We tested the hypothesis that neonatal maternal separation (NMS), a form of stress that affects hypothalamo-pituitary-adrenal axis (HPA) function in adult rats, alters development of the respiratory control system. Pups subjected to NMS were placed in a temperature and humidity controlled incubator 3 h per day for 10 consecutive days (P3 to P12). Control pups were undisturbed. Once they reached adulthood (8-10 weeks old), rats were placed in a plethysmography chamber for measurement of ventilatory and cardiovascular parameters under normoxic and hypoxic conditions. Measurement of c-fos mRNA expression in the paraventricular nucleus of the hypothalamus (PVH) combined with plasma ACTH and corticosterone levels confirmed that NMS effectively disrupted HPA axis function in males. In males, baseline minute ventilation was not affected by NMS. In contrast, NMS females show a greater resting minute ventilation due to a larger tidal volume. The hypoxic ventilatory response of male NMS rats was 25% greater than controls, owing mainly to an increase in tidal volume response. This augmentation of the hypoxic ventilatory response was sex-specific also because NMS females show an attenuated minute ventilation increase. Baseline mean arterial blood pressure of male NMS rats was 20% higher than controls. NMS-related hypertension was not significant in females. The mechanisms underlying sex-specific disruption of cardio-respiratory control in NMS rats are unknown but may be a consequence of the neuroendocrine disruption associated with NMS. These data indicate that exposure to a non-respiratory stress during early life elicits significant plasticity of these homeostatic functions which persists until adulthood.

Genest S-E, Gulemetova R, Laforest S, Drolet G, Kinkead R. Neonatal maternal separation induces sex-specific augmentation of the hypercapnic ventilatory response in awake rat. J Appl Physiol 102: 1416 -1421, 2007. First published December 21, 2006; doi:10.1152/japplphysiol.00454.2006.-Neonatal maternal separation (NMS) is a form of stress that exerts persistent, sex-specific effects on the hypoxic ventilatory response. Adult male rats previously subjected to NMS show a 25% increase in the response, whereas NMS females show a response 30% lower than controls (8). To assess the extent to which NMS affects ventilatory control development, we tested the hypothesis that NMS alters the ventilatory response to hypercapnia in awake, unrestrained rats. Pups subjected to NMS were placed in a temperature-and humidity-controlled incubator 3 h/day for 10 consecutive days (P3 to P12). Control pups were undisturbed. At adulthood (8 to 10 wk old), rats were placed in a plethysmography chamber for measurement of ventilatory parameters under baseline and hypercapnic conditions (inspired CO2 fraction ϭ 0.05). After 20 min of hypercapnia, the minute ventilation response measured in NMS males was 47% less than controls, owing to a lower tidal volume response (22%). Conversely, females previously subjected to NMS showed minute ventilation and tidal volume responses 63 and 18% larger than controls respectively. Although a lower baseline minute ventilation contributes to this effect, the higher minute ventilation/ CO2 production response observed in NMS females suggests a greater responsiveness to CO2/H ϩ in this group. We conclude that NMS exerts sex-specific effects on the hypercapnic ventilatory response and that the neural mechanisms affected by NMS likely differ from those involved in the hypoxic chemoreflex.control of breathing; plasticity; hypercapnia; sexual dimorphism THE NEONATAL ENVIRONMENT is critical to proper development of neurophysiological function. The formation and fine tuning of neural circuits during early life require adequate sensory guidance, and conditions providing excessive or insufficient levels of stimulation can disrupt system development and compromise their subsequent performance throughout life. The olfactory, tactile, and auditory stimuli that the mother provides her offspring following birth are among the most potent environmental factors contributing to the "neonatal programming" of neural circuits (12,22). Although the life-long consequences of disrupting motherpup interactions have been mainly associated with behavioral and neuroendocrine dysfunction (1, 2, 20), less is known about the impact of mother-pup interaction on other homeostatic functions such as cardiorespiratory regulation. Accordingly, we showed that neonatal maternal separation (NMS) disrupts cardiorespiratory responses to moderate hypoxia in a persistent and sex-specific fashion (8,16). In addition to eliciting the well described enhancement of basal hypothalamo-pituitary-adrenal axis function in rats (34; for review see Ref. 5), we...

