T. Schwerte | University of Innsbruck (original) (raw)
Papers by T. Schwerte
Physiological and Biochemical Zoology, 2010
Technical advances that have made it possible to perform physiological measurements on very small... more Technical advances that have made it possible to perform physiological measurements on very small organisms, including those in embryonic and larval stages, have resulted in the formation of the discipline of developmental physiology. The transparency and size of developing organisms in some areas permit insights into physiological processes that cannot be obtained with opaque, adult organisms. On the other hand, it is widely accepted that without eggs, there are no chickens, so physiological adaptations during early life are just as important to species survival as those manifested by adults. Physiological adaptations of early developmental stages, however, are not always the same as patterns known in adults; they often follow their own rules. The adaptability of early developmental stages demonstrates that development is not stereotyped and a phenotype is not just the result of genetic information and the expression of a certain series of genes. Environmental factors influence phenotype production, and this in turn results in flexibility and plasticity in physiological processes. This article comprises exemplary studies presented at the Fourth International Conference in Africa for Comparative Physiology and * This article was prepared as an overview of a symposium at "Molecules to Migration: Pressures of Life," the
Acta Myologica, 2016
Paraneoplastic neurological syndromes (PNS) are remote effects of cancer. Lambert-Eaton myastheni... more Paraneoplastic neurological syndromes (PNS) are remote effects of cancer. Lambert-Eaton myasthenic syndrome (LEMS) is a well-defined PNS, affecting NMJ. It is characterised by proximal muscle weakness and autonomic symptoms, caused by antibodies interacting with the voltage gated calcium channels (VGCCs) of P/Q-, N-and R-type, in over 50% of the cases associated to an underlying tumour, most often small-cell lung cancer (SCLC). However the molecular mechanisms of LEMS remains largely unknown. Almost all patients will benefit initially from symptomatic treatment with 3,4-diaminopyridine (3,4-DAP), a potassium channel blocking agent. Often, additional treatment is required in the form of immunosuppression Voltage-gated potassium channels (VGKC), or Kv channels, are membrane channels able to open selectively for potassium ions in response to changes in membrane polarity. Antibodies against VGKC have been associated to peripheral nerve hyperexcitability disorders, characterized by constant muscle fiber activity, linked in some cases, to an underlying tumor, most frequently thy-PROCEEDINGS Of thE XVI CONGRESS Of thE ItALIAN SOCIEtY Of MYOLOGY
Journal of Experimental Biology, 1997
The influence of the vagus nerve on swimbladder blood flow in the European eel (Anguilla anguilla... more The influence of the vagus nerve on swimbladder blood flow in the European eel (Anguilla anguilla) was characterized by recording the changes in blood flow rate and blood pressure following stimulation of the vagus nerve. After electrical stimulation, blood flow in the swimbladder artery increased from 0.9 ml min-1 to 2.1 ml min-1. Video recordings of small vessels on the caudal side of the rete mirabile revealed an increase in erythrocyte velocity combined with a small vasodilation. This effect could not be blocked by injection of the -adrenergic antagonist phentolamine, the ss-adrenergic antagonist propranolol or the muscarinic cholinoceptor antagonist atropine. In all preparations with a high initial flow rate (>1.9 ml min-1), vagotomy resulted in a marked decrease in blood flow (by approximately 80 %). This effect was not observed in preparations with a low initial swimbladder blood flow. Stimulation of the vagus nerve produced a decrease, and vagotomy produced an increase, i...
Journal of Experimental Biology, 2000
The analysis of perfusion parameters using the frame-to-frame technique and the observation of sm... more The analysis of perfusion parameters using the frame-to-frame technique and the observation of small blood vessels in transparent animals using video microscopy can be tedious and very difficult because of the poor contrast of the images. Injection of a fluorescent probe (fluorescein isothiocynate, FITC) bound to a high-molecular-mass dextran improved the visibility of blood vessels, but the gray-scale histogram showed blurring at the edges of the vessels. Furthermore, injection of the fluorescent probe into the ventricle of small zebrafish (Danio rerio) embryos (body mass approximately 1 mg) often resulted in reduced cardiac activity. Digital motion analysis, however, proved to be a very effective tool for analysing the shape and performance of the circulatory system in transparent animals and tissues. By subtracting the two fields of a video frame (the odd and the even frame), any movement that occurred within the 20 ms necessary for the acquisition of one field could be visualise...
