Bulbospinal Pathways Research Articles

Therapeutic efficacy of adrenergic agents on systemic and spinal hemodynamics in an acute cervical spinal cord injury rodent model

BACKGROUNDCervical spinal cord injury usually results in cardiorespiratory dysfunctions due to interruptions of the bulbospinal pathways innervating the cervical phrenic motoneurons and thoracic sympathetic preganglionic neurons. PURPOSEThe present study aimed to evaluate the therapeutic effects of adrenergic agents on systemic and spinal hemodynamics during acute cervical spinal cord injury. STUDY DESIGNIn vivo animal study. METHODSThe cardiorespiratory function and spinal cord blood flow and oxygenation level were monitored in response to cervical spinal cord contusion and intravenous infusion of three types of adrenergic agents (phenylephrine, dobutamine, and norepinephrine). RESULTSCervical spinal cord contusion resulted in immediate reduction of respiratory airflow, arterial blood pressure, and spinal cord blood flow. The arterial blood pressure and spinal cord blood flow remained lower than the preinjury value in contused animals infused with saline at 60 min postinjury. Infusion of phenylephrine (500, 1000, and 2000 μg/kg) and norepinephrine (125, 250, and 500 μg/kg) significantly increased the arterial blood pressure, while only norepinephrine augmented the spinal cord blood flow. Conversely, dobutamine (1000 and 2000 μg/kg) reduced both arterial blood pressure and spinal cord blood flow. Notably, administration of adrenergic agents tended to increase spinal cord hemorrhage in contused animals. CONCLUSIONSInfusion of norepinephrine can effectively maintain the blood pressure and improve spinal cord blood flow during acute spinal cord injury. CLINICAL SIGNIFICANCENorepinephrine may be a superior medicine for hemodynamic management; however, the potential hemorrhage should be considered when utilizing the vasopressor to regulate systemic and spinal hemodynamics at the acute injured stage.

Chapter 15 - The phrenic neuromuscular system

The phrenic neuromuscular system consists of the phrenic motor nucleus in the mid-cervical spinal cord, the phrenic nerve, and the diaphragm muscle. This motor system helps sustain breathing throughout life, while also contributing to posture, coughing, swallowing, and speaking. The phrenic nerve contains primarily efferent phrenic axons and afferent axons from diaphragm sensory receptors but is also a conduit for autonomic fibers. On a breath-by-breath basis, rhythmic (inspiratory) depolarization of phrenic motoneurons occurs due to excitatory bulbospinal synaptic pathways. Further, a complex propriospinal network innervates phrenic motoneurons and may serve to coordinate postural, locomotor, and respiratory movements. The phrenic neuromuscular system is impacted in a wide range of neuromuscular diseases and injuries. Contemporary research is focused on understanding how neuromuscular plasticity occurs in the phrenic neuromuscular system and using this information to optimize treatments and rehabilitation strategies to improve breathing and related behaviors.

Ampakines Stimulate Diaphragm Activity after Spinal Cord Injury.

Respiratory compromise after cervical spinal cord injury (SCI) is a leading cause of mortality and morbidity. Most SCIs are incomplete, and spinal respiratory motoneurons as well as proprio- and bulbospinal synaptic pathways provide a neurological substrate to enhance respiratory output. Ampakines are allosteric modulators of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors, which are prevalent on respiratory neurons. We hypothesized that low dose ampakine treatment could safely and effectively increase diaphragm electromyography (EMG) activity that has been impaired as a result of acute- or sub-acute cervical SCI. Diaphragm EMG was recorded using chronic indwelling electrodes in unanesthetized, freely moving rats. A spinal hemi-lesion was induced at C2 (C2Hx), and rats were studied at 4 and 14 days post-injury during room air breathing and acute respiratory challenge accomplished by inspiring a 10% O2, 7% CO2 gas mixture. Once a stable baseline recording was established, one of two different ampakines (CX717 or CX1739, 5 mg/kg, intravenous) or a vehicle (2-hydroxypropyl-beta-cyclodextrin [HPCD]) was delivered. At 4 days post-injury, both ampakines increased diaphragm EMG output ipsilateral to C2Hx during both baseline breathing and acute respiratory challenge. Only CX1739 treatment also led to a sustained (15 min) increase in ipsilateral EMG output. At 14 days post-injury, both ampakines produced sustained increases in ipsilateral diaphragm EMG output and enabled increased output during the respiratory challenge. We conclude that low dose ampakine treatment can increase diaphragm EMG activity after cervical SCI, and therefore may provide a pharmacological strategy that could be useful in the context of respiratory rehabilitation.

