanger group, which initially demonstrated the photocontrol over the enzyme chymotrypsin [57]. Here, the authors synthesized a soluble photochromic ligand (PCL) composed of a light‐sensitive azobenzene‐core linked to diphenylcarbamyl‐chloride (DPCC—a specific inactivator of the enzyme) they termed PAPC (p‐azophenyldiphenylcarbamyl‐chloride; Fig 1G, purple). PCLs are advantageous as they do not require any genetic alterations of their target protein, enabling the regulation of wt proteins, wt‐chymotrypsin in this case. The authors envisioned that the light‐dependent isomerization of the azobenzene in PAPC could be used to gate (i.e., enable or disable) the ability of the pharmacophore (DPCC) to act on its site of action, a serine residue located deep within the enzyme. Indeed, they then showed that the –cis isomer of PAPC is five times more potent in inactivating chymotrypsin than the –trans isomer, suggesting that the DPCC has different pharmacological properties when the attached azobenzene is in the –trans or –cis conformation [57].
This report paved the way for other groups to render their favored enzyme photolabile. To this end, other groups have replaced DPCC with other specific compounds from the toolbox of available pharmacophores. This mix and match quickly yielded photoregulated acetylcholinesterases [58], [59], trypsin [60], aldolases [61], [62], urease, amylase, and more (see list in [63]). Interestingly, the engineering of photoregulated enzymes subsided in the following years. However, the approach was not neglected, rather shifted from targeting soluble proteins to membrane proteins. The first demonstration of the photoswitching of membrane receptors was shown to involve nicotinic acetylcholine receptors (nAChR) [64]. Here too, the authors exploited the changes in geometry of the azobenzene so that the PCL, here containing a carbachol headgroup, would differently interact and affect nAChR. When in –trans, the photoswitch inhibited the receptor more potently than when in –cis. This report established that the approach was not limited to soluble proteins, namely enzymes, and further showed its advantages over conventional pharmacological reagents. Indeed, though several specific drugs acting on nAChRs were available at the time, this approach enabled the study of nAChRs in their native environment with unprecedented temporal resolution, resulting in a flurry of publications [65], [66], [67], [68], [69].
During these years, another development laid the groundwork for what will become one of the most employed strategies in synthetic optogenetics: conjugating PCLs to their target. This involved the design of tethered photoswitches or photoswitchable tethered ligands (PTLs). PTLs are designed to irreversibly bind to their protein target, creating a high local concentration of the drug. To the best of our knowledge, the first tethered photoswitch was the cysteine‐reactive QBr photoswitch (3‐(alpha‐bromomethyl)‐3′‐[alpha‐(trimethylammonium)methyl] azobenzene bromide) [70]. This azobenzene‐based photoswitch included an additional chemical moiety at the other end of the azobenzene that would covalently conjugate the photoswitch onto existing outer cysteine residues in nAChRs [70]. Once bound, QBr selectively activated the channels when in its extended –trans form, and much less so when in –cis (e.g., Fig 1D). The use of PTLs is preferable over PCLs in certain instances as it keeps the photoswitch near the receptor at all times, granting a high local concentration of the chemical headgroup. This can enable efficient competition of the photoswitch with the soluble ligand (e.g., Fig 1E, black filled triangles). It also enables persistent perfusion of the preparation without necessitating constant supplementation of the photoswitch. Owing to these advantages, the photoregulation of enzymes was revisited in the following years with the use of PTLs (e.g., [61], [71], [72]). Collectively, these pioneering experiments demonstrated the potential of the approach, laid the framework (i.e., toolbox) for the design of soluble (PCLs) or tethered (PTLs) photoswitches, and, importantly, hinted at the potential of the approach to render any blind protein light‐sensitive [73], [74], [75].
Synthetic optogenetics revisited
Despite the method's first appearance almost 50 years ago, the approach was gradually ignored and almost completely neglected for many years. However, in 2004, the spark was re‐ignited by a group of scientists from the University of California at Berkeley (amusingly befitting as UCB carries the motto "let there be light" on its seal). The Isacoff–Trauner–Kramer alliance (see perspective by [76]) designed a Synthetic Photoisomerizable Azobenzene‐Regulated K+ channel based on the Shaker channel—dubbed SPARK—to control neuronal excitability [77]. This was the first report using a PTL and a genetically modified channel to control mammalian neuron excitability, only two years after the first optogenetic demonstration using rhodopsin to "chARGe" (depolarize) neurons ([33], but also see perspective by [78]).
To confer light sensitivity to the Shaker channel, Banghart and colleagues synthesized a PTL consisting of a maleimide, azobenzene and a quaternary amine (QA—a K+ channel blocker) denoted MAQ [77]. When in –trans, the end‐to‐end distance of MAQ spans ~17 Å. Knowledge of this distance was then used to locate an outer residue in the Shaker channel that was situated at a similar distance from the pore. This residue was mutated to a cysteine to conjugate the photoswitch (via its maleimide) so that when in –trans, the quaternary amine of MAQ would reach the pore and block the channel. In turn, this would inhibit membrane hyperpolarization resulting in AP firing by the neuron (Fig 2A and B). This process could be rapidly reversed by near‐UV light, as isomerization of MAQ to –cis shortened the molecule (to ~10 Å), physically removing the blocker from the pore, which opened the channel and electrically silenced the neuron. By toggling between green and near‐UV illumination, the authors were able to prevent or enable AP firing repeatedly and remotely with a high spatiotemporal resolution (Fig 2B). Importantly, the authors note that the molecular design of the channel was possible mainly because of the rise in the availability of detailed structural and molecular information of ion channels, and K+ channels in particular (e.g., [79], [80]). In point of fact, SPARK emerged at the time when the Nobel prize in Chemistry was attributed to Roderick MacKinnon for solving the structures of K+ channels [81]. Notably, the genetic manipulation of the channel enabled the exclusive expression of the channel in neurons, thereby limiting PTL conjugation to this defined population, unlike PCLs that cannot be typically confined to selected cells (but see below).
In parallel, two other groups were also toiling over the design of light‐gated channels. Interestingly, both groups made use of the bacterial mechanosensitive channel of large conductance (MscL). The group of Bert Poolman synthesized a new type of an azobenzene‐based photoswitch containing lipophilic phosphate tails, named 4‐Azo‐5P [82]. This photoswitch would incorporate within the leaflets of the membrane so that during trans‐to‐cis isomerization, the photoswitch would exert lateral pressure in the membrane. This would mechanically gate the closely situated MscL channel (Fig 1H). In 2005, the Feringa group also designed a photoregulated MscL, using another clever approach [83]. MscL, though mechanosensitive, also exhibits spontaneous channel opening when polar or charged amino acids are introduced next to the pore [84]. Koçer exploited this phenomenon and synthesized a spiropyran‐based PTL. To note, in contrast to azobenzenes, the isomerization of spiropyrans involves the interconversion between closed‐ and open‐ring structures, which is accompanied by a large polarity change of the molecule [55]. Thus, when this photoswitch was accurately tethered next to the pore (via an iodoacetate tether), its isomerization by light absorption resulted in a local polarity change that induced channel opening (Fig 1F) [83].
Contemporary light‐gated receptors and channels
The early developers of the method have provided the blueprint and building blocks for the development of the approach, which researchers are still using to date to devise more sophisticated photoswitches and light‐gating mechanisms. Of the different light‐sensitive cores (for detailed list see [55]) and protein tethering methods available [85], azobenzenes and maleimides are used most in the synthesis of PCL and PTL photoswitches, for good reasons. Azobenzenes are easily synthesized, exhibit high photostationary states, undergo fast photoisomerization, and photobleach slowly. Analogously, maleimides are commercially available, thiol conjugation is covalent, and the reaction is moderately specific under physiological conditions (pH and temperature). From a protein‐engineering point of view, maleimide conjugation is highly favorable as it requires very minimal protein modification—a single amino acid substitution to cysteine. Even though maleimides can potentially interact with other existing cysteines, free cysteines (i.e., unconjugated and water accessible) are naturally of low abundance in proteins (< 2% [86]). Nevertheless, it should be noted that maleimides hydrolyze rapidly (tens of minutes) in aqueous conditions, offering a narrow time‐window for labeling proteins expressed on the membranes of live cells. Thus, as cysteine conjugation will mostly occur in the first hour after addition of the maleimide–PTL, this process will label only the receptors located at the membrane during that time. This could be both a curse and a blessing (see Box 1). Lastly, the use of maleimide–PTLs requires ectopic expression of the modified, cysteine‐substituted protein. Despite the benefit of genetic‐targeting (e.g., Fig 2B, a single highlighted cell), expression of ectopic genes can result in unwarranted overexpression of the receptors and their promiscuous cellular localizations. With these advantages and limitations in mind, below we describe several families of neuronal receptors and channels rendered photosensitive by either azobenzene‐based PCLs or PTLs. We provide several examples where PCLs and PTLs target the same receptor, with the intent to showcase the alternatives the approaches offer, such as remaining under physiological conditions or enabling genetic‐targeting of the tool. We regret that owing to space constraints, we cannot list all available examples.
