Parabrachial–lateral pontine neurons link nociception and breathing

We investigated the role of the parabrachial complex in cutaneous nociceptor-induced respiratory stimulation in chloralose-urethane anesthetized, vagotomized rats. Noxious stimulation (mustard oil, MO) applied topically to a forelimb or hindlimb enhanced the peak amplitude of the integrated phrenic nerve discharge and, with forelimb application, increased phrenic nerve burst frequency. Bilateral inactivation of neural activity in the parabrachial complex with injection of the GABA agonist muscimol (3 nl) markedly attenuated the response to MO application. Injection of the retrograde tracer FluoroGold within the medullary ventral respiratory column labeled neurons in dorsolateral pontine regions known to receive nociceptive inputs (i.e., Kölliker-Fuse, lateral crescent, and superior lateral subnuclei of the parabrachial complex). Extracellular recordings of 65 dorsolateral parabrachial neurons revealed about 15% responded to a noxious cutaneous pinch with either an increase or a decrease in discharge and 40% of these exhibited a phasic respiratory-related component to their discharge. In conclusion, parabrachial pontine neurons contribute to cutaneous nociceptor-induced increases in breathing.     

Central histamine contributes to the inspiratory off-switch mechanism via H1 receptors in mice

Central histaminergic neurons are distributed in areas of the medulla and pons concerned with respiratory rhythm generation, but their effects on breathing pattern are unknown. We examined breathing pattern during hypercapnic responses in wild type (WT) and H1 receptor knockout (H1RKO) mice at 9-10 weeks of age before and after vagotomy. Minute ventilation increased with PaCO(2) increase equally in both genotypes; respiratory rate response was lower and tidal volume (V(T)) response higher in H1RKO mice than in WT mice. The V(T)-inspiratory time (T(I)) relation during hypercapnia was hyperbolic in both groups, with the curve in H1RKO mice shifted right-upward. After vagotomy, the V(T)-T(I) relation was a vertical line, which shifted right in H1RKO mice. We conclude that alterations of inspiratory off-switch and respiratory rhythm generation change breathing pattern without affecting central chemosensitivity in H1RKO. Histamine might affect breathing pattern centrally via H1 receptors.     

Effect of medroxyprogesterone on inspiratory flow shapes during sleep in postmenopausal women

We previously showed that medroxyprogesterone acetate (MPA) effectively decreases the arterial CO(2) levels in postmenopausal women with partial upper airway obstruction. The aim of the present study was to analyze the effects of MPA on the inspiratory flow shapes during sleep. Eight postmenopausal women with hypoxemia and partial upper airway obstruction during sleep (patients) received MPA 60 mg daily for 14 days. Four matched postmenopausal women without MPA treatment served as controls. Sleep and nasal pressure were recorded on each visit. Each breath was analyzed for duration, volume and inspiratory flow shape class. MPA shortened inspiration and prolonged expiration. The inspiratory volumes increased consistently in all flow shape classes. The inspiratory shapes with single late peak were transformed to those with double peak. MPA also decreased shapes with mid-peak or mid-plateau. MPA did not have an effect on sleep. Sleep modified the flow shape distribution only in patients but in a similar fashion in stages S2, SWS and REM. The results suggest that postmenopausal women present with a significant proportion breaths with poor initial inspiratory flow, which is reversed with MPA-induced respiratory stimulation.     

