[Characterisation of the protective role of erythropoetin in a murine model of acute lung injury]. / Caractérisation du rôle protecteur de l'érythropoïétine dans un modèle murin d'agression pulmonaire aiguë.

In addition to its role in erythropoiesis, erythropoietin (Epo) plays a role in tissue protection, which includes cardioprotective, nephroprotective and neuroprotective effects. The presence of Epo and its receptor (Epo-R) in pulmonary tissue also suggests a cytoprotective effect of Epo in the lung. Our project aims to document this role in a murine model under-expressing Epo. The obtained results will lead to a better understanding of the cytoprotective effects of Epo and will also give an appreciation of its beneficial effects in cases of lung injury.

The kreisler mutation leads to the loss of intrinsically hypoxia-activated spots in the region of the retrotrapezoid nucleus/parafacial respiratory group.

Acute hypoxia elicits a biphasic respiratory response characterized in the newborn by a transient hyperventilation followed by a severe decrease in respiratory drive known as hypoxic respiratory depression. Medullary O(2) chemosensitivity is known to contribute to respiratory depression induced by hypoxia, although precise involvement of cell populations remains to be determined. Having a thorough knowledge of these populations is of relevance because perturbations in the respiratory response to hypoxia may participate in respiratory diseases in newborns. We aimed to analyze the hypoxic response of ponto-medullary cell populations of kreisler mutant mice. These mice have defects in a gene expressed in two rhombomeres encompassing a part of the medulla oblongata implicated in hypoxic respiratory depression. Central responses to hypoxia were analyzed in newborn mice by measuring respiratory rhythm in ex vivo caudal pons-medullary-spinal cord preparations and c-fos expression in wild-type and kreisler mutants. The homozygous kreisler mutation, which eliminates most of rhombomere 5 and mis-specifies rhombomere 6, abolished (1) an early decrease in respiratory frequency within 10 min of hypoxia and (2) an intrinsic hypoxic activation, which is characterized by an increase in c-fos expression in the region of the ventral medullary surface encompassing the retrotrapezoid nucleus/parafacial respiratory group expressing Phox2b. This increase in c-fos expression persisted in wild-type Phox2b-negative and Phox2b-positive cells after blockade of synaptic transmission and rhythmogenesis by a low [Ca(2+)](0). Another central response was retained in homozygous kreisler mutant mice; it was distinguished by (1) a delayed (10-30 min) depression of respiratory frequency and (2) a downregulation of c-fos expression in the ventrolateral reticular nucleus of the medulla, the nucleus of the solitary tract, and the area of the A5 region. Thus, two types of ponto-medullary cell groups, with distinct anatomical locations, participate in central hypoxic respiratory depression in newborns.

Early abnormalities of post-sigh breathing in a mouse model of Rett syndrome.

Rett syndrome is a neurodevelopmental disease accompanied by complex, disabling symptoms, including breathing symptoms. Because Rett syndrome is caused by mutations in the transcriptional repressor methyl-CpG-binding protein 2 (MeCP2), Mecp2-deficient mice have been generated as experimental model. Males of Mecp2-deficient mice (Mecp2(-/y)) breathe normally at birth but show abnormal respiratory responses to hypoxia and hypercapnia from postnatal day 25 (P25). After P30, Mecp2(-/y) mice develop breathing symptoms reminiscent of Rett syndrome, aggravating until premature death at around P60. Using plethysmography, we analyzed the sighs and the post-sigh breathing pattern of unrestrained wild type male mice (WT) and Mecp2(-/y) mice from P15 to P60. Sighs are spontaneous large inspirations known to prevent lung atelectasis and to improve alveolar oxygenation. However, Mecp2(-/y) mice show early abnormalities of post-sigh breathing, with long-lasting post-sigh apnoeas, reduced tidal volume when eupnoea resumes and lack of post-sigh bradypnoea which develop from P15, aggravate with age and possibly contribute to breathing symptoms to come.

Hypoxia-sensing properties of the newborn rat ventral medullary surface in vitro.

The ventral medullary surface (VMS) is a region known to exert a respiratory stimulant effect during hypercapnia. Several studies have suggested its involvement in the central inhibition of respiratory rhythm caused by hypoxia. We studied brainstem-spinal cord preparations isolated from newborn rats transiently superfused with a very low O(2) medium, causing reversible respiratory depression, to characterize the participation of the VMS in hypoxic respiratory adaptation. In the presence of 0.8 mM Ca(2+), very low O(2) medium induced an increase in c-fos expression throughout the VMS. The reduction of synaptic transmission and blockade of the respiratory drive by 0.2 mM Ca(2+)-1.6 mM Mg(2+) abolished c-fos expression in the medial VMS (at the lateral edge of the pyramidal tract) but not in the perifacial retrotrapezoid nucleus/parafacial respiratory group (RTN/pFRG) VMS, suggesting the existence of perifacial RTN/pFRG hypoxia-sensing neurons. In the presence of Ca(2+) (0.8 mM), lesioning experiments suggested a physiological difference in perifacial RTN/pFRG VMS between the lateral VMS (beneath the ventrolateral part of the facial nucleus) and the middle VMS (beneath the ventromedial part of the facial nucleus), at least in newborn rats. The lateral VMS lesion, corresponding principally to the most rostral part of the pFRG, produced hypoxia-induced stimulation, whereas the middle VMS lesion, corresponding to the main part of the RTN, abolished hypoxic excitation. This may involve relay via the medial VMS, which is thought to be the parapyramidal group.

Diencephalic and mesencephalic influences on ponto-medullary respiratory control in normoxic and hypoxic conditions: an in vitro study on central nervous system preparations from newborn rat.

We investigated the effects of the diencephalon and mesencephalon on the central respiratory drive originating from ponto-medullary regions in normoxic and hypoxic conditions, using central nervous system preparations from newborn rats. We used two approaches: 1) electrophysiological analysis of respiratory frequency and the amplitude of inspiratory C4 activity and 2) immunohistochemical detection of Fos protein, an activity-dependent neuronal marker. We found that, in normoxic conditions, the mesencephalon moderated respiratory frequency, probably by means of an inhibitory effect on ventral medullary respiratory neurons. Diencephalic inputs restored respiratory frequency. Moreover, O(2)-sensing areas in the diencephalon (caudal lateral and posterior hypothalamic areas) and mesencephalon (ventrolateral and dorsolateral periaqueductal gray) seem to increase the amplitude of respiratory bursts during adaptation of the central respiratory drive to hypoxia. In contrast, decrease in respiratory frequency during hypoxia is thought to be mediated by a cluster of ventral hypothalamic neurons.