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Marine and aquatic environmental fluctuations in temperature, salinity, oxygen, pH, and other factors over daily and seasonal time scales can be difficult to replicate in lab settings. As a result, experimental animal physiology has often used mean values and static conditions to research organismal processes. While informative for basic science, this approach is likely a severe over simplification of what is occuring in nature. In order to ensure lab-based approaches are meaningful to real-world scenarios, researchers need to consider the mangnitude and regularity of environmental fluctuations, as well as the presense of co-stressors. Here, I&#39;ll showcase some recent data illustrating examples where variability was and was not incorporated into experimental design effectively, as well as discuss technological advancements that can assist in bringing the real environment into the lab.

SEB Conference 2024

Incorporating Variability into Physiological Experiments

fluctuation

fish

temperature

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SEB’s Annual Conference is celebrated for creating the perfect composition of research, new discoveries and collaborative connections. Be creative and innovative by exchanging ideas and knowledge with masters of biology from all over the world.

Marine and aquatic environmental fluctuations in temperature, salinity, oxygen, pH, and other factors over daily and seasonal time scales can be difficult to replicate in lab settings. As a result, experimental animal physiology has often used mean values and static conditions to research organismal processes. While informative for basic science, this approach is likely a severe over simplification of what is occuring in nature. In order to ensure lab-based approaches are meaningful to real-world scenarios, researchers need to consider the mangnitude and regularity of environmental fluctuations, as well as the presense of co-stressors. Here, I'll showcase some recent data illustrating examples where variability was and was not incorporated into experimental design effectively, as well as discuss technological advancements that can assist in bringing the real environment into the lab.

technical paper

Blackwater systems, characterized by low pH, low ion concentrations, and a high concentration of dissolved organic carbon (DOC) are found globally. The most notable are the blackwaters of the Amazon River, one of the most biodiverse ecosystems worldwide. The harsh water chemistry is toxic to most freshwater fish, while endemic species including the dwarf cichlid, thrive due to the protective effects provided by DOC. Natural DOCs were collected in the Amazon basin: a DOC-rich, low pH, ion-poor “blackwater” from the Rio Negro and a water lower in DOC but higher in pH and ion concentration from the Rio Solimoes. Due to the complexity and heterogeneity of DOCs, three model compounds; tannic acid, bovine serum albumin and sodium dodecyl sulfate, with known chemical properties were chosen based on the criteria that they structurally resemble or functionally behave like certain chemical moieties of humic acid, a major component of natural DOCs. Dwarf cichlids were exposed to control water, or one of the five DOCs at pH 7 or pH 4 and net Na+ and Cl- fluxes, ammonia-N and urea-N excretion rates and gill paracellular permeability were measured. Our results demonstrate that the effects of the natural DOCs are related to their chemical structure and water pH. This contributes to understanding the properties of DOC that are interacting with the gill and how this interaction influences ion and water transport in acidic waters (NSERC, ADAPTA, CNPQ). 



The protective ionoregulatory effects of dissolved organic carbon (DOC) and model compounds in a native Amazonian fish, the dwarf cichlid (Apistogramma agassizii).

A muscle’s physiological properties determine its force-length (F-L) and force-velocity (F-V) relationships, and thus the fibre length and contraction speed at which maximal force and power can be produced. Here, we investigated the relationship between F-L and F-V relationships and in vivo muscle function in the guinea fowl (Numida Meleagris) main ankle extensor, the lateral gastrocnemius (LG) muscle during walking and running. To do so, we map each individual birds’ in vivo work loops to their individual force-length and force-velocity relationships. We hypothesize that muscles operate near optimal conditions for force and power production, respectively, during walking and running tasks to optimize efficiency during locomotion. We found that force development occurs entirely on the ascending limb of the F-L relationships curve in both walking and running and thus not at lengths that would maximize force output. LG muscles operate at low velocities at peak force and at optimal intermediate velocities during lower forces. These in vivo LG velocities are consistent with optimizing force and power according to the F-V relationship. Hence, a muscle’s operating length and velocity are not solely driven by efficiency. A potential explanation is that muscles act on the ascending limb of the F-L relationship to improve robustness against perturbations. We are currently performing model-based simulations of guinea fowl locomotion to explore how a muscle’s physiological properties affect its operating length and velocity. Our central hypothesis is that we will need to account for robustness against uncertainty in the simulations to capture the experimentally observed relationship.

