Wissenschaft

New insights into sensory representations-Lessons from the auditory cortex

noreply@blogger.com (Varun) — Sun, 09 May 2010 14:06:00 +0000

In the brain, there are highly ordered representations of sensory input. The existence of orientation columns in the visual cortex where columns of neurons situated next to each other respond to slightly different stimulus orientations and the barrel cortex in S1 where each barrel faithfully receives inputs from one whisker are testimony to this. Recently two papers in the same issue of Nature Neuroscience dealt with the fidelity of sensory representations in the auditory cortex. Rothschild et al. and Bandyopadhyay et al. used in-vivo two photon microscopy to map tone evoked activity in the primary auditory cortex (A1). This is done by bulk loading a large area of the cortex with a membrane permeable calcium dye. When a neuron that has taken up the dye fires an action potential, there is also a transient influx of calcium (related to synaptic transmission). The interaction of the calcium ions with the calcium indicator can be visualized under a two-photon microscope.

Both studies showed, that unlike the visual and barrel cortices, the auditory cortex appears to have a tonotopic map that is fractured. Firstly, less than half the neurons were responsive, and even if they did respond, neurons with similar tuning curves were as likely to be located next to each other as neurons with very different tuning curves. Fig 1 (taken from a review by Castro and Kandler, Nature Neuroscience 2010) elaborates on this.

Fig1a would be the classic tonotopic map where there is a smooth change in frequencies along the rostrocaudal axis with more rostral neurons coding for higher frequencies while more caudal neurons code for lower ones. Here, tonotopy is maintained on the local as well as the global scale. Fig 1b summarizes the results of Rothschild et al. and Bandyopadhyay et al. Although the tonotopic organisation is maintained on a more global scale, locally the map appears to be fractured (From Castro and Kandler, 2010)

So why is the tonotopic map fractured? One possibility is that the thalamocortical projections from the auditory thalamus to A1 become scattered en route. Alternately, thalamocortical axons may be arranged tonotopically but resulting intra-cortical processing may result in the fractured nature of A1. To distinguish between these two possibilities, Bandyopadhyay et al labeled cells with two different calcium indicators, Fluo 4, a low affinity indicator that responds only to spikes in the cells, and OGB-1, a high affinity indicator that responds to subthreshold synaptic inputs into the cells. They found that subthreshold maps were more ordered in comparison to suprathreshold maps based on spiking (Fig 2).

Fig2; Subthreshold and Suprathreshold maps (From Castro and Kandler, 2010)

So what does all of this mean? Bandyopadhyay et al suggest that although two neighboring neurons may receive similar , correlated inputs, they may be part of different fine-scaled assemblies that could process inputs differently. Two adjacent cells may be selective for different input features or different stimulus attributes. Taken together, both studies indicate that frequency is perhaps not the most important feature coded by A1 neurons. Neither is intensity tuning or bandwidth. A1 neurons probably respond to meaningful stimuli and not just to simple sound parameters. This is supported by studies that have shown that A1 neurons are best driven by spectrally and temporally rich stimuli.

References:

Castro JB, & Kandler K (2010). Changing tune in auditory cortex. Nature neuroscience, 13 (3), 271-3 PMID: 20177415

Rothschild, G., Nelken, I., & Mizrahi, A. (2010). Functional organization and population dynamics in the mouse primary auditory cortex Nature Neuroscience, 13 (3), 353-360 DOI: 10.1038/nn.2484

Bandyopadhyay S, Shamma SA, & Kanold PO (2010). Dichotomy of functional organization in the mouse auditory cortex. Nature neuroscience, 13 (3), 361-8 PMID: 20118924

Why it pays to be a biologist

noreply@blogger.com (Varun) — Sat, 24 Oct 2009 10:15:00 +0000

I have nothing against theoretical physicists or mathematicians, but it's a lot safer being a biologist. Here's why

For more geeky humor, visit xkcd

Deep Sea Killers-C. megalodon

noreply@blogger.com (Varun) — Sun, 18 Oct 2009 08:41:00 +0000

Many consider the Great White Shark (Carcharodon carcharias) to be among the most incredible creatures to roam the oceans today. Growing up to a length of 6m (20ft), it is the infamous creature made famous by the success of the movie Jaws.

The Great White Shark as a species can be traced back to the early Pliocene (5 million years ago). It's fossilized teeth (Fig.1) can be found almost anywhere, worldwide, in marine sediments of the correct age. Teeth found in the Miocene (15 to 13.5 million years ago) are slightly rolled and have an eroded edge. Common synonyms for these teeth are Carcharodon rondeletti and Carcharodon sulcidens, but the teeth are identical to those of the living species not regarding intraspecific variation.

Fig. 1. Fossilized tooth of Carcharodon carcharias. Height: 5 cm. Early Pliocene, Sacaco/Peru © L. Andres

About 16 million years ago (during the Miocene), a distinct species appeared in the world's oceans. Carcharodon megalodon (or C. megalodon) was possibly the biggest shark species to have inhabited the oceans. It may have attained an astonishing maximum length of 15 m (50ft) and weighed as much as 50 tonnes (Fig.2)

Fig. 2. Comparing C. megalodon (13 m) and the Great White Shark (6.5 m) © L. Andres

Such estimates are obtained from teeth and certain skeletal components (as sharks have a skeleton made out of cartilage that does not fossilize easily; the teeth however are very durable). Traditional research holds C. megalodon as an ancestor of the white shark. Recent research suggests that it may have been a close relative. It's triangular teeth may have reached a maximum height of 17 cm (Fig.3). It may have hunted in the same stealthy manner as white sharks do, stalking beneath it's prey and rising upwards at great speeds to deliver a forceful, and often fatal, first bite. It's prey probably included primitive whales and other large marine mammals.

Fig. 3. Fossilized tooth of Carcharodon megalodon. Height: 13 cm. Middle to Late Miocene, Florida/USA © L. Andres

Around 1.5 million years ago (towards the end of the Pliocene), C. megalodon became extinct. The development of megatooth sharks can be traced back until the Cretaceous period. It is directly linked with the development of other animals. In the Cretaceous not only sharks, but also marine reptiles ruled the waters. This condition changed, however, after the extinction of the dinosaurs (65 million years ago) in favour of the sharks. Sharks now occupied the ecological niches for predators. Additionally, the basis for a more energy-rich nutrition was created by the rise of marine mammals (such as the cetoheriids; ancestors of baleen whales) in the Eocene (55 to 33 million years ago). During the Late Oligocene (30-25 mya) the climatic conditions were much more favourable. Temperatures were significantly higher than they are today, tropical and subtropical waters reached much further into the higher latitudes of the polar regions. During the following epoch, the Miocene (25-5 million years ago), modern baleen whales (Mysticeti) developed and spread more and more. There was an increase in size in whales and simultaneously in megatooth sharks. During the spreading out of the whales they presumably reached cooler polar waters that provided them with a richer food supply to which the whales adapted themselves. The whales were migrating between cool water feeding grounds and warm water breeding grounds. The climate became colder at the end of the Miocene and the beginning of the Pliocene (about 5 million years ago). The ice cover of the Antarctic polar region grew bigger and the mean sea level dropped. Living conditions and habitat for C. megalodon, who probably loved warm waters, obviously were restricted to such a degree that the species became extinct during the Pliocene.

Reference:

C. megalodon-Megatooth Shark by Lutz Andres

Roesch, Ben S. 1998. A Critical Evaluation of the Supposed Contemporary Existence
of Carcharodon megalodon. The Cryptozoology Review 3 (2): 14-24.

Evolution of scorpion venom

noreply@blogger.com (Natasha) — Tue, 29 Sep 2009 10:55:00 +0000

The predominant pharmacologically active components in scorpion venoms are small polypeptide molecules, usually basic in nature. Scorpion venoms and their component elements have been studied for over 35 years; but lately the focus in these studies has shifted from their pharmacological and electrophysiological properties to their molecular structures. This came about due to the realization that many classes of peptides seem to bind to characteristic spots on their targets, which for the most part are ion channels. The peptide toxins also show considerable identity in their arrangement of cysteine residues within the polypeptide chain. These cysteine residues are terribly important because disulphide bonds are one of the major contributors to conformational stability in small peptides.

The first pharmacological studies on scorpion toxins concentrated on their effects on mammalian model systems, so mice were the first ones to draw the short end of the hypodermic syringe, for a while. Peptides were consequently classified into the alpha- and beta-types, where each type bound to its own special site on voltage-gated sodium channels. Eventually, however, it was discovered that some peptides worked against ion channels in insects, and now Sarcophaga argyrostoma blowfly larvae were adopted as the victim model system of choice. All anti-insect toxins induced paralysis, but one class of toxins was found to induce a contraction paralysis, while the other induced a depressant paralysis.

For a while it seemed that all peptides could be neatly docketed in this way, but continuing studies unearthed peptide toxins that were primarily anti-mammal but showed some anti-insect activity too. Subsequently peptides were found that showed comparable anti-insect and anti-mammal activities, one of which – showing anti-insect as well as alpha-type and beta-type anti-mammal activities – was first reported in the paper (Loret et al., 1991) referenced below. This sparked off a line of thought on how the types of peptides found in scorpions in different parts of the world could be used to put forward theories about where scorpions first appeared and how they diverged into the large number of species we see today.

The reason this cross-reactivity of toxins against insect and mammal receptors was such a big deal was this: Before this was discovered, toxins were found to be easily classifiable not only on the basis of their primary peptide sequences and pharmacological activities, but also based on geography. Alpha-type anti-mammal toxins came only from Old World scorpions and beta-type anti-mammal toxins came only from New World scorpions. All anti-insect toxins had been purified from Old World scorpions only. Deviations from this geographical structure brought to light interesting ideas about venom evolution in scorpions.

For example, the toxin mentioned above which had a high effect on mammals but a low effect on insects was a beta-type toxin from Centruroides (a New World species) which was toxic to insects, but 50 times weaker than Old World anti-insect toxins (these studies happened in the 80s). This could indicate that anti-insect activity is just starting to evolve in New World venoms, but has already become established in Old World ones, indicating that the Old World venoms appeared first in their evolutionary history.

The paper referenced below is a report of a toxin from Androctonus australis Hector (pictured below), called AaH IT4. The toxin was purified by successive steps of chromatography, on gel filtration, DEAE-Sephadex and C8 HPLC columns. Toxicities were tested on S. argyrostoma larvae and male C57/BL6 mice, and the ED₅₀ values (for larvae) and LD₅₀ values (for mice) were recorded. Binding assays using ¹²⁵I-iodinated toxins on synaptosomal preparations from cockroaches and rats; radioimmunoassay assays to check for cross-reactions with rabbit antisera against known anti-insect, alpha-type and beta-type toxins; and circular dichroism analyses for structural data were carried out. In addition, sequence analysis and sequence alignment against various other scorpion venoms were carried out.

The experimental data showed that AaH IT4 competed with all three types of toxins for target-binding, and that it cross-reacted with the antibody against a beta-type toxin, indicating some structural similarity with the beta-toxin class. The dendrogram generated from the sequence alignment showed that AaH IT4 is more closely related to beta-type toxins than to either alpha-type or anti-insect toxins (which supports the result of the RIA experiment), although the divergence between AaH IT4 and the beta-type toxin lineage took place a long time ago. This an important point because beta-type toxins come from New World scorpions while AaH IT4 was purified from an Old World toxin, so any sequence similarity suggests that some relationship may exist between the two.

Another point to be made about AaH IT4 is that the sequence analysis shows an absence of the amino acid proline. Proline had previously been found in every purified and studied peptide scorpion toxin, and was suspected to play a role in the stability of their conformations, since proline is more conformationally restricted as compared to the other amino acids. One explanation for the binding of AaH IT4 to three different kinds of binding sites is that the absence of proline allows a certain amount of backbone flexibility which may allow the peptide to switch between conformational states that preferentially bind to each of the target sites.