American Journal of Physiology-Regulatory, Integrative and Comparative Physiology, 2000
The minnow ( Phoxinus phoxinus) was raised up to the stage of swim bladder inflation at temperatu... more The minnow ( Phoxinus phoxinus) was raised up to the stage of swim bladder inflation at temperatures between 10°C and 25°C, and the time of development significantly decreased at higher temperatures. Accordingly, initiation of cardiac activity was observed at day 2 in 25°C animals and at day 4 in 12.5°C animals. Only a minor increase in body mass was observed during the incubation period, and, at the end of the incubation period, animals raised at 25°C did not have a significantly lower body mass compared with animals raised at 15°C. Metabolic activity, determined as the rate of oxygen consumption of a larva, increased from 3.3 to 19.5 nmol/h during development at 15°C and from 5.6 to 47.6 nmol/h during development at 25°C. Heart rate showed a clear correlation to developmental stage as well as to developmental temperature, but at the onset of cardiac activity, diastolic ventricular volume and also stroke volume were higher at the lower temperatures. Furthermore, stroke volume incre...
The Journal of clinical investigation, Jan 7, 2015
The Popeye domain-containing 1 (POPDC1) gene encodes a plasma membrane-localized cAMP-binding pro... more The Popeye domain-containing 1 (POPDC1) gene encodes a plasma membrane-localized cAMP-binding protein that is abundantly expressed in striated muscle. In animal models, POPDC1 is an essential regulator of structure and function of cardiac and skeletal muscle; however, POPDC1 mutations have not been associated with human cardiac and muscular diseases. Here, we have described a homozygous missense variant (c.602C>T, p.S201F) in POPDC1, identified by whole-exome sequencing, in a family of 4 with cardiac arrhythmia and limb-girdle muscular dystrophy (LGMD). This allele was absent in known databases and segregated with the pathological phenotype in this family. We did not find the allele in a further screen of 104 patients with a similar phenotype, suggesting this mutation to be family specific. Compared with WT protein, POPDC1S201F displayed a 50% reduction in cAMP affinity, and in skeletal muscle from patients, both POPDC1S201F and WT POPDC2 displayed impaired membrane trafficking. ...
American Journal of Physiology - Regulatory, Integrative and Comparative Physiology, 2003
Larval zebrafish ( Danio rerio) of two different age classes (“swim-up” larvae, 9 days old; “free... more Larval zebrafish ( Danio rerio) of two different age classes (“swim-up” larvae, 9 days old; “free-swimming” larvae, 21 days old) were exposed to either an endurance/continuous training or interval training. Control animals were kept in stagnant water. A comparison of cardiac activity of trained (either endurance or interval) and untrained animals at the end of the training regime revealed no differences in heart rate, end-diastolic and end-systolic ventricular volume, and cardiac output. Training also had no influence on the concentration of erythrocytes in the blood. Thus, at the level of total oxygen transport in the blood, training did not provoke any improvement during the first 32 days of development. Significant changes, however, were observed at the tissue level. In free-swimming larvae [i.e., between 21 and 32 days postfertilization (dpf)] endurance training increased the capillarization of both axial muscle caudal to the anus and the tail fin. In addition, mitochondrial den...
Journal of Experimental Biology, 2011
SUMMARYPermeability of rainbow trout gill pavement cells cultured on permeable supports (single s... more SUMMARYPermeability of rainbow trout gill pavement cells cultured on permeable supports (single seeded inserts) changes upon exposure to freshwater or treatment with cortisol. The molecular components of this change are largely unknown, but tight junctions that regulate the paracellular pathway are prime candidates in this adaptational process. Using differential display polymerase chain reaction we found a set of 17 differentially regulated genes in trout pavement cells that had been exposed to freshwater apically for 24 h. Five genes were related to the cell–cell contact. One of these genes was isolated and identified as encoding claudin 28b, an integral component of the tight junction. Immunohistochemical reactivity to claudin 28b protein was concentrated in a circumferential ring colocalized to the cortical F-actin ring. To study the contribution of this isoform to changes in transepithelial resistance and Phenol Red diffusion under apical hypo-or hyperosmotic exposure we quanti...
Journal of Experimental Biology, 2011
SUMMARY β-Adrenergic receptors (βARs) are crucial for maintaining the rate and force of cardiac m... more SUMMARY β-Adrenergic receptors (βARs) are crucial for maintaining the rate and force of cardiac muscle contraction in vertebrates. Zebrafish (Danio rerio) have one β1AR gene and two β2AR genes (β2aAR and β2bAR). We examined the roles of these receptors in larval zebrafish in vivo by assessing the impact of translational gene knockdown on cardiac function. Zebrafish larvae lacking β1AR expression by morpholino knockdown displayed lower heart rates than control fish, whereas larvae deficient in both β2aAR and β2bAR expression exhibited significantly higher heart rates than controls. These results suggested a potential inhibitory role for one or both β2AR genes. By using cultured HEK293 cells transfected with zebrafish βARs, we demonstrated that stimulation with adrenaline or procaterol (a β2AR agonist) resulted in an increase in intracellular cAMP levels in cells expressing any of the three zebrafish βARs. In comparison with its human βAR counterpart, zebrafish β2aAR expressed in HEK2...
