Memory and Location, Location, Location

When you remember first meeting the love of your life, do you also have a strong memory of where you both were and also where you were in relation to objects in the scene? When I first met my wife, Doris, it was at a party and she was at a piano surround by “bird dog” males, who I saw from an adjacent room. In my mind’s eye, I still see both rooms and where everybody was.

Do you remember where you were on 9/11? I was in the waiting room of a hospital, looking over a series of lounge chairs at a large-screen TV program that was reporting the news.

It seems that many people remember not only events but where they were at the time of the event. But how does this happen? We do know that a new experience may be “consolidated” into a lasting memory, especially if it stirs emotion and you replay it in your mind. That is certainly the case when you meet the love of your life or see a terrible event.

If you were there, you would surely remember what you were doing.

Back in the 1970s I was studying the part of the brain known as the hippocampus, and it was known at the time that this structure is crucial for consolidating memories. I and others were focused on an EEG rhythm (theta rhythm of 4-7 waves per second) that was especially prominent when an animal moves around in an enclosure EEG signals are summed over dozens of neurons, and therefore to get more precise data some investigators put microelectrodes into the hippocampus so they could monitor the nerve impulse activity of single neurons as the animal moved around.

It was quickly discovered that some hippocampal neurons fired impulses selectively when an animal was in a special location within the enclosure. Collectively, these “place” neurons were actually mapping the enclosure space and tracking the animal’s position as it moved around in this space.

New insight on an additional role for place neurons has come from a new research report on human epileptics with electrodes implanted in their hippocampus to locate the diseased tissue. These patients played a virtual-reality game in which their avatar drove through a virtual town and delivered items to stores. Their task was to memorize the layout and what was delivered at each store. Meanwhile, place cells in the hippocampus were monitored and their place coding was noted. Then when participants were asked to recall the memory of what went where, the place-responsive activity was reinstated even though the subjects were not actually playing the game but recalling it from memory. And the activity of place cells was similar to that during the learning stage.

In other words, neural representations of the content of the experience had become linked with the spatial and temporal context. Such evidence provides strong evidence for the theory that memory formation and recall involve association of event with context, especially spatial and temporal context. This linkage creates a mutually reinforcing interaction of event and location. We tend to remember both or neither.

Can we apply these findings to improving everyday learning and memory situations? Of course, we can. The key elements for making it easier to learn something new are to:

1.  Identify a context that stirs emotions, preferably positive emotions like meeting someone you are attracted to.

2.  Be especially aware of where you are at the time and where you are in relation to the location of various objects.

The hippocampus uses these emotional and spatial cues to facilitate the consolidation of memory. We know that memory is promoted by making associations. Emotions and spatial cues are probably the most effective kinds of cues.

Sources: Miller, J. F. et al. (2013) Neural activity in human hippocampal formation reveals the spatial context of retrieved memories. Science. 342, 1111-1114

END NOTE: If you find these posts helpful, you are not alone. I am gratified to have so many readers. My reader views here and at a cross-posting site now total over 800,000. Thank you so much for wanting to read what I write. You might also want to read some of my books: see http://thankyoubrain.com

The Role of Research Funding in Education Practice

If we learn anything about educational policy, we should learn that what we have tried over the last couple of decades to improve student achievement doesn’t work very well. I won’t bore you with all the statistics showing that academic ability of U.S. students lags that of most other developed countries and our ranking is not improving. I would like to explore one reason for lack of progress besides faulty federal policy. And that reason is research funding. We don’t know enough about how brains learn and remember, nor how to apply what we do know to educational policy.

The recently announced winners of the Nobel Prize included nine from the U.S. In a recent joint interview by leaders of the American Association of Science, the winners spent little time discussing their research achievements, preferring to expound on some serious concerns about how research is now funded in the U.S.

Of course, some of these concerns could be considered whining. Like much of the general public, the scientific community has become dependent on government. When federal funds don’t grow at high rates, scientists can be cry-babies too. Nobody seems to care much about the growing federal debt and a very real threat of looming financial disaster in this country. Interest groups, including those in science, think if we have to cut funding, it should come out of somebody else’s piece of the pie. As a result, nothing much gets cut anywhere.