Box 1:In need of answers
Labeling efficiency—Whereas naturally occurring photoreceptors are found at the membrane at a 1:1 stoichiometry with their chromophores, synthetic optogenetics tools are not. Following expression of modified receptors and incubation with the relevant PTL, it is difficult to determine the number of modified receptors that have been labeled by the PTL. Labeling efficiency will determine the effectiveness of the tool. The extent of labeling can be approximated by equating the photocurrent with the maximal ligand‐induced current. However, this method does not comply with photoswitches that exhibit partial agonism (e.g., GluAzo [131]). As a result, methods to increase labeling efficiency, means to measure it, and alternative labeling schemes are still under development.
Narrow labeling time—Maleimides hydrolyze rapidly (tens of minutes). This presents the user with a narrow time‐window, during which the photoswitch would need to be applied as close as possible to the site expressing the receptors. For prolonged in vivo experiments, this is limiting as the photoswitch may need to be reapplied during the course of the experiment to maintain a constant amount of labeled receptors. This is cumbersome and possibly damaging, as it entails multiple applications of the photoswitch. Photoswitches with better stability would broaden the labeling time‐window and would also allow the user to apply them at a larger distance from the site of expression. This type of photoswitch could also be slowly and steadily administered to the brain via an implemented cannula, lessening the burden of multiple injections.
Solubility—Azobenzene are lipophilic compounds, typically solubilized in DMSO prior their application onto the preparation [180]. Even more challenging are azobenzenes further decorated with lipophilic groups (such as fatty acids, e.g., [181]). Therefore, photoswitches with improved solubility are still under pursuit [182].
Near‐UV absorption—Most azobenzene‐based photoswitches absorb near‐UV (< 380 nm, see Fig 1B), to isomerize from –trans to –cis, wavelengths that may be harmful to cells. This justifies the synthesis of red‐shifted photoswitches. Indeed, several red‐shifted photoswitches have been synthesized (e.g., MAG460 [171]), though this shift comes at the expense of bistability, resulting in the spontaneous relaxation of the photoswitch back to –trans. This may lower the photostationary population of the –cis isomers and would require longer or repetitive illumination to obtain continuous photoresponses. Therefore, better red‐shifted photoswitches that maintain excellent switching behavior are necessary.
Broad absorption—Azobenzenes exhibit broad absorption, in particular the –cis isomer (Fig 1B [55]). This complicates the use of photoswitches with additional optical tools (such as fluorescent proteins and probes), because the photoswitch will be isomerized back to –trans during the imaging of the probes. Thus, narrower absorbing photoswitches are needed to achieve all‐optical interrogation of cells (see [183]).
Clinical use—Whereas several research groups have shown the utility of photoswitches in vivo (e.g., fish and rodents), there are still questions pertaining to the safety of the use of azobenzene‐based photoswitches. This concern stems from their poor solubility and metabolic stability, both of which may lead to potential long‐term toxicity.
Availability—Most photoswitches are not commercially available, rather are typically obtained directly from the research groups. There are few exceptions, such as several MAG variants sold by Aspira Scientific (www.aspirasci.com). The lack of a chemical depository hinders the broad dissemination of the photoswitches. Unlike DNA depositories (e.g., Addgene), the synthesis process is much more expensive and laborious.
GABA receptors
GABA is the major inhibitory neurotransmitter of the brain. It binds to ionotropic GABAA or metabotropic GABAB receptors found at the membrane of neurons. When activated, GABAA receptors open and negatively charged chloride ions flow into the cell and inhibit neuronal firing [87]. The importance and variety of these receptors, and lack of specific pharmacology, has motivated several groups to tackle activation or antagonism of these receptors by light.
A couple of examples have shown that GABAA receptors can be photoregulated by azobenzene‐based PCLs employing propofols, compounds known to potentiate GABAA currents, denoted azo‐propofols [88], [89]. For example, in the report by Stein et al, azo‐propofol in –trans potentiates the GABA‐induced currents by almost twofold, and this effect is fully reversed by near‐UV light. Notably, azo‐propofols interact with unmodified GABAA receptors; thus, with restricted light illumination, these enable to map the localization of the receptors within neurons.
Examples of tethered photoswitches used with modified GABAA receptors include a large toolbox of azobenzene‐based PTLs containing various ligands as headgroups, namely muscimol, GABA, or its guanidinium analogs [90], [91]. Their mechanism of action is postulated to resemble that of MAQ and SPARK (see above and Fig 1E), wherein the PTL is attached to an introduced cysteine on the receptor and the azobenzene modulates the pharmacophore's ability to physically reach its site of action on the receptor when in one conformation, but not when in the other. Notably, these photoswitches, and their cognate‐modified GABAA subunits, were the first to be used in mammalian brains in vivo [91], demonstrating the viability of this approach for manipulating receptors in living brains of behaving animals.
Noteworthy, the metabotropic GABAB receptor family has not been addressed to date by the technique, though other PTL‐gated GPCRs have been described (see below). This is unfortunate, as light‐gated GABAB receptors could help unravel their unique molecular roles at the post‐synapse, where they are suggested to modulate the function of ionotropic glutamate receptors, for example by reducing calcium permeability through NMDA receptors [92]. Other GABA‐binding proteins have also been the target of light‐aficionados. The most abundant GABA transporter in the brain, mGAT1, was recently made photoantagonizable via PCLs containing tiagabine, a potent mGAT1 inhibitor [93]. We note this, although beyond the scope of this review, as this is the first time that the synthetic optogenetic approach was applied on membrane transporters. Attractively, there are numerous other photochromic agents described in the literature that could, in principle, target GABA‐binding proteins, but to date remain untested [94]. These are ready for the picking by laboratories interested in studying GABA‐mediated synaptic processes by light.
Glutamate receptors
On the other side of the spectrum, there is glutamate. Glutamate is the major excitatory neurotransmitter that activates members of the glutamate‐receptor superfamily, which includes 18 ionotropic receptors [95] and eight GPCRs [96]. These receptors are expressed at pre‐, post‐, and extra‐synaptic regions, regulate essential processes in the brain, spinal cord, and peripheral nervous system, and are implicated in a myriad of diseases [97], [98]. A large repertoire of pharmacological agents has been developed to probe the roles of glutamate receptors, but these agents attend to but a few receptor subtypes [95].
To address some of the limitations of pharmacological agents and of other methods used to study glutamate receptors, numerous synthetic tools have been developed, with the light‐gated kainate receptor—LiGluR—as the prominent representative of this group (e.g., [99], [100], [101], [102], [103], [104], [105], [106], [107], [108]). LiGluR tethers L‐MAG (maleimide–azobenzene with a 4'L stereochemistry glutamate) by a cysteine introduced into the receptor's LBD (Fig 1E) [106]. As noted above, this residue was located by analyzing the structures of several LBDs of kainate receptors (e.g., Fig 3 and [109]). In particular, it is located at the surface of the protein and next to the opening of the LBD. To note, here the photoswitch does not interact directly with the pore of the receptor (as in SPARK, compare Fig 1D and E), but rather with allosteric elements of the receptor, explicitly the LBD. When bound at this site, the elongated –trans form of MAG does not allow the glutamate headgroup to reach into the glutamate‐binding pocket located deep within the LBD. Isomerization to –cis by near‐UV light propels the glutamate end into this groove, enabling LBD closure and, subsequently, channel opening (Fig 1E). Green‐light illumination isomerizes L‐MAG back to –trans, pulling the glutamate away from the receptor and triggering channel closure. LiGluR was initially expressed in neurons for the purpose of driving neuronal excitability in vitro [105] and extended to in vivo studies more recently [103], further validating the technique's applicability in the live mammalian brain. LiGluR was also used to study the desensitization process of the kainate receptor, showing the utility of the tool for biophysical studies of the receptor [110]. In the degenerating retina, LiGluR was expressed in RGCs where, following in vivo injections of L‐MAG into the retina, the now light‐responsive LiGluR restored light sensitivity to the retina [100], [102]. LiGluR has also been expressed in non‐neuronal cells, namely astrocytes, to stimulate glutamate exocytosis owing to its relatively high Ca2+‐permeability [104].