Journal of Fish Biology, 2003
For the first time, spontaneous intestinal activity was demonstrated and quantified before the on... more For the first time, spontaneous intestinal activity was demonstrated and quantified before the onset of exogenous feeding in zebrafish Danio rerio embryos and larvae in vivo, using digital motion analysis. At 3 days post fertilization (dpf), erratic and spontaneous contraction waves were observed in the gut. Later (4-7 dpf), more distinct contraction patterns were distinguished, and anterograde and retrograde contraction waves projecting anally and orally along the intestine, respectively, as well as local rectal contraction waves could be identified and quantified. The frequency of both anterograde intestinal and local rectal contractions increased significantly during the first days of development. There was a tendency towards shorter anterograde contraction waves in the first dpf stage investigated, but the velocity of the waves did not differ significantly between the different dpf stages. The presence of developing neurones in the gut of zebrafish was established using immunohistochemistry, staining for a suite of marker proteins (Hu C/D, HNK-1 and acetylated tubulin). Structural neurones were present in the developing gut from the first dpf stage investigated (2 dpf). In conclusion, during the period (3-7 dpf) when erratic contraction waves turn into a more organized pattern of motility there is also a pronounced development of the innervation, suggesting a correlation in time of the development of gut motility and its neuronal control.
Journal of Biological Chemistry, 2008
The paralyzed zebrafish strain relaxed carries a null mutation for the skeletal muscle dihydropyr... more The paralyzed zebrafish strain relaxed carries a null mutation for the skeletal muscle dihydropyridine receptor (DHPR)  1a subunit. Lack of  1a results in (i) reduced membrane expression of the pore forming DHPR ␣ 1S subunit, (ii) elimination of ␣ 1S charge movement, and (iii) impediment of arrangement of the DHPRs in groups of four (tetrads) opposing the ryanodine receptor (RyR1), a structural prerequisite for skeletal muscletype excitation-contraction (EC) coupling. In this study we used relaxed larvae and isolated myotubes as expression systems to discriminate specific functions of  1a from rather general functions of  isoforms. Zebrafish and mammalian  1a subunits quantitatively restored ␣ 1S triad targeting and charge movement as well as intracellular Ca 2؉ release, allowed arrangement of DHPRs in tetrads, and most strikingly recovered a fully motile phenotype in relaxed larvae. Interestingly, the cardiac/neuronal  2a as the phylogenetically closest, and the ancestral housefly  M as the most distant isoform to  1a also completely recovered ␣ 1S triad expression and charge movement. However, both revealed drastically impaired intracellular Ca 2؉ transients and very limited tetrad formation compared with  1a. Consequently, larval motility was either only partially restored ( 2a-injected larvae) or not restored at all ( M). Thus, our results indicate that triad expression and facilitation of 1,4-dihydropyridine receptor (DHPR) charge movement are common features of all tested  subunits, whereas the efficient arrangement of DHPRs in tetrads and thus intact DHPR-RyR1 coupling is only promoted by the  1a isoform. Consequently, we postulate a model that presents  1a as an allosteric modifier of ␣ 1S conformation enabling skeletal muscle-type EC coupling.
![Research paper thumbnail of Corrigendum to “The Popeye domain containing 2 (popdc2) gene in zebrafish is required for heart and skeletal muscle development” [Dev. Biol. 363 (2012) 438–450]](https://attachments.academia-assets.com/91466564/thumbnails/1.jpg)
](Developmental Biology, 2012
Corrigendum to ''The Popeye domain containing 2 (popdc2) gene in zebrafish is required for heart ... more Corrigendum to ''The Popeye domain containing 2 (popdc2) gene in zebrafish is required for heart and skeletal muscle development'' [Dev. Biol. 363 (2012) 438-450]
Developmental Biology, 2012
The Popeye domain containing (Popdc) genes encode a family of transmembrane proteins with an evol... more The Popeye domain containing (Popdc) genes encode a family of transmembrane proteins with an evolutionary conserved Popeye domain. These genes are abundantly expressed in striated muscle tissue, however their function is not well understood. In this study we have investigated the role of the popdc2 gene in zebrafish. Popdc2 transcripts were detected in the embryonic myocardium and transiently in the craniofacial and tail musculature. Morpholino oligonucleotide-mediated knockdown of popdc2 resulted in aberrant development of skeletal muscle and heart. Muscle segments in the trunk were irregularly shaped and craniofacial muscles were severely reduced or even missing. In the heart, pericardial edema was prevalent in the morphants and heart chambers were elongated and looping was abnormal. These pathologies in muscle and heart were alleviated after reducing the morpholino concentration. However the heart still was abnormal displaying cardiac arrhythmia at later stages of development. Optical recordings of cardiac contractility revealed irregular ventricular contractions with a 2:1, or 3:1 atrial/ventricular conduction ratio, which caused a significant reduction in heart frequency. Recordings of calcium transients with high spatiotemporal resolution using a transgenic calcium indicator line (Tg(cmlc2:gCaMP) s878) and SPIM microscopy confirmed the presence of a severe arrhythmia phenotype. Our results identify popdc2 as a gene important for striated muscle differentiation and cardiac morphogenesis. In addition it is required for the development of the cardiac conduction system.
