text:
How and where, in the human brain, are memories stored?
Background
I am a computer programmer who is fascinated by artificial intelligence and artificial neural networks, and I am becoming more curious about how biological neural networks work.
Context & what I think I understand
In digesting all I have been reading, I am beginning to understand that there are layers to neural networks.  A front-line layer of neurons may receive, for example, a visual stimulus such as a bright light.  That stimulus is taken in by the front-line neurons, each of which produce a weighted electro-chemical response that results in a binary decision to pass an electrical charge through its axon to the dendrites of the tens of thousands of neurons to which it is connected.
This process repeats through layers channeling the electrical signals and focusing them based on their permutations until ultimately a charge is passed to a focused response mechanism such as the nerves that control shrinking of the pupils.
Hopefully I got that correct.
Preamble to the question ;)
Assuming that I am not completely off-base with my basic understanding of how a biological neural network operates, I am beginning to grasp how an input (stimulus) results in an output (response) such as motor movement or reflexes.  That would just seem to be basic electricity of open and closed circuits.
HOWEVER, what confuzzles me still is how a memory is stored.  The analogy to an electrical circuit breaks down here, for in a circuit I can't really stop the flow of electrons unless I dam up said electrons in a capacitor.  If I do that, once the electrons are released (accessed), they are gone forever whereas a memory endures.
So. . .
How the heck are memories constructed and stored in the human brain?  Are they stored in a specific region?  If so, where?

Regarding pupil size alterations it is the circular muscle of the iris which controls pupil size.  Effectors are always either muscles or glands - not nerves.  Otherwise spot on :)

“the analogy to an electrical circuit breaks down here” – it’s not really an analogy, and it doesn’t break down. This is how the brain works: not an analogy, but rather the actual mechanism. The only part missing is the dynamics in structure.

@Rory - But the nerves control the muscles, no? That is what Matthew certainly means.

What do you mean by "focusing them based on their permutations"? Permutation usually means a bijection of a set onto itself or plainly rearrangement. What do you mean by rearranging electrical signals?

Unfortunately, we are all still "confuzzled" by how memory works. We are far from a complete understanding of how memory is stored and recalled. Nonetheless, we do know a little, so read on.
Your understanding of basic neural function is almost correct. First, an individual neuron will signal through its single axon onto the dendrites of many downstream neurons, not the other way around. Second, I am not sure what you mean by "focusing them based on their permutations," but it is true that neural information can undergo many transformations as it propagates through a circuit. Third, if there is a behavioral outcome of the network activity like a muscle response or hormone release, those effects are mediated by nerves communicating with muscles and hormone-releasing cells. I'm not sure if that is what you meant by "focused response mechanism."
Finally, as you have discovered, the analogy of neural circuits to electrical circuits is relatively poor at any reasonably sophisticated level of analysis. My opinion is that biological systems are often poorly served by being framed as engineering problems. Others will disagree with that, but I think understanding a biological system on its own terms makes many things much clearer.
The key thing missing from the electrical circuit analogy turns out to be one of the keys to understanding information storage in neural circuits--the synapse, the site where one neuron communicates with another. The synapse transforms the electrical signal from the upstream neuron into a chemical signal. That chemical signal is then converted back into an electrical signal by the downstream neuron.
The strength of the synapse can be adjusted in a long-term way by changing the level of protein expression--this is called long-term potentiation (LTP) or long-term depression (LTD). LTP and LTD therefore can regulate the ease with which information can flow along a particular path. As a basic example (that should not be taken too seriously), imagine a set of neurons that represents "New York City" and another set of neurons that represents "My Friend John." If you then happen to be in New York City with your friend John, both of those groups of neurons will be active and synapses between these two networks will be strengthened because they are co-active (see Hebbian plasticity). In this way, the idea of NYC and the idea of John are now bound together.
Where are these neurons that represent NYC and John? We are still not totally clear on this, and the question is complicated because there are many different types of memory. For instance, your memory of how to ride a bike (procedural memory) is not treated the same as your memory of what you ate for breakfast (episodic memory). However, a best current answer is that the hippocampus and its associated regions are important for the initial encoding of memories and the neocortex is where longer term memories are stored. There is substantial communication between these two areas so that memories can be effectively adjusted over time.

Update
In response to Jule's comment asking for some resources, I realize it is important to make the point that the Hebbian model I outlined hasn't been definitively shown. Like with all aspects of neuroscience, there is a lot of good work at the molecular and cellular level and good work at the behavioral level, but the causal link between the two is not so clear. Nonetheless, Hebb's idea is still the mainstream working model for how memory works. Some reading might include:
1) Neves, G., Cooke, S.F., Bliss, T.V.P., 2008. Synaptic plasticity, memory and the hippocampus: a neural network approach to causality. Nature Reviews Neuroscience 9, 65–75.
 A review on hippocampal memory and its relation to LTP/LTD and Hebbian theory. Notes the general difficulty of proving the theory and some ways for experiments to move forward.
2) Lisman, J., Grace, A.A., Duzel, E., 2011. A neoHebbian framework for episodic memory; role of dopamine-dependent late LTP. Trends in Neurosciences 34, 536–547.
 A review proposing an elaboration of the Hebbian model that includes neuromodulatory influence on plasticity and memory process.
3) Johansen, J.P., Cain, C.K., Ostroff, L.E., LeDoux, J.E., 2011. Molecular Mechanisms of Fear Learning and Memory. Cell 147, 509–524.. An excellent review on fear learning and memory with an extensive section on Hebbian theory.
4) Liu, X., Ramirez, S., Pang, P.T., Puryear, C.B., Govindarajan, A., Deisseroth, K., Tonegawa, S., 2012. Optogenetic stimulation of a hippocampal engram activates fear memory recall. Nature. A research article which is perhaps a realization of some of suggestions in the Neves et al review. They use light to reactivate a fear memory. This suggests that activation of the hippocampal network that was active during memory formation is sufficient to elicit the memory.

Do you have a reference to a paper that summarizes the experimental evidence for this theory?

Thank you once again yamad for your thorough answer on this fascinating subject.

For a little more on ways in which the nervous system is plastic, see my answer here http://biology.stackexchange.com/a/1359/72

Memories are not stored in synapses: https://www.scientificamerican.com/article/memories-may-not-live-in-neurons-synapses/

I would like to point out some ways your understanding is wrong. "Neural networks" are usually a computer science term, only very, very loosely based on actual neural networks. The idea of layers in a neural network is pretty much an invention of computer science, it doesn't really reflect the reality. Also, neurons are not binary switches. It isn't so much about on/off, so much as a temporal rate code of action potentials. The actual brain sort of stochastic, and does not really work on the level of the single action potential.
The adaptation of the term "neural networks" by those devious computer scientists is unfortunate. Their work is virtually unrelated to actual neural networks.

I think you overstate the difference from neural networks. For instance, they are often not binary but rather have a smooth transition function (an idea copied from biology), are stochastic (an idea copied from biology) and doesn’t work on the level of a single AP. The adaptation of the term isn’t unfortunate, it’s a consequence of where the inspiration came from. Almost every CS concept has such an inspiration, and many non-CS concepts have as well (ever seen a crane?).

@KonradRudolph They simply don't approach the complexity of an actual neuronal network. Even the computational neuroscientists, who are explicitly trying to model neurons, fall short. It's an unfortunate adaptation of terms because now "neural network" no longer refers to a network of neurons. It refers to a crude simulacrum of a neural network.
The degree to which computer scientists have neglected the biology is disturbing and has stunted the field of AI.

@Preece I agree with all of your comment except the last sentence. Biology isn’t being neglected, it’s simply not relevant: neural networks in CS solve a problem, and that problem isn’t “try to model biology as accurately as possible”, nor is it trying to create a “classical” AI. It’s simply a statistical tool for pattern matching / clustering. That said, modelling the biology accurately would be interesting in its own right; it isn’t done for one simple reason: it’s not computationally feasible.

@KonradRudolph there is more to looking at the Biology then trying to model the nervous system. Neural structures are implementations of algorithms. I think analyzing these algorithms is at the heart of AI research. You don't need to model neurons to model neural algorithms. Trying to get AI without looking into them will (has) provided many interesting things, but the potential of the field is barely tapped.

@Preece But much of the work with neural networks isn’t really done in a context of AI research, it’s simply a tool in statistics, and here people are only interested in statistical properties, and it’s easy to show that a single hidden layer ANN is just as mathematically expressive as any more complex version so more complex versions are not just impractical, they are unnecessary. And on the AI / comp. neuroscience side of things there is a lot of research done in this area.

I can make a rough analogy in terms of digital media storage.
Our memories exist as a relationship between our perceptions and our sensations. Computers store input readily. However, humans store memories perceptually. This means who we are and how we remember an event permanently changes our recollection.
If you look at the progression of lossy video codecs as resolution has increased you'll see the algorithms have changed. Each generation has become accustomed to seeing certain types of video artifacts: from film, to analog static, to vhs, to h.264. Think of the algorithms as individual modes of perception.
We have ancient basal circuits which allow visual and frontal cortex areas to collaborate in the formation of memories. It's not an objective process. We all have our own algorithms.


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Evolution in 37 years, is it possible?
I am confused, can evolution ( speciation ) really occur in such a short time ?

In 1971, biologists moved five adult pairs of Italian wall lizards from their home island of Pod Kopiste, in the South Adriatic Sea, to the neighboring island of Pod Mrcaru. Now, an international team of researchers has shown that introducing these small, green-backed lizards, Podarcis sicula, to a new environment caused them to undergo rapid and large-scale evolutionary changes."

Here is a short video featuring the scientist.
Two related articles:

http://www.sciencedaily.com/releases/2008/04/080417112433.htm
http://news.nationalgeographic.com/news/2008/04/080421-lizard-evolution.html

Edit: I asked a similar question previously about two people having 44 chromosomes and their possibility of creating a new species. You might like to read/answer that as well:
Can two humans with 44 chromosomes produce viable offspring?

Evolution just depends on the amount of selective pressure.  The classic selection for antibiotic resistance is just a single generation. It may seem artificial, but it is adaptation in a single generation.  Also wanted to add this link: yeast become multicellular in a 60 generations. http://www.scientificamerican.com/article.cfm?id=test-tube-yeast-evolve

I am confused, can evolution ( speciation ) really occur in such a short time?