The authors themselves hypothesize that since the evolution of insect-specific toxins is clearly advantageous to scorpions (for whom insects form a major part of the diet), it is possible that AaH IT4 represents the closest approximation available right now to some kind of ancestral toxin sequence/structure, which later diverged into anti-mammal and anti-insect varieties.

I think it’s worth mentioning, in addition, that the isolated AaH IT4 corresponds to 0.06% of the total protein in the venom, so it’s possible that the production of this particular venom component may have undergone some down-regulation over the years, as a result of the development of other, more specialised toxins.

Post-1991, of course, the issue of toxin classification has become even more complicated, and the venoms being studied now include those with anti-arthropod specificity, and the set of recognised targets have grown to include voltage- and ligand-gated potassium- as well as calcium-channels instead of just the initial emphasis on voltage-gated sodium-channels. Peptides with microbial activity have also been reported.

References:

Loret, E., Martin-Eauclaire, M., Mansuelle, P., Sampieri, F., Granier, C., & Rochat, H. (1991). An anti-insect toxin purified from the scorpion Androctonus australis Hector also acts on the .alpha.- and .beta.-sites of the mammalian sodium channel: sequence and circular dichroism study Biochemistry, 30 (3), 633-640 DOI: 10.1021/bi00217a007

Innate Immune Cells: Mediators of the Angiogenic Switch?

noreply@blogger.com (Varun) — Sat, 26 Sep 2009 13:01:00 +0000

The role of Matrix metalloprotease type 9 (MMP-9) in the activation of Vascular Endothelial Growth Factor (VEGF), the induction and maintenance of chronic angiogenesis and early stage tumor growth has been well established. But what is the source of MMP-9 ? In a study published in 2006, Nozawa and co-workers, using the RIP1-Tag2 transgenic mouse as a model, identified two inflammatory cell types, neutrophils and macrophages, as the major sources of MMP-9.

The RIP1-Tag2 transgenic mouse is a well characterized model of multistep carcinogenesis involving the pancreatic islets. Nozawa and co-workers first isolated constituent cell types from a tumor and cell-sorted Gr-1⁺ and Mac-1⁺ cells and then carried out a semi-quantitative RT-PCR (see Fig.1)

Fig. 1: Semi-quantitative RT-PCR. MMP-9⁺ is expressed by Gr-1⁺ and Mac-1⁺cells innate immune cells (Adapted from Nozawa et. al, 2006)

Clearly, MMP-9 was expressed predominantly by innate immune cells positive for the Gr-1 and Mac-1 markers.

In order to determine the localization of these cells , a double label immunohistochemical staining of various leukocyte markers in normal and neoplastic tissue was performed. CD68 and F4/80 are two macrophage markers. Cells positive for both markers were observed inside as well as on the periphery of angiogenic islets and tumors (see Fig. 2). However, the cells inside were MMP-9^- while those along the periphery were MMP-9⁺

Fig. 2: MMP-9⁺ macrophages located along the periphery of the angiogenic islet.Macrophages inside the islet were MMP-9^- (Adapted from Nozawa et. al, 2006)

Along similar lines, an antibody against the "7/4" (a neutrophil marker) was used to localize neutrophils. Suprisingly, the situation, this time, was the opposite. 7/4⁺ cells inside the lesions were also MMP-9⁺ while 7/4⁺ cells along the periphery were not (see Fig. 3). All 7/4⁺ cells also displayed polymorphic nuclei, strongly indicating that these cells were neutrophils.

Fig. 3: MMP-9⁺ neutrophils located inside the angiogenic islet. Neutrophils along the periphery of the islet were MMP-9^- (Adapted from Nozawa et. al, 2006)

Neutrophils (7/4⁺ and Gr-1⁺) represented only 0.4% of the total tumor cells and expressed high levels of MMP-9. In contrast, macrophages (CD68⁺) represented 2% of the total tumor cells and expressed low levels of MMP-9.

The researchers then tried to determine the role of these MMP-9⁺ Gr-1⁺ cells that infiltrate angiogenic islets and tumors. An experimental regimen involving daily inoculation of anti-Gr-1 at 7 weeks (when hyperplastic islets start to undergo angiogenesis) was followed.

After 1 week of injection, most neutrophils disappeared from the pancreatic islets of the transgenic mice (see Fig.4)

Fig. 4: Left: Control; Right: Following 1 week of treatment with anti-Gr-1, MMP-9⁺ neutrophils located inside the angiogenic islet disappeared (Adapted from Nozawa et. al, 2006)

After two weeks of injection, the neutrophil population rebounded in neoplastic islets. Incredibly, none of the rebound neutrophils were MMP-9⁺ (see Fig. 5)!

Fig. 5: Left: Control; Right: Following 2 weeks of treatment with anti-Gr-1, neutrophils rebounded back into the islets. However, none of the rebound neutrophils were MMP-9⁺ (Adapted from Nozawa et. al, 2006)

In a short 2 week prevention trial (week7-week9), carried out to assess angiogenic switching frequency, the anti-Gr-1 antibody regimen, reduced the number of angiogenic islets by 57%!

Furthermore, immunostaining for VEGF:VEGF-R2 complex with GVM39 (red) and for endothelial cells with Meca-32 (green) showed a decrease in bioactive VEGF:VEGF-R2 interaction, consequent to the depletion of infiltrating MMP-9⁺ neutrophils (see Fig.6)

Fig. 6: Left: Control; Right: Short intervention trials lead to reduced VEGF:VEGF-R2 interactions. Endothelial cells are stained with an antibody directed against Meca-32 (a pan-endothelial cell marker) (Adapted from Nozawa et. al, 2006)

Taken together, the results from this paper indicate that subtle infiltration by innate immune cells (such as neutrophils) may play a role in the progression of neoplasias towards angiogenic tumors.

Reference:

Nozawa H, Chiu C, & Hanahan D (2006). Infiltrating neutrophils mediate the initial angiogenic switch in a mouse model of multistage carcinogenesis. Proceedings of the National Academy of Sciences of the United States of America, 103 (33), 12493-8 PMID: 16891410

Alan Turing-May He Finally Rest in Peace

noreply@blogger.com (Varun) — Sun, 13 Sep 2009 20:02:00 +0000

Gordon Brown's official statement on Alan Turing

2009 has been a year of deep reflection - a chance for Britain, as a nation, to commemorate the profound debts we owe to those who came before. A unique combination of anniversaries and events have stirred in us that sense of pride and gratitude which characterise the British experience. Earlier this year I stood with Presidents Sarkozy and Obama to honour the service and the sacrifice of the heroes who stormed the beaches of Normandy 65 years ago. And just last week, we marked the 70 years which have passed since the British government declared its willingness to take up arms against Fascism and declared the outbreak of World War Two. So I am both pleased and proud that, thanks to a coalition of computer scientists, historians and LGBT activists, we have this year a chance to mark and celebrate another contribution to Britain’s fight against the darkness of dictatorship; that of code-breaker Alan Turing.

Turing was a quite brilliant mathematician, most famous for his work on breaking the German Enigma codes. It is no exaggeration to say that, without his outstanding contribution, the history of World War Two could well have been very different. He truly was one of those individuals we can point to whose unique contribution helped to turn the tide of war. The debt of gratitude he is owed makes it all the more horrifying, therefore, that he was treated so inhumanely. In 1952, he was convicted of ‘gross indecency’ - in effect, tried for being gay. His sentence - and he was faced with the miserable choice of this or prison - was chemical castration by a series of injections of female hormones. He took his own life just two years later.

Thousands of people have come together to demand justice for Alan Turing and recognition of the appalling way he was treated. While Turing was dealt with under the law of the time and we can’t put the clock back, his treatment was of course utterly unfair and I am pleased to have the chance to say how deeply sorry I and we all are for what happened to him. Alan and the many thousands of other gay men who were convicted as he was convicted under homophobic laws were treated terribly. Over the years millions more lived in fear of conviction.

I am proud that those days are gone and that in the last 12 years this government has done so much to make life fairer and more equal for our LGBT community. This recognition of Alan’s status as one of Britain’s most famous victims of homophobia is another step towards equality and long overdue.

But even more than that, Alan deserves recognition for his contribution to humankind. For those of us born after 1945, into a Europe which is united, democratic and at peace, it is hard to imagine that our continent was once the theatre of mankind’s darkest hour. It is difficult to believe that in living memory, people could become so consumed by hate - by anti-Semitism, by homophobia, by xenophobia and other murderous prejudices - that the gas chambers and crematoria became a piece of the European landscape as surely as the galleries and universities and concert halls which had marked out the European civilisation for hundreds of years. It is thanks to men and women who were totally committed to fighting fascism, people like Alan Turing, that the horrors of the Holocaust and of total war are part of Europe’s history and not Europe’s present.

So on behalf of the British government, and all those who live freely thanks to Alan’s work I am very proud to say: we’re sorry, you deserved so much better.

Gordon Brown

Aging by Epigenetics

noreply@blogger.com (Manasi) — Sun, 13 Sep 2009 11:18:00 +0000

We know that organismal longevity and aging is caused by a lot of interacting factors such as nuclear and mitochondrial genome mutations, shortened telomeres, oxidative damage to DNA and other macromolecules, senescence, apoptosis and many more. This review discusses another possible determinant of aging that is 'epigenetics'. We have seen in the earlier post that epigenetics refers to changes in the DNA and histones which are heritable through cell divisions, but do not involve any change in the sequence of the DNA.

Chromatin is broadly divided into two types, euchromatin and heterochromatin. Euchromatin is decondensed during interphase and is relatively transcriptionally active. Heterochromatin on the other hand remains compact and condensed and is transcriptionally inactive. It is further divided into constitutive and facultative. Constitutive heterochromatin is present in the telomer and centromere and appears to be fixed or irreversible throughout the life time of an organism. Facultative heterochromatin on the other hand, is made as a part of regulated cell differentiation process or other changes in cell phenotype. For example, a single X chromosome is silenced in female mammalian cells for dosage compensation.

However, despite this apparent clear distinction between euchromatin and heterochromatin, it is now being understood that chromatin structure is highly dynamic and stochastic. Essential processes, such as DNA replication, transcription and repair all involve disruption of the very compact structure of DNA. The proteins bound to the chromatin are also not static but exhibit relatively high 'on' and 'off' binding states even in the 'fixed' heterochromatin. It has also been showed that, formation of heterochromatin depends on a degree of transcription, which contributes to heterochromatinization through the RNAi pathway. Thus the telomeric heterochromatin is also shown to be transcribed. Thus heterochromatin is not a static entity.

Is aging associated with epigenetic changes?

The enzymes Histone acetyl teransferases (HATs) and Histone deacetylases (HDACs) respectively determine the steady-state level of histone acetylation. In S.cerevisiae, inactivation of a HDAC, Sir2, decreases replicative lifespan while activation extends it. The anti aging effects of Sir2 in yeast are due to translocation of a Sir2 containing protein complex from telomeres to ribosomal DNA (rDNA) repeats. These repeats are prone to recombination to form extrachromosomal rDNa circles (ERCs), which curtail yeast lifespan. However, Sir2 mediated histone deactylation and heterochromatization, prevents formation of ERCs and thus extends lifespan of yeast. Thus, this epigenetic redistribution counteracts organismal aging. Orthologs of Sir2 have been found in many species like nematodes, flies and even mice. Thus its anti aging role seems to have been conserved throughout evolution.

In mammals, it has been seen that there is a reduction in genomic DNA methylation, with age. It occurs mostly at repetitive DNA sequences, predominantly in regions of constitutive heterochromatin. Since methylation induces silencing of genes, this change will promote deheterochromatinization of these regions. However DNA methylation increases at specific sites called the CpG islands. These are CG rich sequences, some of them present in the promoter regions, which can get methylated. Methylation also increases in the histone 4 on the lysine 20 residue (H4K20) in rat liver and kidney, with age. Like DNA methylation H4K20 methylation is also linked to gene repression, supporting the notion that heterochromatin accumulates in some sites atleast with aging.

