Multi-Level Regulation of Opioid-Induced Respiratory Depression.

Opioids depress minute ventilation primarily by reducing respiratory rate. This results from direct effects on the preBötzinger Complex as well as from depression of the Parabrachial/Kölliker-Fuse Complex, which provides excitatory drive to preBötzinger Complex neurons mediating respiratory phase-switch. Opioids also depress awake drive from the forebrain and chemodrive.

Dose-dependent Respiratory Depression by Remifentanil in the Rabbit Parabrachial Nucleus/Kölliker-Fuse Complex and Pre-Bötzinger Complex.

BACKGROUND: Recent studies showed partial reversal of opioid-induced respiratory depression in the pre-Bötzinger complex and the parabrachial nucleus/Kölliker-Fuse complex. The hypothesis for this study was that opioid antagonism in the parabrachial nucleus/Kölliker-Fuse complex plus pre-Bötzinger complex completely reverses respiratory depression from clinically relevant opioid concentrations. METHODS: Experiments were performed in 48 adult, artificially ventilated, decerebrate rabbits. The authors decreased baseline respiratory rate ~50% with intravenous, "analgesic" remifentanil infusion or produced apnea with remifentanil boluses and investigated the reversal with naloxone microinjections (1 mM, 700 nl) into the Kölliker-Fuse nucleus, parabrachial nucleus, and pre-Bötzinger complex. In another group of animals, naloxone was injected only into the pre-Bötzinger complex to determine whether prior parabrachial nucleus/Kölliker-Fuse complex injection impacted the naloxone effect. Last, the µ-opioid receptor agonist [d-Ala,2N-MePhe,4Gly-ol]-enkephalin (100 µM, 700 nl) was injected into the parabrachial nucleus/Kölliker-Fuse complex. The data are presented as medians (25 to 75%). RESULTS: Remifentanil infusion reduced the respiratory rate from 36 (31 to 40) to 16 (15 to 21) breaths/min. Naloxone microinjections into the bilateral Kölliker-Fuse nucleus, parabrachial nucleus, and pre-Bötzinger complex increased the rate to 17 (16 to 22, n = 19, P = 0.005), 23 (19 to 29, n = 19, P < 0.001), and 25 (22 to 28) breaths/min (n = 11, P < 0.001), respectively. Naloxone injection into the parabrachial nucleus/Kölliker-Fuse complex prevented apnea in 12 of 17 animals, increasing the respiratory rate to 10 (0 to 12) breaths/min (P < 0.001); subsequent pre-Bötzinger complex injection prevented apnea in all animals (13 [10 to 19] breaths/min, n = 12, P = 0.002). Naloxone injection into the pre-Bötzinger complex alone increased the respiratory rate to 21 (15 to 26) breaths/min during analgesic concentrations (n = 10, P = 0.008) but not during apnea (0 [0 to 0] breaths/min, n = 9, P = 0.500). [d-Ala,2N-MePhe,4Gly-ol]-enkephalin injection into the parabrachial nucleus/Kölliker-Fuse complex decreased respiratory rate to 3 (2 to 6) breaths/min. CONCLUSIONS: Opioid reversal in the parabrachial nucleus/Kölliker-Fuse complex plus pre-Bötzinger complex only partially reversed respiratory depression from analgesic and even less from "apneic" opioid doses. The lack of recovery pointed to opioid-induced depression of respiratory drive that determines the activity of these areas.

Characteristics of breathing rate control mediated by a subregion within the pontine parabrachial complex.

