Belgium

Calcification is a widespread process in marine organisms and depends on tight control of pH and carbonate chemistry. Sea urchin larvae have been extensively used to study the transport mechanisms underlying the formation of an elaborate calcitic endoskeleton. The generation of this complex mineral structure is initiated in primary mesenchyme cells (PMCs), where amorphous calcium carbonate (ACC) is formed in intracellular vesicles. The intracellular precipitation of ACC requires a specific chemical environment in which pH plays a critical role. However, the pH dynamics within these vesicles and their role in carbon transport are still poorly understood. We developed in vivo methods to characterize vesicular pH in PMCs of Stongylocentrotus purpuratuslarvae to unravel the underlying mechanisms of proton and carbon transport. Inverted confocal microscopy in combination with the ratiometric fluorescent dye SNARF-1 allowed us to accurately measure the pH of intracellular vesicles in PMCs. Simultaneous tracking of endocytosed calcein accompanied by pH recordings allowed a high spatial and temporal resolution of pH dynamics in Ca2+  rich vesicles. This system provides a unique model for live- tissue imaging and sheds light on pH regulatory mechanisms in intracellular calcifying vesicles. Preliminary results showed the presence of acidic and alkaline vesicle populations in PMCs, which can regulate their luminal pH to maintain a favorable chemical environment forCaCO3  formation. Future studies are planned to understand how the larvae can cope with environmental stressors such as ocean acidification, where the decrease in seawater pH is accompanied by a decrease in carbonate ion concentration.

SEB Conference 2025

The calcification process in sea urchin larvae: insights into pH regulation mechanisms in intracellular vesicles of primary mesenchyme cells (PMCs)

intracellular vesicles

ph regulation

calcification

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Understanding the mechanisms that mediate fish responses to the climate change stressors of high temperature and hypoxia is crucial for identifying environmental thresholds and implementing effective conservation and management strategies. Studies have examined the effects of hypoxia on fish upper thermal tolerance and cardiorespiratory physiology. In contrast, in this study, we fitted 10o C-acclimated Atlantic salmon (Salmo salar) with blood flow probes, placed them into respirometers, and measured their cardiorespiratory response to decreasing water oxygen levels (5% air sat. every 15 minutes) at 10o C and after temperature was first increased to 14 and 18o C overnight. No clear critical oxygen threshold (Pcrit ) was identified at 10o C, and the % air sat. at which the salmon went bradycardic was 42.4. Whereas values for Pcrit  and the oxygen level at which bradycardia were initiated in 14 and 18o C fish were 52.1 vs. 57.2 and 61.0 vs. 77.1, respectively. Fish at 10o C lost equilibrium (LOE) at 21.0 % air sat. vs. 27.5 at 14o C and 41.9 at 18o C. Interestingly, their LOE was inversely related to heart rate (fH , 29.1, 36.6 and 42.8 beats min-1  at 10, 14 and 18o C, respectively), and it was the inability of salmon at 18o C to increase stroke volume when fH  fell, and to increase blood oxygen extraction (MO2 /Q; which was much higher in normoxia at 18o C), that limited hypoxia tolerance. These data are consistent with recent studies which show that higher fHs  are detrimental to hypoxia tolerance, whereas the heart’s pumping capacity and increases in MO2 /Q are critical.

Constraints on Cardiac Pumping Capacity and Tissue Oxygen Extraction, Not Heart Rate, Limit Hypoxia Tolerance of Salmon at Warm Temperatures

