Rajita Sinha: The Stressed Brain: Hijacking Cognition, Emotion, Behavior, and Health

The video below showed up in my feed a couple of days before the article I'm sharing on how stress generates enzymes that attack the brain. Together, this information highlights how destructive stress can be on our brains and our cognitive function.

How stress tears us apart: Enzyme attacks synaptic molecule, leading to cognitive impairment

Date: September 18, 2014
Source: Ecole Polytechnique Fédérale de Lausanne

Summary:
Why is it that when people are too stressed they are often grouchy, grumpy, nasty, distracted or forgetful? Researchers have just highlighted a fundamental synaptic mechanism that explains the relationship between chronic stress and the loss of social skills and cognitive impairment. When triggered by stress, an enzyme attacks a synaptic regulatory molecule in the brain, leading to these problems.

Carmen Sandi's team at EPFL discovered an important synaptic mechanism in the effects of chronic stress. It causes the massive release of glutamate which acts on NMDA receptors, essential for synaptic plasticity. These receptors activate MMP-9 enzymes which, like scissors, cut the nectin-3 cell adhesion proteins. This prevents them from playing their regulatory role, making subjects less sociable and causing cognitive impairment. Credit: EPFL  

Why is it that when people are too stressed they are often grouchy, grumpy, nasty, distracted or forgetful? Researchers from the Brain Mind Institute (BMI) at EPFL have just highlighted a fundamental synaptic mechanism that explains the relationship between chronic stress and the loss of social skills and cognitive impairment. When triggered by stress, an enzyme attacks a synaptic regulatory molecule in the brain. This was revealed by a work published in Nature Communications.

Carmen Sandi's team went to look for answers in a region of the hippocampus known for its involvement in behavior and cognitive skills. In there, scientists were interested in a molecule, the nectin-3 cell adhesion protein, whose role is to ensure adherence, at the synaptic level, between two neurons. Positioned in the postsynaptic part, these proteins bind to the molecules of the presynaptic portion, thus ensuring the synaptic function. However, the researchers found that on rat models affected by chronic stress, nectin-3 molecules were significantly reduced in number.

The investigations conducted by the researchers led them to an enzyme involved in the process of protein degradation: MMP-9. It was already known that chronic stress causes a massive release of glutamate, a molecule that acts on NMDA receptors, which are essential for synaptic plasticity and thus for memory. What these researchers found now is that these receptors activated the MMP-9 enzymes which, like scissors, literally cut the nectin-3 cell adhesion proteins. "When this happens, nectin-3 becomes unable to perform its role as a modulator of synaptic plasticity" explained Carmen Sandi. In turn, these effects lead subjects to lose their sociability, avoid interactions with their peers and have impaired memory or understanding.

The researchers, in conjunction with Polish neuroscientists, were able to confirm this mechanism in rodents both in vitro and in vivo. By means of external treatments that either activated nectin-3 or inhibited MMP-9, they showed that stressed subjectscould regain their sociability and normal cognitive skills. "The identification of this mechanism is important because it suggests potential treatments for neuropsychiatric disorders related to chronic stress, particularly depression," said Carmen Sandi, member of the NCCR-Synapsy, which studies the neurobiological roots of psychiatric disorders.

Interestingly, MMP-9 expression is also involved in other pathologies, such as neurodegenerative diseases, including ALS or epilepsy. "This result opens new research avenues on the still unknown consequences of chronic stress," concluded Carmen Sandi, the BMI's director.

Story Source:
The above story is based on materials provided by Ecole Polytechnique Fédérale de Lausanne. Note: Materials may be edited for content and length.

Journal Reference:
Michael A. van der Kooij, Martina Fantin, Emilia Rejmak, Jocelyn Grosse, Olivia Zanoletti, Celine Fournier, Krishnendu Ganguly, Katarzyna Kalita, Leszek Kaczmarek, Carmen Sandi. (2014). Role for MMP-9 in stress-induced downregulation of nectin-3 in hippocampal CA1 and associated behavioural alterations. Nature Communications; 5: 4995 DOI: 10.1038/ncomms5995

The article referenced here is open access, but it is highly technical. For those who want to read more, I am including the Discussion section below the video (at the bottom of the page).

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Rajita Sinha: The Stressed Brain

Published on Sep 16, 2014


A Stockholm Psychiatry Lecture given by Professor Rajita Sinha, Yale University, at Karolinska Institutet Aug 27 2014. Title of the lecture: The stressed brain: hijacking cognition, emotion, behavior and health.

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Here is the discussion section of the article summarized above.