The role of the dorsolateral pons in the control of expiratory duration (Te) and breathing frequency is incompletely understood. A subregion of the pontine parabrachial-Kölliker-Fuse (PB-KF) complex of dogs was identified via microinjections, in which localized pharmacologically induced increases in neuronal activity produced increases in breathing rate while decreases in neuronal activity produced decreases in breathing rate. This subregion is also very sensitive to local and systemic opioids. The purpose of this study was to precisely characterize the relationship between the PB-KF subregion pattern of altered neuronal activity and the control of respiratory phase timing as well as the time course of the phrenic nerve activity/neurogram (PNG). Pulse train electrical stimulation patterns synchronized with the onset of the expiratory (E) and/or phrenic inspiratory (I) phase were delivered via a small concentric bipolar electrode while the PNG was recorded in decerebrate, vagotomized dogs. Step frequency patterns during the E phase produced a marked frequency-dependent decrease in Te, while similar step inputs during the I phase increased inspiratory duration (Ti) by 14 ± 3%. Delayed pulse trains were capable of pacing the breathing rate by terminating the E phase and also of triggering a consistent stereotypical inspiratory PNG pattern, even when evoked during apnea. This property suggests that the I-phase pattern generator functions in a monostable circuit mode with a stable E phase and a transient I phase. Thus the I-pattern generator must contain neurons with nonlinear pacemaker-like properties, which allow the network to rapidly obtain a full on-state followed by relatively slow inactivation. The activated network can be further modulated and supplies excitatory drive to the neurons involved with pattern generation.NEW & NOTEWORTHY A circumscribed subregion of the pontine medial parabrachial nucleus plays a key role in the control of breathing frequency primarily via changes in expiratory duration. Excitation of this subregion triggers the onset of the inspiratory phase, resulting in a stereotypical ramplike phrenic activity pattern independent of time within the expiratory phase. The ability to pace the I-burst rate suggests that the in vivo I-pattern generating network must contain functioning pacemaker neurons.

A Subregion of the Parabrachial Nucleus Partially Mediates Respiratory Rate Depression from Intravenous Remifentanil in Young and Adult Rabbits.

BACKGROUND: The efficacy of opioid administration to reduce postoperative pain is limited by respiratory depression. We investigated whether clinically relevant opioid concentrations altered the respiratory pattern in the parabrachial nucleus, a pontine region contributing to respiratory pattern generation, and compared these effects with a medullary respiratory site, the pre-Bötzinger complex. METHODS: Studies were performed in 40 young and 55 adult artificially ventilated, decerebrate rabbits. We identified an area in the parabrachial nucleus where α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid microinjections elicited tachypnea. Two protocols were performed in separate sets of animals. First, bilateral microinjections of the µ-opioid receptor agonist [D-Ala, N-MePhe, Gly-ol]-enkephalin (100 µM) into the "tachypneic area" determined the effect of maximal µ-opioid receptor activation. Second, respiratory rate was decreased with continuous IV infusions of remifentanil. The opioid antagonist naloxone (1 mM) was then microinjected bilaterally into the "tachypneic area" of the parabrachial nucleus to determine whether the respiratory rate depression could be locally reversed. RESULTS: Average respiratory rate was 27 ± 10 breaths/min. First, [D-Ala, N-MePhe, Gly-ol]-enkephalin injections decreased respiratory rate by 62 ± 20% in young and 45 ± 26% in adult rabbits (both P < 0.001). Second, during IV remifentanil infusion, bilateral naloxone injections into the "tachypneic area" of the parabrachial nucleus reversed respiratory rate depression from 55 ± 9% to 20 ± 14% in young and from 46 ± 20% to 18 ± 27% in adult rabbits (both P < 0.001). The effects of bilateral [D-Ala, N-MePhe, Gly-ol]-enkephalin injection and IV remifentanil on respiratory phase duration in the "tachypneic area" of the parabrachial nucleus was significantly different from the pre-Bötzinger complex. CONCLUSIONS: The "tachypneic area" of the parabrachial nucleus is highly sensitive to µ-opioid receptor activation and mediates part of the respiratory rate depression by clinically relevant administration of opioids.

Opioid-induced Respiratory Depression Is Only Partially Mediated by the preBötzinger Complex in Young and Adult Rabbits In Vivo.

