Effect of temperature, age and the pons on respiratory rhythm in the rat brainstem-spinal cord.

Neonatal animals are extremely tolerant of hypothermia. However, cooling will ultimately lead to ventilatory arrest, or cessation of respiratory movements. Upon rewarming, ventilation can recover spontaneously (autoresuscitation). This study examined the effect of age (P0-P5) and the pons on respiratory-related output during hypothermic ventilatory arrest and recovery using a brainstem-spinal cord preparation of neonatal rats. As temperature fell, burst frequency slowed, burst duration increased, burst shape became fragmented and eventually respiratory arrest occurred in all preparations. Removing the pons had little effect on younger preparations (P0-P2). Older preparations (P4-P5) with the pons removed continued to burst at cooler temperatures compared to pons-intact preparations and burst durations were significantly longer. Episodic breathing patterns were observed in all preparations (all ages, pons on or off) at lower temperatures. At 27 °C, however, episodic breathing was only observed in younger preparations with the pons on. These data suggest that developmental changes occurring at the level of the pons underlie the loss of hypothermic tolerance and episodic breathing.

The conditional nature of the "Central Rhythm Generator" and the production of episodic breathing.

Episodic breathing patterns have been observed in species of all vertebrate classes under certain conditions and/or at certain times in development. This breathing pattern can be considered part of a continuum between no breathing and continuous breathing. In birds and mammals it is also generally part of a developmental continuum in which episodic breathing occurs early in development and rarely in adults. Production of this pattern appears to be an intrinsic property of the medullary rhythm generating mechanism (possibly due to interactions between different rhythm generating sites) that is stabilized by pontine or midbrain inputs and, in intact animals, is primarily regulated by afferent inputs from chemoreceptors and pulmonary stretch receptors; i.e. there is a hierarchy of control. In all cases, episodes appear to be produced by quantal expression of a fundamental rhythm. At present NO, GABA(A) and glycine mediated processes, and possibly mu-opioid receptor mediated processes, are implicated in the clustering of breaths into episodes. The inter-breath interval, which may occur at either the end of the inspiratory or the expiratory phase in different species, is the primary regulated variable in this pattern. The biological significance of clustering breaths into episodes may relate to reducing the oxidative cost of breathing, enhancing gas exchange or minimizing oxidative damage to tissues.

Respiratory rhythm of brainstem-spinal cord preparations: Effects of maturation, age, mass and oxygenation.

We examined the effect of age, mass and the presence of the pons on the longevity (length of time spontaneous respiratory-related activity is produced) of brainstem-spinal cord preparations of neonatal rodents (rats and hamsters) and the level of oxygenation in the medulla respiratory network in these preparations. We found the longevity of the preparations from both species decreased with increasing postnatal age. Physical removal of the pons increased respiratory frequency and the longevity of the preparation. However, tissue oxygenation at the level of the medullary respiratory network was not affected by removal of the pons or increasing postnatal age (up to postnatal day 4). Taken together, these data suggest that the effect of removing the pons on respiratory frequency and the longevity of brainstem-spinal cord preparations with increasing postnatal age are primarily due to postnatal development and appear to be unrelated to mass or changes in levels of tissue oxygenation.

Effect of spinal cord injury on the neural regulation of respiratory function.

Injury at any level of the spinal cord can impair respiratory motor function. Indeed, complications associated with respiratory function are the number one cause of mortality in humans following spinal cord injury (SCI) at any level of the cord. This review is aimed at describing the effect of SCI on respiratory function while highlighting the recent advances made by basic science research regarding the neural regulation of respiratory function following injury. Models of SCI that include upper cervical hemisection and contusion injury have been utilized to examine the underlying neural mechanisms of respiratory control following injury. The approaches used to induce motor recovery in the respiratory system are similar to other studies that examine recovery of locomotor function after SCI. These include strategies to initiate regeneration of damaged axons, to reinnervate paralyzed muscles with peripheral nerve grafts, to use spared neural pathways to induce motor function, and finally, to initiate mechanisms of neural plasticity within the spinal cord to increase motoneuron firing. The ultimate goals of this research are to restore motor function to previously paralyzed respiratory muscles and improve ventilation in patients with SCI.

Effect of spinal cord injury on the respiratory system: basic research and current clinical treatment options.

Spinal cord injury (SCI) often leads to an impairment of the respiratory system. The more rostral the level of injury, the more likely the injury will affect ventilation. In fact, respiratory insufficiency is the number one cause of mortality and morbidity after SCI. This review highlights the progress that has been made in basic and clinical research, while noting the gaps in our knowledge. Basic research has focused on a hemisection injury model to examine methods aimed at improving respiratory function after SCI, but contusion injury models have also been used. Increasing synaptic plasticity, strengthening spared axonal pathways, and the disinhibition of phrenic motor neurons all result in the activation of a latent respiratory motor pathway that restores function to a previously paralyzed hemidiaphragm in animal models. Human clinical studies have revealed that respiratory function is negatively impacted by SCI. Respiratory muscle training regimens may improve inspiratory function after SCI, but more thorough and carefully designed studies are needed to adequately address this issue. Phrenic nerve and diaphragm pacing are options available to wean patients from standard mechanical ventilation. The techniques aimed at improving respiratory function in humans with SCI have both pros and cons, but having more options available to the clinician allows for more individualized treatment, resulting in better patient care. Despite significant progress in both basic and clinical research, there is still a significant gap in our understanding of the effect of SCI on the respiratory system.