Discussion

We tested the hypothesis that MMP gelatinase activity is involved in key proteolytic processing events induced by chronic stress in a hippocampal subfield-dependent manner and in connection with behavioural changes. We show that chronic stress leads to a CA1-specific reduction in the perisynaptic expression of ​nectin-3 and found that this reduction is critically involved in the stress-induced deficits in social exploration, social recognition and CA1-dependent cognition. Interestingly, we found increased ​MMP-9-related gelatinase activity in the hippocampal CA1 in chronically stressed animals and could show that ​MMP-9 itself cleaves recombinant ​nectin-3, a process mediated via the NMDA-receptor. Consistently, intra-CA1 administration of either an ​MMP-9 inhibitor or an NMDA receptor antagonist during stress exposure prevented the development of stress-induced deficits in social exploration, social memory and CA1-dependent cognition. Our findings highlight a fundamental role for ​MMP-9 in the effects of chronic stress on brain function and behaviour.

Nectins are emerging as both targets^{24, 43} and mediators²⁵ of stress actions in hippocampal-dependent memory and structural plasticity. We found molecular-, regional-, cellular compartment- and stress duration-dependent changes, with reduced ​nectin-3 expression after 21 days, but not 1 day, of restraint stress in the CA1 synaptoneurosomal, but not the total fraction. This was paralleled by deficits in several social behaviours and in a CA1-dependent cognitive task. Our results from cell culture experiments suggested that NMDA receptor activation during stress exposure might be implicated in the cleavage of ​nectin-3 in CA1 and its associated behavioural alterations. Previous work has implicated NMDA receptor activation in chronic stress-induced structural alterations in the hippocampus^{12, 44, 45}. Our in vivo study involving the pharmacological administration of the NMDA receptor antagonist ​MK-801, either systemically or directly into the CA1 region, confirmed that this treatment prevented the stress-induced reduction of ​nectin-3 expression in the CA1 synaptoneurosomal fraction as well as the behavioural impairments induced by stress in the sociability and temporal order task.

Using AAV-induced OE of ​nectin-3 either in the whole hippocampus or specifically in the CA1 area, we obtained evidence for a causal role of ​nectin-3 reduction in chronic stress-induced behavioural alterations, with the exception of the aggressive phenotype. We confirmed that the effects of ​nectin-3 OE were not due to altered physiological responses to the stress procedure (for example, body weight changes or ​corticosterone responses) or to changes in anxiety or locomotion. We found increased ​nectin-1 expression associated with AAV-​nectin-3 OE throughout the hippocampus, consistent with evidence in knockout mice indicating that downregulation of either ​nectin-1 or ​nectin-3 induces a parallel decrease in the levels of the other nectins in the hippocampus⁴⁶. ​Synaptophysin levels were not changed by ​nectin-3 OE and/or chronic stress, which is line with findings described for ​nectin-3 knockout mice⁴⁶. In addition, using the same chronic restraint stress protocol as described here, changes in the size of postsynaptic densities were observed but not in synaptic density in the CA1 (ref. 5). Interestingly, consistent with evidence that nectins recruit cadherins to cooperatively promote cell adhesion⁴⁷, we found a reduction in the CA1 perisynaptic ​N-cadherin levels. The specificity of these molecular changes in CA1 was supported by a lack of significant changes in the stressed animals’ synaptoneurosomal compartment of ​SynCAM-1 in the same brain region. To verify that the molecular changes specifically observed in CA1 were associated with well-established CA1-dependent behaviours, we tested animals in the temporal order task that is sensitive to CA1, but not to CA3, lesions⁴⁰. With regard to region-specificity, our findings for CA1 are in contrast with recent evidence in mice showing reduced ​nectin-3 expression in CA3 (refs 24, 25). This disparity may be attributed to differences in the animal species or stress procedures.

MMPs are a family of proteolytic enzymes that degrade components of the extracellular matrix and cleave specific cell-surface proteins⁴⁸, making them particularly suitable to sustain neural remodelling processes¹⁵. The degradation of cell adhesion molecules is one of the main mechanisms whereby MMPs affect neural plasticity^{9, 22} and the synapse-associated ​nectin-3 decrease suggested the potential involvement of proteolytic processing. ​Nectin-1 has been shown to undergo ectodomain shedding by alpha-secretase³²; however, the molecular players involved in ​nectin-3 shedding remained unknown.