One of the histone chaperones, HIRA, shows increased levels of expression in aging baboon skin. This is shown to have a evolutionary conserved role in formation of heterochromatin.

These observations suggest that mammalian aging is associated with chromatin remodelling. In particular, there is a global decrease in DNA methylation, but an increase at specific sites on the genome.

Cellular senescence, is characterized by irreversible proliferation arrest. This may arise due to excessive cell divisions and shortening of the telomere length. Due to this, most human cells have a finite proliferative lifespan. Senescent cells or molecular markers of senescent phenotype increase during aging. Cellular senescence is also well established tumor suppression process, because it can prevent proliferation and neoplastic progression of cells harboring neoplastic lesions. Senescent cells also show chromatin remodeling. Many senescent cells form domains of facultative heterochromatin, called Senescence Associated Heterochromatin Foci (SAHF), which are visibly more condensed that interphase chromatin. These foci have been proposed to silence proliferation promoting genes. Accumulation of SAHF has been associated with aging. Formation of SAHF requires presence of HIRA, which has been shown to be upregulated in aging baboon skin. Also it seems that in cellular senescence too, the constitutive heterochromatin regions are deheterochromatinized. Thus like tissue aging, cellular senescence also is accompanied by redistribution of heterochromatin, from constitutive heterochromatin to other normally euchromatic sites.

Consequences of age-associated epigenetic changes

Studies going back to 1960s indicate that aging is associated with cell aneuploidy. Proper chromosome segregation is dependent on pericentromeric heterochromatin sturcture. Hence cecreased methylation and deheterochromatinization may lead to faulty chromosome segregation and associated aneuploidy. Aneuploidy confers various altered cell phenotypes including a proliferative impairment, which might contribute to decreased tissue renewal capacity with age. It may also give rise to cancer, for which age is the biggest risk factor. Some of the age associated changes in gene expression may also occur due to epigenetics. Methylation of CpG islands is a well described mode of silencing some tumor suppressor genes. A tissue wide age associated methylation of CpG may be an early causative event in the development of neoplasms.

From the discussion it is apparent that chromatin does undergo change with aging in organisms as diverse as yeast and mammals. However with the exception of Sir2 in yeast, the extent to which this impacts the aging process is not yet defined. Some age associated epigenetic alterations in mammals like formation of SAHF, might extend life span by suppressing age associated diseases like cancer. In sum the effects of chromatin on aging are likely to be complex. A pre requisite to properly understanding the contribution of epigenetics to aging is to better understand the specific cell, tissue and system wide malfunctions that are responsible for aging phenotypes like osteoporosis, sarcopenia, cancer and many others. Then it will be feasible to dissect out the contribution to each phenotype of each candidate epigenetic determinant, like global methylation, CpG island methylation and SAHF.

Reference

Sedivy JM, Banumathy G, & Adams PD (2008). Aging by epigenetics--a consequence of chromatin damage? Experimental cell research, 314 (9), 1909-17 PMID: 18423606

Fake Hands and Tables-The Malleability of the Body Image

noreply@blogger.com (Varun) — Tue, 08 Sep 2009 07:47:00 +0000

In " The Classical Rubber Hand Illusion", we discussed the original experiments of Botvinick and Cohen (1998). Their hypothesis for the rubber hand illusion was that vision has higher reliability and spatial acquity than proprioception, so the brain gives more weight to visual information. People would thus localize a body part to it's apparent visual location, particularly when the visible location falls within the possible range dictated by proprioception. Some support for this theory lies in the fact that placing the fake hand perpendicular to the real occluded hand destroys the illusion that the fake hand is one's own.

Armel and Ramachandran (2003) reported a closely related but bizarre illusion. The subject is made to place his or her real hand on a table and the hand is hidden from view. However, instead of stroking a fake rubber hand, the researchers simply stroked and tapped the table in precise synchrony for a minute. Astonishingly, subjects reported sensations arising from the table surface, despite the fact that it bears no physical resemblance to a hand. Whereas Botvinick and Cohen interpret their result in terms of resolving incongruities between visual versus proprioceptive location of the hand, Armel and Ramachandran's experiment would lead one to argue that the illusion arises mainly from Bayesian logic of all perception; the brain's remarkable ability to detect statistical correlations in sensory inputs in constructing useful perceptual representations of the world, including one's own body. It is especially intriguing that the bizarre perceptual representation (assimilating the table into the body image) is resistant to the "top-down" knowledge of the absurdity of the situation!

To measure the extent to which subjects incorporated the external objects into their body image, they were asked to rate the vividness of the illusion. The experimenters also recorded the skin conductance response (SCR), to provide an objective test of whether the table had indeed become informationally coupled with the subject's body image. If the external objects became integrated into the body's image, would they be aroused when the table (or a fake hand for that matter) was 'injured'?

If a finger of the fake hand is bent backwards to seem painful, does the subject register an SCR? To what extent is the fake hand assimilated into the subject's body image? To address this, after ca. 2.5 minutes of the touching procedure, both the real and the fake fingers were lifted, but only the fake finger was bent backwards into a 'painful position'. SCR was recorded at this point and a free response description and intensity rating of the illusion were obtained. The control for this experiment was a 'delayed synchrony' condition wherein touch to the real and fake hands were identical, the only difference being, that the touch to the real hand was delivered 1 sec after the touch on the fake hand. Mean intensity and mean SCRs showed that subjects identified with the fake hand more in the condition where the touch was synchronized rather than the one where the touch was synchronized but delayed (see Fig. 1)

Fig. 1: Mean Intensity ratings (gray bars) and SCR (black circles) in the first experiment where synchronous touch to the real hand was delayed by a second in the control condition. The error bars indicate one SEM (Adapted from Armel and Ramachandran, 2003)

Would subjects still experience the illusion if the form of the external object was manipulated? To explore this, a barren table was stroked and tapped in the same manner and in the same relative location (see Fig.2 ).

Fig. 2: Form manipulation where subjects received the table condition (Adapted from Armel and Ramachandran, 2003)

Band-aids were placed on both the real hand and the table and subjects were told that the band-aid would be pulled off the table but not off their real hands. At the end of the 2.5 min touching period, in lieu of pulling back a fake finger, the band-aid on the table was partially pulled off. In the control condition for this experiment, the real hand was made visible by removing the occluder. Subjects were instructed to look back and forth between their real hands and the table. Furthermore, the real hand and the location where the table was touched were close together so that they could be seen simultaneously even while looking at one or the other. The band-aid was pulled off the table while the subject viewed it. To ensure that the subject was only looking at the table, the experimenter occluded the subject's real finger at this time. In a comparison of the conditions in which the table and real hand were touched with the partition in place or removed, intensity ratings and SCR were significantly different (see Fig. 3). However, the same experiment (using a band-aid) carried out with a fake rubber hand in lieu of the table is more effective at inducing the illusion in terms of intensity ratings but only marginally so for SCR (see Fig. 3)

Fig. 3: Mean Intensity ratings (gray bars) and SCR (black circles) in the second experiment where form of the external object was manipulated. The error bars indicate one SEM (Adapted from Armel and Ramachandran, 2003)

Would subjects still experience the illusion if the form of the external object was manipulated? Each subject viewed touch to a fake hand in a 'realistic' location in one condition and then to a distant fake hand in another (see Fig. 4).

Fig. 4: Location manipulation where subjects received the distant-hand condition (Adapted from Armel and Ramachandran, 2003)

The fake arm was extended so that it lay 3 feet beyond the real hand. In the distant fake hand manipulation, a fake finger was bent back for a painful stimulus. In the control condition, the touch applied to the fake and real hands was asynchronous i.e. touch was random and there was no correlation (see Table 1 at the end for summary of conditions for all experiments). Mean intensity and mean SCR showed that subjects identified with the fake hand and the distant fake hand, more in the conditions where touch was synchronized than when not synchronized. However, the 'anatomically correct' fake hand condition was more effective than the distant fake hand condition (both with synchronous touch, see Fig. 5)

Fig. 5: Mean Intensity ratings (gray bars) and SCR (black circles) in the second experiment where the location of the fake hand was manipulated. The error bars indicate one SEM (Adapted from Armel and Ramachandran, 2003)

The so-called body image appears to be highly malleable. Despite it's appearance of durability, it can be profoundly altered by stimulus contingencies and correlations that one encounters. Taken together, these experiments illustrate an important principle underlying perception: that the mechanisms of perception may be involved in extracting statistical correlations. Further investigations will further our understanding of phenomena such as body-dysmorphic disorder and anorexia nervosa

Table 1: Experimental design for all experiments. For a given experiment, each subject received all conditions in one of different possible orders (Adapted from Armel and Ramachandran, 2003)

Reference:

Armel, K., & Ramachandran, V. (2003). Projecting sensations to external objects: evidence from skin conductance response Proceedings of the Royal Society B: Biological Sciences, 270 (1523), 1499-1506 DOI: 10.1098/rspb.2003.2364

Epigenetics - Implications for behavioral neuroendocrinology

noreply@blogger.com (Manasi) — Sat, 05 Sep 2009 17:07:00 +0000

Individuals vary in their sociosexual behavior and reactivity. How an organism interacts with the environment to produce these variations has been a focus in psychology since its inception as a scientific discipline.There is now no question that cumulative experiences throughout life history interact with genetic predispositions to shape the individual's behavior. Recent evidence suggests that events in the past generations may also influence how an individual responds to events in their own life history. Epigenetics is the study of how the environment can affect the genome of an individual and its descendants all without changing the sequence of the DNA.

The early stages of life, beginning before birth and up to weaning in mammals, are the time of maximum neuronal plasticity.It is during this early period that hormones and genotype predispose an individual's responses to future experiences throughout the life cycle. Suites of genes underlie the fundamental plasticity of an organism, particularly during development and life history transitions. How do these gene networks interact with the experiences that cumulate during an individual's life history?

An important interface between the environment (both internal and external) is that of epigenetic modifications. Exactly how these modifications occur is still relatively unknown, but recent studies indicate that origin of such effects may occur at in the previous generations. That is, experience of previous generations may modify regulatory factors affecting gene expression without changing the sequence of the DNA, but the physiology and behavior of the organism may be substantially influenced. Thus understanding how such events really occur will enhance our understanding of how the environment influences the relationship between genotype and behavior during sensitive periods of development.

Molecular vs Molar epigenetics

There have been several reviews recently as to the origin of the field of epigenetics, all of which recognize the multiple roots of the current tree of research. The debates in the 16th-17th century pitted preformationism against epigenesis, with a central question of how a multicellular organism develops from a single cell, the zygote. The former camp believed that adult characters were present fully formed in the egg and simply unfolded during growth, while the latter held that traits arose due to interactions between the multiple constituent parts of the zygote. Spawned after the resolution of this conflict were two different groups, one rooted in anatomy and geology, which later became the broad science of biology, and the other focused on sensation, perception and mind, which ultimately became the study of psychology. Thus though both have a common origin, they evolved very differently both in perspective and substance. These the author labels respectively as, 'Molecular epigenetics' and 'Molar epigenetics'.

Molar epigenetics arises from historical literature in psychilogy, particularly functionalism. Rather than measuring sensory processes, the functionalists focused on the organism itself and development of behavior in relation to its natural environment. Molecular epigenetics arose from molecular biology and modern genetics and its emphasis was on gene regulation and its developmental significance.Thus the object of study in Molecular epigenetics is gene expression during embryogenesis, while in Molar epigenetics, it is on the individual's interactions with the biotic and physical environment usually after birth.