Ventilatory effects of gap junction blockade in the NTS in awake rats

We tested the hypothesis that focally perfusing carbenoxolone, which blocks gap junctions, into the nucleus tractus solitarius (NTS) would reduce the ventilatory response to CO(2). We measured minute ventilation (V(E)), tidal volume (V(T)) and respiratory frequency (F(R)) responses to increasing concentrations of inspired CO(2) ( [formula: see text] ) in rats during wakefulness. Focal perfusion of acetazolamide (10 microM) into the NTS increased V(E) and V(T) during exposure to room air. Carbenoxolone (300 microM) decreased the V(E) and V(T) response to CO(2) when perfused within, but not adjacent to the NTS in animals less than 10 weeks of age. F(R) was decreased at [formula: see text] in these animals. Carbenoxolone did not decrease V(E), V(T) or F(R) in animals 10 weeks of age and older. Carbenoxolone did not decrease V(E), V(T) or F(R) when focally perfused outside the NTS at any age tested. The NTS is an important CO(2) chemosensory site at all ages, and gap junctions amplify the ventilatory response to CO(2) in animals less than 10 weeks of age.     

Spatiotemporal organization of frog respiratory neurons visualized on the ventral medullary surface

We visualized the spatiotemporal activity of respiratory-related neurons in the frog using the isolated brainstem spinal cord preparation. We recorded optical signals from the ventral surface of the medulla using a voltage-sensitive dye, and calculated cross-correlations with the integrated respiratory activity of the trigeminal nerve. Lung burst-related depolarizing optical signals were observed bilaterally as longitudinal columns in the ventrolateral medulla between the levels of trigeminal and hypoglossal rootlets, mostly caudal to the vagal rootlet. However, we could not differentiate between neurons involved in rhythm generation and motoneurons. The dye weakened the buccal rhythm and slowed the lung rhythm, which might have influenced the results. Extracellular recording of respiratory neurons verified the optically identified area. Strychnine disrupted the spatiotemporal organization of optical signals, although trigeminal periodic bursts persisted. We conclude that the pattern generator but not the rhythm generator of lung burst in the frog involves glycinergic mechanisms and lies as longitudinal columns in the reticular formation of the ventrolateral medulla.     

Heterogeneity of brainstem blood flow response to hypoxia in the anesthetized rat

Cerebral blood flow is strictly regulated during hypoxic stress. Because of the preponderant role of the brainstem in cardiorespiratory controls, blood flow response to hypoxia is stronger in this region than in the cortex. However, the brainstem is made up of various regions which differ in their responsiveness to chemical stimuli. The objective of this study was to evaluate the distribution of blood flow during hypoxia using microsphere deposition methods in three brainstem regions containing key structures in cardiorespiratory controls: the nucleus tractus solitarus (NTS), the ventral respiratory groups (VRG) and the pontine respiratory groups (PRG). Microsphere injections were made during normoxia (FIO2=0.21) and after 15 min of hypoxia (FIO2=0.21). Based on this index, blood flow increase during hypoxia was higher in the VRG than in the dorsal part of the brainstem, containing the NTS and the PRG (P=0.002, n=10). These results suggest that blood flow response to hypoxia favours O(2) delivery in brainstem regions involved in respiratory rhythm generation.     

Commentary on the definition of eupnea and gasping

Despite clear qualitative differences, it has proven difficult to identify criteria that reliably differentiate eupnea and gasping--particularly when multiple species or experimental preparations are considered. From a motor control perspective, this is unsurprising. Three organizational rules are common to nearly all rhythmic activities: (1) the basic rhythm is produced by a small network of cells, (2) the activity of this network in isolation often differs dramatically from the behavior of the whole animal, and (3) the rhythmogenic networks responsible for related behaviors are not fixed and independent but dynamically modifiable and overlapping. In this context, it becomes clear that the definition of a particular pattern and the investigation of the mechanisms underlying its production are inseparable. Rather than attempting to rigidly apply criteria developed using any one experimental preparation, the classification of respiratory patterns must evolve alongside our understanding of how each pattern is produced-a process that is only aided by investigations using a variety of experimental preparations.     