Spinal cord injury in neonates alters respiratory motor output via supraspinal mechanisms.

Upper cervical spinal cord injury (SCI) alters respiratory output and results in a blunted respiratory response to pH/CO2. Many SCI studies have concentrated on respiratory changes in neural function caudal to injury; however few have examined whether neural plasticity occurs rostral to SCI. Golder et al. (2001a) showed that supraspinal changes occur to alter respiratory output after SCI. Furthermore, Brown et al. (2004) showed that neural receptors change rostral to a thoracic SCI. We hypothesized that SCI in neonates will alter supraspinal output, show a blunted response to pH and alter receptor protein levels in the medulla. On postnatal day 0/1, a C2 SCI surgery was performed. Two days later, neonates were anesthetized and brainstem-spinal cords removed. Respiratory-related activity was recorded using the in vitro brainstem-spinal cord preparation and the superfusate pH was changed (pH 7.2, 7.4 and 7.8). The respiratory-like frequency was significantly reduced in SCI rats indicating supraspinal plasticity. Increasing the pH decreased respiratory-like frequency and peak amplitude in injured and sham controls. Increasing the pH increased burst duration and area in sham controls, whereas in injured rats, the burst duration and area decreased. Western blot analysis demonstrated significant changes in glutamate receptor subunits (NR1, NR2B and GluR2), adenosine receptors (A1, A2A), glutamic acid decarboxylase (65) and neurokinin-1 receptors in medullary tissue ipsilateral and contralateral to injury. These data show that supraspinal plasticity in the respiratory system occurs after SCI in neonate rats. The mechanisms remain unknown, but may involve alterations in receptor proteins involved in neurotransmission.

GABA, not glycine, mediates inhibition of latent respiratory motor pathways after spinal cord injury.

Previous work has shown that latent respiratory motor pathways known as crossed phrenic pathways are inhibited via a spinal inhibitory process; however, the underlying mechanisms remain unknown. The present study investigated whether spinal GABA-A and/or glycine receptors are involved in the inhibition of the crossed phrenic pathways after a C2 spinal cord hemisection injury. Under ketamine/xylazine anesthesia, adult, female, Sprague-Dawley rats were hemisected at the C2 spinal cord level. Following 1 week post injury, rats were anesthetized with urethane, vagotomized, paralyzed and ventilated. GABA-A receptor (bicuculline and Gabazine) and glycine receptor (strychnine) antagonists were applied directly to the cervical spinal cord (C3-C7), while bilateral phrenic nerve motor output was recorded. GABA-A receptor antagonists significantly increased peak phrenic amplitude bilaterally and induced crossed phrenic activity in spinal-injured rats. Muscimol, a specific GABA-A receptor agonist, blocked these effects. Glycine receptor antagonists applied directly to the spinal cord had no significant effect on phrenic motor output. These results indicate that phrenic motor neurons are inhibited via a GABA-A mediated receptor mechanism located within the spinal cord to inhibit the expression of crossed phrenic pathways.

Spinal activation of serotonin 1A receptors enhances latent respiratory activity after spinal cord injury.

BACKGROUND/OBJECTIVE: Hemisection of the cervical spinal cord results in paralysis of the ipsilateral hemidiaphragm. Removal of sensory feedback through cervical dorsal rhizotomy activates latent respiratory motor pathways and restores hemidiaphragm function. Because systemic administration of serotonin 1A receptor (5HT1A) agonists reversed the altered breathing patterns after spinal cord injury (SCI), we predicted that 5HT1A receptor activation after SCI (C2) would activate latent crossed motor pathways. Furthermore, because 5HT1 A receptors are heavily localized to dorsal horn neurons, we predicted that spinal administration of 5HT1A agonists should also activate latent motor pathways. METHODS: Hemisection of the C2 spinal cord was performed 24 to 48 hours, 1 week, or 16 weeks before experimentation. Bilateral phrenic nerve activity was recorded in anesthetized, vagotomized, paralyzed Sprague-Dawley rats, and 8-OH-DPAT (5HT1A agonist) was applied to the dorsal aspect of the cervical spinal cord (C3-C7) or injected systemically. RESULTS: Systemic administration of 8-OH-DPAT led to a significant increase in phrenic frequency and amplitude, whereas direct application to the spinal cord increased phrenic amplitude alone. Both systemic and spinal administration of 8-OH-DPAT consistently activated latent crossed phrenic activity. 8-OH-DPAT induced a greater respiratory response in spinal injured rats compared with controls. CONCLUSION: The increase in crossed phrenic output after application of 8-OH-DPAT to the spinal cord suggests that dorsal horn inputs, respiratory and/or nonrespiratory, may inhibit phrenic motor output, especially after SCI. These findings support the idea that the administration of 5HT1A agonists may be a beneficial therapy in enhancing respiratory neural output in patients with SCI.