We have recently described photoswitchable NMDA receptors (LiGluNs) [111]. We created a small library of GluN‐subunits bearing cysteine mutations to tether L‐MAGs, with a similar engineering logic as described for LiGluR, culminating in several photoswitchable NMDAR subunits. This process allowed us to photoagonize and photoantagonize the NMDA current [52], [111]. The difference in activity was obtained by tethering L‐MAG at different locations at the LBD of the subunit. At one position, when in –cis, the glutamate headgroup could properly dock within the glutamate‐binding pocket resulting in proper clamshell closure and channel opening. However, when the photoswitch was tethered to another residue, even if only one amino acid away, the isomerization of L‐MAG would incorrectly position the glutamate headgroup in the pocket, leading to the obstruction of clamshell closure and channel opening. This obstruction persisted in the presence of soluble agonists, acting as a non‐competitive antagonist [111]. We have also used the glutamate‐based L‐MAG photoswitch to photoantagonize the glycine‐binding GluN1 subunit. In this instance, the glutamate moiety of L‐MAG acts as an antagonist when positioned in the glycine‐binding pocket, as it prevents the binding of the native ligand glycine. These examples display the versatility of the method and demonstrate that a single photoswitch can be used with multiple subunits of the glutamate‐receptor family, even if those bind a different ligand than the one used in the photoswitch (e.g., glutamate and glycine).
To obtain modest expression levels of LiGluNs at the PM, we have taken advantage of the fact that GluN2 subunits are retained at the ER, unless bound to the GluN1 subunit. In this manner, the expression levels of LiGluN2 subunits are effectively controlled by the availability of the endogenous pool of GluN1 subunits in the neuron.
A unique example of a light‐gated ionotropic glutamate receptor has been described by Janovjak et al [112]. The authors generated HyLighter (short for hyperpolarizing light‐gated glutamate receptor), a chimeric receptor containing the LBD of the mammalian iGluR6 and the membrane‐spanning domains (including the pore) of sGluR0, a prokaryotic homolog of mammalian iGluRs from cyanobacteria [113]. The interesting feature of sGluR0 is that, unlike all other glutamate receptors, it holds the signature of the selectivity filter of K+ channels (GYG motif [114]) and, thereby, solely conducts K+ ions. Thus, when HyLighter was bound to L‐MAG, near‐UV illumination would open the channel and hyperpolarize the neuron due to K+‐ion outflow.
Multiple light‐gated metabotropic glutamate receptors (LimGluRs) have been engineered [115], [116], [117], [118], [119]. Levitz et al show that photocontrol over their LimGluR version requires 4'D stereochemistry of MAG, as opposed to 4'L (as for LiGluR and LiGluN). Broichhagen et al show that LimGluRs could support the fusion of a SNAP‐tag [120] to tether a benzylguanine‐based photoswitch, denoted BGAG [121]. Whereas the SNAP‐tagging strategy overcomes several limitations of the maleimide chemistry, namely its short viable lifetime and potential off‐target cysteines conjugation (see Box 1), it requires the incorporation of a large protein domain (SNAP, ~20 kDa) into the receptor, which might be more detrimental to the protein's function than a single amino acid substitution.
Photoswitchable tethered ligands, though extremely useful, require genetically modified channels that need to be ectopically expressed (Fig 2B, plasmid). Overexpression of ion channels may limit their usage in certain contexts and preparations (see Box 1). To bypass this drawback, and still make use of PTLs, Izquierdo‐Serra et al devised a method to tether photoswitches to specific endogenous glutamate receptors without the necessity of cysteine mutagenesis. This was achieved by synthesizing a series of new photoswitches—targeted covalent photoswitches (TCPs)—containing a short‐lived, highly reactive anchoring group, such as epoxide and N‐hydroxysuccinimide esters [122]. These anchoring groups covalently react with lysine residues exposed on the protein's surface. The ligand headgroup of the photoswitch affords a kinetically controlled site‐selective conjugation specifically in the vicinity of the LBD, ensuring target specificity (by affinity labeling [123]). Synonymously, there are photoswitchable affinity labels (PALs), which are conceptually identical to TCPs (some PALs even include epoxide as do TCPs) in the sense that they are also tethered to native channels by affinity labeling [124], [125].
In sharp contrast, several soluble PCLs have been engineered to selectively target ionotropic glutamate receptors. For instance, GluAzo is a photoagonizing PCL specific to iGluR5 and ‐6 (Fig 3) [126]. Laprell et al [127] designed a similar caged photoswitch to non‐specifically activate all types of NMDA receptors, termed ATG. Lastly, several potent PCLs that photoregulate AMPA receptors by a similar "uncaging" mechanism have also been synthesized, for example, ATA and ShuBQX‐3 [128], [129]. Importantly, while the ON photoswitching of PCLs can be compared to the photolysis of "conventional" caged compounds, PCLs can be turned OFF reversibly, contrasting the irreversible photolysis of caged drugs (see [130]).
Recently, the Trauner and Schiefner groups succeeded in crystalizing a kainate receptor's LBD bound to the GluAzo photoswitch (Fig 3) [131]. The structures show that the interactions of the glutamate moiety of GluAzo with its binding pocket are essentially the same as with soluble glutamate. In addition, the structure highlights how the LBD can assume a closed conformation with the photoswitch by allowing the azobenzene part of the molecule to protrude from the LBD via an exit tunnel only when in –trans (Fig 3A and B). Thus, because this cavity cannot accommodate the –cis isomer, it explains why only trans‐GluAzo can induce channel opening. Whereas soluble glutamate is almost entirely buried within the LBD after clamshell closure, GluAzo is not and results in a slightly more open LBD conformation (Fig 3C and D). This further elucidates why GluAzo acts as a partial agonist, as the degree of LBD closure correlates with the extent of channel activation [132], [133]. Together, the structures validate many assumptions made in the design of light‐gated glutamate receptors and should further assist in the design of better kainate receptor photoswitches, as well as of photoswitches for other glutamate receptors.
Potassium channels
The K+ channel family is one of the largest ion channel families and the most widely distributed among organisms, consisting of more than 50 genes, found in both excitable and non‐excitable cells [114]. Their roles are diverse (e.g., they control heart rate, insulin secretion, electrolyte transport, and cell volume regulation), but with respect to neuronal physiology, K+ channels are fundamentally responsible for setting membrane potential, decreasing excitability, and shaping action potentials [134], [135].
K+ channels can be divided into several subfamilies, namely voltage‐gated, calcium‐gated, inward rectifying, and two‐pore (2P) channels [136]. Many of these have been targeted by PCLs or subjected to genetic manipulation to tether PTLs. In general, most synthetic optogenetic tools based on K+ channels have been developed for hyperpolarizing neuronal membranes, with a few exceptions. As briefly mentioned above, SPARK is a genetically modified channel based on the Drosophila Shaker potassium channel, belonging to the voltage‐gated family of channels (KV). SPARK, when bound to MAQ, was initially designed to hyperpolarize neurons (H‐SPARK) [77]. However, SPARK was also further mutated and converted into a non‐selective cation channel (as opposed to the exclusive K+ conductance of Shaker) to depolarize neurons (D‐SPARK) [137]. Similarly, several other potassium channels were designed to tether MAQ or MAQ‐like PTLs to control channel gating and membrane excitability, such as Kv1.3, Kv3.1, Kv7.2, and the Ca2+‐activated SK2 channel (Fig 2) [125]. This long list of modified light‐gated K+ channels employing the same photoswitch, and likely photogating mechanism (i.e., channel block by the QA moiety), demonstrates once more the transferability of the photoswitch between members of the same family. Moreover, the MAQ photoswitch was even applied to photogate other ion channels (e.g., Ca2+ and Na+ channels [138]). This is possible with photoswitches that are based on pharmacophores with non‐specific and broad activity such as QX‐314. Together, these demonstrations display the versatility of the technique.