Electrical epidural stimulation of the cervical spinal cord: implications for spinal respiratory neuroplasticity after spinal cord injury.

Traumatic cervical spinal cord injury (cSCI) can lead to damage of bulbospinal pathways to the respiratory motor nuclei and consequent life-threatening respiratory insufficiency due to respiratory muscle paralysis/paresis. Reports of electrical epidural stimulation (EES) of the lumbosacral spinal cord to enable locomotor function after SCI are encouraging, with some evidence of facilitating neural plasticity. Here, we detail the development and success of EES in recovering locomotor function, with consideration of stimulation parameters and safety measures to develop effective EES protocols. EES is just beginning to be applied in other motor, sensory, and autonomic systems; however, there has only been moderate success in preclinical studies aimed at improving breathing function after cSCI. Thus, we explore the rationale for applying EES to the cervical spinal cord, targeting the phrenic motor nucleus for the restoration of breathing. We also suggest cellular/molecular mechanisms by which EES may induce respiratory plasticity, including a brief examination of sex-related differences in these mechanisms. Finally, we suggest that more attention be paid to the effects of specific electrical parameters that have been used in the development of EES protocols and how that can impact the safety and efficacy for those receiving this therapy. Ultimately, we aim to inform readers about the potential benefits of EES in the phrenic motor system and encourage future studies in this area.

Diaphragm Motor-Evoked Potential Induced by Cervical Magnetic Stimulation following Cervical Spinal Cord Contusion in the Rat.

Cervical spinal injury is typically associated with respiratory impairments due to damage to bulbospinal respiratory pathways and phrenic motoneurons. Magnetic stimulation is a non-invasive approach for the evaluation and modulation of the nervous system. The present study was designed to examine whether cervical magnetic stimulation can be applied to evaluate diaphragmatic motor outputs in a pre-clinical rat model of cervical spinal injury. The bilateral diaphragm was monitored in anesthetized rats using electromyogram at the acute, subchronic, and chronic stages following left mid-cervical contusion. The center of a figure-of-eight coil was placed 20 mm caudal to bregma to stimulate the cervical spinal cord. The results demonstrated that a single magnetic stimulation can evoke significant motor-evoked potentials in the diaphragms of uninjured animals when the animal's head was placed 30 mm right or left from the center of the coil. The spontaneous bursting of the diaphragm was significantly attenuated by contusion injury at all-time-points post-injury. However, the threshold of the diaphragmatic motor-evoked potential was reduced, and the amplitude of the diaphragmatic motor-evoked potential was enhanced in response to cervical magnetic stimulation at the acute injury stage. Moreover, the motor-evoked potentials of the bilateral diaphragm in animals with contusions were generally larger when the coil was placed at the left spinal cord at the subchronic and chronic injury stages. These results suggested that cervical magnetic stimulation can be used to examine the excitability of phrenic motor outputs post-injury, and magnetic stimulation applied more laterally may be more effective for triggering diaphragmatic motor-evoked potentials.