But there are legitimate issues about science policy and how federal money is allocated for research. The U.S. leads the world in Nobel Prizes by about 3 to 1. Actually, this understates our prowess. Young scientists from all over the world come here to learn world-class science, but we make most of them go back home to become science superstars in their own country. At the same time, we have little stomach for deporting uneducated illegal aliens. It’s hard to think of a more stupid immigration policy.

Randy Schekman, physiology winner, said that many of our best young foreign scientists are "returning to their countries because those countries, unlike ours, see the promise of investment in basic science. It's actual damage that's occurring right now." Several of the other laureates concurred.

The future of U.S. science may well be in jeopardy. The new Nobelists point out that the buying power of National Institute of Health funding has shrunk by 28% over the last seven or eight years. Moreover, government needs to re-think how that money is distributed. For example:

·         Michael Levitt, who won the award for chemistry, said "A huge change in the last 30 years has been that people under 40 get almost no money, and people over 65 get lots of money. Everyone here would agree that we made our discoveries when we were under 40."

·         James Rothman, winner in physiology, blamed big-science mentality where too much money as being “allocated into pre-determined projects is heading toward bureaucrat-driven science."

I fought the grant wars for years and finally gave up. I got too few grants for all the time I wasted writing proposals. Several of the Nobelists commented on how much time scientists waste writing proposals that don’t get funded.

Another part of the problem, apparently not mentioned in the interview, is that too much money is soaked up by too few grants. Universities have negotiated enormous overhead fees, sometimes exceeding 100% of the grant, and that is money that mostly goes into the institutions’ general fund, not the funded project. Also, too much grant money goes to salary support for the scientists. It used to be that universities were committed to supporting their scientists. Now, universities expect their scientists to hustle money for the university. As a result, too much grant money gets consumed by projects that are scientifically sexy and will sell, not necessarily for the most promising science. An associated problem is that government spends way too much grant money pursuing scientific fads. Far too many areas in science have no chance of getting competitive funding simply because they are currently unfashionable.

Source:

http://membercentral.aaas.org/announcements/us-nobel-prize-winners-honored-dc

Does Music Help Memory?

When I was in veterinary medical school, I could often be found lounging in the fraternity living room listening to jazz records. My classmates were stunned that I was wasting so much time, when most of them had to study while I seemingly had nothing to do. O.K., so maybe I graduated fifth in my class rather than first, but I was not nearly as stressed as my classmates.

My reason for sacrificing study time was that it bolstered my spirits. Veterinary medicine is a lot harder than most people think. Veterinarians learn the same anatomy, physiology, pharmacology, microbiology, and so on as physicians do. In some schools, human and veterinary medical students take many of the same basic science classes. Moreover, veterinary students have to learn about multiple species, learn more public health, and take a year’s worth of surgery.

But back to the music issue: some people, especially students, think that listening to music helps the memory. Historically, supporters of this practice have referred to this as the “Mozart effect.” Most students, of course, listen to pop music rather than Mozart. Students are notorious for listening to music while studying. Why isn’t music a distraction? I have written before about how extraneous stimuli can prevent memory consolidation, which in the case of studying, consumes cognitive resources and prevents the formation of memory that lasts long enough for the next examination.

Because so many students listen to music while studying, formal experiments were recently reported on whether or not that is a good thing. These experiments, conducted in Finland, had a scientific rationale. Prior research had shown that listening to music that people considered pleasurable increased the release of dopamine in the brain, and dopamine is well known as a “feel good” neurotransmitter. Other research had also shown that dopamine promotes learning to approach rewards, while a deficiency of dopamine promotes learning of punishments.

Seventy three subjects, mean age of 27.1 years, listened to a battery of 14 songs and identified three that they really liked and three that were emotionally neutral. One of each was selected for use in the study, in which subjects were grouped in four different listening patterns involving a positive (P) or neutral (N) song during study and the opposite kind of song during testing. Thus, there were four groups, NN, NP, PP, PN. Each group was formed to have an approximately equal number of musicians and non-musicians.

The learning involved memorizing 54 pairs of Japanese characters, in which one character was arbitrarily given a high reward value (a simple smiley face feedback display during training) and the other character a low reward value (frowning face feedback). In the test phase, pairs were shuffled and thus served as a measure of how well the original learning was generalized.