The Kramer group (UC Berkeley) has extensively explored the photoregulation of endogenous K+ channels by a large repertoire of PCLs and PALs ([124], [139], [140], [141], for an extensive list see [138]). Of the many examples, we would like to highlight one interesting case involving a photoswitch denoted AAQ (acrylamide–azobenzene–QA). AAQ includes an acrylamide tethering group intended to react (to tether the photoswitch) with native extracellular residues located near the pore of the channels [124]. Indeed, Fortin et al found that AAQ photoregulated the activity of K+ channels, presumably from the extracellular side of the membrane, meriting its classification as a PAL. Surprisingly, it was later discovered that the hydrophobic acrylamide group facilitated permeation of AAQ into the cell, before it could react with external residues (Fig 4A) [139]. Thus, AAQ is a membrane‐permeable PCL and its photoregulation mechanism involves its entry into the inner vestibule of the channel when in –trans, but not when in –cis. Once in the vestibule, the QA headgroup exerts its block (Fig 4A). Though given lemons, the group was able to make lemonade from the AAQ misfortune by using these observations for the development of new hydrophobic photoswitches (e.g., BzAQ, PrAQ) that acted from the intracellular side of the membrane to block voltage‐gated potassium channels, but also voltage‐gated calcium and sodium channels [138], [139].
Finally, 2P K+ channels and inward rectifiers (Kir) have not escaped the attention of photoengineers. Briefly, to photocontrol endogenous 2P‐TREK channels with the MAQ PTL, Sandoz et al devised a conditional expression scheme. To this end, the authors created a conditional TREK subunit bearing a cysteine and a truncated carboxy‐terminus (ΔCT), required for ER export of the subunit when intact. Thus, only when the ΔCT‐subunit dimerized with a wt subunit (endogenously present in the ER of the neuron) could the heteromeric channel complex exit the ER and reach the PM [142], [143]. To open the GIRK channel, belonging to the family of inward rectifiers [144], the Trauner group designed a PCL denoted LOGO [145], [146]. LOGO is an interesting PCL consisting of an azobenzene‐based photoswitch containing VU0259369, a potent and selective GIRK1 channel activator [147]. Whereas GIRK channels are uniquely opened by the binding of Gβγ molecules derived from activated Gαi‐coupled GPCRs [148], VUO259369, and hence LOGO, can open the GIRK1‐containing channels even when the G‐protein cycle is disrupted by PTX. This photoswitch can thereby bypass the need to activate GPCRs to elucidate the gating mechanisms of the GIRK channel in neurons.
P2X receptors
P2X receptors are trimeric, ATP‐binding cation channels [149]. With regard to the brain, P2X receptors are implicated in several diseases, such as neuropathic pain [150]. To photogate P2X receptors, the team of Grutter has employed a unique strategy consisting of using charge to induce channel opening, reminiscent of the strategy employed by Kocer et al [83]. To this end, the authors have synthesized a maleimide–azobenzene‐based PTL comprising positively charged trimethyl ammonium derivatives at the end of the molecule [151]. When the PTL was tethered to cysteines at different TM domains, light‐dependent isomerization induced changes in the local charge that was sensed by local residues of the TMs and induced channel opening or closing, depending on the tethering location. With the use of this approach, the authors describe a unique ATP‐independent gating mechanism of P2X receptors. Accordingly, the authors suggest the name of opto‐gating to describe this process, as the photoswitchable reagents do not include the ATP ligand, nor target any particular active or known allosteric sites of the receptors [151]. The same group next developed a new type of photoswitch they name photoswitchable tweezers (PXs). PXs are closely related to PTLs, but instead of tethering a single cysteine residue, they tether two cysteine residues resembling molecular rulers [152]. However, unlike the latter, PXs are capable of pulling or pushing on gating elements of the receptor during isomerization (Fig 1J) [153]. With these, the authors induce forceps‐like motion to open the pore of the P2X channels, resembling the motion of a camera's iris.
Nicotinic acetylcholine receptors
Acetylcholine (ACh) is an important neurotransmitter in both central and peripheral nervous systems (CNS and PNS, respectively). Its actions are mediated by two types of receptors—the G‐protein‐coupled muscarinic receptor (mAChR) and the nicotinic AChRs (nAChRs) [154]. nAChRs function as non‐selective, excitatory cation channels with roles in the sympathetic and parasympathetic nervous system, and in skeletal muscle, they mediate contraction. In the brain, however, their roles are much less understood. They are found throughout the neuron's membrane with no obvious concentration at synaptic junctions. In fact, a large proportion of receptors can be found at nerve terminals. These receptors are considered neuromodulatory agents for neurotransmitter release and are implicated in numerous diseases [154], [155].
As noted above, the history of synthetic optogenetics intimately involves these receptors (e.g., PCL‐Bis‐Q [65], [70] and PTL‐Q‐Br [70], [156] and see above). However, Bis‐Q was found to be a muscle‐type photoagonist [157], and Q‐Br could not be targeted to defined subunits, as it conjugated native cysteines located next to the pore of the channel, extant in several subunits [70]. Present studies have now elaborated on these earlier reports and produced several light‐gated nAChRs via azobenzene‐based PCLs and PTLs. Azocholine (azobenzene‐choline) is a photoagonizing PCL that targets and activates α7‐type nAChRs [157]. Two genetically modified β subunits (2 and 4) have been engineered to covalently bind the photoactivating PTL, MAACh (maleimide–azobenzene–acylcholine), or the deactivating PTL, MAHoCh (maleimide–azobenzene–homocholine). The latter enable to photocontrol unique β2/4‐containing nAChRs receptors, denoted LinAChRs [158].
Next‐generation photoswitches
Spatially confine photomanipulation of light‐gated receptors located deep within scattering tissue is best achieved by 2PE [159]. Whereas 2PE is readily used for photolysis of caged compounds [160], [161], [162] or for the photoactivation of photoreceptor‐based optogenetic tools (e.g., [163], [164], [165], [166], [167], [168]), the activation of synthetic optogenetic tools by 2PE is far behind. This stems from the azobenzene's poor 2P absorption properties, owing to the molecule's symmetry [169]. Among the first to tackle this issue was the group of Gorostiza Pau. In order to increase the absorption cross section of the L‐MAG photoswitch, the authors modified it to increase its push–pull characteristics or introduced light‐sensitive antennae to sensitize the isomerization process by resonance. These modifications yielded MAG2p and MAGA2p, respectively [170]. The authors then used 2PE to photoactivate LiGluR in cultured neurons and astrocytes. Comparably, we have used a red‐shifted MAG photoswitch, denoted L‐ and D‐MAG460 [171], to photoactivate LiGluR or LimGluRs, respectively, in neurons [101]. Additional 2P‐compatible photoswitches have also been reported (e.g., [119], [127]), showing that the field is steadily transitioning in the direction of 2PE, which is more suitable for in vivo applications. Notably, the transition from a 2PE‐incompatible to 2PE‐compatible photoswitch requires the design of an entirely new photoswitch by methods as described above. This may result in an altered or non‐functional photoswitch. Therefore, as the number of 2PE‐incompatible photoswitches exceeds, by far, that of compatible ones, it will take some time before we see photoswitches with revisited 2PE‐compatible design.
Bringing synthetic optogenetics to the clinic
Given that the use of photoswitches for clinical applications depends on the ability to deliver light into the tissue, it is not surprising that most of the translational progress made with synthetic optogenetics has mainly focused on the visual system [100], [102], [172], [173], [174]. Collectively, these reports show that a variety of photoswitches and modified receptors can endow surviving neurons of the degenerating retina with sensitivity to environmental light and partially restore visual function to blind mice (see review in [175]). However, there are still several hurdles to pass before this treatment transitions from bench to bedside, such as toxicity and solubility issues (see Box 1).