Effects of five nights of normobaric hypoxia on the ventilatory responses to acute hypoxia and hypercapnia

This study examined the effects of five nights of normobaric hypoxia on ventilatory responses to acute isocapnic hypoxia (AHVR) and hyperoxic hypercapnia (AHCVR). Twelve male subjects (26.6 +/- 4.1 years, standard deviation (S.D.)) slept 8-9 h per day overnight for 5 consecutive days at a simulated altitude of 4,300 m (FiO2= approximately 13.8%). Using the technique of dynamic end-tidal forcing, the AHVR and AHCVR were assessed twice prior to, immediately after, and 5 days following the hypoxic exposure. Immediately following the exposure, AHVR was increased by 1.6 +/- 1.3 L min(-1) %(-1) (P<0.01) when compared with control values. Likewise, after the exposure, ventilation in hyperoxia was increased (P<0.001) and was associated with both an increase in the slope (1.5 +/- 1.4 L min(-1) Torr(-1); P<0.05) and decrease in the intercept (-2.7 +/- 4.3 Torr; P<0.05) of the AHCVR. These results show that five nights of hypoxia can elicit similar perturbations, in both AHVR and AHCVR, as have been reported during more chronic altitude exposures.     

Studies on the ascending pathways from the thermosensitive region of the spinal cord

Summary In young guinea pigs the ascending pathways from the spinal thermosensitive region were studied by means of 1. microelectrode recording and 2. micro-electrocoagulation. In the first series of studies, impulse frequency was recorded from single units of the spinothalamic tract which responded to a temperature rise in the spinal segments C₅-T₂ with an increase of discharge frequency. At a spinal cord temperature of 38–39° C these units showed a firing rate of 1–5 imp./sec; local heating of the spinal cord (dT/dt=0.1° C/sec) to 40–41° C caused an increase in discharge frequency to 20–25 imp./sec. The mean static impulse frequency was 3 imp./sec at a spinal cord temperature of 39° C, and 10 imp./sec at a spinal cord temperature of 42.5° C.In the second series of studies bilateral RF-coagulations were carried out in different sites of the diencephalon. These experiments showed that the ascending fibres from the spinal thermosensitive region connect the thermosensitive spinal region with a hypothalamic temperature control centre.Supported by the Deutsche Forschungsgemeinschaft (Br. 184/10).     

Influence of rubrospinal tract and the adjacent mesencephalic reticular formation on the activity of medullary respiratory neurons and the phrenic nerve discharge in the rabbit

Suprapontine brain sites acting on the central respiratory system have been demonstrated to give rise to inspiratory as well as expiratory facilitatory effects. In the present study the inspiratory inhibitory effect which has been reported in the cat to be elicited consistently by electrical stimulation of the rubrospinal tract and the adjacent mesencephalic reticular formation was examined in the urethane-anaesthetized rabbit. Stimulation of these sites with single electrical shocks of moderate intensity induced a short latency (onset after 3.0 ms) transient (duration: 29 ms) inhibition of the phrenic nerve activity (PHR). Short volleys of stimuli applied in mid- to late-inspiration led to a premature off-switch of inspiration. The extracellularly recorded discharge activity of the different types of medullary respiration-related units (RRU) reflected these alterations, accordingly. Axonal connections of RRU with mesencephalic structures were evaluated. Examination of orthodromic responses of medullary RRU to stimulation of this pathway revealed that most bulbospinal inspiratory neurons (10 out of 13) were paucisynaptically inhibited after short latency (at least 1.2 ms). The conduction time from bulbospinal inspiratory neurons to the recording site of PHR was 1.6 ms. Thus, a disynaptic pathway — including bulbospinal inspiratory neurons — is suggested inducing inspiratory inhibition 3.0 ms after single shock midbrain stimulation. This inhibition results in disfacilitation of phrenic motoneurons. The fact that extensive electrolytic lesions of the pneumotaxic center in rostral pons did not abolish the observed inspiratory inhibitions excludes these structures from being involved. A direct pathway from the red nucleus and the adjacent reticular formation to phrenic nuclei of the spinal cord, however, can not be excluded from being involved in the demonstrated inspiratory inhibition. The described effects may play a role in behavioral or voluntary control of respiration.     