Diaphragm motor evoked potential induced by cervical magnetic stimulation following cervical spinal cord contusion in the rat

Cervical spinal cord injury is usually associated with respiratory impairments due to damage of bulbospinal respiratory pathways and cervical phrenic motoneurons. The magnetic stimulation is a non-invasive and safe approach to evaluate and modulate the central and peripheral nervous system. The purpose of the present study was to examine whether cervical magnetic stimulation can be applied to evaluate the diaphragmatic motor outputs in a pre-clinical rodent model of cervical spinal cord injury. The bilateral diaphragm electromyogram was measured in anesthetized rats at the acute, subchronic and chronic stage following left mid-cervical spinal cord contusion. The center of figure-of-eight coil was placed 20 mm caudal to the Bregma to stimulate the cervical spinal cord. The results demonstrated that cervical magnetic stimulation can evoke a significant diaphragm motor evoked potential in uninjured animals when the coil was placed at the lateral position (3 cm right or left to the midline). The spontaneous bursting of diaphragm was significantly attenuated by cervical spinal cord contusion at all-time points post-injury. However, the threshold of diaphragm motor evoked potential was reduced and the amplitude of diaphragm motor evoked potential was augmented in response to cervical magnetic stimulation at the acute injured stage. In addition, the motor evoked potential of bilateral diaphragm in contused animals was generally greater when the coil was placed at the left spinal cord at the subchronic and chronic injured stages. These results suggested that cervical magnetic stimulation can be used to examine the excitability of phrenic motor outputs, and the lateral cervical magnetic stimulation may be more effective to trigger the diaphragm motor evoked potential.

Developments in Understanding Diffuse Noxious Inhibitory Controls: Pharmacological Evidence from Pre-Clinical Research.

Bulbospinal pathways regulate nociceptive processing, and inhibitory modulation of nociception can be achieved via the activity of diffuse noxious inhibitory controls (DNIC), a unique descending pathway activated upon application of a conditioning stimulus (CS). Numerous studies have investigated the effects of varied pharmacological systems on the expression status of a) DNIC (as measured in anaesthetised animals) and b) the descending control of nociception (DCN), a surrogate measure of DNIC-like effects in conscious animals. However, the complexity of the underlying circuitry that governs initiation of a top-down inhibitory response in reaction to a CS, coupled with the methodological limitations associated with using pharmacological tools for its study, has often obscured the exact role(s) of a given drug. In this literature review, we discuss the pharmacological manipulation interrogation strategies that have hitherto been used to examine the functionality of DNIC and DCN. Discreet administration of a substance in the spinal cord or brain is considered in the context of action on one of four hypothetical systems that underlie the functionality of DNIC/DCN, where interpreting the outcome is often complicated by overlapping qualities. Systemic pharmacological modulation of DNIC/DCN is also discussed despite the fact that the precise location of drug action(s) cannot be pinpointed. Chiefly, modulation of the noradrenergic, serotonergic and opioidergic transmission systems impacts DNIC/DCN in a manner that relates to drug class, route of administration and health/disease state implicated. The advent of increasingly sophisticated interrogation tools will expedite our full understanding of the circuitries that modulate naturally occurring pain-inhibiting pathways.

Probing the neuromodulatory gain control system in sports and exercise sciences

The monoaminergic bulbospinal pathways from the brainstem are central to motor functions by regulating the gains of spinal motoneurons and represent, in that respect, probably the primary control system for motoneuron excitability. Yet, the efficiency of this system is few, if not never, assessed in the fields of sports and exercise sciences. In this review paper, we propose a methodological approach intended to assess how this neuromodulatory system affects motoneuron excitability. This approach is based on the use of tendon vibration which can, in certain circumstances, induce the generation of the so-called tonic vibration reflex through the stimulation of muscle spindles. Force and EMG responses to tendon vibration are indeed indicative of how this descending system modulates the gain of the ionotropic inputs from Ia afferents and thus of the strength of the monoaminergic drive. After a brief presentation of the neuromodulatory system and of the mechanisms involved in the generation of the tonic vibration reflex, we address some important methodological considerations regarding the use of the TVR to probe this neuromodulatory gain control system. Hopefully, this paper will encourage sports and exercise scientists to investigate this system.

Is Innervation of the Neuromuscular Junction at the Diaphragm Modulated by sGC/cGMP Signaling?