Another potentially relevant clinical application of photoswitches pertains to chronic pain. The Kramer group has developed a cell‐impermeant PCL named QAQ, containing two QA groups spanning the azobenzene. QAQ enables photosilencing of neurons by non‐specifically inhibiting several voltage‐gated channels (NaV, KV, and CaV channels [176]). Interestingly, QAQ exerts its block from the intracellular side of the membrane. To reach the intracellular, QAQ must enter the cells via a membrane conduit, such as the pore‐dilated TRPV1 channel—the major heat sensors in nociceptor neurons—that allows permeation of large and charged molecules such as QAQ (Fig 4B). Because pathological pain is characterized by hyperactivity of nociceptor neurons (which may also exhibit increased activity of TRPV1), QAQ can thereby selectively enter hyperactive nociceptors to silence them. While the pore dilation mechanism is currently under debate [177], this mechanism has been established for a lidocaine‐related anesthetic, QX‐314 [178].
As a therapy for cancer, a new class of photoswitches—photostatins—has recently emerged. These PCLs bind to and interfere with microtubule dynamics, inhibit mitosis, and promote apoptosis [179]. These photoswitches are based on combretastatin A‐4, a potential chemotherapeutic drug in clinical trials. The photoswitch consists of a fragmented drug, whose parts span the azobenzene‐core. Following isomerization to –cis, the two fragments are combined, yielding the final active form of the drug (Fig 1I).
Synthetic optogenetics was once considered a highly specialized technique, benefitting but a handful of laboratories. Today, however, the technique has become more approachable, judging by the growing number of laboratories across the world employing and developing the technique. To date, the variety of tools makes it possible to explore the role and signaling mechanisms of numerous receptors subtypes, in their natural environment, in vitro as well as in vivo. Nevertheless, there are still several hurdles to cross before synthetic optogenetics becomes widely accepted and clinically used (see Box 1). Luckily, major steps have been taken in that direction, with better photoswitches and gating mechanisms increasingly appearing in the literature.
It is important to recognize that whereas photoreceptor‐based optogenetic tools are unsurpassed for controlling mechanisms such as cellular excitability, synthetic optogenetic tools are more suited for controlling native receptor function or for modulating endogenous signaling mechanisms. In fact, the two techniques are highly complementary. For example, ChR2 and LiGluNs could be used concomitantly at the synapse to induce LTP. In such an instance, photoactivation of ChR2 would be required to depolarize the synapse and relieve the Mg2+‐block from LiGluNs. Then, precisely timed photoactivation of LiGluN would induce robust Ca2+‐entry, leading to LTP at the synapse.
Nevertheless, despite the numerous photoswitches produced, synthetic optogenetics is still not as broadly employed by the neuroscience community, especially when compared with opsin‐based optogenetic tools. One of the main reasons for this situation is the lack of commercial availability or absence of a chemical depository (but see exceptions in Box 1). It is unlikely that chemical companies will start synthesizing photoswitches for several reasons: the arduous and expensive synthesis process, low demand, therefore narrow profit margin, and patent issues. Secondly, though a chemical depository would certainly help for research groups to gain access to the compounds, such a chemical bank would need to eventually synthesize the photoswitches in‐house, which is expensive and thus highly unlikely. To date, the most common practice is to directly obtain photoswitches from the developing groups, despite potential conflicts of interests or competitions. In fact, as far as we are aware, most research groups are open to distribute their photoswitches and/or start collaborations.
Taken together, we expect the technique to be further perfected and optimized and hope to see many more exciting developments in the near future, such as novel light‐gated receptors, multiphoton‐compatible switches applied in vivo, and perhaps even acoustic and magneto‐switches.
Conflict of interest
The authors declare that they have no conflict of interest.
See the Glossary for abbreviations used in this article.
- Glossary
- two‐photon excitation
- acrylamide–azobenzene–QA
- ACh
- acetylcholine
- action potential
- benzylguanine‐azoglutamate
- ChR2
- channelrhodopsin‐2
- central nervous system
- diphenylcarbamyl‐chloride
- endoplasmic reticulum
- γ‐aminobutyric acid
- G‐protein activated inwardly rectifying potassium channel
- G‐protein‐coupled receptor
- ligand‐binding domain
- LiGluN
- light‐gated NMDA glutamate receptors
- LiGluR
- light‐gated kainate glutamate receptors
- LimGluRs
- light‐gated metabotropic glutamate receptors
- maleimide–azobenzene and 4'L stereochemistry glutamate
- long‐term potentiation
- mAChR
- muscarinic acetylcholine receptor
- maleimide–azobenzene–QA
- MscL
- mechanosensitive channel of large conductance
- nAChR
- nicotinic acetylcholine receptor
- nAChR
- nicotinic acetylcholine receptors
- N‐methyl‐D‐aspartate receptors
- purinergic (ATP‐gated) receptors
- PALs
- photoswitchable affinity labels
- p‐azophenyldiphenylcarbamyl‐chloride
- photochromic ligand
- plasma membrane
- peripheral nervous system
- photoswitchable tethered ligands
- pertussis toxin
- photoswitchable tweezers
- quaternary ammonium
- retinal ganglion cell
- O6‐alkylguanine‐DNA alkyltransferase
- synthetic photoisomerizable azobenzene‐regulated K+ channel
- targeted covalent photoswitch
- transmembrane
- two‐pore‐domain potassium channel
- visible light operated GIRK opener
- wt
- wild type
- © 2017 The Authors


text:
Sporting Goods and the Chief Purchasing Influencer
Youth Athletics are Changing the Ways Families Spend Free Time
Youth Athletics are Changing the Ways Families Spend Free Time
It's no secret that kids influence household purchases.
For sporting goods, that can be significant when your son or daughter has an affinity toward a particular brand. There are a few factors fueling this influence, one being the size of the market and its orientation toward non-school youth sports. Another is the rise to meet demand of the private, non-school programs that increasingly resemble professional leagues, with selection and competition at earlier ages. In the middle are the parents, being asked to continuously invest in the child with deep, do-anything-for-you love and an open wallet.
It's estimated that at least 35 million kids between the ages of 5 and 18 currently play an organized sport each year in the U.S. Twenty-one million are involved in non-school youth sports, which can include baseball, soccer, lacrosse, rowing, volleyball, and gymnastics. The sports are usually organized through local community programs, such as soccer clubs, that are increasingly funded by private business and run by local, regional, and/or national oversight bodies. According to WinterGreen research, the U.S. youth sports market, which includes everything from travel to private coaching to apps that organize leagues, is now $15.3 billion (larger than the NFL's $14 billion). This is giving rise to new aspects of the market. Major media and retail companies are investing in schedule-managing technology; municipalities are banking on youth sports and issuing bonds for complexes to lure equipment-toting youth and their families.
While the primary benefit of youth sports is fun for the kids, others include building strong, healthy bodies and personalities, developing team skills, and preparing them for success in high school, collegiate, or professional sports. Sports clubs have helped create a large market from what were, until only recently, disparate parts: people buying clothes and equipment, and loosely organized groups of youth playing games on community fields coached by volunteer parents. Now, these teams depend on tournament play, video coaching, and advanced communication to compete effectively.
The web has emerged as the key middleman, playing the roles of scheduler, payment engine, and hype machine in one. There are sites now offering scouting reports and rankings for youth sports. Parents and children alike share video via the internet so that family and friends can experience the action from afar. Video capture of game and practice play provides coaches with yet another tool to help players improve on the field. Some of this video even makes its way to being broadcast on the internet (see https://www.basketballislifekids.com/).
Travel teams are gaining traction in part because they can provide some family weekend vacation fun in the process. As an emerging industry, there's potential for the sale of not only equipment, but also for ancillary services like hotels, airline fares, and car rentals.
The heightened attention and structure surrounding the pursuit of youth athletics, for entire households, can increase the potential for and necessity of more frequent and expensive athletic-related purchases.