Spontaneous crossed phrenic activity in the neonatal respiratory network.

Hemisection of the cervical spinal cord causes paralysis of the ipsilateral hemidiaphragm in adult rats. Activation of a latent crossed phrenic motor pathway can restore diaphragmatic function, although structural changes take place before the pathway can be activated. Since mechanisms are employed to eliminate non-functional projections during development, we predicted that this latent neural pathway might be active during development. Therefore, we examined the effect of spinal hemisection (C2) on respiratory-like activity bilaterally using the brainstem--spinal cord preparation from neonatal rats (0-4 days). Spontaneous crossed phrenic activity (respiratory-like activity recorded from the ipsilateral C4 or C5 ventral roots following C2 hemisection) was observed in an age-dependent manner; younger preparations exhibited more than older preparations. Increasing drive (increasing [K+] or superfusion of theophylline) either increased or induced crossed phrenic activity. Hemisection caused no change in the frequency, the burst area, duration or peak amplitude contralateral to hemisection. Unlike adult rats, this study shows that crossed phrenic activity is present in the in vitro respiratory network of neonatal rats suggesting that a crossed neural pathway may be functionally active in neonates.

Pontine influences on respiratory control in ectothermic and heterothermic vertebrates.

Respiratory rhythm generators appear both evolutionarily and developmentally as paired segmental rhythm generators in the reticular formation, associated with the motor nuclei of cranial nerves V, VII, IX, X, and XII. Those associated with the Vth and VIIth motor nuclei are "pontine" in origin and in fishes that employ a buccal suction/force pump for breathing the primary pair of respiratory rhythm generators are associated with the trigeminal nuclei. In amphibians, while the basic respiratory pump remains the same, the dominant site of respiratory rhythm generation has been assumed by the facial, glossopharyngeal and vagal motor nuclei. In reptiles, birds and mammals, in general there is a switch to an aspiration pump driven by thoraco-lumbar muscles innervated by spinal nerves. In these groups, the critical sites necessary for respiratory rhythmogenesis now sit near the ponto-medullary border, in the parafacial region (which may underlie expiratory-dominated, intercostal-abdominal breathing in non-mammalian tetrapods) and in a more caudal region, the preBotzinger complex (which may underlie inspiratory-dominated diaphragmatic breathing in mammals).

Effect of hypothermia on respiratory rhythm generation in hamster brainstem-spinal cord preparations.

This study examined the effect of hypothermia on respiratory neural output from brainstem-spinal cord preparations of a cold tolerant rodent, the Syrian hamster. Brainstem-spinal cords from neonatal hamsters (0-6 days) were placed in a recording dish and respiratory-like neural activity was recorded from roots of the first cervical nerve. The preparations were cooled and warmed in a continuous or stepwise fashion. Inputs from the pons completely inhibited neural activity under steady state conditions. With the pons removed, fictive breathing was robust. Cooling caused respiratory arrest, followed by spontaneous resumption of activity on re-warming. Preparations from older hamsters (4-6 days old) were more cold tolerant than younger preparations (0-3 days old). Motor discharge was episodic during continuous cooling, and seizure-like discharge was observed during continuous warming. These phenomena were not observed with stepwise temperature changes suggesting that transient temperature effects on membrane properties may be involved. These preparations were not as cold tolerant as hamster pups in vivo but they retained the ability to autoresuscitate at all ages studied.

Ventilatory pattern and chemosensitivity in unanesthetized, hypothermic ground squirrels (Spermophilus lateralis).

The present study examined the effects of severe hypothermia in the absence of anesthesia on breathing pattern, ventilatory control and chemosensitivity in a cold tolerant species capable of seasonal hibernation. Hypothermia was induced in ground squirrels and ventilation and heart rate were recorded in animals breathing air at a body temperature (Tb) of 5 and 10 degrees C. The animals were then exposed to hypercapnic (2, 4 and 6% CO(2)) and hypoxic (12, 10, 8 and 4% O(2)) gas mixtures. We found that severe hypothermia in ground squirrels caused the breathing pattern to change from a continuous pattern to patterns that are commonly observed during hibernation. This suggests that temperature and metabolism alone are important factors in producing these patterns. The relative ventilatory sensitivity to hypercapnia was retained in the ground squirrel during hypothermia while ventilatory sensitivity to hypoxia was totally abolished. This is in contrast to hibernation where a small but significant hypoxic ventilatory response is present along with an enhanced relative response to hypercapnia. This suggests that changes in Tb alone can not account for the changes seen in ventilatory sensitivity during hibernation.