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Making sense of stress in the wild

Imagine leaning forward over the edge of a precipice. Lurching back to safety, you picture the forest hundreds of metres below. Is your heart racing? Are your palms sweating? Our body’s stress response to an ever-changing environment enables us to survive and flourish.

Physiologists play a crucial role in developing our understanding of the mechanisms involved. To highlight the exciting work that they do, our 2017 theme is ‘Making Sense of Stress’. Follow the conversation on Twitter using #YearOfStress.

Launching the theme will be Dr Kimberley Bennett’s talk, ‘Making sense of stress in the wild’, at the Association for Science Education’s (ASE’s) Annual Conference on 6 January 2017. Read a teaser to her talk below!

Coping with stress is a major issue in modern society, but it’s easy to forget that wildlife experiences stress too. Without enough water, plants wilt and die and whole crops fail; without the right habitat, a small population of rare animals dwindles and dies out, causing extinction of the species; a whole coral reef bleaches when the water temperature gets too high, causing catastrophe for the ecosystem, and massively increasing flooding risk for people living by the coast. We really need to pay attention to stress in the wild because the consequences can herald disaster.

Stress is the biological response to a major challenge, whether it’s at the whole organism or cell level. A gazelle in the Serengeti chased by a lion experiences the same stress responses that we do – a surge of adrenaline and cortisol that cause increased heart rate and blood pressure and a release of glucose. These changes make sure there is enough fuel and oxygen to cope with increased demand at the tissue and cell levels. Sudden change or mismatch in the supply of oxygen and fuel leads to increased production of reactive molecules called ‘free radicals’ that can damage cells. If the temperature gets too hot too fast or if the acidity of the cell changes too much, proteins (the molecules that catalyse reactions, transport substances and provide structure) can fall apart or unravel. So cells have to increase their defence mechanisms too. Cellular defences include antioxidants that mop up the free radicals, and heat shock proteins, which refold damaged proteins and stop them forming a sticky mess inside the cell.

The old adage that what doesn’t kill you makes you stronger is often true: short term ‘good stress’ builds up these defences and makes organisms better able to deal with stress later on. However, sometimes defences can be overwhelmed or can’t be maintained for long periods. The organism then experiences the same sorts of problems as people under chronic stress: lower immunity, altered metabolism, anxiety and tissue damage (like ulcers). In wildlife, this can have major consequences for breeding success or even survival. By affecting whether organisms survive and thrive, stress dictates which individuals contribute to the next generation. Stress shapes population dynamics, lifestyle and adaptations, and is therefore a powerful agent of natural selection.

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I work on seals, top marine predators that are used to stress as a normal part of their existence. Their individual and population level health is an indicator of ecosystem health. Seals are air breathing mammals that feed underwater, but need to come to the surface to breathe, and to come ashore to rest, breed and moult. Diving on a single breath hold means they need to conserve oxygen; to do this, blood flow is restricted mostly to the heart and brain, so that other tissues may experience free radical production while oxygen levels are low. On land, seals need to fast, often while they are doing energy-demanding activities i.e. shedding and replacing hair, producing milk, defending pups or territory, or undergoing rapid development. Injury and infection can occur from skirmishes or trampling. Seals may have to reduce their defences to deal with all these demands on their energy when food is not available. In addition to their ‘lifestyle stressors’, seals face stress from competition for access to fish, disturbance on haul out or displacement from foraging grounds as a result of human activity, and the accumulation of contaminants in their blubber.

We need to understand natural and man-made causes of stress in wild populations, distinguish good stress from bad stress, and understand how multiple stressors at the same time can create problems. That means we have to have effective tools to measure stress and its consequences in organisms that can’t tell us how they feel. But can we measure stress responses in wildlife? What do they mean in context? And can they help in managing stress in the wild?

I will address all these questions and more at the ASE’s Annual Conference on Friday 6 January 2017, as part of the annual Biology in the Real World (#BitRW) lecture series. Please drop by the Knight Building, LT 135, at the University of Reading, at 1.30pm to find out more!

Dr Kimberley Bennett, Abertay University

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Danger! High Voltage!

Standing between school students and their weekends was always going to be a tough gig, but it was #PhysiologyFriday and we were determined to put on a show. Together with Orla and Ioannis, PhD students from the University of Reading, I arrived at East Ham Town Hall and was greeted by over 200 excited students and their teachers.