Well, Evolution and Speciation are not the same. Evolution is the adaptation of an existing species to an environment over generations. Speciation is the development of a new species, and the definition of "species" can vary depending on who you talk to - but a very commonly accepted one is that two individuals cannot produce fertile offspring.
However, the answer is still Yes. Evolution can occur in as little as two generations because that's all the time it takes for a change in the DNA to be exhibited in the newest generation. For instance, Lactase Persistence - which allows the bearers to ingest dairy products without digestive stress into adulthood - has evolved at least twice in humans in separate groups. Both are the result of either one or two point mutations (single changes in a Nucleotide within the DNA). The result is that at one point in time for each appearance of lactase persistence, a child was born that could drink milk into adulthood while their parents and cousins could not.
That is evolution; the child was better adapted to the resources available, and because lactase persistence either...

Provided a significant advantage over their kin in the environment (which is likely, as being able to digest milk in an agrarian society opens up a new source of energy)...

or

Didn't hinder the child's ability to grow and produce fertile offspring of their own...

...it has persisted in the populations where it initially evolved.
Speciation is a little trickier, but that's because there isn't a known qualitative way to differentiate species at the genetic level quite yet. It could be a few key gene changes, it could be a whole genome difference of 10%, or a million other factors. Practically speaking, though, Speciation is simply the result of accumulated genome changes that at some point prevent the members of one group from producing fertile offspring with their ancestral group.
Because we simply don't know how much of or where the genome has to change, it's certainly possible that speciation could occur in the same time period as it takes to evolve: Two generations.
Since I'm doubting the wall lizards have a generational time of >37 years (don't mistake it for lifespan, humans can reproduce in our teens - even if we live into our 80's), it's definitely possible that the wall lizards could evolve (and potentially speciate) in such a short amount of time.
With organisms that have a very, very short generational time - like E. coli or other bacteria - you can observe the Evolution of populations in days or hours.

Clarified with your correction.

I want to underline 1 fact about speciation, although you don't object to this. Speciation doesn't necessarily mean the lack of ability to produce healthy offsprings. For instance, Lions and Tigers ( Two different species) can reproduce healthy offsprings (Liger and Tiglons), but they don't mate in the nature. So speciation also requires the lack of inclination to mating with the other.

@MCM I have updated the question. You might like to answer the referenced question as well, please do :)

@Comptrol: the problem lies in the hazy definition of species. Often you would hear that two different species cannot produce fertile offspring, for instance.

@Comptrol - Regarding "ligers" and other such combinations, usually the "Do not reproduce and produce fertile offspring" also runs with the caveat "in the wild" when you're talking about more 'official' definitions of species in Biology. Since Tigers and Lions do not reproduce in the wild the definition fits for them, but it doesn't work for other creatures. Hope that cleared it up a bit.

@MCM - would you be against editing "Evolution is the adaptation of an existing species to an environment over generations." to "Evolution is a change in the characteristics of a population over generations." The existing sentence describes the response to selection, but evolution occurs by drift, mutation, and migration

Evolution can occur in just one full generation
Strong selection will rapidly reduce the gene frequencies of genes which cause negatively selected phenotypes. This reduces the likelihood of unfavourable genotypes occurring in the next generation.
(I regard generation here as the complete cycle of one individual being born to the point at which they successfully give birth/sire young).
Population genetics explanation:
Imagine a population in which LOC1 is a locus affecting a phenotypic marker allowing perfect identification of genotype at that locus. There are two alleles at LOC1, A and a. This gives rise to three different genotypes, AA, Aa and aa  which occur in the population in equal proportions. Those individuals with AA exhibit a certain trait, whilst Aa and aa individuals do not. If only the AA genotypes are allowed to mate then the following generation will only contain AA genotypes at LOC1 (assuming no mutation).
For more population genetics read Principles of Population Genetics and then Introduction to Quantitative Genetics.
Genetic drift caused by using 5 Pairs
The new population was started with just 5 pairs of individuals. This means there is huge potential for fixation of alleles via drift with in the very first generation. I ran a (basic) genetic drift simulation in R just now. Out of 5000 replicates 4709 went to fixation (Single locus, two alleles, using 30 generations of 10 individuals).
Environmental change can cause hidden genetic variation to be exhibited
Molecular regulators which affect the expression of traits can respond differently depending on environmental influences. This point is summarized nicely by the opening lines of this paper- thanks to @leonardo for this point.

Hsp90 is a molecular chaperone for many signal transducers and may
  influence evolution by releasing previously silent genetic variation
  in response to environmental change. In fungi separated by 800
  million years of evolution, Hsp90 potentiated the evolution of drug
  resistance in a different way, by enabling new mutations to have
  immediate phenotypic consequences.

Evolution and speciation are different things
These are just the simple wikipedia definitions, for better ones then consult the literature:

Evolution is the change in the inherited characteristics of biological
  populations over successive generations.

For a good thorough introductory text on evolution I recommend Evolution by Mark Ridley (not to be confused with Matt Ridley - popular evolution science writer).

Speciation is the evolutionary process by which new biological species arise.

Speciation, thanks to a long fought debate over species concepts, is also an ambiguous process. What some species concepts would define as two distinct species, another would call one species, and another may call them 10 species! Seemingly everyone who specializes in the study of speciation would have there own twist on a species concept so don't expect that argument to resolve any time soon! It's not advanced much since Dobzhansky wrote this.

The species concept is one of the oldest and most fundamental in
  biology. And yet it is almost universally conceded that no
  satisfactory definition of what constitutes a species has ever been
  proposed.

When does evolution occur? and when is change not evolution in such a short time?
I've put this bit in because what I find is a strange, but common, question people ask is

"species X has changed/adapted to novel selection very rapidly, has
  evolution really occurred?"

The way people ask this makes it sound like they want a certain amount of evolution to occur before we can say something has evolved. To me saying evolution has occurred means that some change has occurred, via any of the mechanisms of evolution, which results in one subset of individuals being different from another it is compared to (spatially or temporally separated). You could ask if such strong evolution can really occur over a short time period, but the answer is obvious - yes it can. If it has occurred then why do you think it is exceptional? what is your alternative explanation for the differences seen?

also; the fact they only introduced five pairs means there could have been a lot of change in the first generation via genetic drift - I'll edit that, and some examples of rapid evolution, in to the answer soon - thanks @leonardo

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Why was polyploidy not lethal in certain octodontid rodents?
As discussed in Why is polyploidy lethal for some organisms while for others is not?, polyploidy is normally lethal in mammals.
However, two species of Octodontidae (South American rodents), are tetraploid due to a recent doubling of all chromosomes:

Tympanoctomys barrerae: 4x = 102
Pipanacoctomys aureus: 4x = 92 (apparently some chromosomes were lost after the polyploidy event)

See the paper discussing T. barrerae, the first of these tetraploid rodents discovered.
According to the Wikipedia article, both are believed descended from the same species, Octomys mimax (or possibly now-extinct close relatives thereof), which has 2x = 2n = 56 chromosomes, half those of T. barrerae.
What was special about Octomys that allows it to survive polyploidy, unlike most mammals?
Interesting:

The sperm head of Tympanoctomys is is by far the largest naturally occurring in mammals, and its size is causally related to the double genome size it has to accomodate.

Actually, that is not what is discussed in the question you linked to. The following is a quote from the very comprehensive accepted answer (emphasis mine):

Polyploidy arises easily in both animals and plants, but reproductive
  strategies might prevent it from propagating in certain circumstances,
  rather than any reduction in fitness resulting from the genome
  duplication.

In fact, try rereading that answer and the references therein, it answers your general question.
There is a popular theory (mostly accepted) that there have been whole genome duplication events (WGDs) in the vertebrate ancestry. If true, it means that all mammals are the descendents of polyploid ancestors. For a very nice review of WGDs in vertebrate evolution, see here [1]. Octomys is simply the only known mammal with a more recent WGD event. As for what makes it special (taken from [2]):

Our data demonstrate that parental-specific silencing of at least one
  gene and normal X chromosomal dosage mechanism are conserved in the
  tetraploid genome. We hypothesize a concerted action of genetic and
  epigenetic mechanisms during the process of functional diploidization
  of this tetraploid genome.

1) Van de Peer Y, Maere S, Meyer A., The evolutionary significance of ancient genome duplications., Nat Rev Genet. 2009 Oct;10(10):725-32.
2) Bacquet C, et al., Epigenetic processes in a tetraploid mammal, Mamm Genome. 2008 Jun;19(6):439-47.

I would say in practice that it is more complex than this. Trichrisomy (extra chromosome) is often (not always) fatal or causes significant negative phenotype (Downs syndrome for instance) in humans.  Genome duplication has occurred several times in the course of evolution, but it is still very rare in animals.  Plants are much more resilient to polyploidy than animals.

@shigeta, true but the main difference here is that polyploidy at the species level (such as WGD) is very different to individual aberrations such as Downs'. Individuals are very sensitive to changes in ploidy, however, changes in gene or even chromosome copy number have occurred multiple times in multiple lineages. Clearly they were accompanied by compensatory changes elsewhere to allow the species to deal with the change in ploidity. I think it is not reasonable to compare trichrisomy or Down's to WGD events.

the point is taken that while its not disputed that occasionally animals tolerate genome duplication, its still not clear to me that the whole genome duplication is tolerated by all animals.  there may be something particularly permissive in these rodents given the frequency seen in that branch...

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Is it known how the first viruses formed?
The oldest known virus is known to have infected prehistoric insects 300 million years ago. A virus is basically a parasitic strand if DNA or RNA encapsulated in a protein coat. It enters cells by "wearing" a protein coat made out of proteins needed by a certain cell (e.g. Lung cells and flu virus).
Now, my question:

Do we know how viruses naturally formed? To simplify the question, if we went back to the first virus, how would it form?

Note that I am looking for a scientific answer, and not wild-ass speculation. While I can accept some degree of speculation the accepted answer will NOT be based entirely on speculation if it is avoidable.