We found that decreased ​nectin-3 expression in the hippocampal CA1, but not in the CA3, synaptoneurosomes of the stressed animals is accompanied by increased gelatinase activity. This suggested an increase in ​MMP-2 and/or ​MMP-9 activity, as these two MMPs are the most prominent gelatinases expressed in the brain. Our cell culture experiments also indicated that NMDA receptor stimulation leads to increased ​nectin-3 proteolytic cleavage that is ​MMP-9 dependent. The involvement of ​MMP-9 and not ​MMP-2 is consistent with a previous study showing that ​MMP-2 does not interact with ​nectin-3 (ref. 49). Furthermore, we provide direct evidence that ​MMP-9 cleaves recombinant ​nectin-3. Interestingly, ​MMP-9 cleaves several postsynaptic proteins involved in trans-synaptic adhesion via their interaction with presynaptic proteins. The list of such ​MMP-9 targets includes ​β-dystroglycan that binds to neurexins⁴² as well as ​neuroligin-1 also binding neurexins⁵⁰. Our findings are in line with previous reports implicating hippocampal ​MMP-9 in changes in dendritic spine morphology⁵¹ as well as in the cellular processes that contribute to a stressful learning task²⁰. Importantly, we show that intra-CA1 treatment with a specific ​MMP-9 inhibitor prevented the emergence of chronic stress-induced effects in social exploration and CA1-dependent cognition. Therefore, our results are consistent other findings that indicate a crucial role for extracellular proteolysis in the stress-induced behavioural alterations, with former studies highlighting the role of serine proteases, including the ​tissue-plasminogen activator¹² and ​neuropsin¹⁰.

Although deregulated social behaviour is a hallmark of many psychiatric disorders⁵², studies focusing on the link between chronic stress and psychopathology has mainly concentrated on studies in mood and cognition^{2, 7}, whereas the effects of stress on social behaviours are much less known. In agreement with our previous study⁸, we confirm here that chronic restraint stress for 21 days leads to clear alterations in the social domain, including reduced sociability, impaired social memory and increased aggressive behaviours. The hippocampus has been implicated in social behaviours both in rodents⁵³ and in humans⁵⁴. Consistent with our findings, social recognition in rats was disrupted by CA1 damage⁵⁵. However, although the effects of stress on sociability and social memory were rescued with ​nectin-3 OE, increased aggressive behaviours were not modified by this treatment. We have recently found that targeting the cell adhesion molecule ​neuroligin-2 expression or function in the hippocampus alters aggressive behaviour^{8, 56}, suggesting the involvement of the hippocampus in the regulation of aggression. However, it should be noted that those treatments were not confined to the CA1 area, which, on its own, might not modulate aggressive behaviours.

In summary, our findings identify a key role for ​MMP-9 proteolytic processing of ​nectin-3 in the hippocampal CA1, through a mechanism that engages NMDA receptors, among the processes leading to chronic stress-induced changes in social and cognitive behaviours. In addition to ​nectin-3, recently identified as potential mediators in stress-related disorders²⁵, our study highlights ​MMP-9 activity as a novel target for the treatment of stress-related neuropsychiatric disorders, in particular depression, which is typically characterized by deficits in the social and cognitive domains.

An Engineered Bacterium Is Able to Copy DNA that Contains Unnatural Genetic Letters

For all of recorded history, billions of years, the blueprint of life has been written with just four letters — adenine (A), thymine (T), cytosine (C) and guanine (G), the DNA subunits contained in all organisms. Now that has changed.

Scientists at Scripps Research Institute in La Jolla, California, have identified a pair of bases, known as d5SICS and dNaM, that are compatible with the enzymatic machinery that reproduces DNA. They succeeded in getting their new base pair to copy itself and be transcribed into RNA, which required that the enzymes that had evolved to use A, T, C and G bases recognize the "alien" base pair.

It's not quite like they are building alien lifeforms in the petri dish, but this is a huge step forward in our understanding how DNA functions and if it can be hijacked.

First life with 'alien' DNA

An engineered bacterium is able to copy DNA that contains unnatural genetic letters.

Ewen Callaway
07 May 2014

Synthorx

The addition of new letters to the 'alphabet of life' could enable biologists to vastly expand the range of proteins that they could synthesize.

For billions of years, the history of life has been written with just four letters — A, T, C and G, the labels given to the DNA subunits contained in all organisms. That alphabet has just grown longer, researchers announce, with the creation of a living cell that has two 'foreign' DNA building blocks in its genome.

Hailed as a breakthrough by other scientists, the work is a step towards the synthesis of cells able to churn out drugs and other useful molecules. It also raises the possibility that cells could one day be engineered without any of the four DNA bases used by all organisms on Earth.

“What we have now is a living cell that literally stores increased genetic information,” says Floyd Romesberg, a chemical biologist at the Scripps Research Institute in La Jolla, California, who led the 15-year effort. Their research appears online today in Nature1.

Each strand of the DNA's double helix has a backbone of sugar molecules and, attached to it, chemical subunits known as bases. There are four different bases: adenine (A), thymine (T), cytosine (C) and guanine (G). These letters represent the code for the amino-acid building blocks that make up proteins. The bases bind the two DNA strands together, with an A always bonding to a T on the opposite strand (and vice versa), and C and G doing likewise.