Our mission was to deliver an engaging session around electricity and pass on an innovative human-human interface educational resource that the students had won as part of a Physiological Society competition, The Science of Life 2016. After a brief introduction to human electricity and how we can detect it, we took the students on a journey from the first electroencephalogram (EEG) to the potential of neuroprosthetics in treating paralysis. We then tackled a live demonstration of the kit and the students were intrigued to see the interface in action.

 

Surely, one human cannot control another’s arm using only their brain activity? The wonders of neuroscience said it should be possible, but we sensed some doubters in the audience. Using the human-human interface Ioannis was going to harness his brain power to move Orla’s arm against her wishes……

Thankfully the demonstration was up to scratch with the students requesting several encores, and even asking for the voltage to be turned up! Normally the complexities of electrophysiology remain in the lab due to expensive and static equipment, however this kit gave us the ability to be mobile and take the dark art of electrophysiology to young minds keen to explore neuroscience. It was well worth it. The students left school for the weekend with a sense of excitement and thirst to learn more. Orla, Ioannis and I left East Ham Town Hall feeling that we had sparked the interest of the next generation of physiologists.

Dr Mark Dallas

@drmarkdallas

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Myth-busting: Do heart cells really have their own ‘pulse’?

By Dr. Andy James, University of Bristol

The comedian, Jake Yapp, has been presenting on BBC Radio 4’s website a series of short comic histories of science entitled, “Everything We’ve Ever Known About …” Informative and entertaining, these are invariably funny. A recent edition concerned the heart, neatly covering the development of our understanding of heart function through the millennia, from the ancient Egyptians to the present day – where Jake concludes by commenting that if you isolate a heart muscle cell it will have its own pulse. Actually, strictly speaking, a pulse is the increase in pressure within the arteries as a result of beating of the heart. ‘Rhythm’ is a better word.

http://www.bbc.co.uk/programmes/p04bwbxl/player

It’s a commonly held misconception, even amongst specialists in cell biology, that all heart muscle cells have their own rhythm that synchronises when the cells couple together (meaning they join and allow electricity to flow between them). Beautiful as this idea is, it simply isn’t true. The heart’s rhythm originates as electrical activity in a specialised area of the heart known as the sinus node- the heart’s physiological pacemaker. The heart muscle cells of the sinus node generate rhythmic electrical activity so that, when isolated, they contract regularly.

In the intact heart, a conduction system ensures that the electrical activity is passed to the chambers to produce a well-coordinated contraction. Many of the cells of the conduction system would also beat when isolated. But the cells from the walls of the chambers that produce the pressure to – as the English anatomist, William Harvey, would have it – propel the blood, do not. Chamber cells only need to receive the signal, but not generate it themselves.

The persuasive and attractive idea that all heart muscle cells show their own rhythm may have arisen from attempts to maintain heart muscle cells for several days in a petri dish. Heart muscle cells from adult mammalian hearts do not generally multiply under these conditions and while it is possible to keep the cells alive for a short time, after a few days, the cells die. However, cells from embryonic, or even neonatal, hearts do multiply in culture. They often show rhythmic contractions and, yes, they synchronise once they form connections with their neighbours.

So do heart cells really have their own pulse? It depends! Cells from adult hearts are programmed to behave in a fixed way- as a pacemaker cell, a conducting cell or a contractile cell from a chamber wall. Cells from immature hearts, on the other hand, retain the ability to change their properties and tend to develop pacemaker-like function in the petri dish.

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Non-invasive imaging of transient biomolecular machines in vivo

pic2(This article was originally published in Physiology News 104, page 24-26)

While homeostasis is considered a steady state in physiology, it is by no means static. Instead, it is the sum of a concerted effect of sub-cellular events driven by molecular machines. These machines range from large enzymes to reactive small molecules, all of which exert an effect on the cell to alter its function for better or for worse.

Indeed, the over- or under-activity of these molecular machines can result in disease or injury (e.g. reactive oxygen species (ROS) and inflammation), or in an intended response to applied therapy (e.g. tumor casapase-3 (apoptosis) activation following radiation or chemotherapy). These active biomolecules rarely work in isolation, but are rather parts of integral networks within which any single sub-cellular machine is only transiently active or transiently present, often rapidly giving way to its successor in the signaling chain. This transiency makes these biomolecular machines elusive to analysis by traditional ex vivo methods in the context of the living organism.