And most likely the first viruses would be much older than the 300 million year old you mention, and probably infected bacteria or whatever was floating around back then. I suppose even an RNA world could have virus like parasites that manage to hijack some replication machinery, but now I'm getting into the speculation you didn't want.

After further research I came upon this question which seems closely related.....do you think mine is a duplicate?

I'm not sure any of the answers will be proven. You're talking about an age we simply do not know much about. 300 million years ago is a very long time to be sure about such things.

One of the main points of contention in the study of virus evolution is whether or not they appear before the last universal cellular ancestor (LUCA) or afterwards (commonly accepted: genes that "escaped" from host organisms aka the escape hypothesis or vagrancy hypothesis).  Basically though, the LUCA is the most recent ancestor that all organisms living on Earth are derived from.
Increasing evidence support that ancestry of viruses may predate the LUCA, for example an idea is that there "was an 'ancient virus world' of primordial replicators that existed before any cellular organisms and that both RNA (first) and DNA (later) viruses originated at this time, donating some features to the first cellular organisms.  However, "a competing theory is that RNA cells existed before the LUCA and that RNA viruses were parasites on these RNA cells that later evolved DNA as a way of escaping host cell responses."
So basically, some theories support the pre-LUCA ancient virus origin and some support post-LUCA.  Because it's impossible to replicate conditions from billions of years ago, several approaches using evolutionary genetics and probability models are recommended by the author of the paper, referenced below.  He also discusses the specific supporting criteria for the theories that you can read in greater detail.
Holmes, E. C. (2011). "What does virus evolution tell us about virus origins?" J Virol 85(11): 5247-5251.

Also good to keep in mind that viruses are not a monophyletic group but have multiple origins. Some viral clades like Caudovirales probably predate the divergence of the cellular domains whereas e.g mimiviridae have much later (but still quite ancient origins). Also, neither of these groups is in any way related to e.g. ssDNA viruses

Great question. There are several hypotheses, but in reality no one really "knows" because this is incredibly difficult to prove. We may never know for certain.
Anyway, on to the three main hypotheses, I got this from http://en.wikipedia.org/wiki/Viral_evolution, I think you'll find it to be helpful.
"There are three classical hypotheses on the origins of viruses: Viruses may have once been small cells that parasitized larger cells (the degeneracy hypothesis or reduction hypothesis); some viruses may have evolved from bits of DNA or RNA that "escaped" from the genes of a larger organism (the vagrancy hypothesis or escape hypothesis); or viruses could have evolved from complex molecules of protein and nucleic acid at the same time as cells first appeared on earth (the virus-first hypothesis)."
However, as you may have guessed, there are plenty of questions that these theories do not answer. The next paragraph:
"None of these hypotheses was fully accepted: the regressive hypothesis did not explain why even the smallest of cellular parasites do not resemble viruses in any way. The escape hypothesis did not explain the complex capsids and other structures on virus particles. The virus-first hypothesis was quickly dismissed because it contravened the definition of viruses, in that they require host cells. Virologists are, however, beginning to reconsider and re-evaluate all three hypotheses."
There is a little more information on the Wikipedia page, I would recommend that you read that and do a little research on the different theories if you would like to know more.
I hope this helps,
CDB


text:
Cytochalasin B in cell movement
In an experiment by Pollard, T.D. and R.R. Weihing, amoeba cells injected with  Cytochalasin B were inhibited of movement, while the control group amoeba cells did move. A hypothesis was made that Cytochalasin B blocked polymerization of actin monomers into actin filaments, thus preventing the cell from moving. To validate this hypothesis the experiments were made again with cycloheximide (inhibits new protein synthesis), and dinitrophenol (inhibits new ATP formation), and colchicine (inhibits polymerization of microtubules Here are the results:$\\$
$\begin
Condition:                                  % of cells that didn't move
No drug                                      3
Cycochalasin B                            95
Colchicine                                   4
Cycloheximide                             3
Cycloheximide + Cycochalasin B    94
Dinitrophenol                               5
Dinitrophenol + Cycochalasin B      85$
$\\$
Question: Why were these control important? How did they help validate the hypothesis?
Note: this is for self-study.

The other inhibitors work on other processes, as you already noticed. There is almost no difference between the conditions when they are added and the corresponding condition when they are missing, suggesting those other processes play no role in the observed phenomenon. Eliminating those other processes makes it more believable that actin polymerization is important, rather than being one of multiple factors.
Moreover, often cell will adapt to an inhibitor by increasing or decreasing other kinds of activity. For example, it could be that actin polymerization inhibition prevents amoeba from eating. In this case, because any imaginable motive mechanism requires energy, starving amoeba may become unable to move, even though it was not using actin for movement. In order to refute such scenarios, where actin polymerization is not directly inducing movement, and where cytochalazine does not inhibit movement directly mechanism, supplementary experiments were performed.
DNP induces something similar to starvation, yet, in these experiments, did not abolish movements when given alone. Nor did DNP rescue movement, because the combination (DNP + cytochalazine) is virtually identical to cytochalazine alone. In both circumstances, DNP did nothing, thus showing that ATP depletion is not influencing movement. This is proof that actin polymerization is directly causing amoeba movement, and not an indirect ATP-mediated effect. Similarly, cytochalazin's effect is independent of protein synthesis, once more supporting the hypothesis that actin polymerization is important on its own.

When you work with living cells, there are usually more pathways which can be affected, when you start treating the cell a drug. To make sure, which pathway is specifically inhibited, you need these controls, as an inhibition of any of them could theoretically have the same effect:

Protein synthesis (cycloheximide) as this provides new actin monomers which are important for the enlongation.
ATP synthesis (dinitrophenol) as ATP is necessary for the movement
along actin filaments.
Inhibition of microtubuli (colchicine) inhibits the microtubuli found
in the flagella of amoeba, which also provide a form of movement

The controls show that only the addition of Cytochalasin B to the culture inhibits the movement by inhibiting the enlongation of the actin filaments which proves the hypothesis of the group.


text:
How stable is linear DNA transfected in eukaryotic cells?
I would like to know, with references from the literature, what is the half life of a linear dsDNA transfected in a mammalian cells.
For example, if I transfect human cells with a PCR product (500-2000 bp), that does not contain any replication or transcription signal, how fast will it be degraded by the cell (hours, days, weeks...)? Is there any difference in the half life if the DNA is in the cytosol or in the nucleus?

I would assume that it won't be degraded but will be just diluted out.

I am looking around, I cannot find a clear answer. I would expect foreign DNA to be actively digested by some enzyme rather than having it flowing around...

I don't know about linear DNA, but plasmid DNA is often detectable in cells after they stop producing detectable amounts of protein. They tend to get transcriptionally silenced, but aren't degraded as quickly. CpG methylation probably affects half-life as well, as normal PCR product DNA is not methylated, and normal eukaryote DNA is methylated.

I am specifically looking for more info about linear DNA especially PCR products. Thanks for your comment, I will search about the effects of CpG methylation as well.

Why do you want to know about linear DNA and PCR products rather than plasmids? Have you tried any literature searches so far?

I am inducing homologous recombination in cells and I am using PCR products as donor templates, but (without getting into details) I need to be sure that the donor template is not in the cells anymore if not integrated. I have looked around but I could not find any paper with clear data. To detect the recombination event I want to amplify back the donor templates from my pool of cells. Now, if the linear DNA is still around I cannot be sure if I am amplifying that one or the integrated one.


text:
Restriction endonucleases are found in?
Quoting from : Scientific American July 1975 The Manipulation of genes by Stanley Cohen :

Restriction endonucleases (and modification methylases) are widespread
  in microorganisms; genes for making them were found on viral
  chromosomes and extrachromosomal plasmid DNA as well as on many
  bacterial chromosomes.

Why would the genes for making RE be found on viral chromosomes ? Also, could you give some examples where they are found on plasmids ?

There are many proposals for the ecological role of restriction-modification (RM) systems, and why they would exist on mobile genetic elements (e.g. plasmids and viruses). In this case, I am specifically talking about the viruses that infect bacteria (aka, bacteriophage).
1) RM systems may have an anti-viral function. Normally we would think of such systems as being part of the host chromosome, but as Alan Boyd suggested, once a temperate phage has integrated itself into the chromosome, its fitness is tied to its host. Therefore, a RM system found in a prophage could prevent infections by additional phage. See here for a discussion of this issue:
http://www.ncbi.nlm.nih.gov/pubmed/23471617
2) RM systems may have "addictive" properties by acting as toxin-antitoxin systems. Basically, this means that if the genes for the RM system are lost, the host cell dies. This can provide selection for the maintenance of plasmids and prophage inside of the host cell. See here for a discussion of this issue:
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3874152/
3) Finally, viruses need to destroy the host genome -- both to suppress any anti-viral response and to release nutrients for viral replication. While this is typically achieved with nucleases other than restriction endonucleases, there is at least one situation where a RE seems to be involved. This is discussed in the first link above (see the section entitled "Role in Nutrition")

The familiar restriction enzyme EcoRI is plasmid encoded.

Betlach et al. (1976) A restriction endonuclease analysis of the bacterial plasmid controlling the EcoRI restriction and modification of DNA. Fed. Proc. 35:2037 - 43.

For an example of a phage encoded system see:

Dempsey et al. (2005) Sau421, a BcgI-like restriction-modification system encoded by the Staphylococcus aureus quadruple-converting phage Phi42. Microbiology 151: 1301-1311

This second paper suggests an answer to why a phage would have such a system: lysogens of Phi42 are resistant to infection by all 23 members of a standard set of S. aureus phages. Once a phage has lysogenised it is in its interest to prevent lysis of its host.

If you have a dsDNA genome and replicate it by rolling circle like most bacteriophages and many plasmids do, an endonuclease is pretty much a necessity.

Could you elaborate your answer and include some references ?

this is a nice question and post!  the RE nicks the phage genome so that it can be replicated by the polymerase.  Without it there would be no copying off of the chromosome.

But the nicking isn't carried out by a restriction endonuclease but by a specific protein which remains linked to the 5' end of the DNA. If this was accomplished by an R-M system on the phage the site would presumably be modified to protect it from double-stranded cutting.

I generally think of REs as producing double-stranded cuts. If I understand correctly, this answer only applies to those that create single-stranded cuts. Are they also called RE?

i just wonder which has an older origin- CRISPR-Cas or RE?

