Plasma ionized magnesium during acute hyperventilation in humans.

1. Respiratory alkalosis accompanies the clinical syndrome of tetany, precipitates cardiac arrhythmias and predisposes to coronary vasoconstriction. Magnesium plays a critical role in the maintenance of membrane function, and magnesium depletion is often associated with cardiac arrhythmias or vasoconstriction. 2. As technology for detecting circulating ionized magnesium (the most interesting form with respect to physiological and biological properties) is now available in the form of new magnesium-selective electrodes, the effect of respiratory alkalosis induced by voluntary overbreathing for 30 min on circulating ionized magnesium was studied in eight healthy subjects. 3. The total plasma magnesium concentration was not modified by hyperventilation. On the contrary, hyperventilation was associated with a significant reduction in the ionized magnesium concentration of 0.05 (0.02-0.15) mmol/l (median and range) and in the free magnesium fraction of 0.06 (0.01-0.19). During hyperventilation the relative intravascular magnesium mass, calculated from changes in total plasma magnesium concentration and haematocrit, decreased significantly. 4. It is concluded that acute overbreathing reduces the circulating ionized magnesium concentration and the intravascular magnesium mass. It is therefore conceivable that extracellular magnesium deficiency is at least a subsidiary cause of the syndrome of tetany and the cardiac complications that are precipitated by hyperventilation.

Mechanisms of the pressor response to intravenous thyrotropin-releasing hormone in the rat.

1. The purpose of this study was to examine the contribution of the sympatho-adrenomedullary system to the blood pressure response to an intravenous bolus of thyrotropin-releasing hormone (TRH) in conscious medullectomized and sham-operated rats. 2. The peak pressor effect of 0.5 mg TRH was significantly increased in rats having no adrenal medulla (+24.2 +/- 1.6 mmHg, mean +/- s.e.m., P < 0.01) as compared to sham-operated animals (+12.2 +/- 3.0 mmHg). 3. Blockade of alpha-adrenergic receptors with phentolamine abolished the pressor effect of TRH in control rats (+2.1 +/- 1.9 mmHg) but did not attenuate the blood pressure response of medullectomized rats (+21.5 +/- 4.7 mmHg). In contrast, beta-blockade with propranolol blunted the blood pressure responsiveness of rats subjected to adrenal medullectomy (+12.4 +/- 2.6 mmHg) but did not modify the effect of TRH in sham-operated controls (+10.9 +/- 2.9 mmHg). 4. The direct in vitro effect of TRH on isolated mesenteric rat arteries was also evaluated. TRH did not induce contractions of isolated arteries. 5. These results suggest that in rats with intact adrenals, the pressor effect of intravenous TRH is mediated primarily by a stimulation of alpha-adrenergic receptors. Adrenal medullectomy appears to enhance the blood pressure response to intravenous TRH. Activation of cardiac beta-adrenoceptors seems to contribute to the blood pressure increasing effect of intravenous TRH in medullectomized animals.

Alteration in neuromuscular function after a 5 km running time trial.

The aim of this study was to characterize the effect of a 5 km running time trial on the neuromuscular properties of the plantar flexors. Eleven well-trained triathletes performed a series of neuromuscular tests before and immediately after the run on a 200 m indoor track. Muscle activation (twitch interpolation) and normalized EMG activity were assessed during maximal voluntary contraction (MVC) of plantar flexors. Maximal soleus H-reflexes and M-waves were evoked at rest (i.e. H (MAX) and M (MAX), respectively) and during MVC (i.e. H (SUP) and M (SUP), respectively). MVC significantly declined (-27%; P < 0.001) after the run, due to decrease in muscle activation (-8%; P < 0.05) and M (MAX)-normalized EMG activity (-13%; P < 0.05). Significant reductions in M-wave amplitudes (M (MAX): -13% and M (SUP): -16%; P < 0.05) as well as H (MAX)/M (MAX) (-37%; P < 0.01) and H (SUP)/M (SUP) (-25%; P < 0.05) ratios occurred with fatigue. Following exercise, the single twitch was characterized by lower peak torque (-16%; P < 0.001) as well as shorter contraction (-19%; P < 0.001) and half-relaxation (-24%; P < 0.001) times. In conclusion, the reduction in plantar flexors strength induced by a 5 km running time trial is caused by peripheral adjustments, which are attributable to a failure of the neuromuscular transmission and excitation-contraction coupling. Fatigue also decreased the magnitude of efferent motor outflow from spinal motor neurons to the plantar flexors and part of this suboptimal neural drive is the result of an inhibition of soleus motoneuron pool reflex excitability.

