[Use of murine models for the study of obstructive sleep apnea syndrome in Down syndrome]. / Utilisation des modèles murins pour l'étude du syndrome d'apnées obstructives du sommeil dans le syndrome de Down.

Down syndrome (DS), or trisomy 21, is a genetic disorder caused by the presence of an extra copy of chromosome 21, leading to various characteristic physical features as well as developmental and cognitive delays. Obstructive sleep apnea syndrome (OSAS) is a common disorder in both adult and pediatric patients with DS. Several characteristics of DS may contribute to the development or worsening of OSAS. Numerous murine models of DS exist. A number of studies have explored apneas and the risk of upper airway obstruction in these models, but up until now, only in adulthood.

Maternal dietary omega-3 deficiency worsens the deleterious effects of prenatal inflammation on the gut-brain axis in the offspring across lifetime.

Maternal immune activation (MIA) and poor maternal nutritional habits are risk factors for the occurrence of neurodevelopmental disorders (NDD). Human studies show the deleterious impact of prenatal inflammation and low n-3 polyunsaturated fatty acid (PUFA) intake on neurodevelopment with long-lasting consequences on behavior. However, the mechanisms linking maternal nutritional status to MIA are still unclear, despite their relevance to the etiology of NDD. We demonstrate here that low maternal n-3 PUFA intake worsens MIA-induced early gut dysfunction, including modification of gut microbiota composition and higher local inflammatory reactivity. These deficits correlate with alterations of microglia-neuron crosstalk pathways and have long-lasting effects, both at transcriptional and behavioral levels. This work highlights the perinatal period as a critical time window, especially regarding the role of the gut-brain axis in neurodevelopment, elucidating the link between MIA, poor nutritional habits, and NDD.

Melatonin prevents learning disorders in brain-lesioned newborn mice.

Perinatal brain injuries often result in irreversible learning disabilities, which manifest in early childhood. These injuries are chiefly ascribable to marked susceptibility of the immature brain to glutamate-induced excitotoxicity. No treatments are available. One well-characterized model of perinatal brain injuries consists in injecting the glutamate analog ibotenate into the brain of 5-day-old mice. The resulting excitotoxic lesions resemble the hypoxic-ischemic gray-matter lesions seen in full-term and near-term newborns, as well as the white-matter lesions of preterm newborns. We previously reported that these lesions disrupted odor preference conditioning in newborn mice. The aim of this study was to assess the effectiveness of the neuroprotector melatonin in preventing learning disabilities in newborn mice with ibotenate-induced brain injury. In postnatal day (P) 6-P7 pups, we tested psychomotor reflexes, spontaneous preference for maternal odors as an index of memory, ultrasonic vocalization responses to stroking as an index of sensitivity to tactile stimuli, and conditioned preference for an odor previously paired with stroking as an index of learning abilities. Without melatonin, conditioning was abolished, whereas spontaneous odor preference, psychomotor reflexes, and sensitivity to tactile stimuli were normal. Thus, abolition of conditioning was not associated with sensorimotor impairments. Histological analysis confirmed the efficacy of melatonin in reducing white-matter lesions induced by ibotenate. Furthermore, treatment with melatonin protected the ability to develop conditioning. Thus, melatonin, which easily crosses the blood-brain barrier and has been proven safe in children, may be effective in preventing learning disabilities caused by perinatal brain injuries in human preterm infants.

Effects of temperature on ventilatory response to hypercapnia in newborn mice heterozygous for transcription factor Phox2b.

Congenital central hypoventilation syndrome (CCHS) is a rare disease with variable severity, generally present from birth and chiefly characterized by impaired chemosensitivity to hypercapnia. The main cause of CCHS is a mutation in the PHOX2B gene, which encodes a transcription factor involved in the development of autonomic medullary reflex pathways. Temperature regulation is abnormal in many patients with CCHS. Here, we examined whether ambient temperature influenced CO(2) sensitivity in a mouse model of CCHS. A weak response to CO(2) at thermoneutrality (32 degrees C) was noted previously in 2-day-old mice with an invalidated Phox2b allele (Phox2b+/-), compared with wild-type littermates. We exposed Phox2b+/- pups to 8% CO(2) at three ambient temperatures (TAs): 29 degrees C, 32 degrees C, and 35 degrees C. We measured breathing variables and heart rate (HR) noninvasively using a novel whole body flow plethysmograph equipped with contact electrodes. Body temperature and baseline breathing increased similarly with TA in mutant and wild-type pups. The hypercapnic ventilatory response increased linearly with TA in both groups, while remaining smaller in mutant than in wild-type pups at all TAs. The differences between the absolute increases in ventilation in mutant and wild-type pups become more pronounced as temperature increased above 29 degrees C. The ventilatory abnormalities in mutant pups were not associated with significant impairments of heart rate control. In both mutant and wild-type pups, baseline HR increased with TA. In conclusion, TA strongly influenced the hypercapnic ventilatory response in Phox2b+/- mutant mice. These findings suggest that abnormal temperature regulation may contribute to the severity of respiratory impairments in CCHS patients.