We previously reported NO/sGC signaling in the upper respiratory pathway, receiving input from the respiratory neurons of the brainstem to phrenic motoneurons in the C3–C6 spinal cord. In order to assess whether innervation of the neuromuscular junction (NMJ) at the diaphragm is modulated by sGC/cGMP signaling, we performed unilateral 8-day continuous ligation of the phrenic nerve in rats. We examined sGCβ1 within the lower bulbospinal pathway (phrenic motoneurons, phrenic nerves and NMJs at the diaphragm) and the cGMP level in the contra- and ipsilateral hemidiaphragm. Additionally, we characterized the extent of phrenic nerve axonal degeneration and denervation at diaphragm NMJs. The results of our study show that continuous 8-day phrenic nerve ligation caused a marked increase in sGCβ1 (immunoreactivity and the protein level) in the ipsilateral phrenic motor pool. However, the protein sGCβ1 level in the phrenic nerve below its ligation and the cGMP level in the ipsilateral hemidiaphragm were evidently decreased. Using confocal analysis we discovered a reduction in sGCβ1-IR boutons/synaptic vesicles at the ipsilateral MNJs. These findings are consistent with the marked axonal loss (∼47%) and significant NMJs degeneration in the ipsilateral diaphragm muscle. The remarkable unilateral decrease in cGMP level in the diaphragm and the failure of EMG recordings in the ipsilateral hemidiaphragm muscle can be attributed to the fact that sGC is involved in transmitter release at the diaphragm NMJs via the sGC-cGMP pathway.

Sympathetic Preganglionic Neurons (SPNs): a Promising Target to Regain Cardiorespiratory Control in Spinal Cord Injured (SCI) Patients

BackgroundThe consequences of spinal cord injuries (SCI) are massive and become more severe with higher levels of injury. Lesions located on cervical portions, specifically between C1‐C3 cause an interruption of the descending bulbospinal pathways. These pathways include a key component of the neural control of respiration, linking the brainstem breathing rhythm generator with phrenic motor neurons that activate the diaphragm. Disruption to these pathways result in respiratory muscle paresis and/or paralysis, requiring mechanical ventilation. Respiratory complications, such as pneumonia and atelectasis complicate the process of weaning patients from ventilators, contributing to patient mortality. Previous data collected in the Wilson lab demonstrated that sympathetic preganglionic neurons (SPNs) located at the thoracic spinal cord have remarkable oxygen and pressure‐sensitive capabilities. Here, we hypothesized that SPNs can stimulate phrenic motor neurons independent of the brainstem.AimTo study the novel direct influence of SPNs on the phrenic motor nucleus.MethodsAn in situ artificially‐perfused cervico‐thoracic rat spinal cord preparation was used to record phrenic and splanchnic nerve activity while having the cervical and thoracic spinal cord separately perfused. This approach allowed us to selectively expose the thoracic segment to acute hypoxia (30 Torr; 10 min) in order to observe if cardiorespiratory responses elicited by the SPN’s could stimulate phrenic motor neurons located at the cervical spinal cord. Additionally, we recorded the electromyography activity from the diaphragm during hypoxia and ischemia (0 mmHg; 10 min).ResultsThe results demonstrated that stimulation of the SPNs with hypoxia evoked phrenic nerve and diaphragm activity. This indicates that even without inputs from the brainstem, SPNs are responsive to hypoxia and ischemia and might have capability to stimulate phrenic motor neurons.ConclusionThe implications of these findings point to the thoracic SPNs as a unique and promising target to regain cardiorespiratory control in SCI patients. Further pharmacological investigations may reveal effective treatment strategies to improve life quality and reduce mortality of SCI patients.Support or Funding InformationAlberta Innovates Health Solutions (LL), Eyes High (NB), NSERC (HY), Canadian Institute of Health Research (RW).

The Contribution of Serotonergic Receptors and Nitric Oxide Systems in the Analgesic Effect of Acetaminophen: An Overview of the Last Decade.