The relationship between muscle strength and physiological age: a cross-sectional study in boys aged from 11 to 15.

OBJECTIVE: To investigate the relationships between isokinetic knee flexor and extensor muscle strength and physiological and chronological age in young soccer players. MATERIAL AND METHODS: Seventy-nine young, healthy, male soccer players (mean+/-standard deviation age: 12.78+/-2.88, range: 11 to 15) underwent a clinical examination (age, weight, height, body mass index and Tanner puberty stage) and an evaluation of bilateral knee flexor and extensor muscle strength on an isokinetic dynamometer. Participation in the study was voluntary. RESULTS: The peak torque increased progressively (by 50%) between the ages of 11 and 15 and most significantly between 12 to 14. The knee flexor/extensor ratios only decreased significantly between 14 and 15 years of age. Puberty stage was the most important determinant of the peak torque level (ahead of chronological age, weight and height) for all angular velocities (p<0.0001). Muscle strength increased significantly between Tanner stages 1 and 5, with the greatest increase between stages 2 and 4. CONCLUSION: The present study showed that isokinetic muscle strength increases most between 12 and 13 years of age and between Tanner stages 2 and 3. There was strong correlation between muscle strength and physiological age.

Quand les normes font loi : topologie intégrée et processus différenciés de la normalisation internationale

International standardisation refers to voluntary technical specifications pertaining to the production and exchange of goods and services across borders. The paper outlines a theoretical framework which spells out the contention of emerging hybrid forms of non state authority in the global realm. It argues that international standardisation is confronted with a deep rift between promoters of further socialisation of international standards (i.e. a transfer of the universal scope of law into the official framework of standard-setting bodies) and multinational corporations in favour of globalisation of technical standards (i.e. universal recognition of minimal sectorial market-based standards). The problems related to the development of a possible ISO standard of system management in corporate social responsibility provides evidence of the argument.

Spinal modulations accompany peripheral fatigue during prolonged tennis playing.

To examine the time course of alteration in neural process (spinal loop properties) during prolonged tennis playing, 12 competitive players performed a series of neuromuscular tests every 30 min during a 3-h match protocol. Muscle activation (twitch interpolation) and normalized EMG activity were assessed during maximal voluntary contraction (MVC) of plantar flexors. Spinal reflexes and M-waves were evoked at rest (i.e., H(max) and M(max) , respectively) and during MVC (i.e., H(sup) , V-wave, M(sup) , respectively). MVC torque declined significantly (P<0.001) across the match protocol, due to decrease (P<0.001) in muscle activation and in normalized EMG activity. The impairment in MVC was significantly correlated (r=0.77; P<0.05) with the decline in muscle activation. H(max) /M(max) (P<0.001), H(sup) /M(sup) (P<0.01) and V/M(sup) (P<0.05) ratios were depressed with fatigue and decreased by ∼80%, 46% and 61% at the end of exercise, respectively. Simultaneously, peak twitch torque and M-wave amplitude were significantly (P<0.01) altered with exercise, suggesting peripheral alterations. During prolonged tennis playing, the compromised voluntary strength capacity is linked to a reduced neural input to the working muscles. This central activation deficit partly results from a modulation in spinal loop properties.

Is accurate perception of body image associated with appropriate weight-control behavior among adolescents of the Seychelles

BACKGROUND: We examined body image perception and its association with reported weight-control behavior among adolescents in the Seychelles.METHODS: We conducted a school-based survey of 1432 students aging 11-17 years in the Seychelles. Perception of body image was assessed using both a closed-ended question (CEQ) and Stunkard's pictorial silhouettes (SPS). Voluntary attempts to change weight were also assessed.RESULTS: A substantial proportion of the overweight students did not consider themselves as overweight (SPS: 24%, CEQ: 34%), and a substantial proportion of the normal-weight students considered themselves as too thin (SPS: 29%, CEQ: 15%). Logistic regression analysis showed that students with an accurate weight perception were more likely to have appropriate weight-control behavior.CONCLUSIONS: We found that substantial proportions of students had an inaccurate perception of their weight and that weight perception was associated with weight-control behavior. These findings point to forces that can drive the upwards overweight trends.