BACKGROUND: The preBötzinger Complex (preBC) plays an important role in respiratory rhythm generation. This study was designed to determine whether the preBC mediated opioid-induced respiratory rate depression at clinically relevant opioid concentrations in vivo and whether this role was age dependent. METHODS: Studies were performed in 22 young and 32 adult New Zealand White rabbits. Animals were anesthetized, mechanically ventilated, and decerebrated. The preBC was identified by the tachypneic response to injection of D,L-homocysteic acid. (1) The µ-opioid receptor agonist [D-Ala2,N-Me-Phe4,Gly-ol]-enkephalin (DAMGO, 100 µM) was microinjected into the bilateral preBC and reversed with naloxone (1 mM) injection into the preBC. (2) Respiratory depression was achieved with intravenous remifentanil (0.08 to 0.5 µg kg(-1) min(-1)). Naloxone (1 mM) was microinjected into the preBC in an attempt to reverse the respiratory depression. RESULTS: (1) DAMGO injection depressed respiratory rate by 6 ± 8 breaths/min in young and adult rabbits (mean ± SD, P < 0.001). DAMGO shortened the inspiratory and lengthened the expiratory fraction of the respiratory cycle by 0.24 ± 0.2 in adult and young animals (P < 0.001). (2) During intravenous remifentanil infusion, local injection of naloxone into the preBC partially reversed the decrease in inspiratory fraction/increase in expiratory fraction in young and adult animals (0.14 ± 0.14, P < 0.001), but not the depression of respiratory rate (P = 0.19). PreBC injections did not affect respiratory drive. In adult rabbits, the contribution of non-preBC inputs to expiratory phase duration was larger than preBC inputs (3.5 [-5.2 to 1.1], median [25 to 75%], P = 0.04). CONCLUSIONS: Systemic opioid effects on respiratory phase timing can be partially reversed in the preBC without reversing the depression of respiratory rate.

Changes in pontine and preBötzinger/Bötzinger complex neuronal activity during remifentanil-induced respiratory depression in decerebrate dogs.

Introduction: In vivo studies using selective, localized opioid antagonist injections or localized opioid receptor deletion have identified that systemic opioids dose-dependently depress respiratory output through effects in multiple respiratory-related brainstem areas. Methods: With approval of the subcommittee on animal studies of the Zablocki VA Medical Center, experiments were performed in 53 decerebrate, vagotomized, mechanically ventilated dogs of either sex during isocapnic hyperoxia. We performed single neuron recordings in the Pontine Respiratory Group (PRG, n = 432) and preBötzinger/Bötzinger complex region (preBötC/BötC, n = 213) before and during intravenous remifentanil infusion (0.1-1 mcg/kg/min) and then until complete recovery of phrenic nerve activity. A generalized linear mixed model was used to determine changes in Fn with remifentanil and the statistical association between remifentanil-induced changes in Fn and changes in inspiratory and expiratory duration and peak phrenic activity. Analysis was controlled via random effects for animal, run, and neuron type. Results: Remifentanil decreased Fn in most neuron subtypes in the preBötC/BötC as well as in inspiratory (I), inspiratory-expiratory, expiratory (E) decrementing and non-respiratory modulated neurons in the PRG. The decrease in PRG inspiratory and non-respiratory modulated neuronal activity was associated with an increase in inspiratory duration. In the preBötC, the decrease in I-decrementing neuron activity was associated with an increase in expiratory and of E-decrementing activity with an increase in inspiratory duration. In contrast, decreased activity of I-augmenting neurons was associated with a decrease in inspiratory duration. Discussion: While statistical associations do not necessarily imply a causal relationship, our data suggest mechanisms for the opioid-induced increase in expiratory duration in the PRG and preBötC/BötC and how inspiratory failure at high opioid doses may result from a decrease in activity and decrease in slope of the pre-inspiratory ramp-like activity in preBötC/BötC pre-inspiratory neurons combined with a depression of preBötC/BötC I-augmenting neurons. Additional studies must clarify whether the observed changes in neuronal activity are due to direct neuronal inhibition or decreased excitatory inputs.

Pontine µ-opioid receptors mediate bradypnea caused by intravenous remifentanil infusions at clinically relevant concentrations in dogs.