Comparative Biochemistry and Physiology Part A: Molecular & Integrative Physiology, 1999
Comparative Biochemistry and Physiology Part A: Molecular & Integrative Physiology, 2006
Increased ventilation frequency (f V) in response to hypoxia in adult fish depends on ionotropic ... more Increased ventilation frequency (f V) in response to hypoxia in adult fish depends on ionotropic N-methyl-D-aspartate (NMDA) receptors. Nonetheless, the ontogeny of central control mechanisms mediating hypoxic ventilatory chemoreflexes in lower vertebrates has not been studied. Therefore, the aim of this study was to determine when the hypoxic ventilatory response during zebrafish (Danio rerio) development is mediated via NMDA receptors, by performing physiological experiments and western blot analysis of NMDA receptor subunits. Zebrafish larvae at stages 4-16 days post-fertilisation (dpf) were exposed to an hypoxic pulse in control groups and in groups treated with MK801 (NMDA receptor antagonist). The hypoxic increase in f V was present at all larval stages, and it matured during development. The reflex became MK801 sensitive at 8 dpf, but did not completely rely on a glutamatergic transmission until 13 dpf. This, together with changing subunit composition during the different stages (increasing amounts of NMDAR1 subunits and appearance of NMDAR2A subunits in adults), suggests that the amount of functional NMDA receptors needed to achieve a fully developed reflex is not attained until later stages. Furthermore, our results suggest that other non-NMDA receptor mechanisms are responsible for the hypoxia-induced increase in f V during the earlier developmental stages.
Comparative Biochemistry and Physiology Part A: Molecular & Integrative Physiology, 2009
Time PCR revealed a reduced expression of clock, period and timeless. These alterations in the mR... more Time PCR revealed a reduced expression of clock, period and timeless. These alterations in the mRNA concentration of clock genes were also reflected in a modified activity profile comparing wdt and bre larvae. In conclusion cardiac impairment, i.e. a heart rate reduction of more than 50%, ultimately affected circadian rhythms and in turn altered cell cycle, developmental and activity profiles. Further results indicated that in bre zebrafish tissue hypoxia and/or changes in metabolism link heart failure to the circadian clock.
Comparative Biochemistry and Physiology Part B: Biochemistry and Molecular Biology, 2000
Journal of experimental …, 2006
Cardiovascular Research, 2012
Aim: Opening of the mitochondrial permeability transition pore (mPTP) occurs during cardiac ische... more Aim: Opening of the mitochondrial permeability transition pore (mPTP) occurs during cardiac ischemia and reperfusion and is an irreversible step on the pathway towards cardiomyocyte death. In isolated mitochondria, mPTP opening is triggered by calcium overload facilitated by oxidative stress, although the chronology has been suggested to be different in isolated cells. Our objective was to establish the events that cause mPTP opening in the intact heart. Methods: We performed multiphoton imaging of Langendorff-perfused mouse hearts expressing an inducible, Ca 2+-sensitive reporter (GCaMP2), to examine the response to a hypoxia-reoxygenation protocol of individual cardiomyocytes within an intact, perfused heart. Results: We found that during reperfusion, spontaneous calcium waves arose and propagated across multiple cells at 3.3 mm/s. In cells in which [Ca 2+ ] returned to normal there was no effect on mitochondrial membrane potential, measured using the dye TMRM. However, in~56% of cells the calcium waves were followed after 100 + 27 s (N=10 hearts) by loss of TMRM fluorescence due to opening of the mPTP. During reoxygenation, coordinated redox changes also occurred across large regions, and preceded mPTP opening on average by 122 + 38 s (N=9 hearts). Significantly fewer calcium waves resulted in mPTP opening in the presence of mPTP inhibitor cyclosporin A, or mitochondrial-targeted ROS scavenger, MitoQ. Only MitoQ, however, reduced the number of Ca 2+ waves. Conclusion: These experiments define the spatiotemporal relationship between changes in Ca 2+ , redox state, and mPTP opening during reoxygenation in the intact heart. Tissue oxidation coincident with localized calcium waves together conspire to cause mPTP opening and subsequent cell death. 437 FRET-based imaging of local cAMP pools reveals altered phosphodiesterase activity and cAMP compartmentalisation in an in vitro model of cardiac hypertrophy
Physiological and Biochemical Zoology, 2010