Acetaminophen is a widely used analgesic and antipyretic agent. It is also available in over the counter formulations, which has increased its wide use. There have been many studies to date that have aimed to evaluate the mechanism of the analgesic action of acetaminophen. Additional to the inhibition of the cyclooxygenase pathway in the central nervous system, the involvement of opioidergic, cannabinoidergic, dopaminergic, cholinergic, and nitrergic systems as well as the contribution of descending pain inhibitory systems like the bulbospinal serotonergic pathway has been proposed as possible mechanisms of the analgesic action of acetaminophen. In this review, we aimed to collect the data from studies revealing the contribution of the central serotonergic system and the role of central nervous system-located serotonergic receptor subtypes in the analgesic effect of acetaminophen. While doing this, we mainly focused on the research that has been performed in the last ten years and tried to link the previous data with the lately added results. In addition to serotonergic system involvement, we also reviewed the role of nitric oxide in the analgesic action of acetaminophen, especially with the new findings reported over the last decade.

Plasticity Induced Recovery of Breathing Occurs at Chronic Stages after Cervical Contusion.

Severe midcervical contusion injury causes profound deficits throughout the respiratory motor system that last from acute to chronic time points post-injury. We use chondroitinase ABC (ChABC) to digest chondroitin sulphate proteoglycans within the extracellular matrix (ECM) surrounding the respiratory system at both acute and chronic time points post-injury to explore whether augmentation of plasticity can recover normal motor function. We demonstrate that, regardless of time post-injury or treatment application, the lesion cavity remains consistent, showing little regeneration or neuroprotection within our model. Through electromyography (EMG) recordings of multiple inspiratory muscles, however, we show that application of the enzyme at chronic time points post-injury initiates the recovery of normal breathing in previously paralyzed respiratory muscles. This reduced the need for compensatory activity throughout the motor system. Application of ChABC at acute time points recovered only modest amounts of respiratory function. To further understand this effect, we assessed the anatomical mechanism of this recovery. Increased EMG activity in previously paralyzed muscles was brought about by activation of spared bulbospinal pathways through the site of injury and/or sprouting of spared serotonergic fibers from the contralateral side of the cord. Accordingly, we demonstrate that alterations to the ECM and augmentation of plasticity at chronic time points post-cervical contusion can cause functional recovery of the respiratory motor system and reveal mechanistic evidence of the pathways that govern this effect.

Differences between flexion and extension synergy-driven coupling at the elbow, wrist, and fingers of individuals with chronic hemiparetic stroke

ObjectiveThe flexion and extension synergies were quantified at the paretic elbow, forearm, wrist, and finger joints within the same group of participants for the first time. Differences in synergy expression at each of the four joints were examined, as were the ways these differences varied across the joints. MethodsTwelve post-stroke individuals with chronic moderate-to-severe hemiparesis and six age-matched controls participated. Participants generated isometric shoulder abduction (SABD) and shoulder adduction (SADD) at four submaximal levels to progressively elicit the flexion and extension synergies, respectively. Isometric joint torques and EMG were recorded from shoulder, elbow, forearm (radio-ulnar), wrist, and finger joints and muscles. ResultsSABD elicited strong wrist and finger flexion torque that increased with shoulder torque level. SADD produced primarily wrist and finger flexion torque, but magnitudes at the wrist were less than during SABD. Findings contrasted with those at the elbow and forearm, where torques and EMG generated due to SABD and SADD were opposite in direction. ConclusionsFlexion and extension synergy expression are more similar at the hand than at the shoulder and elbow. Specific bulbospinal pathways that may underlie flexion and extension synergy expression are discussed. SignificanceWhole-limb behavior must be considered when examining paretic hand function in moderately-to-severely impaired individuals.

Compensatory Function of the Diaphragm after High Cervical Hemisection in the Rat.