In tiny biological systems, the storage and release of elastic energy during ultrafast movements is modulated by how structural materials deform in space and time. Trap-jaw ants deform their springy head capsules, a geometrically complex 3D structure, to power mandible strikes ranking among the fastest biological movements known. Visualizing deformation of these tiny, curved exoskeletons is challenging, requiring imaging tools to resolve micron-scale deformations across millimeter-scale structures (~2 mm, anterior–posterior) of strikes occurring in ~70 µs. To address this, we developed an ultra-high-speed Fourier light field microscope (FLFM) optimized for capturing real-time deformation of the trap-jaw ant head capsule with high spatial and temporal resolution. Our FLFM integrates an ultra-fast sensor (Photron E980S, achieves 1 MHz+ fps) with 3-6 array lenses to capture unique perspectives of the ant head capsule at 100,000 frames per second. White paint speckled on the ant heads provided optical features tracked using optical flow with sub-pixel precision in each perspective. A calibrated model of the optical system was then used to compute the 3D motion of the dots from the flow vectors. We resolved z-axis (in and out of the ant head capsule) recoil displacements averaging 33.3 ± 11.4 µm with a standard error of 2.5 μm— the first time such dynamic and distributed elastic recoil has been fully quantified in an ultrafast biological spring-latch system. This modular hardware and software approach can be adapted to other small-scale, high-speed systems, offering a new workflow for integrating high-speed imaging with 3D deformation analysis.

Ultrafast fourier light-field microscopy reveals 3D recoil of tiny elastic structure

The ability to live in trees is a ubiquitous adaptation in mammals, but the specific mechanisms underlying the evolution of arboreal locomotion are variable among groups. For example, primates evolved in an arboreal context and acquired unique traits such as opposable first digits and nails instead of claws, which are still shared by the majority of extant species. However, the initial function of these specificities are still unclear. Studying the biomechanics of movements and forces exerted by the autopods during arboreal locomotion in primates and other arboreal mammals provides insight into the impact of the arboreal lifestyle on their morpho-functional adaptations. Here we developed an innovative force sensor technology that mimics the morpho-functional adaptation of distal pads of arboreal mammals. This patented technology measures the direction and intensity of contact forces, with a spatially resolved and dynamic resolution. Its design enables to make sensors of variable shapes and sizes and can be used in natural environments. We designed a version of this system in the shape of a branch to study the biomechanics of grasping during arboreal locomotion in various mammals in zoological parks. These data will notably allow to elucidate whether nails of primates allow them to apply more force compared to the claws of other mammals, by quantifying for the first time the variation of forces exerted by the different anatomical regions of the phalanges in those species. This technology and experimental methodology have multiple applications, ranging from biomechanics to haptics, health and robotics.

An innovative force sensor technology mimicking distal pads' functional morphology to study the evolution of locomotion in arboreal mammals

A bird’s beak has several functions, from feeding and drinking to singing, leading to a compromise in morphology. Adaptive evolution in relation to feeding ecology is an important driver in beak shape and size. To crack hard seeds, several species evolved a strong beak. Singing on the other hand is an important factor in sexual selection. Females usually prefer more complex songs and high trill rates, reached by fast beak movements. This leads to a trade-off between strong and fast beaks. The biomechanical principles underlying this force-velocity trade-off are, however, still unclear. In a first step to fill this knowledge gap, the anatomy of the jaw muscles was compared for a series of estrildid and fringillid finches varying in beak morphology and singing capacities. Using manual dissections, muscle fascicle lengths were measured and physiological cross-sectional area (PCSA) was calculated. Fascicle length and PCSA indicate the specialization of a muscle in either speed or force. As expected, the closing muscles are well developed in the strong biting species, but the imbalance with the opener muscles also increases. Conversely, for fast singers, a smaller imbalance between opening and closing is expected to facilitate singing. The weak correlations between metrics for singing performance and muscle architecture indicate that other factors are more important in determining song trill rates.

Need for speed: are jaw muscle fascicle length and PCSA different in species varying in singing capacities?