"Decision-making within families today is almost entirely collaborative — and as kids become more influential, they're impacting purchasing decisions,"
-Christian Kurz, vice president of research at Viacom International Media Networks.
A 2012 study by Nickelodeon, which Kurz oversaw, found that 71% of parents say they solicit opinions from their kids regarding purchases for the kids themselves, and more than 60% when making family purchases.
The typical 13- to 17-year-old shopper lists quality products, brands offered, and price as three of the top reasons for patronizing a given retailer. In a study conducted by Touchstone Research, teens said that owning or using brand-name products is particularly important to them in apparel, footwear, and sports equipment categories. As youths mature and develop their own sense of fashion and personal aesthetic, product design can also play a part. Think of the youth basketball player who idolizes Lebron James. Anthony G. Lee of Saint John Fisher College cites Noble, Haytko, and Phillips' study (2009) and how they use the Socialization Theory to help explain the way the younger generations choose consumer goods, incorporating the ideas of "freedom, finding yourself, blending in/out, brand personality, fashion knowledge, value seeking, and comfort of brands" as intrinsic and extrinsic motivators.
Approximately 79 percent of teen girls and 76 percent of teen boys shop online, which accounts for 18 percent of their total spending. Integrating ads into non-traditional media content like social media, events, and sponsorships can provide cost-effective, high-impact ways to reach this target. Youth want to make consumer choices that reflect — and help define — the identities they are creating for themselves. Many reason that "if it's good enough for a pro athlete, then it's more than good enough for me." Equipment endorsed by pro athletes can be a key early indicator of a product's performance. At ICF Olson, we understood this when beginning our work for Bauer hockey equipment, where we used both professional hockey players and youth influencers in showing devotional and passionate, yet jaunty hockey life in a series of short films custom-built for the web and social media. This viral, true to life messaging helped us push Bauer from the #4 player in the category to #1. View our Bauer case study here: http://www.icfolson.com/case-study/hockey-never-sleeps
The sporting goods industry has opportunities to grow commensurate with the passion of youth athletes and their families. Let's get going!


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Lorraine really understood what we were looking for, offered great pre-marketing advice and worked very hard for us and keep us very well informed.
Stella and Mark
She was friendly, helpful and seemed caring and passionate about what she does. Was a great communicator. So grateful!
Lorraine was amazing with a very trying property that had a lot of negative aspects but also positive aspects too and Lorraine made the most of it and we were very happy with the outcome.
Annabelle & Alexander
Professional, extremely helpful, great communication, personal service
Andrew and Joanne - William Brittan Avenue
Lorraine did everything in her power to help us through our first home purchase. She was very accommodating,helpful and patient with us through the biggest step of our lives.
Shaun & Lisa
Very pleased with all the process. Impressed with the help Lorraine has given us.
Lionel & Rachelle
Lorraine , was very helpful,friendly and made the purchase of our house stress free.
Simon & Corina - Seaforth Place
Lorraine Is Quick to Respond to Any Request and is Really helpful with the whole selling process.
Neil and Susan - Saddleback Green
The Agents were very pleasant to deal with, and we did feel they did their very best to sell our home as quickly as they could. We must add they did all they could to make it stress free for us. We do wish to thank them for this.
SR & SJ Munn - Burbank Drive
Impressed with the service received
Lorraine was professional, accommodating, friendly and efficient. We spoke with agent's from three companies prior to making a decision on who we would list with. Lorraine's initial presentation was by far the best. She explained things thoroughly and professionally and we developed a good rapport with her right from the start. She kept us informed right throughout the process and was open and honest.
Professional. Great knowledge, excellent communicator.
Gail & Brian
Good communication, great advice.
Chris & Kelly - Isobel Street
Lorraine was extremely friendly and professional. The process was very stressful for us but she was re-assuring and accommodating the whole way through :)
Felicity & Andrew - Sabys Road
Lorraine has always kept me well informed and up to date with everything.
Very happy with the agent. Communication very good keeps us up to date with progress.
G & S Smith - Sabys Road
We sold our house using Ray White and once again the process was seamless, informative, friendly and professional.
G & L Besley - Gillatt Gardens
Awesome service, friendly hard worker.
M & C McGimpsey - Gillatt Gardens
Very professional, kept us fully informed every step of the way. Was sure we were realistic in our price expectations, so we could set our budget. Went the extra mile following up and encouraging purchasers.
K, T & P Studholme & C Lamb - Kilbride Gardens
Lorraine was very approachable and business like and worked very hard to achieve the sale.
B & J Morrison - Hoon Hay Road
The documents are well presented. Communications and details are prompt and delivered well. We recommend Ray White. Thank you so much.
A & K Lee - Craven Street
Went over and beyond her job description to make the tenant feel OK during the process. Very patient, and always answered her phone.
S Yaxley & M Tully - Craven Street
Excellent professional service. Result was higher than expectation.
S Griffiths - Kowhai Terrace
Excellent professional service. Result was higher than expectation.
C/- Sue Gardener
Pleasant and very helpful.
D May & C Martin - Green Street
Lorraine's attention to detail from the very first meeting to view the house and her thorough understanding through out the selling process.
Bruce & Anna
Lorraine was professional, friendly, easy to contact and discuss any matters with. Nothing was a problem, she went beyond expectations even to helping stage the house with some of her own furnishings.
Mr & Mrs Edens - Boon Street
She listened to us, then advised us and came up with a strategy that met our hopes and expectations.
J & A Wilson - Waimairi Road
Professional, listened, outstanding, understanding, awesome experience, trustworthy.
E & A Bool - Kumara Place
I am a repeat customer of Lorraine's and have already recommended her services this week.
Winifred & Rob - Hoon Hay Road
Lorraine was incredibly professional, and she achieved the desired outcome.
K McWilliams - Cliffton Terrace
What a lovely girl Lorraine is.
P Harwood - Pages Road
Really happy with the marketing of the property as we had lots of potential bidders at auction, this meant we got the price for sale we were after.
R Grant - Centaurus Road
Lorraine did a fantastic job of getting people through the door, following up on potential buyers and advising on our offer. She is wonderfully positive and approachable and has kept us well informed on the progress of our sale.
J Holt - Nicholls Street
Really good service, really easy process and the house sold quickly.
J Stringer - Poulson Street
We were really pleased with the whole sale process, we were kept well informed all the way through.
M & L Mckenzie - Cradock Street
Awesome service. Great advice and goes above and beyond to help us in making the right decisions.
B Sangster - Salmond Street
Thank you so much for all your support and effort into the selling of our home. It has been so much appreciated.
P & M Chamberlain - Hoon Hay Road
Lorraine deserves a medal for her unflagging efforts against the enemies of the insurance cos and a reluctance of Christchurch people to buy real estate in the city.
R Linklater - Nottingham Avenue
We both felt you were excellent to work with and quick to action any queries. You also stood out, as you were easy to get on with and enabled us to feel as though we were your only clients! I believe we had a good working relationship and wouldn't hesitate to recommend you! We love our house.
V Halliday & G O'Brien - Broken Run
I engaged the services of Lorraine to sell my Christchurch property in September 2012. Lorraine went the full mile - all of the various aspects of the sale process were well explained, and the best possible strategy adopted. Lorraine achieved a very good result. She always kept in regular contact and was positive at all times. I therefore have no hesitation in recommending Lorraine to all prospective home property buyers and sellers.
R & P Savage - Antonia Place
We both found you excellent to work with & quick to action any queries. You also stood out, as you were easy to get on with & enabled me to feel as though we were your only clients! I believe we had a good working relationship & wouldn't hesitate to recommend you. We love our house & feel pretty organised, now it's time to plan garden boxes & getting into the lawns. Many thanks.
V Halliday & G O'Brien - Broken Run
Lorraine is adorable to work with.
R Summers - Francis Avenue
Lorraine is enthusiastic, committed, direct, thoroughly professional, with a warm and friendly manner. I thoroughly enjoyed working with Lorraine. Her business-like approach combined with personal interest and a sense of humour was very refreshing - will happily recommend her to all my friends and associates.
K Somervell - Deloraine Street
We feel that we can trust her. Easy to talk to, approachable and can listen to what we wanted. Pleased to have sold and think Lorraine worked hard for us. Thank again!