NanoImpactNet - building a multi-stakeholder dialogue on the health and environmental impact of nanomaterials

NanoImpactNet (NIN) is a multidisciplinary European Commission funded network on the environmental, health and safety (EHS) impact of nanomaterials. The 24 founding scientific institutes are leading European research groups active in the fields of nanosafety, nanorisk assessment and nanotoxicology. This 4-year project is the new focal point for information exchange within the research community. Contact with other stakeholders is vital and their needs are being surveyed. NIN is communicating with 100s of stakeholders: businesses; internet platforms; industry associations; regulators; policy makers; national ministries; international agencies; standard-setting bodies and NGOs concerned by labour rights, EHS or animal welfare. To improve this communication, internet research, a questionnaire distributed via partners and targeted phone calls were used to identify stakeholders' interests and needs. Knowledge gaps and the necessity for further data mentioned by representatives of all stakeholder groups in the targeted phone calls concerned: • the potential toxic and safety hazards of nanomaterials throughout their lifecycles; • the fate and persistence of nanoparticles in humans, animals and the environment; • the associated risks of nanoparticle exposure; • greater participation in: the preparation of nomenclature, standards, methodologies, protocols and benchmarks; • the development of best practice guidelines; • voluntary schemes on responsibility; • databases of materials, research topics and themes, but also of expertise. These findings suggested that stakeholders and NIN researchers share very similar knowledge needs, and that open communication and free movement of knowledge will benefit both researchers and industry. Subsequently a workshop was organised by NIN focused on building a sustainable multi-stakeholder dialogue. Specific questions were asked to different stakeholder groups to encourage discussions and open communication. 1. What information do stakeholders need from researchers and why? The discussions about this question confirmed the needs identified in the targeted phone calls. 2. How to communicate information? While it was agreed that reporting should be enhanced, commercial confidentiality and economic competition were identified as major obstacles. It was recognised that expertise was needed in the areas of commercial law and economics for a wellinformed treatment of this communication issue. 3. Can engineered nanomaterials be used safely? The idea that nanomaterials are probably safe because some of them have been produced 'for a long time', was questioned, since many materials in common use have been proved to be unsafe. The question of safety is also about whether the public has confidence. New legislation like REACH could help with this issue. Hazards do not materialise if exposure can be avoided or at least significantly reduced. Thus, there is a need for information on what can be regarded as acceptable levels of exposure. Finally, it was noted that there is no such thing as a perfectly safe material but only boundaries. At this moment we do not know where these boundaries lie. The matter of labelling of products containing nanomaterials was raised, as in the public mind safety and labelling are connected. This may need to be addressed since the issue of nanomaterials in food, drink and food packaging may be the first safety issue to attract public and media attention, and this may have an impact on 'nanotechnology as a whole. 4. Do we need more or other regulation? Any decision making process should accommodate the changing level of uncertainty. To address the uncertainties, adaptations of frameworks such as REACH may be indicated for nanomaterials. Regulation is often needed even if voluntary measures are welcome because it mitigates the effects of competition between industries. Data cannot be collected on voluntary bases for example. NIN will continue with an active stakeholder dialogue to further build on interdisciplinary relationships towards a healthy future with nanotechnology.

First evaluation of human sperm quality in various geographic regions of Switzerland