Results indicated that people with more musical experience 

learned better with neutral music but tested better

with pleasurable music. The opposite was true for people without music training. My explanation is that pleasurable music is a distraction for a musically trained person who could be expected to pay more attention and devote more cognitive resources to pleasurable music’s inherent structure in the process of analyzing and realizing its pleasing quality. Neutral music is more easily ignored. A central tenet of learning is that any kind of distraction impairs formation of memory. The musically untrained people learned better with positive music, presumably because of the positive emotions it generated without the complication of analyzing it and thus interfering with memory formation. Clearly, the role of music listening in learning differs among individuals.

I looked at their song list and found no jazz–all of it was either concert-type music or pop songs.  That is a serious oversight, in my view. What the researchers may have missed is the possible positive effect of the unique rhythms and syncopation of jazz. I am reminded of a study I reported in my book, Memory Power 101, showing that chewing gum helps learning.

I am musically untrained, and maybe my listening to jazz improved my learning in vet school by creating positive emotions. A great deal of research has shown that positive emotions have an indirect enhancing effect on forming memories. Negative emotions impair memory. No solid neuroscience explanation exists, but it is no doubt highly relevant that the same brain structure, the hippocampus, mediates both emotions and memory formation.

Source:

Gold, B. P. et al. (2013) Pleasurable music affects reinforcement learning according to the listener. Frontiers in Psychology. Vol. 4 Article 541.  Doi: 10.3389/fpsyg.2013.00541

Turning Off Depression Triggers

It’s normal to get depressed. Let’s face it, a lot of life experience IS depressing. Depression that is severe enough to be considered a clinical malady is a state that persists for long periods. It’s not the initial depression that is the problem, but rather its sustained nature.

So, the issue is what sustains the depression. I contend that continual rehearsal of negative emotions, which can be done explicitly or implicitly, is the driver of clinical depression. I don’t know if psychologists agree or not, but as a neuroscientist I know that rehearsal of thoughts and feelings strengthens the mediating synapses and circuits.

Obviously, consciously rehearsing bad events and our depressive response will help to cement depression in neural circuitry. But even implicit rehearsal can have the same effect. This being the case, it seems important to focus on the triggers that activate recall, explicit or implicit, of stored representations of depressed feelings. Bad events and their associated feelings are stored in memory. As long as such memory is stored and not activated in recall, little harm is done. But environmental and mental cues can dredge depression from its hidden stores. Repeated retrieval is what makes depression pervasive and persistent.

So, it would seem important to focus on ways to block the retrieval cues. Yet, part of the problem is in recognizing what these cues are, as they are frequently buried do deep in memory that the cues are no longer available for explicit recognition. Yet, subconsciously, the cues still wrench the sadness from memory storage.

One solution that sometimes works is to change environments. Even if you don’t know what the depression cues are, you know they can somehow be embedded in the current environment and lifestyle. Maybe the problem is with some of the people you run around with. People who drag you down are not all that hard to spot. Avoid them. Maybe the problem is with your career or work environment, which has saddled you with depressing experiences from time to time. Staying in that environment assures that depression triggering cues will be encountered again.

It is not always feasible to change environment or lifestyle. You can’t abandon those you love just because

they depress you. You may not be able to change jobs or careers for economic or other practical reasons. In those cases, it helps to promote recall of happy experiences as a substitute.

Common experience and a great deal of formal research have shown the usefulness of “happy thoughts” as a way to boost positive mood. Here, the trick is to enhance recall of the buried memories of happy experiences. The same neural mechanisms involved in rehearsal and recall of depressing experiences are involved. But, you can’t be sad and happy at the same time. Thus, triggers that recall happy experiences do so at the expense of triggers that would trigger depressive feelings.

Recent research emphasizes the importance of memory cues as therapy for depression. In this study, clinically depressed patients were asked to recall 15 positive memories. Patients in one group, the controls, were asked to rehearse these positive experiences by grouping them according to similarities. Patients in the experimental group were taught to use a mnemonic technique to rehearse the positive memories. This technique, the ancient “method-of-loci” method, entails associating a mental image of an item to be remembered to an image of a well-known physical location. For example, to remember a grocery list, you might use location of objects in your car. You might picture a banana in your side view mirror, apples on the steering wheel, bread on the dashboard, cereal in the rear-view mirror. As you drive to the grocery store, you rehearse the items in terms of their mental images and locations within the car. There are many powerful permutations of this method that I explain in my recent book, Memory Power 101.  