T Hamilton - Whincops Road
Lorraine was very helpful and made everything as smooth as possible. Thank you very much for everything.
C Beecroft & N Webber - Oakhampton Street
Lorraine was very professional and friendly and I would have no hesitation in recommending her to anyone either buying or selling.
K Andrew - Kedleston Drive
Lorraine helped to make the buying process very simple and straight forward, not to mention quick. Her friendly, professional manner made the purchase a pleasure.
J Holt - William Brittan Avenue
I found Lorraine to be fabulous. I knew everything I needed to know. Selling my house was a new experience for me. Lorraine made it easy. She expected everything & helped me to get the best price I had already consulted other agents. What I got for my house was way above other quotes.
R Kelly - Edinburgh Street
Lorraine was very professional and committed to selling our home. Great feedback, contactable 24hrs! Lorraine worked very hard to sell our home and to also sell our buyers home in order for us to be able to sell this house. It is a dream come true for us to be able to move on from this house and start our new life together over in Bishopdale. Thank you Lorraine 'you did it'. Lorraine is a wonderful agent. I hope she gets many more sales in the future!
T Graham - Halswell Junction Road
A huge thank you to Lorraine, she has made the processing of my house stress free. Lorraines verbal and written feedback has been fab and I can see that she is commited to providing a great service. I look forward to recommending Lorraine to family and friends.
M McArthur - Sabys Road
Lorraine kept us informed and involved throughout the process. It felt like we were a team, with the goal of selling. This happened sooner and with a better outcome than expected.
D & D Gorst - Rollesby Street
Lorraine is friendly and approachable.
R & A Stein - Calverton Place
Fantastic professional service—highly recommend Lorraine.
C & A Gallagher - Frances Avenue
I would like to thank you for your help and support, you are more than just OUR real estate agent, I consider you a wonderful friend and I will keep in touch. Again Lorraine you have out shined yourself, Paul & I think your most likely the best real estate person there is.
P & D Porter - Cridland Place
Lorraine has been great. She is friendly, approachable and is a great sales agent. We would definitely recommend Lorraine to anyone.
O Bandaranaike & U Kudaliyanage - Somerville Crescent
Lorraine was an amazing salesperson, always fully informed us of any enquiries etc and kept us realistic throughout the marketing and sale of our property - thanks heaps. Ray White is a very well run friendly business - thanks for all your hard work in these difficult times.
M Withers & J Zamitt-Withers - Main Race Road
Exceptionally good close contact throughout sale including auction process. Lorraine was a great to work with, very considerate, good communicator.
A Simmonds - Halswell Road
Lorraine was a great agent to work with, very supportive, good suggestions for presentation and always positive that we would have a successful outcome. We have no hesitation in recommending her to our friends or colleagues. Thank you for everything.
K & R White - Patterson Terrace
We think Lorraine did a good job, had good open home attendances and the end result was great.
C & R Forrest - Halswell Road
Lorraine was professional, friendly, easy to contact and discuss any matters with. Nothing was a problem, went beyond expectations even to helping stage the house with some of her own furnishings.
Mr & Mrs Edens - Boon Street
She listened to us, then advised us and came up with a strategy that met our hopes and expectations.
J & A Wilson - Carruthers Street
Professional, listened, outstanding, understanding, awesome experience, trustworthy.
Eileen & Alan
Lorraine and Shala were incredibly professional, and they achieved the desired outcome.
K McWilliams - Clifton Terrace
I am a repeat customer of Lorraines and have already recommended her services this week.
Rob and Dianne
Awesome service. Great advice and goes above and beyond to help us in making the right decisions.
This agent goes out of her way to give the best support for all parties. Nothing is too much trouble and her bright manner is comforting at this stressful time.we could not have done this without her knowledge of everything.
Alan and Catherine
Very helpful in getting everything ready for sale, plus ensuring any hurdles thrown up by others were dealt with OK.
V Halliday & G O'Brien - Broken Run
Lorraine managed to achieve an excellent price for the house that we sold.
N Smith and M Fallows - Geraldine Street
I found Lorraine professional in her approach to selling my property. She was honest and approachable.
Very happy with Lorraine's work even though my situation wasn't the easiest we managed to get through it all and sell the house I could not fault Lorraine.


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I have a little card I made tonight for a few challenges. I've had the Camping Critters cart for a while now and had never used it. I was really excited to open it up tonight. Hope you like it!!
|Skunk: Campin'Critters
Sun: Simply Charmed
Waves: Life is a Beach
Thank you: Inkadinkado-Happy Everything stamps
PKS face: Everyday Character Face Assortment
Paper Playtime-use water
Clear It Out-Summer is here
My Craft Spot-It's Hot out there
Cricut Cardz-Thank you, use hearts, no pink or red
Cricut Circle-Create a Scene
Also, if you haven't entered my GIVEAWAY, click HERE for a chance to win...


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History is rife with stories of how people bursting with creativity were forced to take traditional safe jobs in deadening organizations while they toiled at creative pursuits on the side.
Early American writer Nathanial Hawthorne was a customs agent. The ultra creative Franz Kafka worked for insurance companies. I had a relative who became one of Holland's greatest regional writers, but spent his entire life working as an office manager. He would get up at 4 am each day to write before he went to work.
This imperative to take a safe but boring job popped into my head during a recent discussion with someone who was tasked with reinventing a business operation that has been running sleepily in well-grooved tracks for some time.
My friend knew his industry very well, and was a good analyst, so could see what was needed to put some life back into the organization, and revive the business.
But he couldn't figure out how to make the conversion. The organization was operating in the creative field but had long ago lost its creativity. Working there had become an annuity job for most of the people in it, providing them with a very nice life, even though they knew the business wasn't working.
They had made the choice: Shelve their innate creativity, keep their mouths shut, and let the place lumber along. This is not an unfamiliar situation. The rise of entrepreneurship aside, society generally encourages us to take the "safe" steady job that provides us with a good consumer-oriented life.
My friend correctly deduced that as an outsider, he had to find some champions from within the company to make change happen. But it was clear that those champions were not going to be among the central management who had a strong incentive to maintain the status quo. So where would he find them?
Find the secret rebels, I said. And let them do their thing.
All organizations have rebels, but the organizational imperative is to usually kill them, or at least neutralize them so they don't harm the smooth running of the operational machinery. Over time organizations can form antibodies – conventional thinking, a unique language, a socially-imposed groupthink – with a sole purpose of destroying anything that upsets the system.
So the rebels usually hide, sometimes quietly nurturing their creativity in their outside world. I'm convinced this is one reason for the rise of social networking – it allows people to express their creativity. Online, everyone can reinvent themselves.
In the 21st century no organization can remain content to operate in familiar grooves that have become ruts. Change is the norm today, and if you don't sieze it and work with it, you probably won't survive.
So, since your organizational life may depend on it, you might want to make an extra effort to find those secret rebels. Then let them apply their creativity inside instead of outside where that creative power is lost to you.


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by Cindy Pena
The women's right to vote amendment was passed and implemented almost 100 years ago. The fight to get this passed was definitely not an easy one. It required time, determination and most importantly, unity. Unity with all women to fight for what they deserved; it was a fight for political representation.
However, until this day, women are still extremely underrepresented in politics. According to the Center for American Women and Politics, nationally, women make up 19.6 percent of the 535 seats in Congress: 21 percent of the 100 seats in the U.S. Senate and 19.3 percent of the 435 seats in the House.
In 2016, Connecticut was listed as one of the top states for women in politics with 28 percent of elected state officials being women and 53 of female elected officials in the state legislature, according to CT Post. InsideGov used information from the National Conference of State Legislatures and ranked Connecticut the fifteenth highest state of women in elected office. Although this is a good sign, there is still room for improvement.
The underrepresentation of women in politics means laws pertaining to women's rights, like paid family and medical leave, are created mostly by men. That needs to change. We need more women to enter the political arena to not only to represent womens issues, but also to inspire the younger generation to do so as well.
Mayor of New Britain Erin Stewart is one woman leader breaking that mold. Stewart is the 40th mayor of New Britain and is the youngest mayor in the United States. She went to New Britain High School and is a Central Connecticut State University alumna. She is also a role model to women and girls in New Britain who may have political aspirations.