A decline in human sperm quality and quantity has been reported in numerous Western countries. This observation was also accompanied by an increase in urogenital malformations. The need for epidemiological studies dealing with unbiased populations in order to understand the causes of these observations is obvious. In Switzerland, the large majority of young men are asked to attend a military camp to be drafted into the army. A few weeks before this camp, conscripts were contacted and invited to participate in a large national study on semen quality. The participation was totally voluntary and anonymous. From September 2005 to June 2007, 770 volunteers filled out a questionnaire, underwent a clinical examination and provided sperm, blood and urine samples. Using self-rated health assessments, the observed cohort could be considered as healthy and no testicular cancer was found. Moreover, the testicular volumes, measured using Prader's orchidometry and ultrasonography, were comparable to those already published for young male populations. The median sperm concentration was 47 x 10(6)/ml, which is close to the concentration reported in Denmark, known to have the highest incidence of testicular cancer in Europe. Statistically significant differences were observed between regions with a lower sperm concentration for men residing in the Alps (43 x 10(6)/ml) and in the Zurich area (36 x 10(6)/ml) compared to men from West Plateau (54 x 10(6)/ml) and from the Jura (54 x 10(6)/ml). Such a regional discrepancy could be related to environmental factors, including endocrine disruptors. In order to confirm such regional differences more volunteers from the already studied regions should be studied and other parts of the country should be investigated. The rather low sperm concentration of Swiss young volunteers should be considered as a national health issue and investigated further.

Primary motor cortex inhibition in spinal cord injuries.

OBJECTIVES AND METHODS: Excitability changes in the primary motor cortex in 17 spinal-cord injured (SCI) patients and 10 controls were studied with paired-pulse transcranial magnetic stimulation. The paired pulses were applied at inter-stimulus intervals (ISI) of 2 ms and 15 ms while motor evoked potentials (MEP) were recorded in the biceps brachii (Bic), the abductor pollicis brevis (APB) and the tibialis anterior (TA) muscles. RESULTS: The study revealed a significant decrease in cortical motor excitability in the first weeks after SCI concerning the representation of both the affected muscles innervated from spinal segments below the lesion, and the spared muscles rostral to the lesion. In the patients with motor-incomplete injury, but not in those with motor-complete injury, the initial cortical inhibition of affected muscles was temporarily reduced 2-3 months following injury. The degree of inhibition in cortical areas representing the spared muscles was observed to be smaller in patients with no voluntary TA activity compared to patients with some activity remaining in the TA. Surprisingly, motor-cortical inhibition was observed not only at ISI 2 ms but also at ISI 15 ms. The inhibition persisted in patients who returned for a follow-up measurement 2-3 years later. CONCLUSION: The present data showed different evaluation of cortical excitability between patients with complete and incomplete spinal cord lesion. Our results provide more insight into the pathophysiology of SCI and contribute to the ongoing discussion about the recovery process and therapy of SCI patients.

Assessing patient's belief on HIV testing before elective surgery: room for improvements in Switzerland !

Background: ln Switzerland no HIV test is performed without the patient's consent based on a Voluntary Counseling and Testing policy (VCT). We hypothesized that a substantial proportion of patients going through an elective surgery falsely believed that an HIV test was performed on a routine basis and that the lack of transmission of result was interpreted as being HIV negative. Method: All patients with elective orthopedic surgery during 2007 were contacted by phone in 2008. A structured questionnaire assessed their belief about routine preoperative blood analysis (diabetes, coagulation function, HIV test and cholesterol level) as well as result awareness and interpretation. Variables included age and gender. Analysis were conducted using the software JMP 6.0.3. Results: 1123 patients were included. 130 (12 %) were excluded (Le. unreachable, unable to communicate on the phone, not operated). 993 completed the survey (89 %). Median age was 51 (16-79). 50 % were female. 376 (38 %) patients thought they had an HIV test performed before surgery but none of them had one. 298 (79 %) interpreted the absence of result as a negative HIV test. A predictive factor to believe an HIV test had been done was an age below 50 years old (45 % vs 33 % for 16-49 years old and 50-79 years old respectively, p < 0.001). No difference was observed between genders. Conclusion: ln Switzerland, nearly 40 % of the patients falsely thought an HIV test had been performed on a routine basis before surgery and were erroneously reassured about their HIV status. These results should either improve the information given to the patient regarding preoperative exams, or motivate public health policy to consider HIV opt-out screening instead of VCT strategy.

MR-based follow-up of the superior cerebellar artery after radiosurgery for trigeminal neuralgia.