Chemical lesions of the nucleus isthmi increase the hypoxic and hypercarbic drive to breathing of toads

The nucleus isthmi (NI) is a mesencephalic structure of the amphibian brain that has recently been reported to participate in the hypoxic and hypercarbic drive to breathing. However, previous studies used electrolytic and kainic lesions, which causes diffuse and nonspecific destruction. Thus, in the present study, we assessed the participation of NI in the respiratory response to hypoxia and hypercarbia using lesions produced with ibotenic acid (a substance that selectively destroys cell bodies but spares fibers of passage) into the NI of toads (Bufo paracnemis). Our results demonstrated that, under resting breathing, NI plays no role in pulmonary ventilation. Hypoxia and hypercarbia caused hyperventilation in all groups. Chemical lesions in the NI elicited an increase in ventilatory response to hypoxia and hypercarbia, due to a higher tidal volume. We conclude that NI cell bodies do not participate in the control of pulmonary ventilation under resting conditions, but exert an inhibitory modulation of hypoxic and hypercarbic drive to breathing, acting on tidal volume.     

Expression of HIF-1alpha, VEGF and VEGF receptors in the carotid body of chronically hypoxic rat

We examined the protein expression and localization of HIF-1alpha, VEGF, VEGF receptors in the carotid body (CB) of rats breathing 10% inspired oxygen for up to 4 weeks. The immunoreactivity (IR) of HIF-1alpha was distributed numerously in the nuclei of glomus (type-I) and other cells since hypoxia for 1 day, but was faint and scattered in the normoxic CBs. Cytoplasmic staining of the VEGF was intense in glomus cells of the hypoxic but not the normoxic group. The IR levels of HIF-1alpha and VEGF reached plateau at 4 weeks, and the IRs of VEGFR-1 and VEGFR-2 were strongly positive in the hypoxic group. Yet, the expression of VEGFR-1-IR was mild, whereas the VEGFR-2-IR was intense in normoxic CBs, suggesting an upregulation of VEGFR-1 but not VEGFR-2 in hypoxia. Hence, HIF-1 may activate the expression of VEGF and VEGFR-1 in the CB and the expression of VEGF in the chemoreceptors may play a paracrine role in the vascular remodeling during chronic hypoxia.     

Effects of warm and cool thermal conditions on ventilatory responses to hyperoxic test in neonates

Body temperature interacts with respiratory control, but it is unclear what sites or mechanisms mediate those interactions. We hypothesized that warm and cool thermal conditions affect the decrease in ventilation (VE) seen during the hyperoxic test (HT), a breathing response believed to reflect the strength of the peripheral chemoreceptor drive. A breath-by-breath analysis during a 30 s HT was performed in eight premature neonates (postconceptional age: 36 +/- 1 weeks) under neutral, warm, and cool thermal conditions. Quiet sleep (QS) and active sleep (AS) were scored by neurophysiological criteria. The VE fall was higher in AS than in QS, and warm and cool conditions significantly enhanced the response only in AS (-24.2 +/- 6.0, -39.1 +/- 9.1, and -37.5 +/- 14.1% in neutral, warm, and cool conditions, respectively). Central control mechanisms of the respiratory chemoreflex may explain the increase in peripheral chemoreceptor drive during AS in response to thermal challenges, which may produce increased breathing instability leading to apnea in early life.     