@WYSIWYG Interesting question. Nearly all bacteria have a restriction modification system yet only about half have CRISPR/Cas. In archaea, CRISPR/Cas is more ubiquitous. Ancient ancestry and horizontal gene transfer make it difficult to make strong conclusions on which came first. CRISPR/Cas is interesting in that it requires an RNA co-factor (crRNA/tracrRNA). There may have existed (or still exists) an ancient CRISPR system that functioned without Cas or other proteins (e.g. an endonucleolytic ribozyme with a targeting domain that existed before the evolution of protein synthesis).


text:
Steroid Hormone Signalling
Can the lipid-soluble hormones (like steroid hormones) go inside every cell, as their particular receptors are located in the cytoplasm or nucleus of the target cells? Is there anything special on the membrane of the target cells which enables the steroid hormone to know that it has to enter that particular type of cells and avoid others?
(Peptide hormones function through cognate receptors located on the cell membrane of the target cell.)


text:
Expression of bidirectional promoters
How are bidirectional promoters expressed ? (Won't RNA Pol have to go in 3'-5' direction?)
Why are they more commonly found in eukaryotes than prokaryotes?

Genes controlled by bidirectionl promoters are in head-to-head configurations, meaning that their 5' ends are facing one-another.  Remember that DNA is double stranded, so this means one gene is on the 'top' strand and one gene is on the 'bottom' strand.  Check out the diagram below, genes are in capitals, bidirectional promoter in parenthesis.  Both genes are transcribed 5'->3'
                                         --> Gene 1
 5'-atgcagtcatga(ctgactaagt...tcagtcatga)CTGACTGACTAGTCAT-3'
    |||||||||||| ||||||||||||||||||||||| ||||||||||||||||
 3'-TACGTCAGTACT(gactgattca...agtcagtact)gactgactgatcagta-5'
      Gene 2 <--

As for why they are more commonly found in eukaryotes, those questions are definitely hard to answer.  It has been shown that genes on bidirectional promoters are expressed together more often than two genes that are next to each other on different promoters.  That means this might be a eukaryotic strategy similar to prokaryote's usage of polycistronic mRNA, an easy way to have similar expression patterns of genes that should often be expressed in similar amounts.

Do you think one of the reason it is more often found in eukaryotes is because they have longer genes and use of bidirectional promoter would decrease the chances of mistake in expressing downstream genes ?


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What is the relationship between the light and dark reactions of photosynthesis?
My main concern is why are the dark reactions (those that produce hexose) needed? As far as I understand, the sole purpose of photosynthesis is to synthesize carbohydrates, which are only used for energy storage. So, why don’t plants store energy directly as ATP (by photophosphorylation) and not as carbohydrates? (Of course, that would be too bad for other creatures.)
Elaborating my question:

Plants use chlorophyll to convert light energy into chemical energy of ATP and NADPH by photophosphorylation and chemiosmosis.
Then plants use the ATP and NADPH to synthesize glucose by the Calvin cycle.

So, what is the need of step 2? Why don‘t plants just use that ATP and NADPH for their metabolism?

PS: although @David has provided a nice answer to the current form of the question, the selected answer by @christian still remains selected since it answered my original query i.e. why excess ATP is not feasible for plants instead of why synthesis of carbohydrates is important. Any further answers/comments shall address the latter one, although none of them can be selected as correct since the latter query is not the original one.

No, photosynthesis is used to assimilate inorganic carbon in order to use it in biosynthesis (that's what "synthesis" stands for). It has little to do with energy. There are quasiphotosyntetic bacteria that only use light to harvest energy, hence they depend on first-level producers for organic carbon (organisms with functioning Calvin cycle, Arnon cycle etc).  You need organic carbon to make proteins, nucleic acids and lipids.

thanks, i was actually misled to the fact that carbohydrates are only synthesized for energy storage, and totally forgot that all our body is carbon-based, which we basically get from carbohydrates & their derivatives ;-)

Plants are Eukaryotes and have Mitochondria, so they perform glycolysis and oxidative phosphorylation as well. Animals don't store energy in ATP any more than plants do. We store it in glycogen and fats.

Yes sir @AMR I've understood it now and thats why I've selected the answer below as correct and said thanks to Eli for the comment ;-)

In response to your PS. If what you say was really your intent, why pick on plants? Animals don‘t store energy as ATP either. And your original title "Why is photosynthesis so complicated?" had a focus on photosynthesis, as if you had just been introduced to the subject and could see the importance of the light reaction (perhaps considered in one chapter of a book such as Berg et al., and were puzzled about the added complexity of a separate chapter on the dark reaction, which you assumed was for storage — as you admitted in your response to @EliKorvigo.

@david you're assuming just too much. I picked plants because animals dont synthesize glucose (well, from scratch). And my question title had a focus on photosynthesis because light and dark reactions are in (quite) direct relation, but no such relation is present elsewhere (especially in animals). Again, why this question was asked is a long story that I dont wish to touch upon.

@another'Homosapien' Regarding: "animals dont synthesize glucose" –  we actually do, that's called gluconeogenesis.

@eli ah, my friend! Thats why I said "from scratch". Comments, having size limit, really aren't places for writing with proper semantics, you just have to understand some things. All right, lemme tell you the story: this account belonged to my son, he asked this (and a few more) questions. I, reluctant to make a new ID, started using his account and answering questions. He barely uses this site anymore. So, please dont think I'm a newbie ;)

Possible duplicate of Why is ATP produced in photosynthesis used to synthesize glucose? or the other way round.

@WYSIWYG this one was actually asked earlier than the other one, so... :D

@another'Homosapien' I realized after tagging. I asked the mods to merge the other question to this one, if possible.

ATP is a bad medium for storing energy as detailed here.

ATP has a molecular weight of 507 Da
Glucose has a molecular weight
  of 180 Da, and contains the same amount of energy as 31 ATP molecules

This answer addresses one aspect of the question "Why store energy as carbohydrates rather than ATP?" It could have also considered the actual storage forms (polymerized hexoses) and factors like their neutrality and osmotic properties. However, it does not dispute the assertion in the question  that the sole purpose of the synthesis of carbohydrate in the dark reaction is for storage. It is not, nor is it even the main purpose, as explained in my own answer.

Apology
This question has a very simple answer which was provided in a comment by @EliKorvigo (and acknowledged by the poster) two years before writing this answer. However the commenter did not post an answer so the poster, instead of withdrawing his question accepted an answer that, although not factually incorrect, is not a full answer to the question. I feel that photosynthesis is an important enough topic that a more comprehensive answer should be provided.
The Fallacy
The poster writes in his question:

“the sole purpose of photosynthesis is to synthesize carbohydrates,
  which is only used for energy storage”

In fact, as is emphasized in simpler accounts than bloated biochemistry text books,

“the essential purpose of photosynthesis is to synthesize the
  carbohydrates — and from them the other organic compounds — that
  constitute living organisms themselves (plants and photosynthetic
  bacteria)”

The Purpose of the two reactions of photosynthesis

Dark Reaction: Converts CO2 to carbohydrate compounds
Light Reaction: Converts light energy to chemical energy to enable the
  synthesis of the carbon–carbon and other bonds of the carbohydrates,
  and the NADPH needed to reduce the CO2.

(For details see e.g. Ch. 20 and Ch 19, respectively of Berg et al.)
Distribution of carbohydrate in plants and other functions
Photosynthesis occurs in those (green) tissues of plants that contain chloroplasts. The carbohydrate product must be distributed to those other tissues that do not contain chloroplasts. This transport is done through the phloem, with the glucose 6-P and fructose 6-P first converted to the neutral transport form, sucrose. (see SparkNotes for a simple summary, or Liu et al. (2012) for a more advanced treatment.)
Although much of the sucrose will be metabolized to provide the structural components that constitute the tissues (especially cellulose), some of it will be used to generate the ATP and NADPH needed for metabolism.
Storage of carbohydrates in plants
The accepted answer can be taken as accepting the fallacy in the question that the purpose of  hexose production in photosynthesis is for storage, and further of implying that glucose is the storage form of carbohydrate in plants. Of course, polymeric hexoses, starch and sucrose are the main storage forms of carbohydrate in plants, together with some fats and oils, especially in seeds.

It is a good answer, but I selected the earlier answer because it was to the point. I do not have any problem with the modified form of the question (which, indeed, takes it to another direction) since it is also a good question for the site, but I cannot accept your answer now because it does not answer the original query (how I came to ask this question is a long story). Anyways, +1 for the effort :)

@another'Homosapien' — No problem about which answer is accepted. Just came across this question by chance and felt that an answer addressing the purpose of the dark reaction was required in addition to one dealing only with energy storage.

all_licenses: Creative Commons - Attribution Share-Alike - https://creativecommons.org/licenses/by-sa/4.0/


text:
How does the body switch between aerobic and anaerobic respiration?
Lets take the case of a person doing heavy exercise. Aerobic respiration is taking place, but oxygen is about to be finished up. Glycolysis occurs, Krebs cycle finishes. Now NADH and FADH2  enter electron transport system (ETS) to form ATP, but at the last step, cytochrome c oxidase cannot get oxygen as it just finished up. Now, what happens to the ongoing ETS? Does it go in reverse order and then arnon cycle to get back Acetyl co-A and respire anaerobically, or does this process get paused and respiration switches over to anaerobic mode till oxygen supply is replenished?