Purpose: To study with a non invasive method any potential radiological change on the superior cerebellar artery (SCA) in patients treated radiosurgically for classic trigeminal neuralgia (CTN).Materials and methods: A retrospective measure of maximal dose received by SCA was performed analyzing the treatment planning in 55 consecutive patients treated by Gamma Knife radiosurgery for an CTN, then, a prospective study was designed using high resolution MR, with T2 SPIR, T1 without and with gadolinium enhancement, Proton density, 3D TONE and MIP reconstructions. Inclusion criteria were: patients followed at our institution, follow-up of one year or more, dose received by the SCA of 15 Gy or more and voluntary patient participation in the study. Patients with repeated Gamma Knife radiosurgery for failure or recurrence were excluded. The end points were: SCA occlusion, stenosis or infarction in the territory supplied by SCA.Results: Sixteen patients were studied, with a mean follow-up of 25.2 months (12-42 months). The mean maximal dose received by the SCA was 57.5 Gy. (15-87 Gy). Among these 16 patients studied, neither obstruction of the SCA nor infarction was demonstrated. In one patient a suspicion of asymptomatic SCA stenosis was visualized distant to the irradiation field.Conclusions: SCA can receive a high dose of irradiation during radiosurgical treatment for CTN. This study does not confirm any vascular damage to the SCA after radiosurgery for CTN. (C) 2011 Elsevier B.V. All rights reserved.

Dépistage du VIH: les médecins aux urgences de Suisse Romande connaissent-­‐ils les recommandations suisses ?