Results of the experiment showed that both memorization methods were equally effective when recall was tested right after the training. But a week later, experimenters made a surprise phone call to each patient and asked them to recall the happy thoughts again. This time, clearly better recall occurred in the patients who had used the method-of-loci method. If we can generalize these results, it means that patients can alleviate their depression if they train their brains to be more effective at remembering positive events. This can be done at any stage of life, as we all have a past and there typically were some good moments in that past. Whether you use a method-of-loci method to remember happy times or some other memory device, your life should be more satisfying and less depressing when you consciously train your brain to remember the good times.

Source:

Dalgleish, T. et al. (2013). Method-of_Loci as a mnemonic device to facilitate access to self-affirming

personal memories for individuals with depression. Clinical Psychological Science. Feb. 12, DOI: 10.1177/21677026112468111.

Weight Gain Hurts Memory in Older Women

The more a woman weighs, the worse her memory. No, I am not a chauvinist pig. This claim comes from actual research—by a woman, no less. Diana Kerwin and her colleagues at Northwestern University studied 8,745 women ages 65 to 79 and found that for every one-point increase in female body mass index, the score on a 100 point memory test dropped by one point.

The problem was greatest in women who had put the weight on around the hips, which is fairly typical for weight gain in women. Nobody knows why this is so. Fat deposits may increase the amount of cytokines, which are hormones that can cause inflammation. In a couple of earlier columns I explained how body inflammation, from sore joints or sore throat, for example, can trigger inflammation in the brain. I explained that brains can get inflamed too, irritated from the release of cytokines and other toxins from the brain’s immune cells in response to inflammation. In both genders, these toxins diminish mental capabilities, especially memory. Remember, everything the brain does affects memory (and everything affecting memory affects the brain).

Another obvious possibility is that excess weight often creates vascular problems, and everybody tends to have a problem with circulation in small arteries as they get older. Excess weight is a risk factor for stroke, as well as Alzheimer’s Disease.

This finding about memory loss is just one of many good reasons to lose weight. There are only two ways to lose weight: eat fewer calories and exercise more. Though exercise doesn’t do much to cause weight loss, it has many other benefits (improved circulation of blood to the brain) that can directly benefit memory and cognitive function.

It is not surprising then to learn of recent studies showing that losing weight can improve thinking and memory, in both men and women. John Gunstad, at Kent State University, compared attentiveness and memory test scores in 150 overweight subjects 109 of whom who had bariatric stomach by-pass surgery and 41 controls who did not. Those who lost weight because of gastric bypass surgery showed mental function improvements within 12 weeks after surgery. Those without hypertension improved more than the bariatric patients who had hypertension. Memory performance of the obese controls actually decreased over this period. Gunstad has a U tube video on his work at www.youtube.com/watch?v=gsFP2zAkStU.

Of course bariatric surgery is not without its problems. This surgery can lead to Wernicke's encephalopathy, a condition associated with thiamin or vitamin B1 deficiency. Symptoms of Wernicke's encephalopathy include loss of short-term memory, vision and muscle coordination. Presumably, vitamin supplements prevent this problem.

Most of us lose weight the old fashioned way: diet and exercise. Will weight loss help mental function, especially in people who are overweight but not to the point of obesity? What Gunstad hopes to test next is the possible mental benefit from losing weight through diet and exercise rather than surgery. I suspect he will see a benefit, but it could come from exercise as such rather than the weight loss. As I have reported in earlier columns, normal-weight people see a mental improvement from aerobic exercise.

But up to a point, you can just sit in your lounge chair and munch potato chips and still improve your memory—if you are learning from my book, Memory Power 101.

Sources:

Gunstad, J. et al. (2013). Improved memory function 12 weeks after bariatric surgery. Surgery for Obesity and Related Diseases. 7 (4): 465-472. http://www.soard.org/article/S1550-7289%2810%2900688-X/abstract

Kerwin, D. R. et al. 2010. The cross-sectional relationship between body mass index, waist–hip ratio, and cognitive performance in postmenopausal women enrolled in the Women's Health Initiative. Journal of the American Geriatrics Society. 58 (8): 1427–1432.