Technical advances that have made it possible to perform physiological measurements on very small... more Technical advances that have made it possible to perform physiological measurements on very small organisms, including those in embryonic and larval stages, have resulted in the formation of the discipline of developmental physiology. The transparency and size of developing organisms in some areas permit insights into physiological processes that cannot be obtained with opaque, adult organisms. On the other hand, it is widely accepted that without eggs, there are no chickens, so physiological adaptations during early life are just as important to species survival as those manifested by adults. Physiological adaptations of early developmental stages, however, are not always the same as patterns known in adults; they often follow their own rules. The adaptability of early developmental stages demonstrates that development is not stereotyped and a phenotype is not just the result of genetic information and the expression of a certain series of genes. Environmental factors influence phenotype production, and this in turn results in flexibility and plasticity in physiological processes. This article comprises exemplary studies presented at the Fourth International Conference in Africa for Comparative Physiology and * This article was prepared as an overview of a symposium at "Molecules to Migration: Pressures of Life," the
Acta Myologica, 2016
Paraneoplastic neurological syndromes (PNS) are remote effects of cancer. Lambert-Eaton myastheni... more Paraneoplastic neurological syndromes (PNS) are remote effects of cancer. Lambert-Eaton myasthenic syndrome (LEMS) is a well-defined PNS, affecting NMJ. It is characterised by proximal muscle weakness and autonomic symptoms, caused by antibodies interacting with the voltage gated calcium channels (VGCCs) of P/Q-, N-and R-type, in over 50% of the cases associated to an underlying tumour, most often small-cell lung cancer (SCLC). However the molecular mechanisms of LEMS remains largely unknown. Almost all patients will benefit initially from symptomatic treatment with 3,4-diaminopyridine (3,4-DAP), a potassium channel blocking agent. Often, additional treatment is required in the form of immunosuppression Voltage-gated potassium channels (VGKC), or Kv channels, are membrane channels able to open selectively for potassium ions in response to changes in membrane polarity. Antibodies against VGKC have been associated to peripheral nerve hyperexcitability disorders, characterized by constant muscle fiber activity, linked in some cases, to an underlying tumor, most frequently thy-PROCEEDINGS Of thE XVI CONGRESS Of thE ItALIAN SOCIEtY Of MYOLOGY
Journal of Experimental Biology, 1997
The influence of the vagus nerve on swimbladder blood flow in the European eel (Anguilla anguilla... more The influence of the vagus nerve on swimbladder blood flow in the European eel (Anguilla anguilla) was characterized by recording the changes in blood flow rate and blood pressure following stimulation of the vagus nerve. After electrical stimulation, blood flow in the swimbladder artery increased from 0.9 ml min-1 to 2.1 ml min-1. Video recordings of small vessels on the caudal side of the rete mirabile revealed an increase in erythrocyte velocity combined with a small vasodilation. This effect could not be blocked by injection of the -adrenergic antagonist phentolamine, the ss-adrenergic antagonist propranolol or the muscarinic cholinoceptor antagonist atropine. In all preparations with a high initial flow rate (>1.9 ml min-1), vagotomy resulted in a marked decrease in blood flow (by approximately 80 %). This effect was not observed in preparations with a low initial swimbladder blood flow. Stimulation of the vagus nerve produced a decrease, and vagotomy produced an increase, i...
Journal of Experimental Biology, 2000
The analysis of perfusion parameters using the frame-to-frame technique and the observation of sm... more The analysis of perfusion parameters using the frame-to-frame technique and the observation of small blood vessels in transparent animals using video microscopy can be tedious and very difficult because of the poor contrast of the images. Injection of a fluorescent probe (fluorescein isothiocynate, FITC) bound to a high-molecular-mass dextran improved the visibility of blood vessels, but the gray-scale histogram showed blurring at the edges of the vessels. Furthermore, injection of the fluorescent probe into the ventricle of small zebrafish (Danio rerio) embryos (body mass approximately 1 mg) often resulted in reduced cardiac activity. Digital motion analysis, however, proved to be a very effective tool for analysing the shape and performance of the circulatory system in transparent animals and tissues. By subtracting the two fields of a video frame (the odd and the even frame), any movement that occurred within the 20 ms necessary for the acquisition of one field could be visualise...