Molecular epigenetics

Conrad H. Waddington proposed the term 'epigenetics' from classical embryology, ascribing it to the study of processes by which genotype gives rise to a phenotype. His concept was central to what became the modern era of epigenetics when Robin Holliday proposed a molecular model of heritable gene activation and inactivation during development by DNA methylation and demethylation respectively. Since then the term epigenetics is used to connote the study of change in gene expression without changing the DNA sequence. There are several mechanisms that can achieve this end, such as DNA methylation and modification of histones by processes of methylation, deacetylation and phosphorylation. While DNA methylation is clearly involved in genomic imprinting, the signal for the imprint is still not known.

Molar epigenetics

Early comparitive psychologists were also interested in epigenetics, but from the perspective of interaction of an organism's heredity and the nature of species typical behaviors or 'instincts'. Work principally by Konrad Lorenz and Niko Tinbergen emphasized that such behaviors were a product of natural selection, the result of genes acting in the brain to generate behaviors that were unlearned and innate. Thus, these studies laid the foundation of psychobiology, a field that focuses on how experiences cumulate throughout life to shape the way in which an individual interacts with its environment.

It is necessary to emphasize that, an approach that integrates both Molecular and Molar epigenetics, will be required to reveal the mechanisms that underlie behavioral evolution.

Context dependent vs Germline dependent epigenetic modification

In context dependent epigenetic modification, the change is transmitted within a generation, within an individual's own lifetime, including the interaction of parent and young. An example of this is exposure to endocrine disrupting chemical in utero during childhood, in which case the disease manifests itself later. The extent to which the modification is perpetuated is by the simple presence of the environmental factor, that brings about the epigenetic modification. The modification is present until the effect of the factor is present. Later it declines and to reestablish the effect the individual has to be exposed to the same agent again. This effect is called context dependent epigenetic modification. In germline dependent modification, the genetic imprint is independent of the original causative agent. Here the epigenetic modification is transferred to the subsequent generations because, the change in the epigenome has been incorporated in the germline. Thus the effect is manifest in each generation without the need for re-exposure. The DNA methylation of heritable epialleles are passed through to subsequent generations without being erased as occurs normally during gametogenesis or shortly after fertilization.

If the context in which an individual is nurtured affects its behavior as an adult, it is likely that the activities of the neural circuitry underlying this behavior is also affected. This should apply to both context dependent and germ line dependent epigenetic modifications.

Transgenerational epigenetic imprint on the nuclear genome and its effect on behavior and brain

Two critical elements of demonstrating a germline dependent epigenetic modification are that, first a single exposure of the environmental factor that is never again repeated, and second the number of generations since that exposure. A new model system was used in which an endocrine disrupting chemical (EDC) reprograms methylation patterns that are then incorporated into the germline and hence transmitted to future generations. In this model system the exposure of gestating female rats to pesticide methoxychlor or fungicide vinclozolin during the period of embryonic sex determination, induces an epigenetic transgenerational phenotype through reprogramming the germline in a sex specific manner. Specifically, in each generation males whose ancestor have been treated underwent progressive spermatogonial apoptosis, decreased sperm count and motility and as the animals aged, adult onset disease is accelerated, including cancer and immune cell defects.

Studies were carried out to determine if this altered epigenome also influences male/partner preference behavior. For these studies F3 descendants of females were used which were treated with dimethylsulphoxide buffer alone (control) or with vinclozolin (EDC treated). Partner preference was carried out by placing an individual (male or female) in a testing arena. At either ends was a small cage containing the stimulus rats separated by a wire mesh barrier to allow exchange of visual, olfactory and tactile cues. All males were tested with both types of females as stimulus and vice versa. Behaviors directed to the stimulus animals included time spent in contact with the wire mesh during which the animals often touched noses through the mesh, grooming, aimless walking and sniffing, standing on hind paws and sniffing with nose pointed upwards, contacting the walls of the test cage and time spent in the center of the test arena.

The results were clear cut and sex specific. The females discriminate and prefer males who do not have a history of exposure, while males do not exhibit such a preference. In social engagements in rodents time is spent in mutual facial investigation. Males investigate females equally, while females spend more time investigating males from the control lineage. It is known that pheromones from the vomeronasal organ and urine are involved in mate recognition in rodents. Methylation analysis revealed that Major Urinary Protein 4 (MUP4) is one of the candidate imprinted like genes in the vinclozolin treated lineage. This MUP group of gene products binds to and releases male specific pheromones in rodents. Also males and females both explored odors of the opposite sex than familiar odors. These behaviors may reflect differences in pattern of gene expression in different brain areas.Using gene micro arrays specific ares of the brain like the hippocampus , amygdala and the whole brain have been studied in both control and test males. Of the altered genes only a limited number shows similar changes in all the three regions an some of these have been implicated in schizophrenia, autism and depression.

Cytoplasmic genes like the mitochondrial genes also have CpG islands and polymorphisms in these genes plays a significant role in adaptive evolution. These genes are important for determining sperm quality and motility and mutations result in decrease in human sperm motility. In this regard males of all five generations stemming from females treated during pregnancy with either methoxychlor or vinclozolin have decreased sperm motilty and numbers. The mitochondrial genome is also vitally involved in aging and mutations are involved in onset of age related phenotypes, including reduced fertility. Genes encoding for cytochrome oxidase were tested in transgenerationally imprinted rats. (CO is used as a measure of brain activity). None of the constituent genes of this mito-nuclear gene product, seem to be altered in the three regions of the brain. However, nuclear respiratory factor 2 which modulates CO activity, shows an increased expression in the whole brain, perhaps accounting for the behavioral differences observed between the mice of the two lineages. Thus the relative fitness of the specific mito-nuclear genotype combinations is dependent on the modified DNA environment in which they persist. Thus EDCs could act via epigenetically modifying mitochondrial DNA as well as nuclear DNA, and influence epistatic interactions between cytoplasmic and nuclear genes.

The linking of Molar with Molecular epigenetics, extensive knowledge of hormones which play a role in organizing and activating brain-behavior mechanisms and the predisposition in neuroscience research to use molecular methods to understand cellular function and development should facilitate the incorporation of epigenetics in neuroendocrinological research.

Reference:

CREWS, D. (2008). Epigenetics and its implications for behavioral neuroendocrinology Frontiers in Neuroendocrinology, 29 (3), 344-357 DOI: 10.1016/j.yfrne.2008.01.003

Polycomb group proteins in Drosophila

noreply@blogger.com (Manasi) — Wed, 26 Aug 2009 15:07:00 +0000

In the earlier post titled ' Role of polycomb and trithorax in developmental regulation' we have seen that the main function of the polycomb group proteins (PcGs) is to repress target gene expression, their major targets being the homeobox (Hox) genes. These proteins bind to Polycomb Response Elements (PREs), their target sequences on the DNA. The recruitment of the PcGs to these sites occurs due to covalent histone modifications in the DNA. Here we look into this aspect in some detail.

Till date three distinct PcG complexes have been discovered in Drosophila Polycomb repressor complex 1 and 2 (PRC1 and 2) and Pho repressive complex (Pho RC). All three contain multiple subunits encoded by PcG genes that are crucial for Hox gene silencing. PRC2 is known to have histone methyl transferase activity. It has H3K27 methylase activity. The PRC1 subunit inhibits nucleosome remodelling and transcription and brings about chromatin compaction. The PRC1 and 2 have various catalytic and non catalytic subunits which play a role in silencing target sites. The Pho repressive complex contains Pho and dSfmbt. Pho subunit has a distinct sequence specific DNA binding activity which is essential for targeting. dSfmbt binds to H3 or H4 tail peptides which are mono or di methylated at H3K9 or H4K20 which is crucial for repression.

The above figure depicts a speculative model for long range interactions between the PRE tethered PcG complexes and the methylated nucleosomes in the flanking chromatin. PRC1, PRC2 and PhoRC are locally bound to the PREs whereas trimethylation at H3K27, H3K9 and H4K20 is observed over an extended domain emcompassing the promoter and the coding regions.

Left : In case of PRC1 the Pc chromadomain (red triangle) would dock onto the nucleosomes that are tri methylated at H3K27 and through such bridging interactions will bring them in proximity to the other PRE bound PcG proteins. This may permit the other PRC1 subunits to block remodelling of the target nucleosomes or facilitate methylation of the neighbouring hypomethylated (pink) nucleosomes by PRC2.

Right : In case of PhoRC, the MBT (Malignant Brain Repeats) of dSfmbt (pink triangle) would interact with mono or dimethylated nucleosomes at H3K9 or H4K20.This bridging interaction would permit PRC2 to efficiently tri methylate H3K27 in hypomethylated nucleosomes.

Its is also possible that as yet unindentified HMTases which tri methylate these histones are also localized at the PREs.Similarly proteins with specificity for binding to other mrthyl-lysine modifications in histones might also be localized to the PREs.Various proteins have been purified which are associated to PcG complexes that are strictly essential for silencing. Pcl (Polycomb like) has been shown to be associated with PRC2 and mutations in this protein affect PRC2 functioning. Another protein zeste is a component of PRC1 and it has DNA binding activity.

As described above biochemical purification of PRC1, PRC2 and PhoRC suggests that these three complexes are separate entities. However since they co localize at the PREs various studies are aimed at finding out the physical interactions between subunits from the different complexes which may explain how PRC1 and PRC2 are localized to the PREs. Different models have been put forward based on various studies. In particular Wang et al (2004), reported that E(z) and Esc (subunits of PRC2) directly interact with Pho, leading to the proposal that Pho directly tethers PRC2 to the PREs. the PRE tethered PRC2 then locally trimethylates H3K27 thereby creating binding sites for the chromodomain of the PRC1 subinut Pc. More recent studies however challenge this model. First quantitative ChIp studies by two different labs suggest that PREs at the Ubx gene are infact devoid of nucleosomes.(V Pirrotta). Moreover PREs constitue of hypersensitive sites providing additional evidence that PRE DNA is not packaged into nucleosomes. Second studies by Mohd Sarip et al (2002), suggested that Pho can directly interact with PRC1 subunits, and that in in vitro studies Pho and PRC1 can co assemble on a naked PRE DNA template in absence of nucleosomes. This assembly assumes a conformation that is difficult to reconcile with the conformation of a nucleosome core particle. Thus available evidence suggests that not only PhoRC, but also PRC1 and PRC2 localize onto the PRE through interactions with Pho and other DNA binding proteins and not through covalent histone modifications or interactions with nucleosomes.

This conclusion then raises a major question with regards to the role of histone modification (trimethylation) in PcG silencing. Quantitative ChIp analysis that compared the Ubx gene in its off and on states in Drosophila larvae suggest that trimethylation in the promoter and coding region is essential for PcG silencing. In the 'off' state extensive trimethylation is present throughout the upstream control, promoter and coding regions but in the 'on' state this methylation is restricted only to the upstream control region and not seen in the promoter and coding regions.This led to the proposal that trimethylation at H3K9, H3K27 and H4K20 in the promoter and coding regions is required to demarcate the chromatin interval that is targeted for repression by the PRE-tethered PcG protein complexes. In this context, an important aspect about the relationship between the PREs and histone modifications of the target genes comes from studies with PRE reporter genes in Drososphila, in which the PRE DNA was flanked with FRT ( Flip Recombinase Target) sites and could thus be deleted from the reporter gene. This study indicated that excision of the PRE DNA from the silenced reporter gene, caused loss of PcG silencing, even when the excision was done late in development. Thus although the PcG proteins appear to repress transcription by methylation in promoter and coding regions, silencing depends on the continuous presence of the PREs and the PcGs tethered to them.

Although much progress has been made towards understanding the mechanism of PcG silencing, much remains to be learned. It is still not understood how the PcG proteins are targeted to the PREs beacuse Pho binding sites alone do not make a PRE. ChIp studies may also help elucidate additional DNA binding PcG proteins.Also there may be many additional PcG complexes and PRC1, PRC2 and PhoRC may represent only the most stable or most abundant PcG complexes. Also it seems likely that these findings of PcG silencing in Drosophila will prove important in inderstanding the mechanism of PcG silencing in mammals, given the recent discovery that mammalian PcG proteins seem to be acting as global repressors of developmental control genes in embryonic stem cells.