Evidence is becoming overwhelming that human activities are driving environmental alterations from multiple perspectives, including chemical, ecological, and physical changes. These environmental alterations can have profound impacts on animals, particularly from a physiological standpoint. However, the cognitive system of animals is highly susceptible to both external environmental influences and internal physiological states, suggesting that cognition could also be significantly affected by these alterations. Here, I present a recent set of studies on freshwater teleost fish, the most endangered group of vertebrates, exploring whether and how anthropogenic stressors affect cognition. Various chemical alterations in aquatic environments induce neurotoxicity, directly impairing cognitive function. Microplastics, although indirectly, also impair cognitive functioning. Physical disturbances, such as light pollution, influence cognitive phenotypes, especially during early development. Lastly, ecological changes, such as the introduction of non-native species, affect cognitive variability through hybridization. All these findings converge on the same conclusion: although more challenging to observe, most anthropogenic stressors significantly impair cognition in fish. These cognitive disruptions should be recognized as major contributors to fitness reduction in altered environments, as cognition governs most adaptive behavioral responses. Considering cognition in future studies will likely reveal that the impact of human activities on animals is greater than previously believed.

Fish minds in troubled waters

p53 is a tumour suppressor conserved from sea anemones, worms, insects to vertebrates, but is not conserved in plants. Upon limited DNA-damage, p53 induces cell cycle arrest activating the DNA repair machinery. Under excessive DNA-damage p53 induces cell death. Mutations inTP53occur in more than 50% of human cancers, indicating its importance in cancer suppression. 
Traditional p53 research involves the use of patient biopsies, (transgenic) animal models, cancer cell cultures, or tedious and expensive cell transfection experiments. While these methods collectively boosted our understanding of how p53 operates and led to identification of effective cancer treatment therapies, each approach has its own practical and ethical considerations that often limit p53 research to specialists’ groups in high-income countries.  
We have developed an alternative to animal-experimentation for the study of p53 using Arabidopsis expressing p53 under a conditional expression promoter.  
In this talk I will introduce our results demonstrating that p53 expression in plants is suitable for investigating central mechanisms of p53 regulation and function. We believe that implementing plants for p53 research will reduce animal experimentation, bureaucratic burden, experimental costs while broadening its study to under-represented countries. Importantly, this has the potential to identify novel aspects of p53 function with applications in cancer prevention and treatment.

Designing plant systems for a more efficient and sustainable investigation of p53 activity

Opportunities for increased rate of progress in the use of phenotyping have greatly been enhanced by improvements in machine-learning (ML) models for both image analytics and for time-series type analyses. These two classes of ML problems have great utility in plant phenotyping where solutions have been constrained by difficulties in generating sufficient training data. In image analytics, training data is expensive as annotation of images is labour-intensive. For this class of ML problems, transfer learning and domain adaptation concepts permit synthetic images to be converted to ‘real’ images at low cost, with these images being used to enhance the ML algorithms to identify plant parts and organs. Synthetic data from bio-physical models also has a role in ML. In our work, we use bio-physical models to train ML algorithms to invert satellite reflectance data and estimate crop traits such as crop leaf area index and biomass over time more efficiently than do benchmark naïve inversions based on brute-force parameterisation. In designing further improvements to ML models, we also apply techniques referred to as ‘physics-informed machine learning’ (PIML) which aim to further constrain the ML inversion models to behave within known biological relationships.

Examples of using synthetic data from biophysical and structural models to train algorithms for plant and crop phenotyping