Neurologic Complications Associated with Novel Influenza A (H1N1) Virus Infection in Children. Center for Disease Control, July 24 2009

New Strategy for More Efficient Learning

In 1913, Ebbinghaus demonstrated that spacing learning out over time creates much more efficient learning than cramming a learning task into a single intense session. Now, a new discovery has been made for a specific spaced-learning strategy that so far is the best of all. In reviewing this new design, Kelley and Whatson (2013) point out experiments showing that this kind of spaced learning is optimal for information encoding and for activation of the genes needed to form long-term memory.

And what is the design? The idea begins with the established notion that a given learning task should be “chunked” so that it can be studied in a short time, on the order say of 20 minutes. What is novel about the new design is that a given chunk is studied three times in a single session, with two intervening “rest” periods of 10 minutes in which there is little mental activity. During the rest periods, physical activity, like shooting hoops or cycling, seem to be ideal. The reason for these intervening rest periods is that thinking about new information or performing mental tasks creates interference with the memory-forming processes already under way.

Of course, like most learning tasks, a single session, even with three repetitions within it, is not likely to be sufficient unless you are really adept at mnemonic techniques (Klemm, 2012). After a day or so, this strategy needs to be repeated one or more times.

This is so simple to do and, if replicated in more studies, should become standard practice in schools. However, very few teachers know about this technique and school curricula are not designed to be taught this way. Changing the educational establishment is probably too much to hope for. But this strategy can be used by all students in homework study. Home schoolers and students taking Internet courses can easily use the technique on their own.

If you try this approach, please add comments to this post to let us know how it works for you.

Kelley, P. and Whatson, T. (2013). Making long-term memories in minutes: a spaced learning pattern from memory research in education. Frontiers in Human Neuroscience. 25 September. Doi: 10.3389/fnhum.2013.00589.

Klemm, W. R. (2012). Memory Power 101. New York: Skyhorse Publishing.

Happy Thoughts Can Make You More Competent

“Life, liberty, and the pursuit of happiness:” some people might argue that the U.S. Constitution endorses hedonism, and indeed many politicians want to ignore or get rid of the Constitution, but not necessarily because of hedonism. We should not be dismissive about encouraging people to pursue happiness. Happiness can be good for your brain. Depression is surely bad for your brain.

Positive mood states promote more effective thinking and problem solving. A recent scholarly report[1] reviews the literature demonstrating that positive mood broadens the scope of attentiveness, enhances semantic associations over a wide range, improves task shifting, and improves problem-solving capability. The review also documents the changes in brain activation patterns induced by positive mood in subjects while solving problems. Especially important is the dopamine signaling in the prefrontal cortex.

Published studies reveal that a variety of techniques are used to momentarily manipulate mood. These have included making subjects temporarily happy or sad by asking subjects to recall emotionally corresponding past experiences or to view film clips or hear words that trigger happy or sad feelings,

The effect of happiness on broadened attentiveness arises because the brain has better cognitive flexibility and executive control, which in turn makes it easier to be more flexible and creative. Happy problem solvers are better able to select and act upon useful solutions that otherwise never consciously surface. Happiness reduces perseverative tendencies for errant problem-solving strategies. The broadened attentiveness, for example, allows people to attend to more stimuli, both in external visual space and in internal semantic space, which in turn enables more holistic processing. For example, in one cited study, experimenters manipulated subjects’ momentary mood and then measured performance on a task involving matching of visual objects based on their global versus local shapes. Happy moods yielded better global matching.

Other experiments report broader word association performance when subjects are manipulated to be happier. For example, subjects in a neutral mood would typically regard the word “pen” as a writing tool and would associate it with words like pencil or paper. But positive mood subjects would think also of pen as an enclosure and associate it with words like barn or pigs. This effect has been demonstrated with practical effect in physicians, who, when in a happy mood, thought of more disease possibilities in making a differential diagnosis.

The review authors reported their own experiment on beneficial happy mood effects on insightfulness, using a task in which subjects were given three words and asked to think of a fourth word that could be combined into a compound word or phrase. For example, an insightful response to “tooth, potato, and heart” might be “sweet tooth, sweet potato, and sweetheart.” Generating such insight typically requires one to suppress dominant “knee jerk” responses such as associating tooth with pain and recognizing that pain does not fit potato while at the same time becoming capable of switching to non-dominant alternatives.