Reference:

MULLER, J., & KASSIS, J. (2006). Polycomb response elements and targeting of Polycomb group proteins in Drosophila Current Opinion in Genetics & Development, 16 (5), 476-484 DOI: 10.1016/j.gde.2006.08.005

Evidence for Systemic Spread prior to the Establishment of Primary Tumors

noreply@blogger.com (Varun) — Sat, 22 Aug 2009 11:36:00 +0000

It is widely believed that metastasis is a late event in cancer progression. This view is based on several clinical and experimental observations. First, most cancer patients die from metastasis and not the primary disease. Second, early surgery is often the only cure. Third, somatic genetic changes accumulate during local progression (Fearon and Vogelstein, 1990) which was extrapolated to systemic progression. Fourth, repeated rounds of in vivo selection led to cell lines with increased metastasis formation (Kang et. al 2003; Minn et. al 2005). However, in a recent study titled Systemic Spread is an Early Step in Breast Cancer, published in Cell in 2008, Hüsemann, Geigl, Schubert et. al report that tumor cells can disseminate systemically from the earliest epithelial alterations in HER-2 and PyMT transgenic mice and from ductal carcinoma in situ in women.

The mouse model they work with is the BALB/c mice transgenic for the activated rat HER-2/neu gene (BALB-Neu T mice). This model mimics progression and gene expression profiles of human breast cancer. Females hemizygous for the rat HER-2 gene under control of the MMTV (mouse mammary tumor virus) promoter develop invasive mammary cancer while their HER-2 negative siblings (wild-type BALB/c mice) remain tumor free. Typically, in the BALB-Neu T model, the mammary epithelia starts to express the oncogene at about weeks 3 to 4 (which coincides with the onset of puberty). Hyperplasia can be detected microscopically at weeks 7-9. Progress to in situ carcinomas occurs between weeks 14-18 and at weeks 23-30, invasive cancers become apparent (see Fig. 1a)

Fig. 1a; Left Panel: Whole mount of the mammary gland at week 9 showing absence of tumor in branching ductal tree; Middle Panel: Histological Section at week 9 showing side buds displaying the morphology of atypical ductal hyperplasia; Right Panel: Histology of invasive cancer at week 30 (Adapted from Hüsemann, Geigl, Schubert et. al (2008))

The principle goal was to determine when cells expressing the HER-2 transgene disseminate. The authors chose the lung and bone marrow as the sites to look out for metastasis since the HER-2 receptor is not expressed in either of these two organs. In addition to HER, they also used anti-cytokeratin (CK) antibodies to check whether disseminated cells were epithelial in origin. Surprisingly in BALB-Neu T mice, CK⁺ and HER-2⁺ cells became detectable 4-9 weeks when meticulous analysis could only detect atypical ductal hyperplasia (ADH) (see Fig. 1b)

Fig. 1b; Left Panel: Increase in tumor area at primary site. Triangles indicate mean value, whiskers indicate 95% confidence interval and solid line indicates best fitted curve; Right Panel: Number of CK⁺ cells (red dots) and HER⁺ cells (blue dots) per 5000 bone marrow cells. Triangles, whiskers and solid line indicate same. Note the presence of disseminated cells in the bone marrow even when the primary tumor area is zero.(Adapted from Hüsemann, Geigl, Schubert et. al (2008))

Further, lung micrometastasis from mammary tissue was confirmed by demonstration of mammary-specific alpha casein and lactalbumin transcripts (see Fig. 1c)

Fig. 1c; Left Panel: Lung micrometastasis at week 27, detected using anti-HER-2 antibody;Right Panel: Eight HER-2 ⁺ and two samples of normal lung tissues, analyzed for mammary gland specific transcripts. "+" indicates normal mammary gland and "-" indicates mock control (Adapted from Hüsemann, Geigl, Schubert et. al (2008))

Despite exponential growth at the primary tumor site, the number of CK⁺ cells and HER-2⁺cells in the bone marrow rose marginally over the course of time. Cells singly positive for HER-2 and CK were not congruent. Not all tumor cells expressed both markers (see Fig. 1d)

Fig. 1d; Left Panel: CK⁺, HER-2^- cell; Middle Panel: CK^-, HER-2⁺ cell; Right Panel: CK⁺, HER-2⁺ cell (Adapted from Hüsemann, Geigl, Schubert et. al (2008))

This suggests, either the existence of heterogeneous tumor cell populations that disseminate to distant sites or different cellular states of the disseminated tumor cells (DTCs)

More Evidence for Early Dissemination

Could the HER-2⁺, CK⁺ cells have disseminated from extra-mammary tissues expressing the transgene? The authors took mammary gland fragments from 3-12 week old transgenic mice (displaying only atypical ductal hyperplasia) and transplanted the them into 3 week old wild-type siblings. Wild type bone marrow was screened at different time points (see Fig. 2)

Fig. 2; Left Panel: Week 14 post-transplantation, CK⁺ cells in bone marrow of wild-type siblings; Right Panel: HER-2⁺ cells in the bone marrow (Adapted from Hüsemann, Geigl, Schubert et. al (2008))

Although CK⁺ and HER-2⁺ cells were observed in the bone marrow, the levels were lower than transgenic BALB-Neu T mice, but were definitely above rare false positives. Additionally, just as in the transgenic mice, there was no significant increase in disseminated cells from ADH stages to invasive cancer, in the wild-type siblings.

The malignant nature of these DTCs was established by Comparative Genomics Hybridization (see Fig. 3)

Fig. 3; Top Panel: Karyogram (left) and CGH profile (right) of sorted mouse metaphase of a single HER-2⁺ cell disseminated from a transplanted mammary gland and subsequently isolated from bone marrow of a recipient wild-type mouse; Bottom Panel: Sorting of mouse metaphase of a single leukocyte in a balanced CGH profile; Green and red bars indicate genomic gains and losses respectively. (Adapted from Hüsemann, Geigl, Schubert et. al (2008))

Evidence that early disseminated cells can grow into metastases

The onset of metastasis relative to primary tumor growth was assessed . Histological sections of the lungs were analyzed and micrometastases could be detected from weeks 20 to 21 onward, a time point at which mostly in-situ carcinomas are present at the primary sites (see Fig. 4)

Fig. 4; Progression of lung metastasis in BALB-Neu T mice, with and without the removal of the primary tumor. Left Panel: Increase in tumor area over time in BALB-Neu T mice (same as Fig. 1b); Right Panel: Size of the largest lung metastasis detected in individual mice. Blue squares indicate average size of metastases from non-operated mice at various time points and red triangles indicate average size from operated animals at 10-15 weeks after surgery. Whiskers indicate 95% confidence intervals (Adapted from Hüsemann, Geigl, Schubert et. al (2008))

Since metastases need time to grow, their increase in size paralleling that of the primary lesion supports the conclusion that, at least in some cases, founder cells of metastasis had disseminated earlier and had started to proliferate.

Some other experiments conducted by the group, demonstrate that bone marrow disseminated cells that do not grow into metastases can be released from growth arrest. Also, there does not appear to be an association between the number of disseminated cells and the stage of the tumor. Large tumors do not necessarily seed more.

Overall the paper presents some compelling evidence for the occurrence of dissemination, even before the primary tumor is established. This would indeed force the field to rethink it's current strategies in combating cancer.

Reference:

Hüsemann, Y., Geigl, J., Schubert, F., Musiani, P., Meyer, M., Burghart, E., Forni, G., Eils, R., Fehm, T., & Riethmüller, G. (2008). Systemic Spread Is an Early Step in Breast Cancer Cancer Cell, 13 (1), 58-68 DOI: 10.1016/j.ccr.2007.12.003

Designed and Designoid Objects

noreply@blogger.com (Varun) — Sat, 22 Aug 2009 09:27:00 +0000

Part 2 of the 5 part Royal Institution Christmas Lectures for Children, 1991. With sheer clarity and brilliance, Richard Dawkins puts forward the concept of objects that are designed and "look designed"

Look Mom, Three Hands -The Classical Rubber Hand Illusion

noreply@blogger.com (Varun) — Fri, 21 Aug 2009 06:59:00 +0000

Most illusions are not only fun to experience but are also interesting to study in depth for what they can reveal about perceptual purposes. One of the most interesting illusions discovered in recent times in the rubber hand illusion. It was first reported in a paper in Nature in 1998, titled "Rubber hands 'feel' touch that eyes see" by Matthew Botvinick and Jonathan Cohen. So how does it work? A subject is seated with the left harm resting on a table while a standing screen is positioned besides the arm to hide it from the subject's view. A life-sized rubber model of a left hand and arm is placed on the table, directly in front of the patient. The experimenter uses two paintbrushes to stroke the rubber hand and the real hidden hand, synchronizing the timing of strokes as closely as possible.

In the original study by Botvinick and Cohen, subjects were asked to complete a two part questionnaire that asked for an open description of their experience and also to affirm or deny the occurrence of nine specific perceptual effects (see Fig. 1).

Fig. 1: Questionnaire includes nine statements presented in random order. Subjects indicated their response on a seven-step visual analogue scale ranging from 'agree strongly (+++)' to 'disagree strongly (---)'. Points indicate mean response and bars indicate response range. Underlined questions show a significant tendency to evoke an affirmative response (Adapted from Botvinick and Cohen (1998))

Most subjects indicated that they seemed to feel the touch not of the hidden brush, but that of the viewed brush, as though the rubber hand had sensed the touch. It was hypothesized that the illusion may arise due to a spurious reconciliation of visual and tactile inputs while distorting position sense (proprioception). In a second experiment, subjects were exposed to the illusion for a prolonged period and were then probed for distortion in proprioceptive information. Before and after the viewing period, subjects completed a series of three intermanual reaches. With eyes closed, the right index finger was drawn along a straight edge, until it was judged to be aligned with the index finger of the hidden left hand. The authors found that the subjects' reaches after experiencing the illusion were displaced towards the rubber hand, the magnitude of displacement varying in proportion to the reported duration of the illusion (see Fig. 2)

Fig. 2: Results of reaching experiment. x-axis indicates the percentage of 30-min viewing period during which the illusion was experienced. The y-axis indicates displacement of the three reaches made after the viewing period from the three made before. Data is fitted with a least-squares regression line (adapted from Botvinick and Cohen (1998))

In more systematic explorations by Tsakiris and Haggard (2005), it was shown that the drift, while indicating the position of the real left hand, in the direction of the fake rubber hand not only depends upon synchrony of strokes, but also on the position of the rubber hand (spatial congruency) as well as visual characteristics of the hand (body top-down effects) (see Fig. 3 for experimental setup)

Fig. 3: Participants saw in different conditions (a) a rubber hand in a congruent position, (b) a rubber hand in incongruent position, or (c) a wooden stick. The participant's left hand was out of view for the whole duration of the experiment (Adapted from Tsakiris and Haggard (2005))

The congruent posture elicits the maximum proprioceptive drift while having the rubber hand in an incongruent position or replacing the rubber hand by a wooden stick does not (see Fig. 4).

Fig. 4: Mean proprioceptive drift towards the rubber hand. Error bars indicate standard error. Asterix indicates significant difference between synchronous and asynchronous stimulation (Adapted from Tsakiris and Haggard (2005))

The body is distinguished from other objects as belonging to the self by participating in inter-modal perceptual correlations. In the experiments of Botvinick and Cohen, subjects who referred the tactile sensation to the rubber hand also reported experiencing the rubber hand as belonging to themselves. Indeed eight out of ten subjects employed terms of ownership in the free descriptions (Botvinick and Cohen (1998))

The rubber hand illusion presents a very intriguing case. It shows that certain forms of inter-modal correlations may be sufficient for self attribution even in the face of contradicting signals from other sensory modalities.