Effects of electrical stimulation of the tooth pulp and phrenic nerve fibers on C1 spinal neurons in the rat

 Effects of electrical stimulation of the ipsilateral tooth pulp (TP) on C₁ spinal neurons were determined in 33 anesthetized rats. One hundred and seven neurons responded to TP stimulation. In 10 rats, the activity of 18 C₁ spinal neurons and the amplitude of a digastric electromyogram (dEMG, n=10) increased proportionally during the TP stimulation at an intensity of 1–3 times the threshold for jaw-opening reflex (JOR). Excitatory receptive somatic fields were examined in 61 neurons. Somatic field locations of many neurons (67.2%) involved the ipsilateral face, neck, and jaw. The activity of 45 neurons was increased by both noxious pinch and brushing hair. Of the 107 C₁ spinal neurons responding to TP stimulation, 55 were tested to determine the effects of electrical stimulation of the ipsilateral phrenic nerve (PN) above the heart. Twenty-eight of 55 neurons tested were excited; no change in activity was seen for the remaining 27 neurons. The activity of six neurons increased as the intensity of PN stimulation was increased. Excitatory receptive somatic fields were determined in 28 neurons, and somatic field locations of 17 neurons (60.7%) included the ipsilateral face, neck, and jaw. Both noxious pinch and brushing hair excited all 28 neurons. These results suggest that there may be the convergence of face, neck, jaw, TP, and PN afferents on the same C₁ spinal neurons in the rat. Received: 23 June 1998 / Accepted: 11 December 1998     

Suppressive effect of vagal afferents on cervical dorsal horn neurons responding to tooth pulp electrical stimulation in the rat

The aim of the present study was to test the hypothesis that vagal afferent (VA) inputs modify the tooth pulp (TP) stimulation-evoked activity of the first cervical dorsal horn (C1) neurons via the activation of endogenous noradrenergic and serotonergic systems. In 30 anesthetized rats, the activity of 56 C1 spinal neurons and the amplitude in a digastric muscle electromyogram (dEMG, n=30) increased proportionally during TP stimulation at an intensity of 1-3.5 times the threshold for the jaw-opening reflex (JOR). The activity in 46 of these C1 neurons (82.1%) was suppressed by VA stimulation (1.0 mAx0.1 ms, 50 Hz for 30 s) of the right vagus nerve. The suppressive effects of VA stimulation on C1 spinal neuron activity were significantly reduced after intravenous administration of either the alpha-adrenergic receptor antagonist phenoxybenzamine (POB, 2.0 mg/kg and 4.0 mg/kg) or the 5-hydroxytryptamine-3 (5-HT(3)) receptor antagonist ICS 205-930 (1.0 mg/kg and 2.0 mg/kg). But the 5-HT(1/2) receptor antagonist methysergide (1.0 mg/kg and 2.0 mg/kg) had no significant effect on VA stimulation-induced inhibition of the C1 spinal neuron activity. These results suggest that VA stimulation inhibits nociceptive transmission in the C1 spinal neuron activity via the activation of both noradrenergic and serotonergic (5-HT(3)) descending inhibitory systems.     

Effects of the dopamine antagonists haloperidol and domperidone on the normoxic ventilatory response to CO2 in cats

We investigated the effects of the dopamine antagonists haloperidol and domperidone on the ventilatory response following square-wave changes in end-tidal CO₂ during normoxia in chloralose-urethane anaesthetized cats. In 7 cats these responses were measured before (control, 28 runs) and after the administration of 1 mg/kg haloperidol i.v. (26 runs) and in 8 other cats before (39 runs) and after 0.5 mg/kg domperidone i.v. (34 runs). Each response was separated into a slow central and a fast peripheral part by fitting two exponential functions to the measured ventilation. These functions have as parameters a CO₂ sensitivity, a time constant, a time delay and an apnoeic threshold B (extrapolated PETCO₂ of the steady-state response curve at zero ventilation). Haloperidol significantly diminished the peripheral (Sp) and the central (Sc) ventilatory sensitivity to CO₂ and the B-value (P<0.001). The ratio Sp/Sc, the time constants and the time delays were not significantly changed. Domperidone only diminished the B-value significantly (P<0.001). Since domperidone does not readily cross the blood-brain barrier, its effect was a CO₂ independent increase of the ventilation mediated by the peripheral chemoreceptors. Haloperidol exhibited, besides the peripheral stimulatory effect a depressant central effect due to an action on the central integrative structure, resulting in a proportional decrease of Sp and Sc.     