American Journal of Physiology-Regulatory, Integrative and Comparative Physiology, 2000
The minnow ( Phoxinus phoxinus) was raised up to the stage of swim bladder inflation at temperatu... more The minnow ( Phoxinus phoxinus) was raised up to the stage of swim bladder inflation at temperatures between 10°C and 25°C, and the time of development significantly decreased at higher temperatures. Accordingly, initiation of cardiac activity was observed at day 2 in 25°C animals and at day 4 in 12.5°C animals. Only a minor increase in body mass was observed during the incubation period, and, at the end of the incubation period, animals raised at 25°C did not have a significantly lower body mass compared with animals raised at 15°C. Metabolic activity, determined as the rate of oxygen consumption of a larva, increased from 3.3 to 19.5 nmol/h during development at 15°C and from 5.6 to 47.6 nmol/h during development at 25°C. Heart rate showed a clear correlation to developmental stage as well as to developmental temperature, but at the onset of cardiac activity, diastolic ventricular volume and also stroke volume were higher at the lower temperatures. Furthermore, stroke volume incre...
The Journal of clinical investigation, Jan 7, 2015
The Popeye domain-containing 1 (POPDC1) gene encodes a plasma membrane-localized cAMP-binding pro... more The Popeye domain-containing 1 (POPDC1) gene encodes a plasma membrane-localized cAMP-binding protein that is abundantly expressed in striated muscle. In animal models, POPDC1 is an essential regulator of structure and function of cardiac and skeletal muscle; however, POPDC1 mutations have not been associated with human cardiac and muscular diseases. Here, we have described a homozygous missense variant (c.602C>T, p.S201F) in POPDC1, identified by whole-exome sequencing, in a family of 4 with cardiac arrhythmia and limb-girdle muscular dystrophy (LGMD). This allele was absent in known databases and segregated with the pathological phenotype in this family. We did not find the allele in a further screen of 104 patients with a similar phenotype, suggesting this mutation to be family specific. Compared with WT protein, POPDC1S201F displayed a 50% reduction in cAMP affinity, and in skeletal muscle from patients, both POPDC1S201F and WT POPDC2 displayed impaired membrane trafficking. ...
American Journal of Physiology - Regulatory, Integrative and Comparative Physiology, 2003
Larval zebrafish ( Danio rerio) of two different age classes (“swim-up” larvae, 9 days old; “free... more Larval zebrafish ( Danio rerio) of two different age classes (“swim-up” larvae, 9 days old; “free-swimming” larvae, 21 days old) were exposed to either an endurance/continuous training or interval training. Control animals were kept in stagnant water. A comparison of cardiac activity of trained (either endurance or interval) and untrained animals at the end of the training regime revealed no differences in heart rate, end-diastolic and end-systolic ventricular volume, and cardiac output. Training also had no influence on the concentration of erythrocytes in the blood. Thus, at the level of total oxygen transport in the blood, training did not provoke any improvement during the first 32 days of development. Significant changes, however, were observed at the tissue level. In free-swimming larvae [i.e., between 21 and 32 days postfertilization (dpf)] endurance training increased the capillarization of both axial muscle caudal to the anus and the tail fin. In addition, mitochondrial den...
Journal of Experimental Biology, 2011
SUMMARYPermeability of rainbow trout gill pavement cells cultured on permeable supports (single s... more SUMMARYPermeability of rainbow trout gill pavement cells cultured on permeable supports (single seeded inserts) changes upon exposure to freshwater or treatment with cortisol. The molecular components of this change are largely unknown, but tight junctions that regulate the paracellular pathway are prime candidates in this adaptational process. Using differential display polymerase chain reaction we found a set of 17 differentially regulated genes in trout pavement cells that had been exposed to freshwater apically for 24 h. Five genes were related to the cell–cell contact. One of these genes was isolated and identified as encoding claudin 28b, an integral component of the tight junction. Immunohistochemical reactivity to claudin 28b protein was concentrated in a circumferential ring colocalized to the cortical F-actin ring. To study the contribution of this isoform to changes in transepithelial resistance and Phenol Red diffusion under apical hypo-or hyperosmotic exposure we quanti...
Journal of Experimental Biology, 2011
SUMMARY β-Adrenergic receptors (βARs) are crucial for maintaining the rate and force of cardiac m... more SUMMARY β-Adrenergic receptors (βARs) are crucial for maintaining the rate and force of cardiac muscle contraction in vertebrates. Zebrafish (Danio rerio) have one β1AR gene and two β2AR genes (β2aAR and β2bAR). We examined the roles of these receptors in larval zebrafish in vivo by assessing the impact of translational gene knockdown on cardiac function. Zebrafish larvae lacking β1AR expression by morpholino knockdown displayed lower heart rates than control fish, whereas larvae deficient in both β2aAR and β2bAR expression exhibited significantly higher heart rates than controls. These results suggested a potential inhibitory role for one or both β2AR genes. By using cultured HEK293 cells transfected with zebrafish βARs, we demonstrated that stimulation with adrenaline or procaterol (a β2AR agonist) resulted in an increase in intracellular cAMP levels in cells expressing any of the three zebrafish βARs. In comparison with its human βAR counterpart, zebrafish β2aAR expressed in HEK2...