References

Botvinick M, & Cohen J (1998). Rubber hands 'feel' touch that eyes see. Nature, 391 (6669) PMID: 9486643

Tsakiris M, & Haggard P (2005). The rubber hand illusion revisited: visuotactile integration and self-attribution. Journal of experimental psychology. Human perception and performance, 31 (1), 80-91 PMID: 15709864

Role of polycomb and trithorax in developmental regulation

noreply@blogger.com (Manasi) — Wed, 19 Aug 2009 17:38:00 +0000

During development, patterns of differential gene expression, defining determined state of cells, needs to be maintained over several generations. A set of genes called polycomb and trithorax group genes (PcG and trxG) respectively have been identified in Drosophila which seem to exert such a memory function. In Drosophila, the earliest development is controlled by the expression of maternal genes while the final segment specificty is governed by the master regulator homeobox (Hox) genes. The hox genes encode various transcription factors which regulate many downstream genes. These factors have to be expressed in appropriate patterns during development. The expression of Hox genes, is regulated initially by activators and repressors encoded by the gap and pair ruled genes. These factors however, have a very short half life and hence decay once the Hox gene expression is initialized. It is during this period, that the PcG and trxG proteins recognize the transcriptionally repressed and active states (respectively) of the Hox genes and maintain their expression throughout development.

The polycomb and trithorax genes therefore seem to be acting antagonistically. Polycomb genes are involved in chromatin based gene silencing while trithorax group genes counteract this silencing effect to maintain gene activity. Also since they maintain the Hox gene expression throughout the process of development they may serve as molecular memory systems central to the process of development.

The exact mechanism by which these genes act is still not clear. However a pathway has been proposed which recruits these genes to their target sites on the DNA called Polycomb and Trithorax Response Elements(PREs and TREs). Once at their sites they regulate transcription by modulating the chromatin structure, in particular, via post translational modification of histones and by regulating the three dimensional organization of the TREs and PREs.

Various questions, as to the exact mechanism by which the PcG genes initially recognize the repressed state of their target genes (Hox) remain unanswered as of now. Also how these genes maintain this silenced state of chromatin and transmit the same through many cycles of cell division is intriguing too.

In the next post we shall look at the post translational histone modifications that happen once these proteins reach their target sites on the DNA.

The Art of Evolving

noreply@blogger.com (Varun) — Sun, 16 Aug 2009 07:58:00 +0000

Irreducible complexity is an argument that was first put forward by Michael Behe, a biochemist and proponent of intelligent design. Behe himself defines an irreducibly complex system as acollection of well-matched and interacting parts that on the whole, contribute to the functioning of the system, but the removal of any one part could cause the system to cease functioning. Most proponents of intelligent design and creationism like to argue in favor of irreducible complexity by using the eye as an example. Even on the surface, the argument appears ridiculous and ignorant. Consider an individual suffering from a cataract. The crystalline lens of the person is affected to varying degrees (from slight to complete opacity). Equally, the vision too is affected to varying degrees. A slight cataract may prevent the individual from seeing as clearly as an individual with no cataract, but he or she can still see.

So what does evolution have to say? Well, having an eye is not a jackpot. It obviously isn't a case of black and white. There are shades of gray. Most evolutionary biologists argue that having something is better than having nothing. Having 10% of a complete eye is better than having no eye. or having 51% of an eye is better than having 49% of an eye. So how could the eye have evolved? Obviously, the earliest ancestors had no eyes (in the way that we know what an eye is), but they may have had a single layer of photo-sensitive cells (see Fig. 1a). Now the only thing such a layer of cells would have been able to do is to detect light and dark. Not too much, but could've still assisted the animal when it had to detect if there were any predators around in the neighborhood. Euglena, for example, has flat patches of photoreceptors.

Now the next stage in evolution would have led to an indentation being formed in the photoreceptor layer, which gradually deepened to give rise to a cup-shaped structure (see Fig. 1b). This was better. Now this animal could not only distinguish light from dark, but also the direction from which the light was coming, since there is a shadow formed. Cup- shaped light sensitive spots are seen in Planaria. As evolution proceeded, the cup got deeper and deeper and with even the slightest increase in depth, the animal was able to make out, with ever increasing accuracy, the direction of light. Later, the cup may have started to close at the other end, ultimately forming a roughly spherical structure with a hole at one end (see Fig. 1c). This structure would have been equivalent to a pinhole camera. One animal that does have a pinhole camera for an eye is Nautilus, a sea mollusk. Now the resolution of a pinhole camera depends on the size of the aperture. Smaller the aperture, sharper the image. But if the aperture is small, then the amount of light passing in would also be less and hence the image would be dim. Hence, in a pinhole camera, one is always compromised over the other.

In any case, Nautilus had a pinhole camera where the aperture was open. Sea water could constantly flow in and out. So as the process continued, the aperture would have been covered with a thin layer of transparent material (which wouldn't have contributed much at first except for protecting the eye), which may have been the earliest form of the cornea (see Fig. 1d). The cornea, as we know it today, has a refractive power of about 43 diopters and does contribute to focussing light (along with the lens). Gradually as evolution went on, the cornea from being just a sheet of transparent protective material may have gained some refractive properties, allowing sharper images to be formed (even when the size of the aperture was large). Although the cornea contributes the most to the focussing power of the eye, it's focus is fixed. What "tunes" the focus in response to objects at different distances is the curvature of the lens. And thus, was born the lens (see Fig 1e)!

We see how evolution beautifully explains the gradual formation of a complex structure and puts to rest two arguments, one being that of irreducible complexity and the other being that of the Watchmaker.

Unconscious Cognition 2-Going Beyond Zero Awareness

noreply@blogger.com (Varun) — Sat, 15 Aug 2009 22:36:00 +0000

In the first part "Unconscious Cognition 1- Simple Dissociation", we established two sets of assumptions for the zero awareness criterion. The exhaustiveness assumption where the direct measure D is a strictly monotonic function of conscious information c and a weakly monotonic function of unconscious information u and the indirect measure I is a weakly monotonic function of both c and u. Under these conditions, D(c,u)=0 necessitates c=0 and I(c,u)>0 implies I(0,u)>0 which in turn implies u>0. In such a case, I is a perfect measure of unconscious processing. The second assumption was the exclusiveness assumption, where I is an exclusive weakly monotonic function of u alone, and is unaffected by c. And so, the exclusiveness assumption abolishes the need for a direct measure altogether

An interesting way to circumvent these assumptions is to let awareness vary over experimental conditions. It may then be possible to establish a double dissociation, which consists of finding an experimental manipulation that changes D and I in opposite directions (see. Fig 2a). In particular, any pair of experimental conditions that leads to opposite ordering of data points in direct and indirect measures gives evidence for double dissociation. One example could be a priming experiment with two (or more) masking conditions where the priming effect (indirect measure) increases while prime identification (direct measure) performance decreases over experimental conditions. It is obvious that two measures of information going in opposite directions cannot be monotonically driven by a single information source (see Schmidt and Vorberg (2006) for a formal proof).

Double dissociations have surprising features. Firstly, they require D to be non-constant. In order to obtain a double dissociation, variations in awareness over experimental conditions must occur so that there is a non-zero awareness of the prime under atleast some conditions. Also, the assumptions are less stringent. The only assumption we make here is that both D and I are weakly monotonic in c (see Fig 2b). One can even drop the weak monotonicity assumption on u, allowing for c and u to produce arbitrary interactive effects on D and I for instance, c and u could be mutually inhibitory).

An example of Simple Dissociation

There are numerous experiments demonstrating simple dissociation. In one such experiment (Vorberg, Mattler, Heinecke, Schmidt and Schwarzbach (2003, 2004)), participants were asked to make speeded keypress responses to the direction of an arrow-shaped masking stimulus that was preceded by an arrow-shaped prime. The mask has a dual purpose. It acts as the target of the response and at the same time, it reduces the visibility of the prime by metacontrast masking (a form of visual backward masking). As the stimulus onset asynchrony (SOA) between prime and mask increased, the priming effect (indirect measure) also increased with primes pointing in the same direction as the mask shortening the response times while primes pointing in the opposite direction lengthening them. strikingly, this priming response was independent of visual awareness of the prime. This was determined by using stimulus conditions that produced different time-courses of metacontrast masking. Participants were unable to perform better than chance when asked to point the direction of the prime.

An example of Double Dissociation

In a second experiment by the same group, all four pairings of short-duration (14 ms) and long-duration (42 ms) primes and masks were compared, yielding very different kinds of masking functions. When 14 ms primes were combined with 42 ms masks (14:42), the prime identification performance was low and increased only slightly with SOA. When mask duration was reduced to 14 ms (14:14), performance was better. When a 42 ms prime was paired with a 14 ms mask (42:14), performance was nearly perfect. But the 42:14 condition yielded an effect called type-B masking where prime identification performance markedly decreases with prime-mask SOA, then increases again, while the priming effect only increases monotonically all throughout producing a strong double dissociation.

Reference

Schmidt, T. (2007). Measuring unconscious cognition: Beyond the zero-awareness criterion Advances in Cognitive Psychology, 3 (1), 275-287 DOI: 10.2478/v10053-008-0030-3

Unconscious Cognition 1-Simple Dissociation

noreply@blogger.com (Varun) — Sat, 15 Aug 2009 09:48:00 +0000

Attempts to demonstrate unconscious processing of visual stimuli are very old and riddled in controversies, but the controversy does not so much concern the existence of unconscious processing (most researchers seem to be convinced of this), but rather the question of how to demonstrate unconscious processing in a given experiment. If one needs to demonstrate it, one has to make sure that a critical stimulus was completely outside of awareness (the so called zero-awareness criterion). Schmidt (2007) proposes two lines of attack for establishing unconscious processing beyond the zero-awareness criterion. The paper deals with different types of dissociation between measures of awareness and measures of processing.

Simple Dissociations

To demonstrate that a critical stimulus was processed unconsciously, one usually produces some dissociation between different behavioral measures of performance. This is done by comparing two measures obtained from different tasks. One measure (called the direct measure, D) signals the observer's awareness of a critical stimulus. For example, consider a visual perception experiment, where the task is to detect the offset of a vernier (a vernier is simply a set of two vertical lines, one below the other, where the lower line can be offset either to the right or left of the upper line). However, before the vernier is shown (let's say for 25 msec), a prime is flashed for a short period of time (say 15 msec). This prime could be, for example, arrows pointing to the right or left. A forced-choice prime discrimination task ("Was the arrow to the right or left" ) would measure if the observer was aware of the stimulus, and hence would comprise the direct measure. The second measure (called the indirect measure, I) would indicate that the primes themselves are not consciously detected, but they are involved in a priming effect, and hence affect the reaction times of responding to whether the target vernier is offset to the right or left. For example, if the prime and vernier are both congruent (i.e. the arrow points to the right and the vernier is also offset to the right) and if the subject cannot consciously detect the direction of the prime but the reaction times during the congruent cases are always shorter than the reaction times during the incongruent cases (prime pointing to the right but vernier offset to the left), then this would comprise the indirect measure.

Schmidt and Vorberg (2006) examined the assumptions required by the zero-awareness criterion and other approaches. They start by assuming that the direct and indirect measures may depend on two sources of information labeled conscious (c)and unconscious (u). In other words, D = D(c,u) and I = I(c,u), where information is non-negative. The dependency is weakly monotonic, which means that if any type of information increases, the measures can only increase or remain constant. An additional constraint is that if D and I are to be modeled as functions of c and u, then both D and I must be functions of the same underlying conscious and unconscious information. Thus the direct and indirect tasks must be designed to use identical stimuli, identical responses and identical stimulus-response mappings. Schmidt gives an example of this mismatch in the study by Dehaene et. al (1998), where the indirect task was to determine as quickly as possible if a target digit was numerically larger or smaller than 5. The target was preceded by a masked prime digit. The response times were much shorter if the prime was consistent with the target (i.e. both were less than 5 or greater than 5) than when the two were inconsistent (one of them was larger than 5 and the other was smaller). The optimal direct task in this experiment would have been to ask the subject "Was the prime greater than or lesser than 5" because then, both measures would've been tapping into the same source of information (conscious or unconscious). Instead, Dehaene et. al chose two different direct tasks where the subject was asked to detect primes against an empty background, and the second where the subject was asked to discriminate primes from random strings of letter, none of which addressed the critical question of whether the prime was larger or smaller than 5.