I think both @David and @SatwikPasani have given nice answers, but none of them seems complete and involving all details. So I am posting another answer which involves much of the required details for benefit of readers as well as myself.
The simplest answer would be that cell switches from aerobic to anaerobic respiration via allosteric inhibition of enzymes from product build-up. The details, in a stepwise manner, go as follows:

In the absence of oxygen, mitochondria activate a factor known as hypoxia-inducible factor-1 (HIF-1)1. HIF-1 helps in slowly shutting down the electron transport chain. This happens as HIF-1 induces subunit switching from cytochrome-c oxidase subunit 4 isoform 1 (COX4-1) to COX4-22. Also, HIF-1 induces activation of micro-RNA 210 (mir-210)3, which blocks expression of iron-sulfur cluster assembly proteins ISCU1/24, which are required for the function of complex I5, COX106, aconitase and subunit D of succinate dehydrogenase. Apart from this, in decreased levels of O2, nitric oxide (NO) causes inhibition of respiration and even cell death7. Also, nitric oxide synthase is a target of HIF-18. You can understand this by this diagram9:

Now, shutting down electron transport chain causes accumulation of NADH (and FADH2) in the mitochondrion. This build up, in turn, shuts another process known as malate-aspartate shuttle (MA shuttle), which is essentially a process to indirectly transfer NADH from cytosol to mitochondrion since mitochondrial membrane is impermeable to NADH10. Now, since malate-aspartate shuttle is not working anymore, thus NADH remains in the cytosol. See this through the diagram below:

Another point is that accumulation of NADH in the mitochondrion shuts down krebs cycle. It is so because two of the main enzymes, isocitrate dehydrogenase and $\alpha$-ketoglutarate dehydrogenase, both of which require the presence of NAD+ to function, are inhibited by the presence of NADH11. Also, aconitase and succinate dehydrogenase are inhibited by mir-210 as already discussed. Thus, inhibition of krebs cycle occurs, which leads to accumulation of acetyl-coA. See the diagram below:

Now, accumulation of both NADH and acetyl-coA inhibit the activity of pyruvate dehydrogenase12, which leads to accumulation of pyruvic acid in the mitochondrion. This is also clear from the above diagram.
Now, since pyruvic acid is not being metabolized, it is hence no more transported from cytosol to mitochondrion. Transportation of pyruvic acid from cytosol (after glycolysis) to mitochondrion (for krebs cycle) requires mitochondrial pyruvate carrier (MPC). Now, presence of malate is shown to promote the activity of MPC13. But since krebs cycle has already stopped, so no more malate is being produced. Also, since malate is not metabolized through MA shuttle (see second point), so it travels back into the cytoplasm (or is not produced at all). Thus, activity of MPC is significantly reduced. Apart from this, $\alpha$-keto acids are known to inhibit the activity of MPC14. Thus, due to accumulation of $\alpha$-ketoglutarate (which is not metabolized by $\alpha$-ketoglutarate dehydrogenase, see third point), activity of MPC is further reduced. Because of these effects, pyruvate remains in the cytosol. See the following diagram for clarification15:

In the last step, accumulation of pyruvic acid as well as of NADH in the cytosol together promote anaerobic respiration since in this process, NADH is converted to NAD+, which is possible due to the presence of NADH. This is also helpful since it not only helps in decreasing the accumulation of cytosolic NADH, but also in metabolizing the NADH generated from glycolysis. Finally, the complete process can be visualized by the following diagram:

P.S.: However, accumulation of lactate does not happen (i.e. a question like What if excess lactate inhibits anaerobic respiration? would not make sense) because lactate is readily removed from the cytosol to outside the cell so that it is transported to the liver, where it is converted back into glucose. This process is known as Cori Cycle16. See this diagram:

Bonus: A novel research has concluded that extracellular acidosis (induced by release of lactic acid), causes a dramatic increase in efficiency of ATP production by mitochondria during hypoxia(!). Under neutral hypoxia (without acidosis), mitochondria get fragmented and lead to impaired ATP production, causing cell death. But under acidosis-hypoxia, mitochondria become unusually elongated (by inhibiting DRP1-mediated mitochondrial fission17 and inducing mitochondrial fusion via Stress-Induced Mitochondrial Hyperfusion (SIMH) pathway18), cristae get remodelled through OPA119 which protects and maintains ATP production in the cell in hypoxia. See this diagram:

Since the whole process is reversible (by bringing extracellular pH to neutral), thus a dynamic equilibrium can be attained in cells with high oxygen demand, like neurons, through which cell can maintain high ATP production even in hypoxia.
References:
 1. Giancarlo Solaini, Alessandra Baracca, Giorgio Lenaz, Gianluca Sgarbi, Hypoxia and mitochondrial oxidative metabolism, Biochimica et Biophysica Acta (BBA) - Bioenergetics, Volume 1797, Issues 6–7, June–July 2010, Pages 1171-1177, ISSN 0005-2728, http://doi.org/10.1016/j.bbabio.2010.02.011.
 2. Fukuda R, Zhang H, Kim JW, Shimoda L, Dang CV, Semenza GL. HIF-1 regulates cytochrome oxidase subunits to optimize efficiency of respiration in hypoxic cells. Cell 129: 111–122, 2007
 3. Huang X, Ding L, Bennewith KL, Tong RT, Welford SM, Ang KK, Story M, Le QT, Giaccia AJ. Hypoxia-inducible mir-210 regulates normoxic gene expression involved in tumor initiation. Mol Cell 35: 856–867, 2009
 4. Chan SY, Zhang YY, Hemann C, Mahoney CE, Zweier JL, Loscalzo J. MicroRNA-210 controls mitochondrial metabolism during hypoxia by repressing the iron-sulfur cluster assembly proteins ISCU1/2. Cell Metab 10: 273–284, 2009
 5. Puissegur MP, Mazure NM, Bertero T, Pradelli L, Grosso S, Robbe-Sermesant K, Maurin T, Lebrigand K, Cardinaud B, Hofman V, Fourre S, Magnone V, Ricci JE, Pouyssegur J, Gounon P, Hofman P, Barbry P, Mari B. miR-210 is overexpressed in late stages of lung cancer and mediates mitochondrial alterations associated with modulation of HIF-1 activity. Cell Death Differ. 2010. October 1
 6. Chen Z, Li Y, Zhang H, Huang P, Luthra R. Hypoxia-regulated microRNA-210 modulates mitochondrial function and decreases ISCU and COX10 expression. Oncogene 29: 4362–4368
 7. Lee VY, McClintock DS, Santore MT, Budinger GR, Chandel NS. Hypoxia sensitizes cells to nitric oxide-induced apoptosis. J Biol Chem 277: 16067–16074, 2002
 8. Jung F, Palmer LA, Zhou N, Johns RA. Hypoxic regulation of inducible nitric oxide synthase via hypoxia inducible factor-1 in cardiac myocytes. Circ Res 86: 319–325, 2000
 9. Wheaton WW, Chandel NS. Hypoxia. 2. Hypoxia regulates cellular metabolism. American Journal of Physiology - Cell Physiology. 2011;300(3):C385-C393. doi:10.1152/ajpcell.00485.2010.
 10. Wikipedia contributors. "Malate-aspartate shuttle." Wikipedia, The Free Encyclopedia. Wikipedia, The Free Encyclopedia, 7 Mar. 2017. Web. 7 Apr. 2017.
 11. Berg JM, Tymoczko JL, Stryer L. Biochemistry. 5th edition. New York: W H Freeman; 2002. Section 17.2, Entry to the Citric Acid Cycle and Metabolism Through It Are Controlled.
 12. Budde RJA, Fang TK, Randall DD, Miernyk JA. Acetyl-Coenzyme A Can Regulate Activity of the Mitochondrial Pyruvate Dehydrogenase Complex in Situ. Plant Physiology. 1991;95(1):131-136.
 13. Mowbray A. Evidence for the role of a specific monocarboxylate transporter in the control of pyruvate oxidation by rat liver mitochondria. FEBS Lett. 1974;44:344–347. doi: 10.1016/0014-5793(74)81174-9
 14. Halestrap AP, Brand MD, Denton RM. Inhibition of mitochondrial pyruvate transport by phenylpyruvate and alpha-ketoisocaproate. Biochim Biophys Acta. 1974;367:102–108. doi: 10.1016/0005-2736(74)90140-0.
 15. Schell JC, Rutter J. The long and winding road to the mitochondrial pyruvate carrier. Cancer & Metabolism. 2013;1:6. doi:10.1186/2049-3002-1-6.
 16. Wikipedia contributors. "Cori cycle." Wikipedia, The Free Encyclopedia. Wikipedia, The Free Encyclopedia, 28 Feb. 2017. Web. 7 Apr. 2017.
 17. Smirnova, E., Griparic, L., Shurland, D. L. & van der Bliek, A. M. Dynamin-related protein Drp1 is required for mitochondrial division in mammalian cells. Mol. Biol. Cell 12, 2245–2256 (2001).
 18. Tondera, D. et al. SLP-2 is required for stress-induced mitochondrial hyperfusion. EMBO J. 28, 1589–1600 (2009).
 19. Frezza, C. et al. OPA1 controls apoptotic cristae remodeling independently from mitochondrial fusion. Cell 126, 177–189 (2006).

I think using end-product inhibition to illustrate the possible allosteric inhibition of early enzymes in aerobic respiration is misleading because 1. it would be the initial reactant (not the final one) — pyruvate that would build up, and 2. There is no evidence that pyruvate inhibits any of the key enzymes here. The NADH/NAD+ ratio might have an effect on pyruvate metabolism, but it is generally other compounds such as citrate that are the regulatory molecules, and these do not fit into that simple reaction pathway model.

The simple answer to your question is that in the absence of oxygen the electron transport chain stops. (It can't go backwards, that would generate oxygen.) However your assumption that it stops at the last step is incorrect, all the cytochromes are converted to their reduced form, and there is feedback that diverts the NADH elsewhere.
The situation is complicated because it depends what tissue you are talking about and what sort of exercise and what is the fuel for respiration. However let us consider the skeletal muscle at the start of a non-sprint run. Let us assume that it will be using glycogen as a source of fuel, which is converted to pyruvate and NADH. The pyruvate will enter the tricarboxylic acid (TCA) cycle in the mitochondria and be oxidized there and also produce metabolites. The NADH from glycolysis (produced by reducing NAD+) will enter the mitochondria and be oxidized; however it does not enter directly but via a shuttle, one of which — the malate-aspartate shuttle, show below — will be considered in a more detail lower down.
Now let us suppose there is a sudden need to sprint so that the muscle cannot get sufficient oxygen to deal with the NADH produced in glycolysis. What happens? The key event is that pyruvate is oxidized to lactate. Why? Not because lactate is useful, but because this is an anaerobic means of oxidizing the NADH back to NAD+ to allow glycolysis to continue and ATP to be produced at the substrate level.
Why doesn't the NADH keep going into the mitochondria (where it would be wasted as the electron transport chain has stopped)? Because the shuttle stops — in the case of the malate-aspartate shuttle mitochondrial malate dehydrogenase stops converting malate to oxaloacetate, malate builds up and therefore the malate transporter stops bringing malate into the mitochondrion.