Life-threatening side effects such as profound bradypnea or apnea and variable upper airway obstruction limit the use of opioids for analgesia. It is yet unclear which sites containing µ-opioid receptors (µORs) within the intact in vivo mammalian respiratory control network are responsible. The purpose of this study was 1) to define the pontine region in which µOR agonists produce bradypnea and 2) to determine whether antagonism of those µORs reverses bradypnea produced by intravenous remifentanil (remi; 0.1-1.0 µg·kg(-1)·min(-1)). The effects of microinjections of agonist [D-Ala(2),N-Me-Phe(4),Gly-ol(5)]-enkephalin (DAMGO; 100 µM) and antagonist naloxone (NAL; 100 µM) into the dorsal rostral pons on the phrenic neurogram were studied in a decerebrate, vagotomized, ventilated, paralyzed canine preparation during hyperoxia. A 1-mm grid pattern of microinjections was used. The DAMGO-sensitive region extended from 5 to 7 mm lateral of midline and from 0 to 2 mm caudal of the inferior colliculus at a depth of 3-4 mm. During remi-induced bradypnea (~72% reduction in fictive breathing rate) NAL microinjections (~500 nl each) within the region defined by the DAMGO protocol were able to reverse bradypnea by 47% (SD 48.0%) per microinjection, with 13 of 84 microinjections producing complete reversal. Histological examination of fluorescent microsphere injections shows that the sensitive region corresponds to the parabrachial/Kölliker-Fuse complex.

Contribution of the caudal medullary raphe to opioid induced respiratory depression.

BACKGROUND: Opioid-induced respiratory depression can be partially antagonized in the preBötzinger Complex and Parabrachial Nucleus/Kölliker-Fuse Complex. We hypothesized that additional opioid antagonism in the caudal medullary raphe completely reverses the opioid effect. METHODS: In adult ventilated, vagotomized, decerebrate rabbits, we administrated remifentanil intravenously at "analgesic", "apneic", and "very high" doses and determined the reversal with sequential naloxone microinjections into the bilateral Parabrachial Nucleus/Kölliker-Fuse Complex, preBötzinger Complex, and caudal medullary raphe. In separate animals, we injected opioid antagonists into the raphe without intravenous remifentanil. RESULTS: Sequential naloxone microinjections completely reversed respiratory rate depression from "analgesic" and "apneic" remifentanil, but not "very high" remifentanil concentrations. Antagonist injection into the caudal medullary raphe without remifentanil independently increased respiratory rate. CONCLUSIONS: Opioid-induced respiratory depression results from a combined effect on the respiratory rhythm generator and respiratory drive. The effect in the caudal medullary raphe is complex as we also observed local antagonism of endogenous opioid receptor activation, which has not been described before.

The effect of caudal vs intravenous morphine on early extubation and postoperative analgesic requirements for stage 2 and 3 single-ventricle palliation: a double blind randomized trial.

BACKGROUND: High-dose single-shot caudal morphine has been postulated to facilitate early extubation and to lower initial analgesic requirements after staged single-ventricle (SV) palliation. METHODS: With Institutional Review Board approval and written informed parental consent, 64 SV children aged 75-1667 days were randomized to pre-incisional caudal morphine-bupivacaine (100 µg·kg(-1) morphine (concentration 0.1%), mixed with 0.25% bupivacaine with 1 : 200,000 epinephrine, total 1 ml·kg(-1)) and postcardiopulmonary bypass (CPB) intravenous (IV) droperidol (75 µg·kg(-1)) ('active caudal group') or pre-incisional caudal saline (1 ml·kg(-1)) and post-CPB IV morphine (150 µg·kg(-1)) with droperidol (75 µg·kg(-1)) ('active IV group'). Assignment remained concealed from families and the care teams throughout the trial. Early extubation failure rates (primary or reintubation within 24 h), time to first postoperative rescue morphine analgesia, and 12-h postoperative morphine requirements were assessed for extubated patients. RESULTS: Thirty-one (12 stage 2) SV patients received caudal morphine and 32 (15 stage 2) received IV morphine. Extubation failure rates were 6/31 (19%) for caudal and 5/32 (16%) for IV morphine. For successfully extubated patients (n = 54), active caudal treatment significantly delayed the need for postoperative rescue morphine in stage 3 patients (P = 0.02) but not in stage 2 patients (P = 0.189) (Kaplan-Meier survival analysis with LogRank test). The reduction in 12-h postoperative morphine requirements with active caudal treatment did not reach significance (P = 0.085) but morphine requirements were significantly higher for stage 2 compared with stage 3 patients (P < 0.001) (two-way anova in n = 50 extubated patients). CONCLUSIONS: High-dose caudal morphine with bupivacaine delayed the need for rescue morphine analgesia in stage 3 patients. All stage 2 patients required early rescue morphine and had significantly higher postoperative 12-h morphine requirements than stage 3 patients. Early extubation is feasible for the majority of stage 2 and 3 SV patients regardless of analgesic regimen. The study was underpowered to assess differences in extubation failure rates.