During digestion, snakes experience large cardiovascular changes to accommodate increased metabolic demands, including tachycardia driven by vagal tone withdrawal and non-adrenergic, non-cholinergic factors. Stroke volume doubles without cardiac growth, while blood pressure remains stable, likely due to reduced peripheral resistance. However, specific modifications transitions in arterial resistance from fasting to digestion remain unclear. Endothelial-derived catecholamines, 6-cyanodopamine and 6-nitrodopamine, are potent endogenous positive chronotropic factors in isolated rat heart. Additionally, 6-nitrodopamine is released by the corn snake’s aorta and promotes vasodilation in aortic rings, but the effects of 6-cyanodopamine in snakes are unknown. This study aimed to analyze the adrenergic regulation of the mesenteric arteries of fasting (2–3 weeks) and digesting (24–48 hours) pythons. Second order arteries were mounted in a wire myograph, and concentration-response curves for adrenaline and noradrenaline were performed, with or without pre-incubation with 100 nM 6-cyanodopamine. We observed a significant reduction in Emax only for adrenaline in digesting snakes compared to fasting ones. This finding suggests that different mechanisms regulate the vascular response to adrenaline and noradrenaline in fasting and digesting snakes. Additionally, 6-cyanodopamine enhanced the maximum response to noradrenaline exclusively in fasting snakes. The absence of this effect in digesting snakes is likely related to the same mechanism underlying the altered response to adrenaline. These findings indicate that arterial responses to catecholamines differ between fasting and digestion, suggesting metabolic state-dependent vascular regulation.

Different vascular responses to catecholamines in fasting and digesting snakes

Biological invasions can cause deleterious impacts for biodiversity, ecosystem health and the economy, yet how individual traits impact invasive population spread and range expansion remain unclear. In this study we investigated the morphological, behavioural and physiological phenotypes of round goby (Neogobius melanostomus), a highly invasive fish, from populations at the edge and the core of an invasion gradient along the Trent Severn River Waterway, in Canada. Our edge (invasion front) population was significantly male biased, with larger and more reproductive fish compared to the fish at the core, an area established over 20 years ago. Using 68 size-matched males from each population we conducted repeated behavioural assays to assess boldness, and activity. In addition, thermal tolerance (CTmax) tests were conducted initially at 19°C, and then again after a 4-week acclimation to either 15°C or 27°C. While boldness and activity were similar between populations, core fish exhibited higher CTmax at 19°C but had similar thermal tolerance after acclimation, suggesting a comparable thermal plasticity. Our findings suggest larger and reproductive individuals are involved in the range expansion of round goby and appear to be in boom or population expansion phase. The results also suggest a more limited role of individual behaviour in shaping the invasion dynamic and raise questions about potential trade-offs between physiological tolerance and dispersal. By offering insights into the mechanisms underlying the invasion process, this study highlights invasion gradients as promising frameworks for invasion research and for developing mitigation strategies.

Investigating the phenotypic drivers of the invasion dynamic in round goby 

Due to the ongoing global warming, extreme marine heat wave events have increased in severity, frequency and duration, setting marine ecosystems at high risk. In this context, the application of methods that can alleviate global warming devastating impacts on marine organisms constitutes a necessity. Repeated time-limited exposure of an organism to an environmental stimulus modifies its response mode, enhancing its resilience and adaptation to environmental stress. This “stress memory” effect is referred to as “hardening”. From this perspective, in the present study, we have applied the heat hardening process onMytilus galloprovincialis mussels. Our aim was to investigate the heat hardening effect on mussels’ cellular pathways and identify the underlying biochemical mechanisms, in terms of apoptosis, that enhance these organisms’ thermal tolerance in the laboratory. Thereafter, heat-hardened mussels were transferred to the field where they were left to acclimatize under increasing ambient sea water temperatures, to evaluate their resilience to constantly changing environmental conditions. Heat hardening resulted in decreased apoptosis at all elevated temperatures (24 °C, 26 °C, and 28 °C) in the heat-hardened mussels compared to the nonhardened individuals. In the field, increased ambient temperature resulted to increasing levels of apoptosis in the nonhardened mussels, with the hardened individuals exhibiting a significant decrease in apoptotic indicators compared to their counterparts. In conclusion, the exposure of mussels to heat hardening in the laboratory seems to provide a “stress memory” response that enhances heat tolerance and increases cell survival through reduced cell death when mussels were exposed to complex natural conditions.

Verification of the enhanced physiological performance of laboratory heat-hardened Mediterranean mussels when acclimatized in the complex field conditions

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