Relating in vivo muscle dynamics to force-length-velocity relationships: from experiments to musculoskeletal models

Escape responses in schooling fish have been researched at length, though little is known regarding how responses may differ as schooling fish grow. To investigate this, we elicited escape responses in a subset of individuals from schools (consisting of 20 individuals) of different size ranges of shiner perch (Cymatogaster aggregata), by triggering a visual stimulus when 3-6 individuals were positioned l within view of the stimulus, with their resulting reaction to a threat ensuing further responses across the rest of school which could not see the stimulus because of an opaque barrier that was suspended above the water level. We analysed individual turning rates, latencies, and the propagation of the reaction among individuals. Our experiments revealed distinct patterns in response behaviour: as expected, turning rate decreased with fish size. Conversely, these smaller individuals exhibited longer latencies in initiating the escape response, suggesting a trade-off between speed of turning and reaction time across sizes. Furthermore, size was found to significantly affect the spread of information among individuals that detected the escape-triggering stimulus, with larger individuals facilitating faster propagation of the response throughout the school. These findings highlight the complex interplay between individual size and collective behaviour in fish schools, suggesting that size not only influences individual escape performance but also the efficiency of information transfer within groups. Our results contribute to a deeper understanding of the mechanisms underpinning escape responses in gregarious fishes, with implications for the study of predator-prey interactions and the evolutionary dynamics of schooling behaviour.

The effect of fish size on the escape responses of schooling shiner perch Cymatogaster aggregata


In recent years, research on spider silk has surged, focusing on its mechanical, medicinal, and commercial applications. Despite this, our understanding of the behavioral utilization of distinct silk types and their functions remains limited. Moreover, there is a notable gap in methodologies to comprehend the use of spider silk glands. This study focuses on Pholcus phalangioides, commonly known as Daddy long-legs spiders, known to build three dimensional irregular webs with unique behaviours. These include using sticky prey wraps to capture airborne prey, and viscous-sticky traps to ensnare ground-dwelling prey and dragline and bridging lines for locomotion. We employed multiple methodologies on live individuals as well as silk fibres collected for in-depth understanding of silk glands uses for these behaviours. Methods included spigot morphology analysis, gland dissections for morphology, light microscopy for identifying silk fibre types, high-speed video recording for behavioral study, shock freezing to observe spider behavior under Scanning Electron Microscopy and mechanical tests for single fibres to understand their role in ecological uses. Results suggest that a multi-methodology approach offers a comprehensive understanding of silk gland behavioral use. This study provides a unique perspective beyond the prevalent focus on orb weavers, offering insights into lesser-known species. The study highlights the limitations of single techniques, emphasizing the need for a diverse methodology. Additionally, it presents a protocol applicable to studying any spider's behavioral use of silk glands.


A multimethod approach to study the behavioral utilization of spider silk glands.

Songbirds use acoustic signals (songs) to defend territories and attract mates. Proximate and ultimate causes of variations in song structure have been extensively studied, but vocal amplitude is often overlooked and rarely measured. This lack of data is unfortunate because signal amplitude is crucial in communication. In bird song, it plays an important role not only for signal transmission, but also in territorial behaviours and mate choice. However, since a long time it has been assumed that differences in song amplitude between species are mainly due to variation in body size, with bigger species producing louder songs. We present comparative data from twelve common European songbird species that challenge this view. We found marked variation in song amplitude but this variation was not related to body size. Instead, song amplitude varied with small-scale social and environmental conditions (such as time of day or the presence of competitors). If body size affects amplitude at all, then the effect is so small that it is buried by variation due to these or other factors. Generally, our data highlight the importance of individual plasticity in song performance, and hint at a complex interplay between the constraints and functions of bird song.