Interaction between the pulmonary stretch receptor and pontine control of expiratory duration.

Medial parabrachial nucleus (mPBN) neuronal activity plays a key role in controlling expiratory (E)-duration (TE). Pulmonary stretch receptor (PSR) activity during the E-phase prolongs TE. The aims of this study were to characterize the interaction between the PSR and mPBN control of TE and underlying mechanisms. Decerebrated mechanically ventilated dogs were studied. The mPBN subregion was activated by electrical stimulation via bipolar microelectrode. PSR afferents were activated by low-level currents applied to the transected central vagus nerve. Both stimulus-frequency patterns during the E-phase were synchronized to the phrenic neurogram; TE was measured. A functional mathematical model for the control of TE and extracellular recordings from neurons in the preBötzinger/Bötzinger complex (preBC/BC) were used to understand mechanisms. Findings show that the mPBN gain-modulates, via attenuation, the PSR-mediated reflex. The model suggested functional sites for attenuation and neuronal data suggested correlates. The PSR- and PB-inputs appear to interact on E-decrementing neurons, which synaptically inhibit pre-I neurons, delaying the onset of the next I-phase.

Clinically relevant infusion rates of mu-opioid agonist remifentanil cause bradypnea in decerebrate dogs but not via direct effects in the pre-Bötzinger complex region.

Systemic administration of mu-opioids at clinical doses for analgesia typically slows respiratory rate. Mu-opioid receptors (MORs) on pre-Bötzinger Complex (pre-BötC) respiratory neurons, the putative kernel of respiratory rhythmogenesis, are potential targets. The purpose of this study was to determine the contribution of pre-BötC MORs to the bradypnea produced in vivo by intravenous administration of clinically relevant infusion rates of remifentanil (remi), a short-acting, potent mu-opioid analgesic. In decerebrate dogs, multibarrel micropipettes were used to record pre-BötC neuronal activity and to eject the opioid antagonist naloxone (NAL, 0.5 mM), the glutamate agonist D-homocysteic acid (DLH, 20 mM), or the MOR agonist [D-Ala(2), N-Me-Phe(4), gly-ol(5)]-enkephalin (DAMGO, 100 microM). Inspiratory and expiratory durations (T(I) and T(E)) and peak phrenic nerve activity (PPA) were measured from the phrenic neurogram. The pre-BötC was functionally identified by its rate altering response (typically tachypnea) to DLH microinjection. During intravenous remi-induced bradypnea (approximately 60% decrease in central breathing frequency, f(B)), bilateral injections of NAL in the pre-BötC did not change T(I), T(E), f(B), and PPA. Also, NAL picoejected onto single pre-BötC neurons depressed by intravenous remi had no effect on their discharge. In contrast, approximately 60 microg/kg of intravenous NAL rapidly reversed all remi-induced effects. In a separate group of dogs, microinjections of DAMGO in the pre-BötC increased f(B) by 44%, while subsequent intravenous remi infusion more than offset this DAMGO induced tachypnea. These results indicate that mu-opioids at plasma concentrations that cause profound analgesia produce their bradypneic effect via MORs located outside the pre-BötC region.

Endogenous glutamatergic inputs to the Parabrachial Nucleus/Kölliker-Fuse Complex determine respiratory rate.