Contexte¦Le VIH reste une des préoccupations majeures de santé publique dans le monde. Le nombre de patients infectés en Europe continue de croître et s'élève, en 2008, à 2.3 millions (1). De plus, environ 30 % des personnes séropositives ignorent leur statut et, de ce fait, contribuent à la propagation de l'épidémie. Ces patients sont responsables de la moitié des nouveaux cas du VIH (2) ; ils transmettent, en effet, 3.5 fois plus l'infection que les patients dont le diagnostic est connu (3).¦Aux USA, en raison de l'épidémiologie actuelle du VIH, les Centers for Disease Control and Prevention (CDC) ont, en septembre 2006, mis le point sur la nécessité d'étendre drastiquement les tests et, de ce fait, ont publié de nouvelles recommandations. Non seulement, le test devra dépasser les groupes à risque dans les zones à grande prévalence mais aussi, être répandu à toute la population adulte de 13 à 64 ans sauf si la prévalence du VIH est en dessous de 0.1 % (4). Cette démarche est appelée routine opt-out HIV screening et plusieurs arguments parlent en faveur d'un dépistage systématique. Cette maladie rempli tout d'abord les 4 critères pour l'introduction d'un dépistage systématique : une maladie grave pouvant être mise en évidence avant l'apparition des symptômes, son diagnostic améliore la survie par une progression moins rapide et diminution de la mortalité, des tests de dépistage sensibles et spécifiques sont disponibles et les coûts sont moindres en comparaison aux bénéfices (5). Aux USA, 73 % des patients diagnostiqués à un stade avancé de l'infection VIH entre 2001 et 2005 avaient eu recours à l'utilisation des systèmes de soins au moins une fois dans les 8 ans précédant le diagnostic (6). Ces occasions manquées font aussi partie des arguments en faveur d'un dépistage systématique. En règle générale, le médecin se basant uniquement sur les symptômes et signes, ainsi que sur l'anamnèse sexuelle sous-estime la population à tester. Ce problème de sélection des candidats n'a plus lieu d'être lors d'un tel screening. Après cette publication des recommandations du CDC, qui introduit le dépistage systématique, il a été constaté que seulement 1/3 du personnel soignant interrogé connaissait les nouvelles directives et seulement 20 % offrait un dépistage de routine à tous les patients concernés (7). Cette étude nous montre alors qu'il est impératif de vérifier le niveau de connaissances des médecins après la publication de nouvelles recommandations.¦Devant le problème de l'épidémie du VIH, la Suisse opte pour une stratégie différente à celle des Etats-Unis. La Commission d'experts clinique et thérapie VIH et SIDA (CCT) de l'OFSP a tout d'abord publié, en 2007, des recommandations destinées à diminuer le nombre d'infections VIH non diagnostiquées, grâce à un dépistage initié par le médecin (8). Cette approche, appelée provider initiated counselling and testing (PICT), complétait alors celle du voluntary counselling and testing (VCT) qui préconisait un dépistage sur la demande du patient. Malheureusement, le taux d'infections diagnostiquées à un stade avancé a stagné aux environs de 30 % jusqu'en 2008 (9), raison pour laquelle l'OFSP apporte, en 2010, des modifications du PICT. Ces modifications exposent différentes situations où le test du VIH devrait être envisagé et apportent des précisions quant à la manière de le proposer. En effet, lors d'une suspicion de primo-infection, le médecin doit expliquer au patient qu'un dépistage du VIH est indiqué, un entretien conseil est réalisé avec des informations concernant la contagiosité élevée du virus à ce stade de l'infection. Si le patient présente un tableau clinique qui s'inscrit dans le diagnostic différentiel d'une infection VIH, le médecin propose le test de manière systématique. Il doit alors informer le patient qu'un tel test sera effectué dans le cadre d'une démarche diagnostique, sauf si celui-ci s'y oppose. Enfin, dans d'autres situations telles que sur la demande du patient ou si celui-ci fait partie d'un groupe de population à grande prévalence d'infection VIH, le médecin procède à une anamnèse sexuelle, suivie d'un entretien conseil et du test si l'accord explicite du patient a été obtenu (10).¦Nous pouvons donc constater les différentes stratégies face à l'épidémie du VIH entre les USA et la Suisse. Il est nécessaire d'évaluer les conséquences de ces applications afin d'adopter la conduite la plus efficace en terme de dépistage, pour amener à une diminution des transmissions, une baisse de la morbidité et mortalité. Aux USA, des études ont été faites afin d'évaluer l'impact de l'approche opt-out qui montrent que le screening augmente la probabilité d'être diagnostiqué (11). En revanche, en Suisse, aucune étude de ce type n'a été entreprise à l'heure actuelle. Nous savons également qu'il existe un hiatus entre la publication de nouvelles recommandations et l'application de celles-ci en pratique. Le 1er obstacle à la mise en oeuvre des guidelines étant leur méconnaissance (12), il est alors pertinent de tester les connaissances des médecins des urgences d'Hôpitaux de Suisse au sujet des nouvelles recommandations sur le dépistage du VIH de l'OFSP de mars 2010.¦Objectifs¦Montrer que les recommandations de l'OFSP de mars 2010 ne sont pas connues des médecins suisses.¦Méthodes¦Nous testerons la connaissance des médecins concernant ces recommandations via un questionnaire qui sera distribué lors d'un colloque organisé à cet effet avec tous les médecins du service des urgences d'un même établissement. Il n'y aura qu'une séance afin d'éviter d'éventuels biais (transmission d'informations d'un groupe à un autre). Ils recevront tout d'abord une lettre informative, accompagnée d'un formulaire de consentement pour l'utilisation des données de manière anonyme. La feuille d'information est rédigée de façon à ne pas influencer les candidats pour les réponses aux questions. Le questionnaire comprend deux parties, une première qui comprend divers cas cliniques. Les candidats devront dire si ces situations se trouvent, selon eux, dans les nouvelles recommandations de l'OFSP en termes de dépistage du VIH et indiquer la probabilité d'effectuer le test en pratique. La deuxième partie interrogera sur la manière de proposer le test au patient. La durée nécessaire pour remplir le questionnaire est estimée à 15 minutes.¦Le questionnaire élaboré avec la collaboration de Mme Dubois de l'UMSP à Lausanne et vont être testés par une vingtaine de médecins de premier recours de Vidy Med et Vidy Source, deux centres d'urgences lausannois.¦Réstulats escomptés¦Les médecins suisse ne sont pas au courant des nouvelles recommandations concernant le dépistage du VIH.¦Plus-value escomptée¦Après le passage du questionnaire, nous ferons une succincte présentation afin d'informer les médecins au sujet de ces recommandations. Aussi, l'analyse des résultats du questionnaire nous permettra d'agir au bon niveau pour que les nouvelles recommandations de l'OFSP de mars 2010 soient connues et appliquées, tout en ayant comme objectif l'amélioration du dépistage du VIH.