However, measuring the activity of these molecules and overcoming the difficulty of target transiency can enable both the detection of disease prior to outward signs and symptoms, and the assessment of the success or failure of therapy prior to disease progression (Aboagye, 2006; Aboagye, 2010). Molecular imaging is a powerful tool for assessing a target biomolecular machine non-invasively and longitudinally over an entire volume of interest in living subjects (James & Gambhir, 2012). The ability to apply molecular imaging to interrogate these highly transient sub-cellular functions of fundamental importance in living subjects is provided by new chemistries for the design of activatable molecular imaging probes over a range of pre-clinical (i.e. fluorescent) to clinically-relevant (PET, MRI) imaging modalities.image1

Reactive oxygen species (ROS) are highly reactive oxygen-centered biomolecules underlying a broad range of diseases (Winterbourn, 2008). In the context of injury, ROS are very early effectors of the tissue response to the source of harm. Novel optical nanoprobes have been developed from semiconducting polymers that can sensitively report on the presence of specific ROS in living mice (Pu et al., 2013; Pu et al., 2014; Shuhendler et al., 2014). Our work with the nanoprobes has enabled the real-time detection of the very early response of tissues to mechanical or chemical injury in real-time and in living subjects.

The very early response to tissue injury (i.e. within the first 30 min) have been imaged with evidence pointing to the mast cell component of the innate immune system as being responsible for the very early ROS signaling (Fig. 1; previously unpublished data). Additionally, a dual optical channel probe has been developed capable of the real-time differentiation of hydrogen peroxide versus peroxynitrite mediated drug-induced hepatotoxicity in living subjects (Shuhendler et al., 2014).

‘The molecular imaging of these key sub-cellular machines is sure to continue to provide an unprecedented level of interrogation of physiology to both enhance our investigations of health and disease, and improve clinical outcomes’

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Caspase-3 is a cysteine-aspartate protease whose activation signals the committal of the cell to die through apoptosis, a common death pathway induced by a variety of cancer chemotherapeutics and radiation. We designed a modular probe to undergo bioorthogonal macrocyclization upon activation by caspase-3, with the macrocycles self-assembling into nanoparticles in situ in tumor tissue (Shen et al., 2013; Shuhendler et al., 2015; Ye et al., 2014a; Ye et al., 2014b). Since caspase-3 activity correlated with the degree of probe retention in dying tumor tissue, this molecular imaging strategy was applicable to imaging through fluorescence, PET, and MRI. Importantly, this self-assembling molecular imaging probe was able to predict the degree of therapy outcome (i.e. the degree of tumor treatment effect) in mice following a single imaging session 48 hours after a single dose of therapy (Fig. 2).

Enhanced contrast production following therapy was not detected using a non-activatable control analog, illustrating the utility of our bioorthogonal self-assembly probe design for therapy response monitoring. Multiple sub-cellular markers for apoptosis have been identified each with molecular imaging probes targeting these sub-cellular changes, including annexin-V detecting phosphatidyl serine flipping from the inner to the outer cell membrane, ML-10 detecting depolarization of the cellular membrane, and fluorodeoxyglucose detecting the loss of metabolic function (i.e. reduced glucose utilization) (Witney et al., 2015). When these targets and respective probes were compared to the macrocyclization probe responding to caspase-3 activation, only the macrocyclizing probe accurately reported tumor response to therapy in living subjects as confirmed by ex vivo measurement of apoptosis (Witney et al., 2015). The ability of the macrocyclization agent to directly probe the effector of apoptotic cell death (i.e. caspase-3) and to significantly amplify the enzyme activity signal by being a substrate of this molecular machine may account for its performance in vivo.

The ability to investigate transient biomolecular machines in living subjects by molecular imaging can provide non-invasive, longitudinal, and predictive data about sub-cellular physiology within the intact organism. The molecular imaging of these key sub-cellular machines is sure to continue to provide an unprecedented level of interrogation of physiology to both enhance our investigations of health and disease, and improve clinical outcomes.