The Kölliker-Fuse Nucleus (KF) has been widely investigated for its contribution to "inspiratory off-switch" while more recent studies showed that activation of the Parabrachial Nucleus (PBN) shortened expiratory duration. This study used an adult, in vivo, decerebrate rabbit model to delineate the contribution of each site to inspiratory and expiratory duration through sequential block of glutamatergic excitation with the receptor antagonists 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo[f]quinoxaline-2,3-dione (NBQX) and d(-)-2-amino-5-phosphonopentanoic acid (AP5). Glutamatergic disfacilitation caused large increases in inspiratory and expiratory duration and minor decrease in peak phrenic activity (PPA). Hypoxia only partially reversed respiratory rate depression but PPA was increased to >200 % of control. The contribution of PBN activity to inspiratory and expiratory duration was equal while block of the KF affected inspiratory duration more than expiratory. We conclude that in the in vivo preparation respiratory rate greatly depends on PBN/KF activity, which contributes to the "inspiratory on- "and "off-switch", but is of minor importance for the magnitude of phrenic motor output.

Inputs to medullary respiratory neurons from a pontine subregion that controls breathing frequency.

Neurons in a subregion of the medial parabrachial (PB) complex control expiratory duration (TE) and the inspiratory on-switch. To better understanding the underlying mechanisms, this study aimed to determine the types of medullary neurons in the rhythmogenic preBötzinger/Bötzinger Complex (preBötC/BötC) and adjacent areas that receive synaptic inputs from the PB subregion and whether these inputs are excitatory or inhibitory in nature. Highly localized electrical stimuli in the PB subregion combined with multi-electrode recordings from respiratory neurons and phrenic nerve activities were used to generate stimulus-to-spike event histograms to detect correlations in decerebrate, vagotomized dogs during isocapnic hyperoxia. Short-time scale correlations were found in 237/442 or ∼54% of the ventral respiratory column (VRC) neurons. Inhibition of E-neurons was ∼2.5X greater than for I-neurons, while Pre-I and I-neurons were excited. These findings indicate that the control of TE and the inspiratory on-switch by the PB subregion are mediated by a marked inhibition of BötC E-neurons combined with an excitation of I-neurons, especially pre-I neurons.

The contribution of endogenous glutamatergic input in the ventral respiratory column to respiratory rhythm.

Glutamate is the predominant excitatory neurotransmitter in the ventral respiratory column; however, the contribution of glutamatergic excitation in the individual subregions to respiratory rhythm generation has not been fully delineated. In an adult, in vivo, decerebrate rabbit model during conditions of mild hyperoxic hypercapnia we blocked glutamatergic excitation using the receptor antagonists 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo[f]quinoxaline-2,3-dione (NBQX) and d(-)-2-amino-5-phosphonopentanoic acid (AP5). Disfacilitation of the preBötzinger Complex caused a decrease in inspiratory and expiratory duration as well as peak phrenic amplitude and ultimately apnea. Disfacilitation of the Bötzinger Complex caused a decrease in inspiratory and expiratory duration; subsequent disfacilitation of the preBötzinger Complex resulted in complete loss of the respiratory pattern but maintained tonic inspiratory activity. We conclude that glutamatergic drive to the preBötzinger Complex is essential for respiratory rhythm generation. Glutamatergic drive to the Bötzinger Complex significantly affects inspiratory and expiratory phase duration. Bötzinger Complex neurons are responsible for maintaining the silent expiratory phase of the phrenic neurogram.

Anesthetic effects on synaptic transmission and gain control in respiratory control.

All volatile and most intravenous general anesthetics currently in clinical use cause respiratory depression at concentrations suitable for surgery. While various in vitro studies have identified potential molecular targets, their contributions to respiratory depression are poorly understood. At surgical concentrations, anesthetics principally affect ligand-gated, rather than voltage-gated ion channels. Here we focus on anesthetic-induced effects on synaptic transmission in brainstem respiratory neurons. The spontaneous discharge patterns of canine respiratory bulbospinal premotor neurons in vivo depend principally on NMDA and non-NMDA receptor-mediated excitation, while GABAA receptors mediate gain modulation and silent-phase inhibition. Studies examining the effects of volatile anesthetics on synaptic neurotransmission to these neurons suggest a primary role for postsynaptic enhancement of GABAA receptor function, partly offset by a reduction in presynaptic inhibition and a presynaptic reduction in glutamatergic excitation. In studies involving canine inspiratory hypoglossal motoneurons in vivo, which are already strongly depressed by low concentrations (< 0.5 MAC) of volatile anesthetics, the role of acid-sensitive, two-pore domain K+ (TASK) channels was found to be minimal at these subanesthetic concentrations. Potentiation of GABAA receptor-mediated inhibition was suggested. These studies on canine respiratory neurons provide valuable insights into mechanisms of anesthetic depression within a respiratory control subsystem; future studies will be required to determine anesthetic effects on sources of respiratory drive, rhythm, and their control.