Unilateral high cervical spinal hemisection (i.e., C2Hx) interrupts the respiratory bulbospinal pathway and results in paralysis of the hemidiaphragm. The ipsilateral diaphragmatic activity can partially recover over weeks to months; however, its contribution to the tidal volume generation is less than 20%. Accordingly, we hypothesized that the contralateral diaphragm exerts a compensatory function to maintain the essential ventilation following C2Hx. The cardiorespiratory pattern and bilateral diaphragm electromyogram (EMG) signals were measured in urethane-anesthetized and spontaneously breathing adult rats at 1 day, and 2 or 8 weeks post-C2Hx or C2 laminectomy. The functional contribution of the diaphragm was assessed by measuring immediate changes of the tidal volume following phrenic nerve section. At 1 day post-injury, the tidal volume was significantly reduced after contralateral phrenicotomy in C2Hx animals (54 ± 3% decline) compared with uninjured controls (20 ± 2% decline). Moreover, the arterial carbon dioxide partial pressure was significantly elevated in C2Hx animals (from 76 ± 8 mmHg to 117 ± 5 mmHg) but not in uninjured animals (from 51 ± 4 mmHg to 55 ± 3 mmHg). By 2 and 8 weeks post-injury, contralateral phrenicotomy still caused a greater reduction in the tidal volume in C2Hx than in uninjured animals, and the percentage decline of the tidal volume was similar to the response at 1 day post-injury. These data suggested that unilateral cervical spinal cord injury induced a persistent compensatory plasticity in the contralateral diaphragm, which plays a critical role in maintenance of essential ventilation.

Contribution of 5-HT2A receptors on diaphragmatic recovery after chronic cervical spinal cord injury

Unilateral C2 spinal cord hemisection (C2Hx) interrupts bulbospinal respiratory pathways innervating ipsilateral phrenic motoneurons, resulting in cessation of ipsilateral diaphragm motor output. Plasticity within the spinal neural circuitry controlling the diaphragm can induce partial recovery of phrenic bursting which correlates with the time-dependent return of spinal serotonin (5-HT) immunoreactivity in the vicinity of phrenic motoneurons. The 5-HT2A receptor subtype is present on phrenic motoneurons and its expression is up-regulated after cervical spinal cord injury; however the functional role of these receptors following injury has not been clearly defined. The present study evaluated the functional role of 5-HT2A receptors by testing the hypothesis that pharmacologic blockade would attenuate diaphragm activity in rats with chronic cervical spinal cord injury. Bilateral diaphragm electromyography (EMG) was performed in vagal-intact and spontaneously breathing rats before and after intravenous administration of the 5-HT2A receptor antagonist Ketanserin (1mg/kg). Intravenous ketanserin significantly attenuated ipsilateral diaphragm EMG activity in C2Hx animals but had no impact on diaphragm output in uninjured animals. We conclude that 5-HT2A receptor activation contributes to the recovery of ipsilateral phrenic motor output after chronic cervical spinal cord injury.

Upper Extremity Motor Impairments and Microstructural Changes in Bulbospinal Pathways in Chronic Hemiparetic Stroke

Following hemiparetic stroke, precise, individuated control of single joints is often replaced by highly stereotyped patterns of multi-joint movement, or abnormal limb synergies, which can negatively impact functional use of the paretic arm. One hypothesis for the expression of these synergies is an increased dependence on bulbospinal pathways such as the rubrospinal (RubST) tract and especially the reticulospinal (RetST) tracts, which co-activate multiple muscles of the shoulder, elbow, wrist, and fingers. Despite indirect evidence supporting this hypothesis in humans poststroke, it still remains unclear whether it is correct. Therefore, we used high-resolution diffusion tensor imaging (DTI) to quantify white matter microstructure in relation to severity of arm synergy and hand-related motor impairments. DTI was performed on 19 moderately to severely impaired chronic stroke individuals and 15 healthy, age-matched controls. In stroke individuals, compared to controls, there was significantly decreased fractional anisotropy (FA) and significantly increased axial and radial diffusivity in bilateral corona radiata and body of the corpus callosum. Furthermore, poststroke, the contralesional (CL) RetST FA correlated significantly with both upper extremity (UE) synergy severity (r = −0.606, p = 0.003) and hand impairment (r = −0.609, p = 0.003). FA in the ipsilesional RubST significantly correlated with hand impairment severity (r = −0.590, p = 0.004). For the first time, we separately evaluate RetST and RubST microstructure in chronic stroke individuals with UE motor impairment. We demonstrate that individuals with the greatest UE synergy severity and hand impairments poststroke have the highest FA in the CL RetST a pattern consistent with increased myelination and suggestive of neuroplastic reorganization. Since the RetST pathway microstructure, in particular, is sensitive to abnormal joint coupling and hand-related motor impairment in chronic stroke, it could help test the effects of specific, and novel, anti-synergy neurorehabilitation interventions for recovery from hemiparesis.