Physiopathologie du coeur embryonnaire soumis à l'anoxie-réoxygénation: rôle protecteur du NO et des canaux KATP

RESUME Introduction : Dans le coeur adulte, l'ischémie et la reperfusion entraînent des perturbations électriques, mécaniques, biochimiques et structurales qui peuvent causer des dommages réversibles ou irréversibles selon la sévérité de l'ischémie. Malgré les récents progrès en cardiologie et en chirurgie foetales, la connaissance des mécanismes impliqués dans la réponse du myocarde embryonnaire à un stress hypoxique transitoire demeure lacunaire. Le but de ce travail a donc été de caractériser les effets chrono-, dromo- et inotropes de l'anoxie et de la réoxygénation sur un modèle de coeur embryonnaire isolé. D'autre part, les effets du monoxyde d'azote (NO) et de la modulation des canaux KATP mitochondriaux (mito KATP) sur la récupération fonctionnelle postanoxique ont été étudiés. La production myocardique de radicaux d'oxygène (ROS) et l'activité de MAP Kinases (ERK et JNK) impliquées dans la signalisation cellulaire ont également été déterminées. Méthodes : Des coeurs d'embryons de poulet âgés de 4 jours battant spontanément ont été placés dans une chambre de culture puis soumis à une anoxie de 30 min suivie d'une réoxygénation de 60 min. L'activité électrique (ECG), les contractions de l'oreillette, du ventricule et du conotroncus (détectées par photométrie), la production de ROS (mesure de la fluorescence du DCFH) et l'activité kinase de ERK et JNK dans le ventricule ont été déterminées au cours de l'anoxie et de la réoxygénation. Les coeurs ont été traités avec un bloqueur des NO synthases (L-NAME), un donneur de NO (DETA-NONOate), un activateur (diazoxide) ou un inhibiteur (5-HD) des canaux mitoKATP un inhibiteur non-spécifique des PKC (chélérythrine) ou un piégeur de ROS (MPG). Résultats : L'anoxie et la réoxygénation entraînaient des arythmies (essentiellement d'origine auriculaire) semblables à celles observées chez l'adulte, des troubles de la conduction (blocs auriculo-ventriculaires de 1er, 2ème et 3ème degré) et un ralentissement marqué du couplage excitation-contraction (E-C) ventriculaire. En plus de ces arythmies, la réoxygénation déclenchait le phénomène de Wenckelbach, de rares échappements ventriculaires et une sidération myocardique. Aucune fibrillation, conduction rétrograde ou activité ectopique n'ont été observées. Le NO exogène améliorait la récupération postanoxique du couplage E-C ventriculaire alors que L'inhibition des NOS la ralentissait. L'activation des canaux mito KATP augmentait la production mitochondriale de ROS à la réoxygénation et accélérait la récupération de la conduction (intervalle PR) et du couplage E-C ventriculaire. La protection de ce couplage était abolie par le MPG, la chélérythrine ou le L-NAME. Les fonctions électrique et contractile de tous les coeurs récupéraient après 30-40 min de réoxygénation. L'activité de ERK et de JNK n'était pas modifiée par L'anoxie, mais doublait et quadruplait, respectivement, après 30 min de réoxygénation. Seule l'activité de JNK était diminuée (-60%) par l'activation des canaux mitoKATP. Cet effet inhibiteur était partiellement abolit par le 5-HD. Conclusion: Dans le coeur immature, le couplage E-C ventriculaire semble être un paramètre particulièrement sensible aux conditions d'oxygénation. Sa récupération postanoxique est améliorée par l'ouverture des canaux mitoKATP via une signalisation impliquant les ROS Ies PKC et le NO. Une réduction de l'activité de JNK semble également participer à cette protection. Nos résultats suggèrent que les mitochondries jouent un rôle central dans la modulation des voies de signalisation cellulaire, en particulier lorsque les conditions métaboliques deviennent défavorables. Le coeur embryonnaire isolé représente donc un modèle expérimental utile pour mieux comprendre les mécanismes associés à une hypoxie in utero et pour améliorer les stratégies thérapeutiques en cardiologie et chirurgie foetales. ABSTRACT Physiopathology of the anoxic-reoxygenated embryonic heart: Protective role of NO and KATP channel Aim: In the adult heart, the electrical, mechanical, biochemical and structural disturbances induced by ischemia and reperfusion lead to reversible or irreversible damages depending on the severity and duration of ischemia. In spite of recent advances in fetal cardiology and surgery, little is known regarding the cellular mechanisms involved in hypoxia-induced dysfunction in the developing heart. The aim of this study was to precisely characterize the chrono-, dromo- and inotropic disturbances associated with anoxia-reoxygenation in an embryonic heart model. Furthermore, the roles that nitric oxide (NO), reactive oxygen species (ROS), mitochondrial KATP, (mito KATP) channel and MAP Kinases could play in the stressed developing heart have been investigated. Methods: Embryonic chick hearts (4-day-old) were isolated and submitted in vitro to 30 min anoxia followed by 60 min reoxygenation. Electrical (ECG) and contractile activities of atria, ventricle and conotruncus (photometric detection), ROS production (DCFH fluorescence) and ERK and JNK activity were determined in the ventricle throughout anoxia-reoxygenation. Hearts were treated with NO synthase inhibitor (L-NAME), NO donor (DETA-NONOate), mitoKATP channel opener (diazoxide) or blocket (5-HD), PKC inhibitor (chelerythrine) and ROS scavenger (MPG). Results: Anoxia and reoxygenation provoked arrhythxnias (mainly originating from atrial region), troubles of conduction (st, 2nd, and 3rd degree atrio-ventricular blocks) and disturbances of excitation-contraction (E-C) coupling. In addition to these types of arrhythmias, reoxygenation triggered Wenckebach phenomenon and rare ventricular escape beats. No fibrillations, no ventricular ectopic beats and no electromechanical dissociation were observed. Myocardial stunning was observed during the first 30 min of reoxygenation. All hearts fully recovered their electrical and mechanical functions after 30-40 min of reoxygenation. Exogenous NO improved while NOS inhibition delayed E-C coupling recovery. Mito KATP, channel opening increased reoxygenation-induced ROS production and improved E-C coupling and conduction (PR) recovery. MPG, chelerythrine or L-NAME reversed this effect. Reoxygenation increased ERK and JNK activities land 4-fold, respectively, while anoxia had no effect. MitoKATP channel opening abolished the reoxygenation-induced activation of JNK but had no effect on ERK activity. This inhibitory effect was partly reversed by mitoKATP channel blocker but not by MPG. Conclusion: In the developing heart, ventricular E-C coupling was found to be specially sensitive to hypoxia-reoxygenation and its postanoxic recovery was improved by mitoKATP channel activation via a ROS-, PKC- and NO-dependent pathway. JNK inhibition appears to be involved in this protection. Thus, mitochondria can play a pivotal role in the cellular signalling pathways, notably under critical metabolic conditions. The model of isolated embryonic heart appears to be useful to better understand the mechanisms underlying the myocardial dysfunction induced by an in utero hypoxia and to improve therapeutic strategies in fetal cardiology and surgery.