Major components of endogenous neurotransmission underlying the discharge activity of hypoglossal motoneurons in vivo.

Multibarrel micropipettes were used to simultaneously record unit activity and apply antagonists on individual inspiratory hypoglossal motoneurons (IHMNs) to determine the endogenous activation levels of NMDA, non-NMDA, GABA(A) and serotonin receptors responsible for the IHMN spontaneous discharge patterns in decerebrate dogs. IHMN activity is highly dependent on glutamatergic phasic and tonic drives, which are differentially mediated by the receptor subtypes. Endogenous serotonin significantly amplifies IHMN activity, while GABAergic gain modulation acts to attenuate activity. Thus, alterations in the neurotransmission of any of these systems could markedly alter neuronal output to target muscles.

Retrograde labeling reveals extensive distribution of genioglossal motoneurons possessing 5-HT2A receptors throughout the hypoglossal nucleus of adult dogs.

Inspiratory hypoglossal motoneurons (IHMNs) innervate the muscles of the tongue and play an important role in maintaining upper airway patency. However, this may be reduced during sleep and by sedatives, potent analgesics, and volatile anesthetics. The genioglossal (GG) muscle is the main protruder and depressor muscle of the tongue and contributes to upper airway patency during inspiration. In vitro data suggest that serotonin (5-hydroxytryptamine, 5-HT), via the 5-HT(2A) receptor (5-HT(2A)R) subtype, plays a key role in controlling the excitability of IHMNs. The distribution of GG motoneurons (GGMNs) within the hypoglossal (XII) nucleus has not been studied in the adult dog. Further, it is uncertain whether the 5-HT(2A)R is located on GGMNs in the adult dog. We therefore used the cholera toxin B (CTB) subunit as a retrograde tracer to map the location of GGMNs in combination with immunofluorescent labeling to determine the presence and colocalization of 5-HT(2A)R within the XII nucleus in adult mongrel dogs. Injection of CTB into the GG muscle resulted in retrogradely labeled cells in a compact column throughout the XII nucleus, extending from 0.75 mm caudal to 3.45 mm rostral to the obex. Fluorescence immunohistochemistry revealed extensive 5-HT(2A)R labeling on CTB-labeled GGMNs. Identification of the 5-HT(2A)R on GGMNs in the XII nucleus of the adult dog supports in vitro data and suggests a physiological role for this receptor subtype in controlling the excitability of GGMNs, which contribute to the maintenance of upper airway patency.

Central pathways of pulmonary and lower airway vagal afferents.

Lung sensory receptors with afferent fibers coursing in the vagus nerves are broadly divided into three groups: slowly (SAR) and rapidly (RAR) adapting stretch receptors and bronchopulmonary C fibers. Central terminations of each group are found in largely nonoverlapping regions of the caudal half of the nucleus of the solitary tract (NTS). Second order neurons in the pathways from these receptors innervate neurons located in respiratory-related regions of the medulla, pons, and spinal cord. The relative ease of selective activation of SARs, and to a lesser extent RARs, has allowed for more complete physiological and morphological characterization of the second and higher order neurons in these pathways than for C fibers. A subset of NTS neurons receiving afferent input from SARs (termed pump or P-cells) mediates the Breuer-Hering reflex and inhibits neurons receiving afferent input from RARs. P-cells and second order neurons in the RAR pathway also provide inputs to regions of the ventrolateral medulla involved in control of respiratory motor pattern, i.e., regions containing a predominance of bulbospinal premotor neurons, as well as regions containing respiratory rhythm-generating neurons. Axon collaterals from both P-cells and RAR interneurons, and likely from NTS interneurons in the C-fiber pathway, project to the parabrachial pontine region where they may contribute to plasticity in respiratory control and integration of respiratory control with other systems, including those that provide for voluntary control of breathing, sleep-wake behavior, and emotions.