Differential innervation of superficial versus deep laminae of the dorsal horn by bulbo-spinal serotonergic pathways in the rat

Convergent data showed that bulbo-spinal serotonergic projections exert complex modulatory influences on nociceptive signaling within the dorsal horn. These neurons are located in the B3 area which comprises the median raphe magnus (RMg) and the lateral paragigantocellular reticular (LPGi) nuclei. Because LPGi 5-HT neurons differ from RMg 5-HT neurons regarding both their respective electrophysiological properties and responses to noxious stimuli, we used anatomical approaches for further characterization of the respective spinal projections of LPGi versus RMg 5-HT neuron subgroups.Adult Sprague-Dawley rats were stereotaxically injected into the RMg or the LPGi with the anterograde tracer Phaseolus vulgaris leucoagglutinin (PHA-L). The precise location of injection sites and RMg vs LPGi spinal projections into the different dorsal horn laminae were visualized by PHA-L immunolabeling. Double immunofluorescent labeling of PHA-L and the serotonin transporter (5-HTT) allowed detection of serotonergic fibers among bulbo-spinal projections.Anterograde tracing showed that RMg neurons project preferentially into the deep laminae V-VI whereas LPGi neuron projections are confined to the superficial laminae I-II of the ipsilateral dorsal horn. All along the spinal cord, double-labeled PHA-L/5-HTT immunoreactive fibers, which represent only 5–15% of all PHA-L-immunoreactive projections, exhibit the same differential locations depending on their origin in the RMg versus the LPGi.The clear-cut distinction between dorsal horn laminae receiving bulbo-spinal serotonergic projections from the RMg versus the LPGi provides further anatomical support to the idea that the descending serotonergic pathways issued from these two bulbar nuclei might exert different modulatory influences on the spinal relay of pain signaling neuronal pathways.

Promoting Respiratory Motor Plasticity After Spinal Cord Injury Through High Frequency Stimulation (HF‐SCS)

More than 50% of the spinal cord injury (SCI) population have injuries at the cervical level. Located at this level are the respiratory bulbospinal pathways which mediate breathing. Therefore, injuries at the cervical level can leave many of these patients unable to breath. One promising intervention to restore breathing is HF‐SCS (300Hz). In previous investigations, we demonstrated that HF‐SCS applied at the T2 level after an experimental complete C2 spinal cord section leads to evoked inspiratory diaphragmatic EMG activity that can enable survival of the animal. However, the extent to which HF‐SCS can promote spinal respiratory motor plasticity in order to promote restoration of breathing function is still unknown. To address this question, we performed a HF‐SCS at the T2 level in six anesthetized rats while recording bilateral diaphragmatic EMG activity one week after C2 hemisection. As expected, C2 hemisection abolished activity in the hemidiaphragm ipsilateral to the hemisection, while spontaneous activity contralateral to the C2 hemisection still remained. HF‐SCS resulted in robust inspiratory activity bilaterally in the diaphragm. Following discontinuation of HF‐SCS bilateral spontaneous activity persisted. Moreover, EMG activity on the hemisected side was synchronized with the contralateral side. Taken together, these results suggest that HF‐SCS after a C2 hemisection can enhance activity and promote plasticity of the crossed phrenic pathways which still remain. Further studies are planned to explore the mechanisms of these observations with the ultimate goal to restore function in patients following SCI.Support or Funding InformationFunded by University of Kentucky College of Medicine Funds & The MetroHeath Foundation.Disclosure: Dr. DiMarco has a significant financial interest in Synapse BioMedical, Inc, a manufacturer of diaphragm pacing systems.

