Intercostal and Abdominal Respiratory Motoneurons in the Neonatal Rat Spinal Cord: Spatiotemporal Organization and Responses to Limb Afferent Stimulation

Abstract

Respiration requires the coordinated rhythmic contractions of diverse muscles to produce ventilatory movements adapted to organismal requirements. During fast locomotion, locomotory and respiratory movements are coordinated to reduce mechanical conflict between these functions. Using semi-isolated and isolated in vitro brain stem-spinal cord preparations from neonatal rats, we have characterized for the first time the respiratory patterns of all spinal intercostal and abdominal motoneurons and explored their functional relationship with limb sensory inputs. Neuroanatomical and electrophysiological procedures were initially used to locate intercostal and abdominal motoneurons in the cord. Intercostal motoneuron somata are distributed rostrocaudally from C(7)-T(13) segments. Abdominal motoneuron somata lie between T(8) and L(2). In accordance with their soma distributions, inspiratory intercostal motoneurons are recruited in a rostrocaudal sequence during each respiratory cycle. Abdominal motoneurons express expiratory-related discharge that alternates with inspiration. Lesioning experiments confirmed the pontine origin of this expiratory activity, which was abolished by a brain stem transection at the rostral boundary of the VII nucleus, a critical area for respiratory rhythmogenesis. Entrainment of fictive respiratory rhythmicity in intercostal and abdominal motoneurons was elicited by periodic low-threshold dorsal root stimulation at lumbar (L(2)) or cervical (C(7)) levels. These effects are mediated by direct ascending fibers to the respiratory centers and a combination of long-projection and polysynaptic descending pathways. Therefore the isolated brain stem-spinal cord in vitro generates a complex pattern of respiratory activity in which alternating inspiratory and expiratory discharge occurs in functionally identified spinal motoneuron pools that are in turn targeted by both forelimb and hindlimb somatic afferents to promote locomotor-respiratory coupling.

Spinal and Pontine Relay Pathways Mediating Respiratory Rhythm Entrainment by Limb Proprioceptive Inputs in the Neonatal Rat

“…), stimulus pulse trains (0.2-2 V, 0.5 ms at 5-10 Hz) were applied to selected hindlimb muscles or dorsal spinal roots via a stimulating tungsten electrode (Frederick Haer) or a homemade glass suction electrode, respectively. DR stimulation followed the procedure described previously (Morin and Viala, 2002;Giraudin et al, 2008). Briefly, the threshold stimulus for so-called "low-threshold" (presumed proprioceptive) DR afferent fibers was determined by increasing the train shock intensity from a subthreshold value until a level at which an effect on the relative timing of the subsequent cycle of ongoing spontaneous respiratory rhythmicity was observed.…”

“…In isolated brainstem/spinal cord preparations of the neonatal rat, we previously reported that phasic electrical stimulation of lumbar (Morin and Viala, 2002) or cervical (Giraudin et al, 2008) DRs in a manner that mimics locomotion-induced activation of somatosensory afferents in vivo can reset and entrain respiratory rhythmicity, indicating that these spinal inputs have access to the brainstem respiratory centers. Moreover, since only low stimulus intensities were required to activate the lumbar and cervical sensory pathways (Morin and Viala, 2002;Giraudin et al, 2008; also see below) it was postulated that larger diameter fibers (most likely proprioceptive and cutaneous afferents) were responsible for the DR stimulation-evoked influences on respiratory rhythmogenesis.…”

“…Moreover, since only low stimulus intensities were required to activate the lumbar and cervical sensory pathways (Morin and Viala, 2002;Giraudin et al, 2008; also see below) it was postulated that larger diameter fibers (most likely proprioceptive and cutaneous afferents) were responsible for the DR stimulation-evoked influences on respiratory rhythmogenesis.…”

“…In this context, advances have been made in more recent studies using the isolated in vitro CNS of the newborn rat (Morin and Viala, 2002;Giraudin et al, 2008) and reduced in situ preparations of juveniles (Potts et al, 2005), where rhythmic activation of limb somatic afferents by direct electrical stimulation of cervical and lumbar dorsal roots (DRs) or forelimb flexor muscles, respectively, was found capable of phase resetting and entraining spontaneously generated respiratory rhythmicity with a 1:1 coordination. Furthermore, chemically inactivating the lateral parabrachial nucleus in the brainstem pontine region abolished the respiratory entraining effect of somatic afferent input, thereby implicating an intervening role for this structure in the locomotory-respiratory coupling process (Potts et al, 2005).…”

“…To date however, the nature of the spinal sensory afferents responsible for respiratory entrainment as well as the ability of somatic inputs at different cord levels to exert entraining influences has been little investigated. Moreover, although there is evidence that long-ascending spinal projections are involved in mediating the somatic afferent-derived respiratory entrainment (Morin and Viala, 2002;Giraudin et al, 2008), whether the sensory information is conveyed to the pontine relay en route to the medullary respiratory centers via direct or polysynaptic spinal pathways has not been documented.…”