What it takes to be heard: morphological and ecological correlates of bird song amplitude

Virtually all animal species live in fluctuating environments. Although physiological and behavioral responses to changes in environmental conditions influence fitness outcomes, individuals from a population do not always respond similarly. To understand how individuals cope with changing environments, and whether their responses can allow populations to evolutionary adapt to global change, we need to quantify among- and within-individual sources of phenotypic variation. Using hand-raised great tits (Parus major) kept in outdoor aviaries, we measured individual responses in circulating corticosterone and hemoglobin concentrations, metabolic rate, and breath rate to a simulated cold spell, and in vertical flight performance to simulated molt. We assessed reaction norms repeatedly, in winter and spring of their first year of life and again at these same times in their second year of life. We focused primarily on trait plasticity, quantified as the reaction norm slope, and tested for its variation at the population-level, among-individual level and within-individual level, between seasons and years. We found that individuals differed in their degree of plasticity in corticosterone, breath rate and flight performance. Furthermore, individuals showed ‘’plasticity of plasticity’’ in most traits across seasons and years. Finally, plasticity was moderately to highly repeatable in all traits investigated. These findings suggest that: a) individuals can tailor their plastic responses to the ecological context; b) there is significant standing phenotypic variation in plasticity that could buffer the population from environmental variation; c) plasticity may be genetically determined and if targeted by selection, could enable evolutionary adaptation to global change.

Physiological and performance reaction norms to ecologically relevant challenges: Within-individual plasticity is consistent and plastic at the same time.

Most species experience some amount of temperature variation at daily and seasonal timescales. Variations in temperature are expected to shape key biological traits of ectotherms, but also the breadth of phenotypes expressed within a population. Here, we combine meta-analytical and experimental data to uncover how daily temperature variation impacts phenotypic means and variation of ectothermic animals. We hypothesised that variation around warm temperatures would decrease growth, locomotion and reproductive success but increase phenotypic variation in these traits. We tested this hypothesis by first assembling the most comprehensive database to date, spanning 165 ectothermic species, and performing meta-analyses. Second, using black bream (Acanthopagrus butcheri) as an experimental model, we assessed the upper thermal limits of fish following acclimation to constant or variable daily temperatures, and compared these data to the thermal limits of field-acclimated fish. Our meta-analysis revealed that daily temperature variation increases upper thermal limits across ectotherms, but reproductive traits were compromised. Surprisingly, phenotypic variation did not increase in response to daily temperature variations. Experimental data showed that upper thermal limits of field-acclimated fish does not mirror that of fish acclimated to a constant temperature under lab conditions. Taken together, these findings demonstrate the importance of considering how daily temperature variation impacts key biological traits. Experiments that expose ectotherms to constant temperatures can over-simplify physiological responses to temperature variation that could misguide conservation and management actions.

Temperature variation can be a blessing and a curse for ectothermic animals

In our final year Immunology and Virology module at NTU, we use an employer informed research question as the topic. Six BSc (Honours) Biomedical Science students work together as a team to create a poster on this broad virology based question: they are required to decide as a team which content is delivered in a twelve minute presentation on this topic, relate this to an industry application /enterprise (either already established or a business idea), identify a career opportunity in that field and relate this/these jobs to their skill set, reflecting on how the course has prepared them for this role and career trajectory. Students are required to identify each team member’s contributions, reflect on their progress and approach regarding teamwork, time management, communication, etc. and identify strengths and weaknesses. 

To ensure fairness and equal contributions, students are required to detail each member's efforts, have the opportunity to flag any professional behaviour issues encountered. In this safe HE setting, the module leader provides constructive advice and if needed intervention. 

We iclude stdeunt feedback who have taken this assessment confirming that the positive impact of this activity on their employment.