Inhibitory effects of repeated hyperoxia on breathing in newborn mice.

Brief oxygen therapy is commonly used for resuscitation at birth or prevention of hypoxaemia before procedures during the neonatal period. However, O(2) may severely depress breathing, especially when administered repeatedly. The aim of the present study was to test the effects of repeated hyperoxia on breathing control in newborn mice. A total of 97 Swiss mouse pups were assigned to O(2) or air on post-natal day 0, 1 or 2. Each pup in the O(2) group was subjected to four hyperoxic tests (100% O(2) for 3 min followed by 12 min normoxia), whereas pups in the air group were maintained in normoxia. Breathing variables were measured using flow-through barometric plethysmography. O(2) significantly decreased minute ventilation as seen in a decrease in respiratory rate. This decrease became significantly larger with repeated exposure and ranged -17- -26% for all ages combined. Furthermore, hyperoxia increased total apnoea duration, as compared with the baseline value. In newborn mice, repeated hyperoxia increasingly depressed breathing. This finding further supports a need for stringent control of oxygen therapy, most notably repeated oxygen administration in the neonatal period for premature newborn infants and those carried to term.

Treatment-induced prevention of learning deficits in newborn mice with brain lesions.

Perinatal brain injuries often result in irreversible learning disabilities, which manifest in early childhood. The molecular and cellular mechanisms of these injuries and potential pharmacological treatments are emerging, chiefly from studies in newborn rodents. In newborn mice, experimentally induced lesions can be dramatically reduced by appropriate neuroprotective treatments. However, the early effectiveness of these treatments in preserving cognition remained unknown. Here, we addressed this issue by using intracerebral ibotenate to induce excitotoxic brain lesions in 5-day-old mice (postnatal day 5). On postnatal days 6-7, we tested spontaneous preference for maternal odors, as an index of odor memory, and conditioned preference for an artificial odor previously paired with stroking, as an index of associative learning. Brain-lesioned newborn mice showed normal general status and preference for maternal odors. In contrast, odor conditioning was severely impaired. A previous study showed that fructose 1,6-biphosphate acted as a neuroprotective agent which significantly reduced neocortical lesion size. In the present study, treating the newborn mice with fructose 1,6-biphosphate 15 min before the ibotenate injection reduced neocortical lesion size and restored conditioning. This demonstrates, for the first time, that neuroprotective treatment can protect some features of early cognition.

Ventilatory response to hyperoxia in newborn mice heterozygous for the transcription factor Phox2b.

Heterozygous mutations of the transcription factor PHOX2B have been found in most patients with central congenital hypoventilation syndrome, a rare disease characterized by sleep-related hypoventilation and impaired chemosensitivity to sustained hypercapnia and sustained hypoxia. PHOX2B is a master regulator of autonomic reflex pathways, including peripheral chemosensitive pathways. In the present study, we used hyperoxic tests to assess the strength of the peripheral chemoreceptor tonic drive in Phox2b+/-newborn mice. We exposed 69 wild-type and 67 mutant mice to two hyperoxic tests (12-min air followed by 3-min 100% O2) 2 days after birth. Breathing variables were measured noninvasively using whole body flow plethysmography. The initial minute ventilation decrease was larger in mutant pups than in wild-type pups: -37% (SD 13) and -25% (SD 18), respectively, P<0.0001. Furthermore, minute ventilation remained depressed throughout O2 exposure in mutants, possibly because of their previously reported impaired CO2 chemosensitivity, whereas it returned rapidly to the normoxic level in wild-type pups. Hyperoxia considerably increased total apnea duration in mutant compared with wild-type pups (P=0.0001). A complementary experiment established that body temperature was not influenced by hyperoxia in either genotype group and, therefore, did not account for genotype-related differences in the hyperoxic ventilatory response. Thus partial loss of Phox2b function by heterozygosity did not diminish the tonic drive from peripheral chemoreceptors.

Automatic classification of activity and apneas using whole body plethysmography in newborn mice.

An increasing number of studies in newborn mice are being performed to determine the mechanisms of sleep apnea, which is the hallmark of early breathing disorders. Whole body plethysmography is the method of choice, as it does not require immobilization, which affects behavioral states and breathing. However, activity inside the plethysmograph may disturb the respiratory signal. Visual classification of the respiratory signal into ventilatory activity, activity-related disturbances, or apneas is so time-consuming as to considerably hamper the phenotyping of large pup samples. We propose an automatic classification of activity based on respiratory disturbances and of apneas based on spectral analysis. This method was validated in newborn mice on the day of birth and on postnatal days 2, 5, and 10, under normoxic and hypoxic (5% O(2)) conditions. For both activity and apneas, visual and automatic scores showed high Pearson's correlation coefficients (0.92 and 0.98, respectively) and high intraclass correlation coefficients (0.96-0.99), supporting strong agreement between the two methods. The present results suggest that breathing disturbances may provide a valid indirect index of activity in freely moving newborn mice and that automatic apnea classification based on spectral analysis may be efficient in terms of precision and of time saved.