References

Aboagye EO (2006). Imaging in drug development. Clin Adv Hematol Oncol 4, 902–904

Aboagye EO (2010). The future of imaging: developing the tools for monitoring response to therapy in oncology: the 2009 Sir James MacKenzie Davidson Memorial lecture. Br J Radiol 83, 814–822

James ML & Gambhir SS (2012). A molecular imaging primer: modalities, imaging agents, and applications. Physiol Rev 92, 897–965

Pu K, Shuhendler AJ, Jokerst JV, Mei J, Gambhir SS, Bao Z & Rao J (2014). Semiconducting polymer nanoparticles as photoacoustic molecular imaging probes in living mice. Nat Nanotechnol 9, 233–239

Pu K, Shuhendler AJ & Rao J (2013). Semiconducting polymer nanoprobe for in vivo imaging of reactive oxygen and nitrogen species. Angew Chem Int Ed Engl 52, 10325–10329

Shen B, Jeon J, Palner M, Ye D, Shuhendler A, Chin FT & Rao J (2013). Positron emission tomography imaging of drug-induced tumor apoptosis with a caspase-triggered nanoaggregation probe. Angew Chem Int Ed Engl 52, 10511–10514

Shuhendler AJ, Pu K, Cui L, Uetrecht JP & Rao J (2014). Real-time imaging of oxidative and nitrosative stress in the liver of live animals for drug-toxicity testing. Nat Biotechnol 32, 373–380

Shuhendler AJ, Ye D, Brewer KD, Bazalova-Carter M, Lee KH, Kempen P, Dane Wittrup K, Graves EE, Rutt B & Rao J (2015). Molecular Magnetic Resonance Imaging of Tumor Response to Therapy. Sci Rep 5, 14759

Winterbourn CC (2008). Reconciling the chemistry and biology of reactive oxygen species. Nat Chem Biol 4, 278–286

Witney TH, Hoehne A, Reeves RE, Ilovich O, Namavari M, Shen B, Chin FT, Rao J & Gambhir SS (2015). A Systematic Comparison of 18F-C-SNAT to Established Radiotracer Imaging Agents for the Detection of Tumor Response to Treatment. Clin Cancer Res 21, 3896–3905

Ye D, Shuhendler AJ, Cui L, Tong L, Tee SS, Tikhomirov G, Felsher DW & Rao J (2014a). Bioorthogonal cyclization-mediated in situ self-assembly of small-molecule probes for imaging caspase activity in vivo. Nat Chem 6, 519–526

Ye D, Shuhendler AJ, Pandit P, Brewer KD, Tee SS, Cui L, Tikhomirov G, Rutt B & Rao J (2014b). Caspase-responsive smart gadolinium-based contrast agent for magnetic resonance imaging of drug-induced apoptosis. Chem Sci 4, 3845–3852pic2

Measuring a moving target: Symposium on Hormone Sensing

What do puberty, doping in athletics, and the meat industry have in common? The answer is hormones. Secreted into the blood by specialized organs and tissues, hormones communicate a bewildering array of signals to a myriad of target sites.

Two weeks ago, The Physiological Society brought together experts in physiology, endocrinology, chemistry, physics and engineering, to discuss how to produce a new generation of tools and methods for detecting hormones inside our bodies

“The biggest hurdle facing basic and clinical endocrinologists is how we can measure hormones inside the body,” said one of the symposium organizers, Timothy Wells, Senior Lecturer in Neuroscience at Cardiff University.

Addressed by UK and international experts in hormone-receptor interactions, light-based sensing and nanocarbon-based sensing, the Society’s symposium explored how these molecular interactions could be exploited to quantify the dynamic changes in circulating hormone levels.

 

Cutting edge work featured

One of the potential approaches was presented by Frank Vollmer and his colleagues from the Max Planck Institute for the Science of Light. He and his team are attempting to reach the ultimate limit of detection, by sensing single molecule interactions and the resultant changes in three-dimensional shape. Their new technique, which was published this week in Nature Photonics, may enable the detection of individual hormone molecules.

Thus, the day of talks highlighted just how far we’ve come since Ernest Starling coined the term hormone in 1905.

The event, titled “Novel approaches to Hormone Sensing, The Inaugural Bayliss-Starling Symposium,” was part of the society’s H3 symposia. The next symposium will be held on 15 November about one of the biggest discoveries in biotech, CRISPR. Visit our website for more info: http://www.physoc.org/crispr/