Increasing spinal 5-HT2A receptor responsiveness mediates anti-allodynic effect and potentiates fluoxetine efficacy in neuropathic rats. Evidence for GABA release

Antidepressants are one of the first line treatments for neuropathic pain but their use is limited by the incidence and severity of side effects of tricyclics and the weak effectiveness of selective serotonin reuptake inhibitors (SSRIs). Serotonin type 2A (5-HT2A) receptors interact with PDZ proteins that regulate their functionality and SSRI efficacy to alleviate pain. We investigated whether an interfering peptide (TAT-2ASCV) disrupting the interaction between 5-HT2A receptors and associated PDZ proteins would improve the treatment of traumatic neuropathic allodynia. Tactile allodynia was assessed in spinal nerve ligation-induced neuropathic pain in rats using von Frey filaments after acute treatment with TAT-2ASCV and/or 5-HT2A receptor agonist, alone or in combination with repeated treatment with fluoxetine. In vivo microdialysis was performed in order to examine the involvement of GABA in TAT-2ASCV/fluoxetine treatment-associated analgesia. TAT-2ASCV (100ng, single i.t. injection) improved SNL-induced tactile allodynia by increasing 5-HT2A receptor responsiveness to endogenous 5-HT. Fluoxetine alone (10mg/kg, five i.p. injections) slightly increased tactile thresholds and its co-administration with TAT-2ASCV (100ng, single i.t. injection) further enhanced the anti-allodynic effect. This effect depends on the integrity of descending serotonergic bulbospinal pathways and spinal release of GABA. The anti-allodynic effect of fluoxetine can be enhanced by disrupting 5-HT2A receptor-PDZ protein interactions. This enhancement depends on 5-HT2A receptor activation, spinal GABA release and GABAA receptor activation.

Phrenic motor outputs in response to bronchopulmonary C-fibre activation following chronic cervical spinal cord injury.

Activation of bronchopulmonary C-fibres, the main chemosensitive afferents in the lung, can induce pulmonary chemoreflexes to modulate respiratory activity. Following chronic cervical spinal cord injury, bronchopulmonary C-fibre activation-induced inhibition of phrenic activity was exaggerated. Supersensitivity of phrenic motor outputs to the inhibitory effect of bronchopulmonary C-fibre activation is due to a shift of phrenic motoneuron types and slow recovery of phrenic motoneuron discharge in cervical spinal cord-injured animals. These data suggest that activation of bronchopulmonary C-fibres may retard phrenic output recovery following cervical spinal cord injury. The alteration of phenotype and discharge pattern of phrenic motoneuron enables us to understand the impact of spinal cord injury on spinal respiratory activity. Cervical spinal injury interrupts bulbospinal pathways and results in cessation of phrenic bursting ipsilateral to the lesion. The ipsilateral phrenic activity can partially recover over weeks to months following injury due to the activation of latent crossed spinal pathways and exhibits a greater capacity to increase activity during respiratory challenges than the contralateral phrenic nerve. However, whether the bilateral phrenic nerves demonstrate differential responses to respiratory inhibitory inputs is unclear. Accordingly, the present study examined bilateral phrenic bursting in response to capsaicin-induced pulmonary chemoreflexes, a robust respiratory inhibitory stimulus. Bilateral phrenic nerve activity was recorded in anaesthetized and mechanically ventilated adult rats at 8-9weeks after C2 hemisection (C2Hx) or C2 laminectomy. Intra-jugular capsaicin (1.5μgkg-1 ) injection was performed to activate the bronchopulmonary C-fibres to evoke pulmonary chemoreflexes. The present results indicate that capsaicin-induced prolongation of expiratory duration was significantly attenuated in C2Hx animals. However, ipsilateral phrenic activity was robustly reduced after capsaicin treatment compared to uninjured animals. Single phrenic fibre recording experiments demonstrated that C2Hx animals had a higher proportion of late-inspiratory phrenic motoneurons that were relatively sensitive to capsaicin treatment compared to early-inspiratory phrenic motoneurons. Moreover, late-inspiratory phrenic motoneurons in C2Hx animals had a weaker discharge frequency and slower recovery time than uninjured animals. These results suggest bilateral phrenic nerves differentially respond to bronchopulmonary C-fibre activation following unilateral cervical hemisection, and the severe inhibition of phrenic bursting is due to a shift in the discharge pattern of phrenic motoneurons.