This can be understood at the most general level by the fact that if NADH in the mitochondrion cannot be oxidized it will accumulate, hence slowing and eventually arresting the malate dehydrogenase reaction by simple mass action.
In fact Nature generally uses more sophisticated regulation that allows switch on and switch off to be performed more rapidly and completely, most notably by allosteric regulators. In the case of malate dehydrogenase the situation is not really clear, but there are reports that suggests citrate is an allosteric activator of the enzyme in some tissues (http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1131027/). The concentration of citrate falls as the TCA cycle stops because of the lack of NAD+ (this is because TCA cycle intermediates get tapped off for other purposes) so the activating effect of this intermediate would be removed.
(There is another shuttle for reducing power that my be more important in skeletal muscle. This is the glycerol 3-phosphate shuttle. However as this requires the oxidized form of FAD directly in the mitochondrial membrane it will clearly shut down in anoxia.)
The net result is that the NADH stays in the cytosol to be oxidized by pyruvate in the lactate dehydrogenase reaction.
This can only happen if the pyruvate stays in the cytoplasm and does not move to the mitochondrion. Obviously the lack of NADH to reduce it to acetyl-CoA in the pyruvate dehydrogenase reaction would cause its concentration to build up in the mitochondrion and reduce transport across the membrane by simple mass action. However it is likely that the specific mitochondrial pyruvate transporter is regulated by metabolites which change their concentration under these conditions. The evidence here is indirect as the transporter was only identified and cloned in 2012.

Nice answer, though some more explanation to "Because the malate-aspartate shuttle is subject to regulatory control" or diagram or at least a reference would be highly appreciated :)

The malate-aspartate shuttle and its regulation is a big topic. You can find more about it at https://en.wikipedia.org/wiki/Malate-aspartate_shuttle. Pasani has gone into regulation in more detail. For those not familiar with the molecular details  of allosteric regulation, try https://en.wikipedia.org/wiki/Allosteric_regulation.

In response to your request for more detail of the malate-aspartate shunt I have provided a diagram of my own which provides this but still focuses on what I think are the key points, that net movement of NADH into the mitochondrial matrix and the role of mitochondrial malate dehydrogenase. I prefer my revised answer to the merged answer as I feel it is more focussed. I'll comment on that answer.

Nice question. Now a perfect answer should include all the regulatory steps involved in this transition (a lot) and all the places where a change of metabolic fluxes occur. But, the question can be answered in essence at least, by a very simple model.
Assume that no other regulations act except direct enzymatic feedback inhibition. This means that substrate being acted upon by an enzyme converting it to a product will stop being acted upon when the concentration of the product rises. Now, for the case in question. Lack of oxygen means that cytochrome c cannot work. Hence, it's substrate will accumulate, which is the product of the preceding enzyme, which will be blocked leading to the accumulation of its substrate and so on. This "waterlogged" system will continue leading to metabolite accumulation till pyruvate starts piling up. Now this pyruvate piling does something different because it had two enzymes that could act upon it. The other enzyme, which converts it to lactate does not have this feedback blockage and hence can function. So, the pyruvate is now converted into lactate, which requires NADH, and hence allows glycolysis to proceed (because the formation of NADH was an indispensable reaction in glycolysis), and hence, you have the anaerobic shift. Lactate doesn't pile as it has a favourable muscle-blood gradient and is quickly transported out the cell, and also because it has to pile up significantly before the enzyme forming it is back-logged.
Now, to add the other regulations. What all other regulatory factors do, is to enhance this waterlogged feedback. Instead of the product piling up to a huge amount before the enzyme is inhibited, these regulations will allow a smaller pile-up to produce the feedback inhibition. So in essence, you can view the shift as a simple consequence of blocking the enzymatic chain at a particular level.

A good source of basic biochemicl information is Stryer online at NCBI: http://www.ncbi.nlm.nih.gov/books/NBK21154/ . It's an old edition and you can only search it (not browse it), but the information and presentation is of a high standard.

I think the treatment of pyruvate build up here is a little simplistic as it neglects the movement of pyruvate between cytosol and mitochondrion and the role of the mitochondrial pyruvate transporter.


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Why do women live longer than men?
According to Wikipedia and many other sources, women live substancially longer than men (over 5 years in the U.S.).
Men can reproduce for much longer than women.  So longer living men would have the most offspring, while the longevity of the women would be more or less irrelevant in reproducing.  These offspring would be the product, on average, of men and women where the men are older than the women.
So why do women live longer than men?  Am I misinformed, or what don't I get?

In my place it is the vice versa :)

I believe reproduction itself has something to do....i seem to recall that links have been seen between sexual reproductivity and shortened lifespan.
See this, for example: Bowen RL, Atwood CS. Living and dying for sex. A theory of aging based on the modulation of cell cycle signaling by reproductive hormones. Gerontology. 2004; 50: 265-290.

Behaviour
I would think that the most important causal factor between this difference is behaviour. Men probably smoke more than women, probably eat more junk food and are eventually more stressed. Men also probably take more risks and have more accidents. This last point might have been very important at an earlier time when men were hunting outside while the females would work inside the cave. Depression is more prevalent in men as well and it is possible that might affect health. These differences in behaviour might be caused by genetic (and epi-genetics) differences (sexual chromosomes) or by culture.
Oestrogen
There was a time where some doctors thought that the different was due to oestrogen but I think that this hypothesis has been ruled out.
The Y chromosome is short
Men have a Y chromosome while women have two X chromosomes. At each mitosis, the chromosomes get shortened (telomere are shortened). It is possible that the presence of the Y chromosome may explain some of the observed difference due to its small size. However, according to @cantona'sCollar the Y chromosome is often lost in somatic cells and therefore the difference of length between the X and the Y chromosome doesn't seem to be a good explanation.
Grandmother hypothesis
While women stop reproducing earlier than males, women often provide more care to their grandchildren than men do. As a result of this behaviour, genes coding for longer lifespan are beneficial by the fact that they help other copies of themselves (found in the grandchildren) to thrive (kin selection).

Depression is more common in women not in men:
http://www.nimh.nih.gov/health/publications/depression/index.shtml?ct=39994
Also, I would say that the roots of different behaviors are most likely to be in the genes since lifespan of men are shorter than women in most cultures. My guess would be the effect of testosterone. Men are more prone to take risks - even the life threatening ones - because of this hormone while women are more likely to choose the safer path.

I edited my answer to basically redraw some claims that were wrong or at least had little support. Thanks for your comment.

In reasonably fit people, men die from cardiovascular events at least 5 times more than women even after adjustment for smoking, cholesterol, high blood pressure, blood glucose, etc. This endocrinology paper of August 2013 attributes some of that to estrogen. I do agree that telomere length and mitochondrial DNA play a role, but isn't telomere length maintained on non-X chromosomes in women longer than men, or am I mistaken?

I have no idea if telomere shorten faster in all chromosomes in men. In this kind of discussion it is important to not forget the cause-consequence framework. It is possible that men died earlier because of more risky behaviour (such as hunting) and in consequence they would have relaxed selection at later age allowing late acting deleterious alleles to accumulate on the Y chromosome and things  like that.

I should say that the bit about Y chromosomes does not seem right. A "-Y" abnormal karyotype in somatic cells is frequently seen in older men (>70 years of age) and it doesn't seem to necessarily correlate with any clinical abnormalities.

@Cantona'sCollar What do you mean by A"-Y"? Do you mean that the Y chromosome end up merging with an autosomal chromosome? That's interesting. However, It doesn't change the fact that if it does not happen to everyone, you will still see a different life expectancy for men and for women. Also, if life expectancy were to differ only slightly due to telomere length of the Y chromosome, then it would reduce selection at later ages and further reduce life expectancy.

"-Y" means a loss of the entire Y chromosome. If there was a translocation with an autosome or something, it would end up being detected in standard karyotyping analyses (usually based on banding patterns).
The Y chromosome has so few genes on it, and most are involved in fertility/fecundity and are not very likely to contribute to long term health. Therefore, I cannot imagine how the presence/absence of a Y has anything to do with explaining different age spans of men vs women.

Oh ok good point. I edited my answer in reaction to your comments. Thank you

The best explanation I have come across so far is that male sex hormones have an adverse affect on longevity.
The effect may simply be that a high testosterone level suppresses the immune response against a variety of infections.
There was a recent study about korean eunuchs from a few hundred years back, that found that "The average lifespan of eunuchs was 70.0 ± 1.76 years, which was 14.4–19.1 years longer than the lifespan of non-castrated men of similar socio-economic status."
That's a pretty hefty difference and a strong argument for testosterone as risk factor for an earlier death. This would also vibe well with reports about increased risk of cardio-vascular death in steroid users.


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Formation of life on other Planets
At the present time it looks as if there is no other life in the Universe other than here on Earth . If this is true then why didn't it occur anywhere else ? Is it possible for something to happen only once ? Hundreds of Billions of  other Planets have formed so whatever happened here why wouldn't it have happened again elsewhere ? It is very , very strange indeed .

Although this is an interesting question, it has few problems with it. 1, I think it's off topic (too broad / unclear what you're asking - in biology topic), and suits astronomy better. 2, there's no proof that life does not exist in some other from somewhere else (as far as I know Mars has signs that it was habitable to some extent in the past - for microbes at least) .

@poka.nandor Most often I post my questions on astronomy but I thought this fitted biology more. I know that there is no proof either way but recently I read that Scientists were scanning millions of Galaxies and could see no indication of life anywhere else. We have to keep looking and to keep an open mind but until we find some evidence it will look like we are alone in the Universe. So what I am asking is how could it , life , occur only on one world out of trillions of worlds and how strange this is if this is actually the case ?

I'M not even sure that with current technology we could tell if Earth has life, even within from our solar system.

@PeterU do you have a reliable (preferably peer-reviewed) reference to back up your statement that "Scientists were scanning millions of Galaxies and could see no indication of life anywhere else"? What were their readouts? $CO_2$? The color green? This completely contradicts the mounting evidence of large numbers of rocky, "Earth-like" planets withing a sun's habitable zone.