Response of embryonic heart to hypoxia and reoxygenation: an in vitro model

OBJECTIVES: To define properly the consequences of oxygen deprivation and readmission for the functioning of the developing heart. METHODS: Spontaneously beating hearts excised from three-day-old chick embryos were loaded with a drop of viscous nontoxic silicone oil and cultured in a special chamber in which variations of PO2 at the tissue level could be strictly controlled. All parts of the hearts were simultaneously submitted to identical changes in PO2. Instantaneous heart rate, myocardial shortening, velocities of contraction and relaxation, and mechanical propagation along the heart tube were determined photometrically. RESULTS: The hearts, submitted to a PO2 ramp (0 to 9.3 kPa) or absolute anoxia, reacted rapidly, reversibly and reproducibly. Under sustained anoxia, ventricular activity stopped after 3.8±0.7 mins (n=4) and then resumed intermittently in the form of tachycardic bursts. Brief anoxia (1 min) provoked tachycardia followed by bradycardia, induced contracture, depressed contractility and retarded atrioventricular propagation. Upon reoxygenation, ventricular contractions ceased suddently for 20±11 s (n=5), whereas a residual atrial activity could persist. The duration of this arrest and the rate of recovery depended on duration of the preceding anoxia. Such a dysfunction constitutes the embryonic analogue of the oxygen paradox observed in adult hearts. Initial impulses, including arrhythmic activity, originated exclusively from the atrium, and no ventricular ectopic beats were detected whatever the conditions of oxygenation. CONCLUSIONS: This in vitro model seems promising for studying the pathophysiological mechanisms associated with hypoxia and reoxygenation in the developing heart.