Journal of Fish Biology, 2003
For the first time, spontaneous intestinal activity was demonstrated and quantified before the on... more For the first time, spontaneous intestinal activity was demonstrated and quantified before the onset of exogenous feeding in zebrafish Danio rerio embryos and larvae in vivo, using digital motion analysis. At 3 days post fertilization (dpf), erratic and spontaneous contraction waves were observed in the gut. Later (4-7 dpf), more distinct contraction patterns were distinguished, and anterograde and retrograde contraction waves projecting anally and orally along the intestine, respectively, as well as local rectal contraction waves could be identified and quantified. The frequency of both anterograde intestinal and local rectal contractions increased significantly during the first days of development. There was a tendency towards shorter anterograde contraction waves in the first dpf stage investigated, but the velocity of the waves did not differ significantly between the different dpf stages. The presence of developing neurones in the gut of zebrafish was established using immunohistochemistry, staining for a suite of marker proteins (Hu C/D, HNK-1 and acetylated tubulin). Structural neurones were present in the developing gut from the first dpf stage investigated (2 dpf). In conclusion, during the period (3-7 dpf) when erratic contraction waves turn into a more organized pattern of motility there is also a pronounced development of the innervation, suggesting a correlation in time of the development of gut motility and its neuronal control.
Journal of Biological Chemistry, 2008
The paralyzed zebrafish strain relaxed carries a null mutation for the skeletal muscle dihydropyr... more The paralyzed zebrafish strain relaxed carries a null mutation for the skeletal muscle dihydropyridine receptor (DHPR)  1a subunit. Lack of  1a results in (i) reduced membrane expression of the pore forming DHPR ␣ 1S subunit, (ii) elimination of ␣ 1S charge movement, and (iii) impediment of arrangement of the DHPRs in groups of four (tetrads) opposing the ryanodine receptor (RyR1), a structural prerequisite for skeletal muscletype excitation-contraction (EC) coupling. In this study we used relaxed larvae and isolated myotubes as expression systems to discriminate specific functions of  1a from rather general functions of  isoforms. Zebrafish and mammalian  1a subunits quantitatively restored ␣ 1S triad targeting and charge movement as well as intracellular Ca 2؉ release, allowed arrangement of DHPRs in tetrads, and most strikingly recovered a fully motile phenotype in relaxed larvae. Interestingly, the cardiac/neuronal  2a as the phylogenetically closest, and the ancestral housefly  M as the most distant isoform to  1a also completely recovered ␣ 1S triad expression and charge movement. However, both revealed drastically impaired intracellular Ca 2؉ transients and very limited tetrad formation compared with  1a. Consequently, larval motility was either only partially restored ( 2a-injected larvae) or not restored at all ( M). Thus, our results indicate that triad expression and facilitation of 1,4-dihydropyridine receptor (DHPR) charge movement are common features of all tested  subunits, whereas the efficient arrangement of DHPRs in tetrads and thus intact DHPR-RyR1 coupling is only promoted by the  1a isoform. Consequently, we postulate a model that presents  1a as an allosteric modifier of ␣ 1S conformation enabling skeletal muscle-type EC coupling.
Developmental Biology, 2012
Corrigendum to ''The Popeye domain containing 2 (popdc2) gene in zebrafish is required for heart ... more Corrigendum to ''The Popeye domain containing 2 (popdc2) gene in zebrafish is required for heart and skeletal muscle development'' [Dev. Biol. 363 (2012) 438-450]
Developmental Biology, 2012
The Popeye domain containing (Popdc) genes encode a family of transmembrane proteins with an evol... more The Popeye domain containing (Popdc) genes encode a family of transmembrane proteins with an evolutionary conserved Popeye domain. These genes are abundantly expressed in striated muscle tissue, however their function is not well understood. In this study we have investigated the role of the popdc2 gene in zebrafish. Popdc2 transcripts were detected in the embryonic myocardium and transiently in the craniofacial and tail musculature. Morpholino oligonucleotide-mediated knockdown of popdc2 resulted in aberrant development of skeletal muscle and heart. Muscle segments in the trunk were irregularly shaped and craniofacial muscles were severely reduced or even missing. In the heart, pericardial edema was prevalent in the morphants and heart chambers were elongated and looping was abnormal. These pathologies in muscle and heart were alleviated after reducing the morpholino concentration. However the heart still was abnormal displaying cardiac arrhythmia at later stages of development. Optical recordings of cardiac contractility revealed irregular ventricular contractions with a 2:1, or 3:1 atrial/ventricular conduction ratio, which caused a significant reduction in heart frequency. Recordings of calcium transients with high spatiotemporal resolution using a transgenic calcium indicator line (Tg(cmlc2:gCaMP) s878) and SPIM microscopy confirmed the presence of a severe arrhythmia phenotype. Our results identify popdc2 as a gene important for striated muscle differentiation and cardiac morphogenesis. In addition it is required for the development of the cardiac conduction system.