The effect of surgery on patellar tendinopathy: Novel use of MRI questions the exploitability of the rat collagenase model to humans

Backgroundpatellar tendinopathy is an overuse condition most commonly affecting jumping athletes. Surgery is reserved for refractory cases; however, it lacks high level clinical evidence and basic science to support its use. The purpose of this study was to determine the biomechanical and histological response of surgical excision on patellar tendinopathy in the rat collagenase tendinopathy model and correlate MRI findings.MethodsForty-eight Long Evans rats were divided into three groups: i) no patellar tendinopathy with surgical excision, ii) patella tendinopathy with surgical excision, and iii) patellar tendinopathy with no surgical excision. Endpoints included histology, mechanical testing, and MRI pre- and post-surgical intervention at one and four weeks.ResultsNo difference in failure load or histological grading was seen between the groups at all time points. MRIs showed initial loss of tendon continuity followed by complete healing with elongated and thickened tendons in all groups.ConclusionsWhile other research has reported immunohistochemistry and histology of collagenase-induced tendinopathy may be correlated with human pathogenesis, the novel MRI findings from our study suggest that the rat collagenase tendinopathy surgical model may be limited when extrapolating to humans. Further work is needed to determine if any correlation exists between the dosing, location, and animal effect of the collagenase injection model with MRI findings. This is needed before any collagenase model can be used to determine the effect of surgery in the pathogenic response to patella tendinopathy.     

Development of CO(2)-response in the early newborn period in rat

We examined the respiratory response to moderate hypercapnia in rat pups during the first 10 days after birth and also studied immediate early gene expression to investigate whether areas described as chemosensitive in the adult rat are activated also in the early postnatal period. Breathing frequency increased in 1- and 3-day-old pups, but decreased in older animals in response to hypercapnia. Tidal volume and ventilation increased significantly in all age groups but relatively more in the 10-day-old pups as compared to younger animals. In situ hybridisation for c-fos mRNA revealed increased expression in several of the areas assigned as chemosensitive in the adult, including the caudal nucleus tractus solitarii and the ventral lateral medulla. In contrast, locus coeruleus and the majority of midline raphe neurons did not exhibit increased expression of c-fos mRNA. We conclude that the hypercapnic respiratory response tends to decrease during the first postnatal week, but thereafter increases on day 10 due to increased tidal volumes rather than changes in respiratory timing. We also speculate that differences in activation of chemosensitive brainstem neurons may be part of the maturation of the hypercapnic ventilatory response.     

Multiple roles of chemokine CXCL12 in the central nervous system: a migration from immunology to neurobiology

Chemotactic cytokines (chemokines) have been traditionally defined as small (10-14kDa) secreted leukocyte chemoattractants. However, chemokines and their cognate receptors are constitutively expressed in the central nervous system (CNS) where immune activities are under stringent control. Why and how the CNS uses the chemokine system to carry out its complex physiological functions has intrigued neurobiologists. Here, we focus on chemokine CXCL12 and its receptor CXCR4 that have been widely characterized in peripheral tissues and delineate their main functions in the CNS. Extensive evidence supports CXCL12 as a key regulator for early development of the CNS. CXCR4 signaling is required for the migration of neuronal precursors, axon guidance/pathfinding and maintenance of neural progenitor cells (NPCs). In the mature CNS, CXCL12 modulates neurotransmission, neurotoxicity and neuroglial interactions. Thus, chemokines represent an inherent system that helps establish and maintain CNS homeostasis. In addition, growing evidence implicates altered expression of CXCL12 and CXCR4 in the pathogenesis of CNS disorders such as HIV-associated encephalopathy, brain tumor, stroke and multiple sclerosis (MS), making them the plausible targets for future pharmacological intervention.