@poka.nandor yes, you can, with listening to radio waves, for one thing. See here for one description of how planets are evaluated for "life as we know it".

Emphasis on life as we know it... what about planets that dont have civ with radio tech...like earth did not have up until like a hundred years..i know that there are molecules you can look for that might indicate presence of life but detecting those fom millions of kms /light years...

This project tried to look for advanced alien civilizations by looking for their waste heat from other galaxies. The recent news is that they haven't found anything, but this project would probably only detect civilizations that had spread across entire galaxies and used a lot of energy. This doesn't say anything about the presence or absence of biological life on other planets. As far a good way to detect life, molecular Oxygen gas would probably be good.

Issues with the post
I agree that the question is too broad. It is not the only issue, it is also primarily opinion-based (see below for explanations). The post is also somehow unclear (see below for explanations). But here is some kind of an answer that may help you.
If you google your question you will get tons and tons of hits. There is a whole lot of videos on youtube that address this question. You may want to have a look at those and come on StackExchange to ask a more accurate question because for the moment the question is too broad.
As a first short answer to the questions...

At the present time it looks as if there is no other life in the Universe other than here on Earth

It sounds wrong (as a consequence your question would be primarily opinion based). We haven't found life. This doesn't mean that life doesn't exist elsewhere. Actually most people think that life exist elsewhere. Part of the issue behind this question is the definition of life. Often, people refers to Drake equation (you might want to read about it) which gives you the number of planets where life should exist. However, Drake equation is a speculation that has no solid grounds. It is actually pretty complicated (or impossible because we need to assume things about what the original conditions are) to calculate a probability that life occurs on a given planet. While it sounds pretty intuitive to many people (me, including) that life should exist somewhere else, there is really not much certainty.

Is it possible for something to happen only once?

Well....yes, the death of Napoleon occurred only once.... I am not quite sure what you mean with this question, it is unclear.

b What I meant with the question is, if it was possible for life to form on this Planet then how could it be that it did not happen again elsewhere ? Since life came about here that shows that it is not an impossible event and yet so far there is no evidence that it ever happened again. Is this not strange ? Maybe we might never be able to find the answer and also we don't know what could have happened in the past, or what will happen again in the future, with regards to life evolving on a Planet.

Take some time to read stuff on the subject, I cannot make a whole overview of the field, your question is too broad. Your comment has been partially answered above but I can't make much more for you. Really, just take some time to read or watch videos on the subject, there are tons of ressources on the question. You should end up understanding the issues related 1) to the definition of life, 2) to the easiness of detecting life (we haven't sent robots all over the universe yet!) and 3) to how complicated (or impossible) it is to calculate any probability of such thing to occur.

No need to down vote this answer, as there is no other practical method to answer this type of question. Another option is to close the question immediately with out trying to answer it. +1 for the effort put towards answering it although its difficult to answer.

@PeterU: May be there are millions of planets swirling with life or may be not. But so far we cant judge whether there is life exists outside Earth. May be life is not such a big deal as we think it is. Scientific and technological knowledge are developing quite faster and we can believe that this will one day guide us to some farther planet were life is going on or some body else will find us in this remote corner. All we have to do is to patiently wait for it. (cont...)

Most of our intelligent ancestors were unaware of many scientific principles and yet now we idiots know much more than them. Many more years later may be our grand grand grand sons may worry that their grand fathers(us) were unaware of the fact that we are living in a Universe full of life. May be this comment doesn't satisfy your urge but all I can offer is this. :)

@Jay , Yes, I understand what you are saying. At the present time there is no way of knowing but perhaps in the future we may know . I was just stating that it would seem to be more logical if life exists elsewhere as well as here. If it turned out that we are alone it would seem very strange somehow ?

Yes, it would "somehow" seems strange indeed. But really, you will easily find all information you need if you just google your questions. You will have an idea of what are the difficulties in estimating the number of independent origin of life and on the likelihood of actually finding life given that it exists somewhere.

@PeterU: Considering the vastness of Universe, we will never find out we are alone even if we are. We will be in constant search for it. If we found out we are not alone our plans for future will still be to found out more. Life is just like that, It will never let us RIP.


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Lifeless Lake in Siberia
There is a lake in Siberia called Lake Pustoye. No life exists in the lake. There are no fishes and no plants and even trees around the lake die . No birds even visit this lake .The lake even has bubbles in it like champagne .The water has been tested and there is nothing in the water that makes it harmful to life.  It even tastes like champagne and yet life cannot exist in it .Fish were introduced into the lake but they quickly died off . So why can't any life form survive in this lake ?

Can you give links with evidence that The water has been tested and there is nothing in the water that makes it harmful to life?  I.e. not just the claim that this has been done, but some specifics saying who has done it, and what they've found? Otherwise, this looks like just another click-bait made-up claim.

Yeah a I agree with Iayork a claim like that needs some serious sourcing otherwise you can assume it is wrong.

Assuming the claim is supported. perhaps the water is saturated with volcanic gasses, like Lake Nyos in Africa: https://en.wikipedia.org/wiki/Lake_Nyos  That would account for the bubbles and possibly the absence of animal life, though I can't see why it would affect plants or microorganisms.

The article said that supposedly several countries around the world have tested the water and found nothing harmful about it . Also if there is no vegetation or other life forms then how did they expect the fish to survive that they attempted to introduce into the lake . What would the fish have ate ?

If you google" Lake Pustoye Mystery" the very first link on the page "The Secret of the Siberian Pond" will take you to the article @iayork

@PeterU You understand that that article is completely devoid of facts, right? It makes claims about 'large numbers of scientists", "experts", and "specialists", but gives absolutely no information on who they are, where they're located, what they are specialists in, what they looked for, and what they found. It's classic click-bait vagueness and obfuscation. If that's all you have, I'm voting to close this question as "Unclear what you're asking", since you have no actual facts to discuss.

Pubmed, which includes over 28 million peer-reviewed articles, has 0 instances of "Pustoye" in it, suggesting that no experts have actually looked at the lake.

And finally, The Geographical Journal, Vol. 19 (1902) refers to villagers living on Lake Pustoye who "make a comfortable living by fishing", which suggests the lake is far from empty.  I think this is some kind of misunderstanding (perhaps based on a translation of the name) that has been mindlessly parroted by the gullible and sensationalist.

Sounds like carbon dioxide, it seeps up from an under ground source. At high levels it will kill all normal life. There is at least one lake in Africa that has this  CO2 saturation. There was a program on TV a few years ago about it. There is some unusual circulation and out gassing but I forgot the mechanisms..CO2 would explain the "tastes like champagne" .

So I guess that is the reason that my Doctor said that I shouldn't believe what Doctor google says, lol . So in other words you are saying that villagers are fishing in this lake, which would mean that the article is false ? @iayork

@iayork - it sounds all like an excellent answer. Combine the comments (mind to include the links) and et voila?

If the water is bubbly and taste like  champagne, it means that the water is saturated with carbon dioxide, and has a pH of about 3. the water probably kills everything (plant, animals and most bacteria) around it by carbon dioxide poisoning and very acidic water.

I am more curious if something can survive in the lake. Lake Pustoye has actually been tested and scientists have found nothing wrong at all with the water, and when they saw that the lake had bubbles and tested those too, again they found nothing, so they did a few tests. First they put the least demanding fish in the lake but soon found that they all died, they tried the same thing with aquatic plants, those just rotted away. They just straight up labeled it as a dangerous lake due to nothing being able to survive in it, what I am personally curious about is if a tardigrade, aka water bears

https://www.researchgate.net/publication/200464654
This is a detailed comparison between two lakes in the Trans-Volga region in Russia, one of them being Lake Pustoye. I'm not sure if the Lake Pustoye in this article is the same lake being referred to by the op as it isn't in Siberia and there's no mention of champagne-like water, but it does bear the same name and has a curious lack of fish. The authors found that while there is an absence of fish, there is by no means an absence of life in the waters of Pustoye. In the article, they catalog the seasonal shifts of biodiversity in the waters of both lakes. The pH was observed to be low in Lake Pustoye which they implicated to be the likely cause for the lack of fish, though they indicated that more research would be needed to determine the exact cause of the acidification.

Impressive detective work, Sherlock. +1

Pustoye, or Пустое, means empty. It's not an uncommon name for a lake. This is certainly not the one the apocryphal story is about.  Search Озеро Пустое in google maps and you'll find many of them.

license: Creative Commons - Attribution Share-Alike - https://creativecommons.org/licenses/by-sa/4.0/


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Length of sleep time
Why do we wake up after 6-8 hours of sleep ? Why don't we sleep for lets say 20-30 hours ? What mechanism is it that controls when to wake us up and how does it determine the moment that we should wake up ?

Related question.

It's considered a requisite on biology.se that at least some reading/research is carried out and presented with your question. Otherwise it will be closed as homework. [For example, have you read about circadian rhythms?] The site tour and the help center  provide guidance on how to use this site. Please take a few minutes to read about the kind of questions which are on topic here.

Think this review from Nature can answer your question. A brief summary  ( with slight simplification):
The ascending activating system (that is responsible for wakefulness) has two branches:
The first one ascends to the thalamus and activates the thalamic relay neurons - these transmit information to the cortex. They get their main input form two acetylcholine producing group of cells:  the pedunculo-
pontine and laterodorsal tegmental nuclei (PPT/LDT). These two group of cells are also the source of main input for reticular nucleus of the thalamus.

The neurons in the PPT/LDT fire most rapidly during wakefulness and rapid eye movement (REM) sleep, which is the stage accompanied by cortical
  activation, loss of muscle tone in the body and active dreams

The second branch, directly activates neurons in the hypothalamus, basal forebrain (BF) and cortex, bypassing the thalamus. This pathway has origins in monoaminergic neurons in the upper brainstem, caudal hypothalamus, as well as the noradrenergic locus coeruleus (LC), serotoninergic dorsal (DR) and median raphe nuclei, dopaminergic ventral periaqueductal grey matter and histaminergic tuberomammillary neurons. Input from this pathway to the cortex is extended /augmented by the lateral hypothalamic peptidergic neurons (that contain melanin-concentrating hormone (MCH) or orexin/hypocretin), and BF neurons(that produce acetylcholine or GABA).
Sleep is promoted by the so called VLPO (ventrolateral preoptic nucleus).
The VLPO send outputs to major group of cells of the arousal system in the hypothalamus and brainstem.