In awake animals, our laboratory recently showed that the hypoxic ventilatory response of adult male (but not female) rats previously subjected to neonatal maternal separation (NMS) is 25% greater than controls (Genest SE, Gulemetova R, Laforest S, Drolet G, and Kinkead R. J Physiol 554: 543-557, 2004). To begin mechanistic investigations of the effects of this neonatal stress on respiratory control development, we tested the hypothesis that, in male rats, NMS enhances central integration of carotid body chemoafferent signals. Experiments were performed on two groups of adult male rats. Pups subjected to NMS were placed in a temperature-controlled incubator 3 h/day from postnatal day 3 to postnatal day 12. Control pups were undisturbed. At adulthood (8-10 wk), rats were anesthetized (urethane; 1.6 g/kg), paralyzed, and ventilated with a hyperoxic gas mixture [inspired O2 fraction (Fi(O2)) = 0.5], and phrenic nerve activity was recorded. The first series of experiments aimed to demonstrate that NMS-related enhancement of the inspiratory motor output (phrenic) response to hypoxia occurs in anesthetized animals also. In this series, rats were exposed to moderate, followed by severe, isocapnic hypoxia (Fi(O2) = 0.12 and 0.08, respectively, 5 min each). NMS enhanced both the frequency and amplitude components of the phrenic response to hypoxia relative to controls, thereby validating the use of this approach. In a second series of experiments, NMS increased the amplitude (but not the frequency) response to unilateral carotid sinus nerve stimulation (stimulation frequency range: 0.5-33 Hz). We conclude that enhancement of central integration of carotid body afferent signal contributes to the larger hypoxic ventilatory response observed in NMS rats.

Recurrent apneas are important causes of hospitalization and morbidity in newborns. Gestational stress (GS) compromises fetal brain development. Maternal stress and anxiety during gestation are linked to respiratory disorders in newborns; however, the mechanisms remain unknown. Here, we tested the hypothesis that repeated activation of the neuroendocrine response to stress during gestation is sufficient to disrupt the development of respiratory control and augment the occurrence of apneas in newborn rats. Pregnant dams were displaced and exposed to predator odor from days 9 to 19 of gestation. Control dams were undisturbed. Experiments were performed on male and female rats aged between 0 and 4 d old. Apnea frequency decreased with age but was consistently higher in stressed pups than controls. At day 4, GS augmented the proportion of apneas with O 2 desaturations by 12%. During acute hypoxia (12% O 2 ), the reflexive increase in breathing augmented with age; however, this response was lower in stressed pups. Instability of respiratory rhythm recorded from medullary preparations decreased with age but was higher in stressed pups than controls. GS reduced medullary serotonin (5-HT) levels in newborn pups by 32%. Bath application of 5-HT and injection of 8-OH-DPAT [(Ϯ)-8-hydroxy-2-di-(n-propylamino) tetralin hydrobromide; 5-HT 1A agonist; in vivo] reduced respiratory instability and apneas; these effects were greater in stressed pups than controls. Sex-specific effects were observed. We conclude that activation of the stress response during gestation is sufficient to disrupt respiratory control development and promote pathological apneas in newborn rats. A deficit in medullary 5-HT contributes to these effects.

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Perinatal stress disrupts normal development of the hypothalamo-pituitary-adrenal (HPA) axis. Adult male (but not female) rats previously subjected to a stress such as neonatal maternal separation (NMS) are characterized by chronic elevation of plasma corticosterone (Cort) levels and an abnormally elevated hypoxic ventilatory response through mechanisms that remain unknown. The present study tested the hypothesis that a chronic increase of plasma Cort levels alone augments the ventilatory response to hypoxia in adult rats. Three groups of Sprague-Dawley male and female rats were used (control, placebo and Cort implants). Rats subjected to chronic Cort elevation received a subcutaneous Cort implant (300 mg) 14 days prior to ventilatory measurements, whereas sham-operated rats received placebo implants. Controls received no treatment. Plasma Cort levels and body weight profiles were measured to assess protocol efficiency. Whole body plethysmography was used to measure ventilatory activity and metabolic indices during normoxia and following a 20 min period of moderate hypoxia (12% O 2 ). Male rats implanted with Cort showed a ventilatory response to hypoxia higher than placebo-treated rats; this effect was mainly due to a larger tidal volume response. In females, Cort treatment increased the breathing frequency response but the effect on minute ventilation was not significant. Taken together, these data show that chronic elevation of Cort alone increases the ventilatory response to hypoxia, but in a sex-specific manner. These data raise important questions regarding the mechanisms underlying the sexual dimorphism of this effect and the potential link between HPA axis dysfunction and respiratory disorders related to abnormal ventilatory chemoreflex.

To test the hypothesis that stress alters the performance of the respiratory control system, we compared the acute (20 min) responses to moderate hypoxia and hypercapnia of rats previously subjected to immobilization stress (90 min/day) with responses of control animals. Ventilatory measurements were performed on awake rats using whole body plethysmography. Under baseline conditions, there were no differences in minute ventilation between stressed and unstressed groups. Rats previously exposed to immobilization stress had a 45% lower ventilatory response to hypercapnia (inspiratory CO(2) fraction = 0.05) than controls. In contrast, stress exposure had no statistically significant effect on the ventilatory response to hypoxia (inspiratory O(2) fraction = 0.12). Stress-induced attenuation of the hypercapnic response was associated with reduced tidal volume and inspiratory flow increases; the frequency and timing components of the response were not different between groups. We conclude that previous exposure to a stressful condition that does not constitute a direct challenge to respiratory homeostasis can elicit persistent (> or =24 h) functional plasticity in the ventilatory control system.