Given that D-I mismatch can be avoided, how can the null model of only conscious processing be disproved. The traditional way is the zero-awareness criterion, which produces what is called a simple dissociation of direct and indirect measure (zero D in presence of non-zero I, see Fig. 1a). In other words, I(c,u)> 0, implies that either c > 0, or u > 0, or c and u > 0. But, does, D(c,u) = 0, imply c = 0 and hence I(c,u) > 0 only because u > 0 ? No, because, D is a weakly monotonic function of c. It is possible that under this assumption, c did change but D was not sensitive enough to pick up this change. To resolve this problem, one must make the stronger assumption (see Fig. 1b) that D is a strictly monotonic function of c, which means that D can detect any changes in c, no matter how small. Hence, D(c,u) = 0 implies c = 0, irrespective of whether u is zero or not, since D is weakly monotonic on u anyway. And now, if I(c,u) > o, then, I(0,u) > 0 which finally implies u > 0.

The exhaustiveness assumption is important. In it's absence, one can always argue that it is only conscious processing c, that is occurring and while I is sensitive enough to detect it (I > 0), D is not (D=0). Finally, there is an alternative set of assumptions that abolishes the need for an direct measure altogether. This is when the indirect measure can be assumed to be an exclusive monotonic function of u and is unaffected by c. In this case, I(c,u) = I(u) > 0 implies u > 0 directly (see Fig. 1c)

In the next post, we shall look at double dissociation and beyond

Reference

Schmidt, T. (2007). Measuring unconscious cognition: Beyond the zero-awareness criterion Advances in Cognitive Psychology, 3 (1), 275-287 DOI: 10.2478/v10053-008-0030-3

On the Integration of Tactile, Proprioceptive and Visual Signals by the Brain

noreply@blogger.com (Varun) — Fri, 14 Aug 2009 08:23:00 +0000

The integration of different sensory inputs in the brain is crucial not only for taking appropriate motor actions but also for body perception and awareness of the bodily self. Integration occurs in higher areas in the brain usually in areas belonging to the parietal lobe

1) Integrating Vision and Proprioception in Area 5 ( Graziano et. al (2000))

Brodmann Area 5 (or Area 5) is part of the parietal cortex in humans, and in monkeys, is a subdivision of the parietal lobe, occupying primarily the superior parietal lobule. Graziano et. al (2000) studied single neuron responses from Area 5 of monkeys. In their experiments, the arm contralateral to the recording site was outstretched. The real arm of the monkey was covered, and instead a realistic fake hand was kept in view. The experimental design was 2 x 2

2 x 2 experimental design. Cross indicates fixation spot. The gray arm is the

fake arm. The real arm is covered in the experiment (Graziano et. al (2000))

Single neuron recordings indicated that the firing rate of individual neurons depends not only on the position of the real arm, but also on the position of the fake arm. The neuron is significantly affected by the position of the real arm, firing more when he real arm is to the left. Additionally, the firing rate further increases when the fake arm is also to the left.

Single neuron recording from Area 5. The firing rate is maximum when

the real and the fake arms are in congruent position (Graziano et. al (2000))

2) Integrating Vision and Touch in Ventral Intraparietal Area (Duhamel et. al (1998))

The Ventral Intraparietal Area (or VIP) is a discrete area in the depths of the intraparietal sulcus. Duhamel et. al carried out single neuron recordings in the VIP. They found the neurons to possess a bimodal receptive field. Not only did they respond to a combination of visual and tactile stimuli, but the bimodal receptive fields were arranged in an orderly manner.

Bimodal receptive fields of VIP neurons (Duhamel et. al (1998))

Small central visual receptive fields were associated with small tactile receptive fields on the muzzle, whereas large peripheral receptive fields were associated with large tactile receptive fields on the side of the head or body. The neurons also demonstrated direction selectivity, in the sense that a visual or tactile stimulus moving in one direction was preferred over the other, and this preferred direction of visual and tactile stimuli coincided in the majority of cells.

Direction selectivity of VIP neurons (Duhamel et. al (1998))

References

Graziano, M. (2000). Coding the Location of the Arm by Sight Science, 290 (5497), 1782-1786 DOI: 10.1126/science.290.5497.1782

Duhamel JR, Colby CL, & Goldberg ME (1998). Ventral intraparietal area of the macaque: congruent visual and somatic response properties. Journal of neurophysiology, 79 (1), 126-36 PMID: 9425183

Scaredy Cat? Blame it on your genes (Again!!)

noreply@blogger.com (Varun) — Thu, 13 Aug 2009 10:48:00 +0000

Serotonin (5-Hydroxytryptamine or 5-HT), as some of us may know, is a neurotransmitter that is involved in modulation of moods such as anger. The level of re-uptake of serotonin by the presynaptic neuron is crucial and abnormal functioning of the serotonin transporter has been implicated in different neurological diseases. The human 5-HT transporter (5-HTT) gene transcription is modulated by a common polymorphism in the upstream regulatory region and the short variant of the polymorphism reduces 5-HTT transcription and hence reduces 5-HT uptake by lymphoblasts. In this paper, the authors (Lesch et. al (1996)) report association studies demonstrating that the presence of the short variant pre-disposes the individual towards anxiety related personality traits.

Neuroticism scores (seperated into eight groups) and percentages of subjects

from L(n=163) and S(n=342)groups in each of the eight T-score groups.

Notice that the S percentage is higher than L percentagein all T-score groups

above 54 (Lesch et. al (1996))

Reference

Lesch, K., Bengel, D., Heils, A., Sabol, S., Greenberg, B., Petri, S., Benjamin, J., Muller, C., Hamer, D., & Murphy, D. (1996). Association of Anxiety-Related Traits with a Polymorphism in the Serotonin Transporter Gene Regulatory Region Science, 274 (5292), 1527-1531 DOI: 10.1126/science.274.5292.1527

Some Thoughts on P-Zombies and the Gap Explanatory Argument

noreply@blogger.com (Varun) — Wed, 12 Aug 2009 18:37:00 +0000

Dedicated to Anna Traußnig...for the several hours of intellectual orgasms, as she likes to call it

Parts of this article have been inspired by the writings of Robert Kirk, William Seager and Daniel Dennett

A philosophical zombie (or p-zombie as it is often called), is a curious creature that is central to the basic idea of the explanatory gap. It is a hypothetical being that is indistinguishable from you and I, or any normal human being for that matter, except for one fundamental detail. Such a zombie, for example, cries in pain (when you kick it), but it feels no pain. In other words it lacks any conscious experience. But, given their definition, this singular fact will have no bearing on the physical processes that the zombie will undergo in it's own world. Put another way, a zombie's behaviour should be indistinguishable from the behaviour and physical state of a genuine human being. This first case is the well known "zombie duplicate" -There exists a philosophical zombie that is physically indistinguishable from me , assuming of course that I can be granted consciousness. Let's stick with that for the moment. We shall come back to this point later.

Now I could argue that a perfectly physical duplicate of me should live in a perfectly duplicate realm of reality. It could also be argued that the only possible world that could have a perfectly physical duplicate of me is one which is perfectly duplicated , which might be the very same possible world. Are zombies nomologically or even physically possible? To state that they are nomologically possible would be to state that in some world that shares all physical and natural laws of this world, there exists a "human being" who lacks consciousness. But wait, what happens if certain natural and physical laws do not bear any causal relation? If all natural laws are physical laws or at least dictate physical laws and if, consciousness is causally related to physical states (and there is some evidence for that), then there cannot exist any world with the same physical and natural laws as ours and yet have a zombie. It's not possible. On the other hand, if physical laws were dissociated from natural laws, then zombies could exist by breaking natural laws but not the physical ones. Remember, we have access only to physical laws (which we assume, by default to be natural laws, although this may not necessarily be true). Going further, if mental states (such as consciousness) were non physical, it seems reasonable to claim that zombies are not only nomologically possible but even logically (or physically) possible. Of course, the question still remains whether such another distinct reality, separate from ours does exist (this in itself is another huge philosophical debate). Now if this were the case, one would have a tough time proving what consciousness really is (or is not). It raises another question. How are we to assume that a zombie that exists in another physical realm sharing the same physical laws as ours but not the natural laws does not have consciousness? It may possess a different kind of consciousness that is drawn from the natural laws that govern it's world.

In 1999, Robert Kirk, presented an argument which attempted to demonstrate that philosophical zombies are logically impossible. He starts by assuming that a zombie suddenly acquires non-physical qualia. A weak definition of qualia would be the property that distinguishes us from zombies (consciousness if you will). Parts of his argument are as follows:

My zombie twin is unaffected by anything non-physical
So he remains unaffected when he acquires non-physical qualia
In order to become conscious following acquisition of this non-physical qualia, he must be affected by it in some way
But nothing non-physical affects him. So he doesn't become conscious.
To tell the difference between the subjective character of two different perceptual experiences, he must be sensitive to them or detect them in some way
In any case, nothing non-physical affects the zombie, so he cannot distinguish non-physical qualia from having no qualia
Friends of the zombie maintain that a "conscious" being is simply the zombie component with non-physical qualia. This position entails that "conscious" humans cannot tell the difference between tea and coffe, for example.
But we can tell the difference between tea and coffee.
Hence, we are nothing of the sort that friends of zombies maintain we are and hence zombies in their sense are not genuinely possible

Kirk's paper has been criticized but it presents an interesting case.

One final word on physicalism. In short, there are no kinds of things other than physical things (including consciousness). Daniel Dennett argues that even if consciousness were purely physical, our own conviction that we are not zombies, is simply a product on the external physical world. But, we could still be zombies who think we're conscious, who think we have qualia and who think we feel pain, in ways that are we could never discover. If such were the case, it wouldn't be careless to say that "consciousness" could never be defined or understood or even proved.

I for one remain hopeful that science may one day provide a good theory on how consciousness works, but Anna and I both think that even if one ends up showing how it works, one would never be able to show what it really is.

References

Kirk, R. (1999). The Inaugural Address: Why There Couldn't Be Zombies Aristotelian Society Supplementary Volume, 73 (1), 1-16 DOI: 10.1111/1467-8349.00046

Seager, W.E., Are Zombies Logically Possible?-And Why it Matters

http://www.scar.utoronto.ca/~

Take that leg off-It ain't mine

noreply@blogger.com (Varun) — Wed, 12 Aug 2009 09:35:00 +0000

So I recently read this paper from V.S Ramachandran's group at the Salk Institute and came to realize how weird the field of cognitive neuroscience can really get. The paper is on Apotemnophilia (phew!! saying it is a task in itself), which yours truly had never heard of before. In any case, the disease is characterized by the desire to amputate one's own limb (Now how about that). These patients are otherwise mentally normal.

Individuals suffering from apotemnophilia always date the desire for amputation since childhood and often term the limb as being over-present or intrusive. Most obtain an amputation and paradoxically report feeling much more 'complete' and happier. Traditional explanations for the disease range from it being sexual paraphilia, related to the phallic resemblance of an amputee's stump (Someone get Freud in here!) or perhaps the mere sight of an amputee is permanently imprinted in the malleable psyche of a child as the "ideal body representation"

In this paper (McGeoch et. al (2009)), they demonstrate that the disease actually has a neurological basis based on the following observations:

1) Sufferers have no other psychological disorders

2) They desire an amputation of a limb at a specific level

3) There is a left sided bias.