Working as a team on an employer informed research question in final year BSc (H) Biomedical Science 

In the recently introduced integrated foundation year for Nottingham Trent University’s undergraduate bioscience courses, students study a range of practical and transferrable skills which are assessed through a professional practice and skills module. The team lab research project is designed to foster the development of a range of professional and work-relevant skills such as problem-solving, use of professional communication platforms for team communication and document sharing, as well as awareness of health and safety and time management. Module assessment focusses on a team presentations of the project and evidence of laboratory and transferrable skills through a skills portfolio.
The project, designed to introduce the students to the scientific method for investigating research questions in the laboratory, was contextualised to a real-world scenario through collaboration with an external industrial partner. In this case, the project was designed for students to design and deliver a lab-based project to explore bacterial resistance to antibiotics and the claims of natural products to have anti-microbial properties. The external partner, a biomedical scientist within the UK National Health Service specialising in microbiology, set the context of the project through reference to the importance of identification of antibiotic resistance within the NHS. Not only this but they were able to give students one-to-one guidance in the laboratory to support the development of their practical skills, and feedback on their team presentation of the project.
Students will reflect on their experience of the benefits and challenges of working on the team project as part of the presentation. 


Developing professional skills and attributes in foundation biology students

Mechanisms of habituation for exogenous auxin-independent proliferation of tobacco cells

Pavel Jelíneka, Karel Müllerb, Petre I. Dobrevb, Roberta Vaculíkováb, Zuzana Vondrákováb, Barbora Svobodováb, Julie Talpováb, Anita Bírošíkováb, Vojtěch Čermáka, Adéla Přibylováa, Eliška Kobercováa, Lukáš Fischera, Jan Petrášeka

a Department of Experimental Plant Biology, Faculty of Science, Charles University, Viničná 5, 128 44 Prague 2, Czech Republic
b Institute of Experimental Botany, the Academy of Sciences of the Czech Republic, Rozvojová 263,165 02 Prague 6, Czech Republic 

Auxins are essential phytohormones that are involved in the regulation of the plant cell cycle. In vitro cultured plant cells require exogenous auxin for proliferation. However, they may acquire the ability to survive and proliferate in auxin-deprived media through the phenomenon of habituation. In this study, we sought to understand the mechanisms behind the auxin habituation of two tobacco cell lines. We compared two pairs of lines derived from two tobacco cultivars; BY-2 x BY-2H and VBI-0 x VBI-2b. In each pair, we had one line dependent on exogenous auxin (BY-2, VBI-0) and one habituated (BY-2H, VBI-2b). To describe the changes between auxin-dependent and habituated lines, we used a set of approaches such as transcriptome analysis (RNAseq), genomic sequence analysis, and phytohormonome analysis. By combining these approaches, we were able to show that the proliferation of both habituated lines is still dependent on endogenously synthesised auxin. However, each culture possesses different mechanisms. The BY-2H line contains an increased number of copies of the auxin receptor TIR1 in its genome resulting in 200-fold higher expression. We were able to localise the copies of TIR1 on different chromosomes. We were also able to detect higher levels of endogenous IAA in BY-2H compared to the BY-2 line. On the other hand, VBI-2b habituation hypothetically depends on a decreased expression of EMF2, a component of the PRC2 complex, which probably leads to an increased expression of a set of MADS-box transcription factors. Our focus now is to provide functional evidence for both hypotheses.

Supported by CSF grant n. 23-07813S and GAUK n. 414022


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The protective ionoregulatory effects of dissolved organic carbon (DOC) and model compounds in a native Amazonian fish, the dwarf cichlid (Apistogramma agassizii).

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Next from SEB Conference 2024

The protective ionoregulatory effects of dissolved organic carbon (DOC) and model compounds in a native Amazonian fish, the dwarf cichlid (Apistogramma agassizii).

Similar lecture

The ups and downs of incorporating environmentally relevant variability in laboratory experiments

Stay up to date with the latest Underline news!

PRESENTATIONS

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COMPANY

RESOURCES

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