Her impact in New Britain is tremendous. She has revitalized the New Britain area, improved the economic state of New Britain, and worked with community members, like CCSU, to better New Britain. She is just one local example of a woman flourishing in the political arena, and I know there are more around the U.S.
Ultimately, we need more women like Erin Stewart to not only motivate women, but create future political leaders. We have made substantial progress since 1920 when the 19th Amendment was passed, but unfortunately not enough. So go vote, talk to your congressperson, or even run for office; let your voice be heard.
As Michelle Obama once stated, "You may not always have a comfortable life and you will not always be able to solve all of the world's problems at once but don't ever underestimate the importance you can have, because history has shown us that courage can be contagious and hope can take on a life of its own."


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KQBR Job Opening: Digital Sales Executive
EXPERIENCE/ SKILLS: As part of an on-going recruitment campaign for our digital advertising sales team, Townsquare Media Lubbock is looking for a Dedicated Digital Seller who has the ability and desire to make a lot of calls to businesses, create relationships with new potential clients and create great on-going digital advertising campaigns. The Townsquare digital initiatives include display, streaming, custom sites, loyalty programs, e-commerce, Seize the Deal, Townsquare Interactive and other digital products.
Candidates must demonstrate dependability, passion, strong communication and problem-solving skills. Business acumen, open-mindedness and a sense of what it takes to win are essential. Previous digital or sales experience a plus.
JOB SUMMARY: A Townsquare Media Dedicated Digital Seller is the primary contact for new prospective digital customers. You will develop business relationships and sell our digital portfolio of products, by performing the following duties:
Essential Duties and Responsibilities
- Promote and sell digital products to new prospective customers
- Consistently be cold-calling to develop new customer leads
- The majority of your time will be spent on the street meeting, inquiring and crafting digital advertising programs for your clients
- Prepares digital advertising schedules, sales literature, proposals and sales contracts.
- Enters new customer data and other sales contract details for digital clients into computer database.
- Follow accountabilities set forth by your Digital Sales Manager to help guide you to success
- Investigates and resolves client issues.
CONTACT: Blake Davis – firstname.lastname@example.com
Townsquare Media is an equal opportunity employer


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Princess Pandora - Queen of Denial
"We are all in the gutter, but some of us are looking at the stars."
Thursday, November 24, 2016
Song a Day November: Take Me To The River
Yesterday's song is late again. Oops. Busy day yesterday, culminating in a mason's meeting.
Which is why this little ditty, penned in the seventies and covered in the early eighties by Talking Heads.
I never tire of this song.
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Originally from France, Février specializes in using firearms, such as submachine guns and shotguns, and also grenades. She fights in the frontlines, although she isn't good in close range combat. She also fears nothing, except for Enero when she is angry.
Street Fighter V delves further into her personality, depicting her as an overall angry individual devoid of manners, perhaps reflected by her preference of weapon. She seems to prefer to solve her problems with insults and a handful of bullets.
A unique trait of Février's is that her hair color changes with each appearance; her hair is pink in Street Fighter Alpha 3, dark orange in the Super Street Fighter IV OVA, and brown in Street Fighter V.
Street Fighter VEdit
A Shadow FallsEdit
In the events of A Shadow Falls, she joins the other Dolls in a counterattack on the Shadaloo base's assailants; she finds her adversary Birdie ugly and, disgusted by his appearance, relentlessly barrages him with insults and bullets but, thanks to Birdie's chains around his arms, he remains unfazed as their fight begins. Février loses her fight horribly and throws one last insult at Birdie before dropping unconscious.
Fighting style Edit
Février's fighting style mirrors that of the other Dolls, being fairly agile and having access to impressive combos, but not entirely impressive on defense.
Most dolls have only one special, and in Février's case, it involves the use of her Psycho Power-infused Uzi to shoot foes at mid range (despite being a gun, its range is actually rather poor). She is able to angle the shot low, medium, or high, perhaps determined by her AI using the light, medium, or heavy "button". She can combo into all versions of this move except the EX version. The EX does much more damage on its own, gradually covers a wide range in front of her, and is safer on block than the move's other versions.
Trivia Edit
- Although unplayable, Février is the only known character in the main games to use a firearm as a weapon.
|Street Fighter V Characters
| Birdie · Cammy · Chun-Li · Dhalsim · F.A.N.G · Karin · Ken · Laura
M. Bison · Nash · Necalli · R. Mika · Rashid · Ryu · Vega · Zangief
|Season 1
|Alex · Balrog · Guile · Ibuki · Juri · Urien
|Season 2
|Abigail · Akuma · Ed · Kolin · Menat · Zeku
|Season 3
|Blanka · Cody · Falke · G · Sagat · Sakura
|Season 4
|CPU Only
| AS · Aprile · Decapre · Enero · Février · Juli · März · Peter
Phantom Bison · Santamu · Satsuki · Two P


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The British singer-songwriter, from South London, reveals her unique style, which is quirky, feminine with a mixture of boyish features. This 23-year-old beauty can always be spotted in massive high heels and wearing sparkling jewelry.
Her personal style and music is already making waves stateside. Below are a few of her favorites at THE OUTNET. To shop Lianne's picks, go to: Lianne's Style
|Erdem top
|Sara Berman skirt
|Diane von Furstenberg top
|Erdem Dress
|Catherine Malandrino skirt
|Tart skirt
Elizabeth Cole
|Stella McCartney


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It's Day 29 of our "31 Days of Smarties" giveaway & count-down to Halloween!
Reason #29 why we think Smarties are a smart choice for Halloween: Smarties have been around since 1949, and the Smarties formula has changed very little over the years. This means that you may enjoy the same Smarties candy in adulthood that you did as a kid. So keep treating yourself to Smarties and the Smarties family will continue to do what they do best – candy making!
Care to win one of 31 Smarties prizes? If so, now it's your turn to tell us why Smarties are your smart choice for Halloween!
Keep reading for the giveaway details:
On each of the 31 days of October, we will post a reason why we think Smarties are the smart choice for Halloween & for each of the 31 days of October, we will give away a bag of Smarties AND a Smarties Theater Box!
That's 31 Smarties prizes for 31 days of Smarties!!! You can enter to win one of 31 prizes in 2 ways:
1) Like Smarties on Facebook and post a comment, photo with caption or video on our wall about why Smarties are YOUR smart choice for Halloween. An entry comment, caption or video may have a message like this: "Smarties are my smart choice for Halloween because…"
2) Follow @Smarties on Twitter and tweet why Smarties are your smart choice for Halloween with the hash tag #SmartHalloween and mention @Smarties. An entry tweet may look like this: "Having a #SmartHalloween with @Smarties because…"
Entries will be accepted from October 1st to October 31st at 11am EST. The 31 winners will be announced on Halloween (Monday, October 31st) at noon!
Up to three entries per person, per day. Entries will be judged for fun-factor, creativity and Smartie-ness. Winners must provide a mailing address within the USA. Good luck!!!
– The Smarties Family


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Justin Bieber's 'Calvin Klein' Ad Gets Spoofed on 'SNL' (Video)
Kate McKinnon channels her best Justin Bieber in this hilarious spoof from last night's (January 17) episode of Saturday Night Live in New York City.
Co-starring Cecily Strong as model Lara Stone, "Justin" acts super immature during the commercial by playing pranks, crying, and trying to harass the model.
PHOTOS: Check out the latest pics of Justin Bieber
He says things like "This tattoo made me say 'Owie" and "I'm not supposed to drink, but I do," later adding about his underwear, "Yo, my peeppe's in there."
Watch the funny skit below!
Justin Bieber Calvin Klein SNL Spoof


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- Christy42213
- 6 years ago
- Wedding: April 2013 - Rhode Island
So we FINALLY got some of our engagement photos back!!!! These aren't the high res ones. Don't have those yet. But I was too excited to wait and had to share with someone. 😉 Let me know what you think! By the way, we're getting married in only 6 days!!!! Eeeeeeeeeehhhhh!!!!!!!!
Photos by Adeline & Grace Photography.