The last observation points a finger to one particular area in the right parietal lobe called the superior parietal lobe (SPL) which receives connections from a host of other areas, namely the visual, primary somatosensory, secondary somatosensory, premotor and motor cortices. Additionally, the right parietal cortex is known to play a vital role in constructing body image and damage can lead to various disorders like somatoparaphrenia (denial of ownership of left arm) and others. The autors postulate that the right SPL may contain a hardwired representation of the body and if a particular limb were missing from the representation, the consequence may be a desire for amputation.

Right hemisphere of averaged control brain (a,b) and subject brain (c,d). Right SPL is outlined in black. Images a and c show touch to the left foot of control and subject respectively while b and d show touch to the right foot of control and subject respectively . This subject wanted right below-knee amputation (McGeoch et. al (2009))

This confirms the hypothesis that there is a congenital failure to represent affected limbs in the body image. Since visual and somatosensory inputs are still intact, but there is no corresponding limb representation, the result would be a mismatch that manifests itself as an intrusive and over-present limb.

References

McGeoch, P.D., Brang, D., Song, T., Lee, R.R., Huang, M., & Ramachandran, V.S. (2009). Apotemnophilia - the neurological basis of a 'psychological' disorder Nature Precedings : 10101/npre.2009.2954.1

Brang, D., McGeoch, P., & Ramachandran, V. (2008). Apotemnophilia: a neurological disorder NeuroReport, 19 (13), 1305-1306 DOI: 10.1097/WNR.0b013e32830abc4d

The Small Network Argument

noreply@blogger.com (Varun) — Tue, 11 Aug 2009 19:35:00 +0000

Not long ago, the problem of consciousness was thought to be scientifically non-tractable. But with a lot more psychophysical and brain mapping data coming in from studies being done on normal subjects and people with brain disorders, the field has opened up. In the last decade, a variety of brain processes were proposed to account for consciousness and several more keep coming in but as of today, no concrete model exists. Typical examples include recurrent computation (Grossberg (1999) and Lamme (2006)), synchronized neural activity (Bachmann (1994), Engel et. al (1999)), winner takes all computation (Grossberg (1999)), closed loop action-perception processing (O'Regan and Noe (2001)). In the midst of the madness, comes a paper by Herzog et. al titled Consciousness and the Small Network Argument (Herzog, Esfeld and Gerstner (2007)). They propose that none of the current models can fully account for consciousness because of what they call the small network argument:

For each of the above models, there exists a very small neural network that fulfills the respective characteristics of the model but does not exhibit consciousness

1) Recurrent Computation (Lamme (2006))

Two mutually connected neurons make up a recurrent systems. Thus if recurrence is sufficient for consciousness, a network of two neurons must create consciousness (to which not may of us may accede).

2) Winner Takes All Computation (Grossberg (1999))

The basic operation of a winner-takes-all (WTA) circuit is to suppress the activity of all neurons except the one having the largest input. Minimal models require only three neurons fully connected to three presynaptic neurons, plus a neuron for vigilance, taking the tally to seven. Such a network can develop states allowing learning and memory. And so are we willing to grant consciousness to a network of seven neurons?

3) Synchronized Neural Activity (Bachmann (1994), Engel et. al (1999))

If synchronized firing in neurons is the main characteristic of consciousness, then a group of three inter connected neurons firing in synchrony is conscious

4) Closed-loop action perception processing (O'Regan and Noe (2001))

Herzog et. al give the example of a thermostat with a temperature sensor ('perception') controlling a heater ('action'). This could be formulated as a two neuron feed-forward network with a sensory neuron connected to an output neuron controlling the switch.

Thus they argue that if one does not want to attribute consciousness to such small networks, other components are needed. Additional characteristics are often imposed. Typical examples being attention, number of neurons and complexity of the network. Yet, none of these additional components necessarily solves the small network argument.

1) O'Regan and Noe (2001) combined their sensorimotor contingency approach with attention based processing. They state that perception occurs in a closed loop of action and information processing but consciousness emerges only when attention comes into play. The most common example of this being a person driving a car in an unconscious and automatic mode (unconsciously perceiving other cars on the road and taking correct measures), but conscious perception, e.g. of a traffic signal, arises only when attention comes into play. However, in this model, attention can be incorporated with just one single extra input arising from a second group containing a very small number of cells

2) It is often proposed that consciousness may emerge if the brain exceeds a certain number of neurons. In a model with a simple linear arrangement of neurons where each neuron is connected only to it left and right neighbors, there is no reason to believe why a network of 10^10 neurons is more capable of generating consciousness than a network of three neurons. hence size alone cannot account for this

3)Other approaches state that a certain level of complexity must be met before a network can yield consciousness. Herog et. al give the example of Tononi and Edelman (1998) who proposed to measure complexity by defining a functional cluster that was loosely connected to the rest of the network but still has a rich repertoire of internal states. But this complexity criterion can be met even by a small network. Herzog et al. propose a network of nine neurons, organized into clusters of three. Assume the following model of interaction. The state of each neuron is described by p bits. Therefore, each neuron has can take up 2^p possible states. Now in the first 100 msec, the three neurons agree on the first bit by a majority vote, in the next 50 msec, on the second bit and so on, until the last 200/(2^p) msec, when they decide on the last bit. Thus for any arbitrary p, three neurons will agree on p bits in 200 msec, showing a large level of complexity in a small cluster.

Hence, Herzog et. al argue that all the above mentioned components are important aspects to understand consciousness but they are only trivially necessary rather than sufficient. Finally, they argue that one way out of the small network argument is to assume that each network, however small, has a vanishingly small consciousness, but this form of "panpsychism" also has it's problems. What happens if two networks, each with it's own vanishingly small consciousness, are connected? Do the two consciousness merge, do they stay separate (as proposed in split brain patients) or are there new coalitions of neurons making up new consciousness?

Overall, the paper seems to have created a bit of a ripple in the community of neuroscientists working on consciousness. Taylor (2007) in "Commentary on 'the small network' argument" states

..paper of this issue (Herzog, Esfeld, & Gerstenr, 2007) has derived an interesting criterion for any neural model of consciousness, with the disturbing result that a whole raft of neural models of consciousness are deficient when looked at by this criterion. The criterion itself is that a neural model of consciousness is suspect if it can be shown to work using only a relatively small network of neurons (of the order of 10–100 or so)

Reference

HERZOG, M., ESFELD, M., & GERSTNER, W. (2007). Consciousness & the small network argument Neural Networks, 20 (9), 1054-1056 DOI: 10.1016/j.neunet.2007.09.001

The Fuzziness of Boundaries-Max Delbrück

noreply@blogger.com (Varun) — Fri, 26 Dec 2008 15:39:00 +0000

In the summer of 1941, a young man-only 34 years old-showed up for the first time at the Cold Spring Harbor Laboratory Campus. He would ultimately become the leader of a generation of scientists who would establish the field of molecular biology. His name was Max Delbrück. Max was born in Berlin. His father Hans, was a professor of history at the University of Berlin and his mother was the grand-daughter of Justus von Liebig (of chemistry fame).

His early interests were directed towards astronomy. However, during the latter part of his graduate studies in Göttingen, the breakthroughs in quantum mechanics caused him to shift to theoretical physics. He obtained his Ph.D in 1930 following which he went back to Berlin and worked as an assistant to Lise Meitner, It was during this time that he discovered the theoretical basis of gamma ray scattering by a Coulomb field, a phenomenon that to this date bears his name, Delbrück scattering.

In 1937, he moved to Caltech, in the United States, where he teamed up with Emory Ellis doing phage research. Together, they demonstrated that viruses reproduce in one step, rather than exponentially (as cellular organisms do). Following the start of World War II, Delbrück chose to remain in the United States, teaching physics at Vanderbilt University while conducting genetic research. His claim to biological fame came with the establishment of the famous "Phage Group" with Salvador Luria and Alfred Hershey at the CSHL . In 1943, he and Luria demonstrated that genetic mutations in bacteria arise in the absence of selection and not as a response to selection. This work was significantly backed up by mathematical models that they developed which were consistent with experimental results. He shared the 1969 Nobel Prize in Medicine and Physiology with Luria.

His name may not be as familiar as Albert Einstein or Charles Darwin, but Max Delbrück was a scientific giant who changed the world through his research in modern biology. He saw no barriers between the sciences. He had with him his tools from physics and mathematics and the problems from biology. In an essay commemorating Delbrück's 60th birthday, Luria wrote "Seldom has a group been so richly rewarded as have we, the molecular biologist, whom the physicist Max Delbrück, more than anyone else, guided to the explorations of the deep mysteries of life".

In the words of the great man himself

A Max Sing-Along

(lyrics by Max Delbrück, 1968)

I was born about 100 years ago

And there is nothing in this world that I don't know

I have Mendel in the garden

And I steered the ship for Darwin

I showed Otto how the neutrons had to go!

I had Watson grab the helix by the tail

I told Morgan that the flies would never fail

And the prize that went to Feynman

Well the theory was mine man,

And my song and inspiration never stale!

And, my buddy, Linus owes a debt to me,

For the structure of his modern chemistry,

Of the phage I am the father,

But I didn't want the bother,

Of collecting data so statistic-al-ly!

I'm a single-minded scientific man....

The Fuzziness of Boundaries-Introduction

noreply@blogger.com (Varun) — Thu, 25 Dec 2008 11:03:00 +0000

How times change!! A few years back, when I announced my plan of pursuing a research career in fundamental biological sciences, I was labeled by many as a mediocre student who did not want to pursue a degree in engineering for want of insight in the fields of physics, mathematics and engineering. Just a week ago, I was with a colleague (he is an electronics engineer studying at the EPFL) in Lausanne and our discussion soon turned to the evergreen topic of biological neural networks. I impressed upon him, the significance of the ongoing Blue Brain project at the Brain Mind Institute in the EPFL.

The project uses two huge IBM Blue Gene supercomputers to simulate a neocortical column (the basic functional unit of the brain, containing 10,000 neurons). At the push of a button, the model could reconstruct biologically accurate neurons and automatically connect them in a biological manner, a task than involves positioning 30 million synapses (connections that one neuron makes with another, edges between nodes, if you happen to be a graph theorist). All very well except for one tiny detail. The brain happens to consist of 100 billion neurons and 100 trillion synapses!!! Heck, the Blue Brain doesn't come even remotely close to the actual situation. And it requires two huge supercomputers just for a mere 10,000.

Is the Blue Brain Project a sham then? The answer is no. It is an engineering feat and it is here to provide more clues on one of the most profound questions in biology, the mammalian brain. The point I'm trying to make here is that physical sciences, engineering sciences and the biological sciences were never mutually exclusive. Biology brings the problems. Big problems, trust me. The physical sciences bring with it the theory, principles and the foundation needed to tackle them. Finally, the engineering sciences bring the tools.

Having said all of this, in the next part of this series, I shall introduce some great men of science and their efforts in breaking the wall between fields, such that today, I have no idea where one ends and the other starts. The boundaries are seemingly fuzzed up and are getting fuzzier by the day.

The International Genetically Engineered Machines Competition

noreply@blogger.com (Varun) — Sat, 06 Sep 2008 20:39:00 +0000

Last November, I was part of the team from the National Center for Biological Sciences (NCBS), Bangalore, India, at iGEM, which is an annual contest held at the Massachussets Institute of Technology (MIT), USA

(From R to L, Krishna, Mukund and Varun with the trophy)

What is iGEM?

iGEM addresses the question: Can simple biological systems be built from standard interchangeable parts and operated in living cells? Or is biology simply too complicated to be engineered this way? The only way to try and answer this question is to actually engineer biological devices. This forms the basis for the new and emerging field of Synthetic Biology

NCBS won the "Best Model and Simulation Award". For more details of our project, visit the NCBS iGEM Team-2007 Wiki

http://parts.mit.edu/igem07/index.php/Bangalore