Comparative Biochemistry and Physiology Part A: Molecular & Integrative Physiology, 1999
Comparative Biochemistry and Physiology Part A: Molecular & Integrative Physiology, 2006
Increased ventilation frequency (f V) in response to hypoxia in adult fish depends on ionotropic ... more Increased ventilation frequency (f V) in response to hypoxia in adult fish depends on ionotropic N-methyl-D-aspartate (NMDA) receptors. Nonetheless, the ontogeny of central control mechanisms mediating hypoxic ventilatory chemoreflexes in lower vertebrates has not been studied. Therefore, the aim of this study was to determine when the hypoxic ventilatory response during zebrafish (Danio rerio) development is mediated via NMDA receptors, by performing physiological experiments and western blot analysis of NMDA receptor subunits. Zebrafish larvae at stages 4-16 days post-fertilisation (dpf) were exposed to an hypoxic pulse in control groups and in groups treated with MK801 (NMDA receptor antagonist). The hypoxic increase in f V was present at all larval stages, and it matured during development. The reflex became MK801 sensitive at 8 dpf, but did not completely rely on a glutamatergic transmission until 13 dpf. This, together with changing subunit composition during the different stages (increasing amounts of NMDAR1 subunits and appearance of NMDAR2A subunits in adults), suggests that the amount of functional NMDA receptors needed to achieve a fully developed reflex is not attained until later stages. Furthermore, our results suggest that other non-NMDA receptor mechanisms are responsible for the hypoxia-induced increase in f V during the earlier developmental stages.
Comparative Biochemistry and Physiology Part A: Molecular & Integrative Physiology, 2009
Time PCR revealed a reduced expression of clock, period and timeless. These alterations in the mR... more Time PCR revealed a reduced expression of clock, period and timeless. These alterations in the mRNA concentration of clock genes were also reflected in a modified activity profile comparing wdt and bre larvae. In conclusion cardiac impairment, i.e. a heart rate reduction of more than 50%, ultimately affected circadian rhythms and in turn altered cell cycle, developmental and activity profiles. Further results indicated that in bre zebrafish tissue hypoxia and/or changes in metabolism link heart failure to the circadian clock.
Comparative Biochemistry and Physiology Part B: Biochemistry and Molecular Biology, 2000
Journal of experimental …, 2006
Cardiovascular Research, 2012
Aim: Opening of the mitochondrial permeability transition pore (mPTP) occurs during cardiac ische... more Aim: Opening of the mitochondrial permeability transition pore (mPTP) occurs during cardiac ischemia and reperfusion and is an irreversible step on the pathway towards cardiomyocyte death. In isolated mitochondria, mPTP opening is triggered by calcium overload facilitated by oxidative stress, although the chronology has been suggested to be different in isolated cells. Our objective was to establish the events that cause mPTP opening in the intact heart. Methods: We performed multiphoton imaging of Langendorff-perfused mouse hearts expressing an inducible, Ca 2+-sensitive reporter (GCaMP2), to examine the response to a hypoxia-reoxygenation protocol of individual cardiomyocytes within an intact, perfused heart. Results: We found that during reperfusion, spontaneous calcium waves arose and propagated across multiple cells at 3.3 mm/s. In cells in which [Ca 2+ ] returned to normal there was no effect on mitochondrial membrane potential, measured using the dye TMRM. However, in~56% of cells the calcium waves were followed after 100 + 27 s (N=10 hearts) by loss of TMRM fluorescence due to opening of the mPTP. During reoxygenation, coordinated redox changes also occurred across large regions, and preceded mPTP opening on average by 122 + 38 s (N=9 hearts). Significantly fewer calcium waves resulted in mPTP opening in the presence of mPTP inhibitor cyclosporin A, or mitochondrial-targeted ROS scavenger, MitoQ. Only MitoQ, however, reduced the number of Ca 2+ waves. Conclusion: These experiments define the spatiotemporal relationship between changes in Ca 2+ , redox state, and mPTP opening during reoxygenation in the intact heart. Tissue oxidation coincident with localized calcium waves together conspire to cause mPTP opening and subsequent cell death. 437 FRET-based imaging of local cAMP pools reveals altered phosphodiesterase activity and cAMP compartmentalisation in an in vitro model of cardiac hypertrophy