Nature Podcast
Ewen Callaway asked Floyd Romesberg how his alien DNA works.

Test-tube letters

Scientists first questioned whether life could store information using other chemical groups in the 1960s. But it wasn’t until 1989 that Steven Benner, then at the Swiss Federal Institute of Technology in Zurich, and his team coaxed modified forms of cytosine and guanine into DNA molecules. In test-tube reactions, strands made of these “funny letters”, as Benner calls them, copied themselves and encoded RNA and proteins2.

 

Nature special: Beyond divisions: The future of synthetic biology

The bases engineered by Romesberg’s team are more alien, bearing little chemical resemblance to the four natural ones, Benner says. In a 2008 paper, and in follow-up experiments, the group reported efforts to pair chemicals together from a list of 60 candidates and screen the 3,600 resulting combinations. They identified a pair of bases, known as d5SICS and dNaM, that looked promising3. In particular, the molecules had to be compatible with the enzymatic machinery that copies and translates DNA.

“We didn’t even think back then that we could move into an organism with this base pair,” says Denis Malyshev, a former graduate student in Romesberg’s lab who is first author of the new paper. Working with test-tube reactions, the scientists succeeded in getting their unnatural base pair to copy itself and be transcribed into RNA, which required the bases to be recognized by enzymes that had evolved to use A, T, C and G.

The first challenge to creating this alien life was to get cells to accept the foreign bases needed to maintain the molecule in DNA through repeated rounds of cell division, during which DNA is copied. The team engineered the bacterium Escherichia coli to express a gene from a diatom — a single-celled alga — encoding a protein that allowed the molecules to pass through the bacterium's membrane.

The scientists then created a short loop of DNA, called a plasmid, containing a single pair of the foreign bases, and inserted the whole thing into E. coli cells. With the diatom protein supplying a diet of foreign nucleotides, the plasmid was copied and passed on to dividing E. coli cells for nearly a week. When the supply of foreign nucleotides ran out, the bacteria replaced the foreign bases with natural ones.

Alien control

Malyshev sees the ability to control the uptake of foreign DNA bases as a safety measure that would prevent the survival of alien cells outside the lab, should they escape. But other researchers, including Benner, are trying to engineer cells that can make foreign bases from scratch, obviating the need for a feedstock.

Romesberg’s group is working on getting foreign DNA to encode proteins that contain amino acids other than the 20 that together make up nearly all natural proteins. Amino acids are encoded by 'codons' of three DNA letters apiece, so the addition of just two foreign DNA 'letters' would vastly expand a cell’s ability to encode new amino acids. “If you read a book that was written with four letters, you’re not going to be able to tell many interesting stories,” Romesberg says. “If you’re given more letters, you can invent new words, you can find new ways to use those words and you can probably tell more interesting stories.”

Potential uses of the technology include the incorporation of a toxic amino acid into a protein to ensure that it kills only cancer cells, and the development of glowing amino acids that could help scientists to track biological reactions under the microscope. Romesberg’s team has founded a company called Synthorx in San Diego, California, to commercialize the work.

Ross Thyer, a synthetic biologist at the University of Texas at Austin who co-authored a related News and Views article, says that the work is “a big leap forward in what we can do”. It should be possible to get the foreign DNA to encode new amino acids, he says.

“Many in the broader community thought that Floyd's result would be impossible,” says Benner, because chemical reactions involving DNA, such as replication, need to be exquisitely sensitive to avoid mutation.

The alien E. coli contains just a single pair of foreign DNA bases out of millions. But Benner sees no reason why a fully alien cell isn’t possible. “I don’t think there’s any limit,” he says. “If you go back and rerun evolution for four billion years, you could come up with a different genetic system.”

But creating a wholly synthetic organism would be a huge challenge. “A lot of times people will say you’ll make an organism completely out of your unnatural DNA,” says Romesberg. “That’s just not going to happen, because there are too many things that recognize DNA. It’s too integrated into every facet of a cell’s life.”

Nature doi:10.1038/nature.2014.15179

References

1. Malyshev, D. A. et al. Nature http://dx.doi.org/10.1038/nature13314 (2014).
2. Switzer, C., Moroney, S. E. & Benner, S. A. J. Am. Chem. Soc. 111, 8322–8323 (1989). Article | ChemPort
3. Leconte, A. M. et al. J. Am. Chem. Soc. 130, 2336–2343 (2008). Article | PubMed | ISI | ChemPort 

Related stories and links from nature.com

• Chemical biology: DNA's new alphabet 21 November 2012
• Enzymes grow artificial DNA 19 April 2012
• Genome-building from the bottom up 10 October 2010
• Augmenting the alphabet 30 August 2000
• Nature Special: Beyond divisions