The VLPO neurons are primarily active during sleep, and contain the inhibitory neurotransmitters, galanin and GABA.
  The VLPO also receives afferents from each of the major
  monoaminergic systems. Both noradrenaline and serotonin inhibit
  VLPO neurons. The latter do not have histamine receptors, but the
  tuberomammillary neurons also contain GABA, which is inhibitory
  to VLPO neurons, as well as several other potentially inhibitory pep
  tides, such as galanin and endomorphin. Therefore, the VLPO can
  be inhibited by the very arousal systems that it inhibits during sleep.

As you can see in the quote from the paper these two systems can mutually inhibit each other and basically work as a seesaw or 'flip-flop switch'. In this system activity in one  subsystem shuts down the inhibition from the other subsystem and disinhibits its own action (selfreinforcing loop). The transition from one state to the other is relatively fast. Orexin plays important role in stabilizing the switch in one position or the other.

Regulation pf sleep:
Homeostatic:
It is unclear how homeaostatic regulation of sleep work, yet it has been proposed, that adenosine accumulation in the brain may signal for 'need for sleep'. The extended wakefulness causes the exhaustion of brain glycogen reserves and depletion of ATP levels. As ATP is degraded to ADP, AMP and eventually adenosine, extracellular adenosine levels rise in some parts of the brain including the BF.

In addition, adenosine might disinhibit the  VLPO via presynaptic A1 receptors by reducing inhibitory GABAergic inputs.

This implies that during sleep ATP and energy sources are regenerated and free adenosine levels drop, thus inhibition of VLPO might take place again.
Also it is important to note that the amount of sleep we need is proportional to the time we spend awake.
Circadian regulation:
It is known that humans have quite precise :circadian rhythm:

The suprachiasmatic nucleus (SCN) serves as the brain’s ‘master
  clock’. Neurons in the SCN fire in a 24-hour cycle that is driven by a
  transcriptional–translational loop, which persists even when the neurons are dissociated in cell culture.

The SCN has moderate output to the VLPO or orexin neurons, but it has more important projections to the adjacent subparaventricular zone (SPZ) and the dorsomedial nucleus of the hypothalamus (DMH).

The SPZ also has relatively limited projections to the VLPO, orexin
  neurons and other components of the wake-sleep-regulatory system. However, a major target is the DMH. This region receives inputs from many more neurons in the SPZ than the SCN, so the SPZ is in a position to amplify the output of the SCN. (...) The DMH is one of the largest sources of input to the VLPO and orexin neurons and is crucial for conveying SCN influence to the wake–sleep-regulatory system. The DMH projection to the VLPO comes largely from GABA-containing neurons (that is, those that promote wakefulness by inhibiting sleep), and the projection to the LHA originates from neurons containing glutamate and thyrotropin-releasing hormone (which should presumably be excitatory and promote wakefulness).

So as you can see our circadian rhythm as direct control over our sleep-wake cycle, and that the SCN can heavily promote awakening through SPZ and DMH.
Thorough review and details can be found in the article I linked at the beginning, I only tried to select key information to provide a (not so ) brief summary. We only sleep about 7-8 hours normally because this is enough for homeostatic regeneration and our circadian rhythm promotes wakefulness in the morning (this is quite simplified).
Based on their abstract these limited access papers might also be useful (unfortunately I do not have full access to them right now):

http://www.eurekaselect.com/87813/article
http://www.sciencedirect.com/science/article/pii/S0074774210930018
http://www.sciencedirect.com/science/article/pii/S0896627315001312

I think what the OP is getting at is less about the mechanism of sleep -> wake transition and more about how the length of sleep time is regulated...
In that regard, it's an interesting question without a definitive answer. In a sense, it is a question of homeostasis and the time when your sleep need/pressure is satisfied determines when you wake up (without an alarm clock).
It is hypothesised that sleep drive is a function of the previous wake period plus any sleep debt you carried from the previous days.  One molecule, adenosine, is thought to represent that and the accumulation of that in the brain makes you want to sleep [Ref. 1].  Thus, the clearance of external adenosine in the brain to a certain level may be part of the homeostatic mechanism signalling enough sleep.
Another hypothesis is that sleep is required to encode memories long term [Ref. 2] and to calibrate emotion/mood [Ref. 3] which themselves can add to the sleep need and those processes would need to be completed to a certain level to release that sleep need.
As for the specific moment for awakening, if there isn't some external arousal stimulus like daylight/alarm or the rush of cortisol cycle driven by your circadian mechanism, then it will likely correlate to the end of the current 90min sleep cycle when your brain is close to the wake state coming out of REM [Ref. 4]. In fact, it's thought you wake a little every 90min through the night, it's just you don't remember it.
[Ref. 1]: Link to adenosine and sleep review
[Ref .2]: Link to sleep and memory review
[Ref. 3]: Link to sleep and mood review
[Ref. 4]: Link to brief summary of sleep staging


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creating life in the laboratory
Originally life came about through non life but Scientists are unable to achieve this in a laboratory . Why is this ? Is it the time factor ? I believe that it took one billion years for life to evolve here on Earth did it not ?

Should this not be tagged abiogenesis rather than evolution?

What attempts to answer this question have you already taken? We ask that all question posters here attempt to search for an answer to their own question and explicitly indicate what research they've already done, what they learned, and what is still confusing or unknown to them. Our goal is not to simply be an answer site, but rather a site that promotes self-learning with some expert help along the way :). Please take a moment to edit your post with this additional detail, and it will likely be received more positively by our community. Thanks!

It seems that science has moved on somewhat, and scientists have created 'living' (self-replicating) artificial bacterial cells. They built the genome
from physical materials and transplanted the engineered genome into an existing bacterial cell.  This site has a timeline of the history of the project https://www.jcvi.org/first-minimal-synthetic-bacterial-cell

So they have only built the genome and not an entire living cell ? How far away are they from creating a self replicating living organism ?

Well, they have built an entirely new self-replicating organism, but needed an empty cell to get it started.  They may not prioritise making a cell from scratch, because bacterial cells are plentiful.


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What is the importance of urea in mass spectrometry?
What is the importance of urea in mass spectrometry? We use 8M urea to FASP our proteins prior to mass spectrometry.  What is the significance of using 8M urea? and how does it affect the proteins?

What does FASP mean? Otherwise high concentrated urea is used to denaturate proteins.

@Chris filter aided sample preparation

I think that the OP was asking about relevance of using urea with respect to the FASP method. In the FASP method, the primary denaturant is SDS . Protein are denatured with a ~4% SDS solution (buffered to pH 7.5 - 8.0). Then 8 M urea solution is used to replace the SDS. Urea serves two purposes here, first it keeps the protein denatured and in solutions as SDS is replaced/washed off. Second, because urea is a chaotropic agent, it reduces the size of SDS micelles which otherwise would block the pores of the membrane filter.
Read the original article here: http://www.nature.com/nmeth/journal/v6/n5/abs/nmeth.1322.html

One more thing I think should be added: it is important to understand that denaturants like urea, GuHCl, or SDS not only unfold/denature protein, but also keep them in solution thereby allowing their biochemical analysis. Otherwise heating to ~100 ºC or using Trichloroacetic acid can also denature protein, but they render protein insoluble.

8M urea is a denaturant, as @Chris mentioned. Denaturing proteins is important for molecular (aka classical) mass spectrometry. If you analyzed a non-denatured protein, you'd get information not only on the primary structure, but also on noncovalent interaction (2+°):

The classical MS approach, so-called “molecular MS,” analyzes, in the gas phase of the mass spectrometer, individual species initially present in solution after destruction of the noncovalent framework. On the other hand, “supramolecular MS” or “nondenaturing MS” aims at transferring intact noncovalent complexes that preexist in solution into the gas phase of the instrument.
(Sanglier et al. 2008, Methods Mol. Biol.)

Denaturing proteins is important for molecular/classical mass spec because it ensures your analysis only reflects the covalent bonds (primary structure) present in your target protein.


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Why is carbonylation added as a fixed modification in mass spectrometry peptide analysis?
When using mass spectrometry analysis software we use carbonylation as a fixed modification. What is the significance of this modification and why doesn't it change?

which MS method are you using ?

We are using  dda on a high  res lc- ms


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Use of salt as preservative
I know that salt is used as preservative as it dehydrates microbes. Is there any other advantage like - altering pH or inactivating microbial enzymes ?

salt cannot change pH directly. Dehydration which causes the loss of bulk water can alter the pH and also affect protein activity.

High salt environments are strongly hypertonic and disrupt the osmotic gradients of cells.  Excess salt gets forced in (disrupting function) and cell water gets drawn out.
In simpler terms, it dehydrates them.
This link has more info regarding what is required to live in high salt environments (and by corollary, why organisms without such features die):
http://en.wikipedia.org/wiki/Halophile

Another factor involved is the effect that high salt concentrations have on the 3-D structure of proteins. At higher than "normal" concentrations of salt, many proteins will fold differently and no longer be in their "active" form. Since all biological reactions involve enzymes, which are protein, you can see why this unfolding might be problematic.

Salt draws water out of cells via the process of osmosis. Essentially, water moves across a cell membrane to try to equalize the salinity or concentration of salt on both sides of the membrane. If you add enough salt, too much water will be removed from a cell for it to stay alive or reproduce. Organisms that decay food and cause disease are killed by a high concentration of salt. A concentration of 20% salt will kill bacteria. Lower concentrations inhibit microbial growth, until you get down to the salinity of the cells, which may have the opposite and undesirable effect of providing ideal growing conditions! Salt can also be used when you are preserving a product via fermentation. The salt will inhibit the growth of moulds and yeasts. While some moulds and yeast are certainly needed during the fermentation process, by using small amounts of salt, it is possible to regulate and stabilise this process.
It is only possible to do this though if you are using salt that does not contain ‘free-flowing’ agents or iodine.
Refer to :https://suite.io/rachel-tsoumbakos/xpx288 for details.