Other cited experiments showed that happy mood improved performance on “Duncker’s candle task.”  Here, subjects are given a box of tacks, a candle, and a book of matches, and are asked to attach a candle to the wall in a way that will burn without dripping wax on the floor. Subjects in a happy mood were more able to realize that the box could be a platform for the candle when the box is tacked to the wall.  

Such effects of happy moods seem to arise from increased neural activity in the prefrontal cortex and cingulate cortex, areas that numerous prior studies have demonstrated as crucial parts of the brain’s executive control network. Similar effects have been observed in EEG studies. Other research suggests that the happiness effect is mediated by increased release of dopamine in the cortex that serves to up-regulate executive control.

The review authors described a meta-analysis of 49 positive-psychology manipulation studies showing that momentary happiness is readily manipulated by such strategies as deliberate optimistic thinking, increased attention to and memory of happy experiences, practicing mindfulness and acceptance, and increasing socialization. The effect occurs in most normal people and even in people with depression, anxiety, and schizophrenia. Biofeedback training, where subjects monitor their own fMRI scans or EEGs, might be an even more effective way for people to train themselves to be happier.

The main point is that people can be as happy as they choose to be.

For more on how positive mood influences memory ability,

see my new book, Memory Power 101 (http://skyhorsepublishing.com).

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[1] Subramaniam, K. and Vinogradov, S. (2013). Improving the neural mechanisms of cognition through the pursuit of happiness. Frontiers in Human Neuroscience. 7 August. Doi: 10.3389/fnhum.2013.00452

Landmark Research: Why We Need to Get Enough Sleep

In other blog posts I have explained why sleep is good for the brain in general and memory formation in particular. Now a new discovery provides another reason for people to get enough sleep. The study examined a type of support cell in the brain, oligodendrocytes–let’s call them oligos for short. These cells wrap their membranes around nerve cells to form what is called myelin, which forms an electrical insulation in a way that speeds up the propagation of nerve impulses through neural networks. You may have heard about oligos in reading about multiple sclerosis, a disease that impairs nerve communication because oligos die and the myelin insulation degrades.

Speed of transmission is important–it influences IQ for example. As you know from buying a new computer, the faster processor speed gives it new capabilities your old clunker could not do. A similar idea applies to the brain.

Anyway, this new study, from the University of Wisconsin, focused on oligos because other research had shown that sleep promoted the expression of several genes that are involved in synthesis of cell membranes in general and those in oligos in particular. Unlike neurons, oligos die, and are replaced in the brain. Thus, anything that affects their turnover is important for brain function. Sleep has been implicated in this turnover because a common neurotransmitter in the brain, glutamate, is known to increase in wakefulness and decline during sleep. Glutamate  suppresses maturation of oligo precursor cells into formation of myelin insulation.

In this particular study, investigators examined a genome-wide profile of oligo gene expression in mice after a 6-7 hour periods of sleep or spontaneous wakefulness, or four hours of forced wakefulness (sleep deprivation). They found that 357 genes were expressed differently, depending on the time of day, in response to normal daily rhythms. More dramatic was the observation that 714 genes changed expression in conjunction with the sleep/wakefulness cycle, independent of the time of day. Of these genes, 310 were “sleep” genes that were selectively activated during sleep.

Many of the sleep genes contribute to maturation of oligos into myelin. In follow up experiments, mice were injected with a radiolabeled tag that marks the birth of new cells. Injection occurred eight hours before mice spent a long period of either of wakefulness or sleep. The number of newly born oligos was almost double in the sleep group compared to the wake group. More detailed analysis showed that this increase was specifically correlated with the amount of REM sleep (dream sleep in humans).

This REM effect may have particular importance in humans. Most REM sleep occurs in the early morning hours and only after substantial time has been spent in non-REM stages of sleep. Thus, cutting a night’s sleep short by getting up early may decrease the amount of REM time and thus the beneficial effects on oligo proliferation. So don’t feel guilty about “sleeping in” from time to time.

We might also think about how these findings could have special relevance to children, whose brains are incompletely myelinated. Getting children up early in the morning to start school at 8 AM may not be such a good idea. Until school districts get around to changing school hours, you might tell you kids about my learning and memory improvement e-book, Better Grades, Less Effort, available at Smashwords.com.

Source:

Bellesi, M., et al. (2013) Effects of sleep and wake on oligodendrocytes and their precursors. J. Neuroscience. 33 (36), 14288-14300.