| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
1Department of Pharmacology, University of Virginia, Charlottesville, Virginia; and 2Department of Physiology, Universidade Federal de São Paulo–Escola Paulista de Medicina, São Paulo, São Paulo, Brazil
Submitted 22 March 2007; accepted in final form 23 April 2007
 |
ABSTRACT |
|---|
|
|
|
INTRODUCTION |
|---|
|
; Mulkey et al. 2004; Okada et al. 2002; Putnam et al. 2004; Takakura et al. 2006). These neurons selectively innervate the pontomedullary regions that contain the respiratory pattern generator [central pattern generator (CPG)] (Cream et al. 2002; Feldman and Del Negro 2006; Mulkey et al. 2004; Rosin et al. 2006). The excitatory input from RTN neurons seems necessary for the CPG to be active under anesthesia and RTN neurons also appear to contribute notably to respiration in the awake state and during sleep (Akilesh et al. 1997; Feldman et al. 2003; Nattie and Li 2000; Stornetta et al. 2006; Takakura et al. 2006).
The activity of RTN neurons is subject to several feedbacks consistent with their postulated role as a pH-regulated source of excitatory drive to the CPG. For example, their intrinsic acid sensitivity combined with their excitatory input from the carotid bodies forms a long-loop feedback that seems appropriate to contribute to the homeostatic regulation of pCO2 and pO2 (Feldman et al. 2003; Li and Nattie 2002; Nattie et al. 2001; Takakura et al. 2006). RTN neurons also receive inhibitory inputs from the CPG (Guyenet et al. 2005) and are inhibited by the activation of slowly adapting lung stretch receptors (SARs) (Moreira et al. 2007). The latter two feedbacks seem appropriate to reduce the intensity of the excitatory input from RTN chemoreceptors to the CPG when the latter network is being adequately driven by other inputs.
According to our prior study, SARs may inhibit RTN neurons by the simplest possible route: by a direct projection from inhibitory second-order relay neurons (pump cells) located in the solitary tract nucleus (NTS) (Moreira et al. 2007). The present study is designed to test this possibility. Using a combination of electrophysiological and anatomical experiments, we show here that GABAergic NTS pump cells do indeed innervate the RTN region.
|
|
METHODS |
|---|
|
The experiments were performed on 19 male Sprague–Dawley rats (Taconic Farms, Germantown, NY) weighing 250–350 g. Procedures were in accordance with National Institutes of Health Animal Care and Use Guidelines and were approved by the University of Virginia's Animal Care and Use Committee.
In vivo recordings
General anesthesia was induced with 5% halothane in 100% oxygen. The rats received a tracheostomy. Artificial ventilation with 1.4–1.5% halothane in 100% oxygen was maintained throughout surgery. The surgical procedures (arterial cannulation, phrenic nerve dissection, dorsal access to the medulla oblongata through the atlanto-occipital membrane) were described previously (Guyenet et al. 2005; Takakura et al. 2006). A bilateral pneumothorax was performed for added stability and both vagus nerves were left intact.
On completion of surgical procedures, halothane concentration was reduced to 1% (still in 100% oxygen). This anesthetic level is sufficient to abolish the corneal reflex and the following reactions to strong nociceptive stimulation of the hindpaw: retraction of distal phalanges, increases in phrenic nerve discharge (PND) amplitude, and rise of arterial pressure (AP). Rectal temperature was maintained at 37°C and end-expiratory CO2 was monitored throughout the experiment with a capnometer (Columbus Instruments, Columbus, OH). The muscle relaxant pancuronium was administered intravenously at the initial dose of 1 mg/kg and the adequacy of anesthesia was thereafter gauged by the lack of increase in AP and PND amplitude to firm toe pinch. No adjustment of the halothane concentration was needed.
Arterial pressure (AP), PND, and tracheal CO2 were recorded as previously described (Mulkey et al. 2004; Stornetta et al. 2006; Takakura et al. 2006). Lung inflation was monitored by measuring tracheal pressure through a side port of the tracheal cannula. Lung mechanoreceptors were activated by transiently elevating positive end-expiratory pressure (PEEP) (5–20 s) from a resting level of +1 cmH2O to +2, +4 or +6 cmH2O. Single-unit recording and juxtacellular labeling of the recorded units with biotinamide were done as described previously (Mulkey et al. 2004; Stornetta et al. 2006; Takakura et al. 2006). All recordings were done within the portion of the NTS that extends from the rostral end to the caudal end (calamus scriptorius) of the area postrema. Pump cells were encountered at a depth of 300 to 700 µm below the dorsal surface of the medulla oblongata within a longitudinally oriented column of at most 300 µm in width with its center 600 to 1,000 µm from the midline depending on the rat. Most recordings were made on the left side of the brain.
All analog data (end-expiratory CO2, PND, unit activity, AP) were stored on a microcomputer by a micro-1401 digitizer from Cambridge Electronics Design (CED, Cambridge, UK) and were processed off-line using version 5 of the Spike2 software (CED). Processing included action potential discrimination and binning, neuronal discharge rate measurement, and PND "integration" (iPND) consisting of rectification and smoothing (
: 0.015 s) (Mulkey et al. 2004; Stornetta et al. 2006; Takakura et al. 2006). The CED software was also used for acquisition of perievent histograms of neuronal activity and perievent averages of iPND, tracheal CO2, or tracheal pressure. The perievent histograms of neuronal single-unit activity were triggered either on iPND or on the tracheal pressure trace. Each histogram represents the summation of 
100 consecutive central respiratory or ventilation cycles (350 to 800 action potentials per histogram). At the end of the physiological experiment the rats were deeply anesthetized with halothane (4% until AP reached 40 mmHg). Then they were perfused through the left cardiac ventricle with phosphate-buffered saline (pH 7.4; 150 ml) followed by paraformaldehyde (4% in 0.1 M phosphate buffer, pH 7.4, 500 ml). The brains were postfixed overnight in the paraformaldehyde solution. The brains were then sectioned in the coronal plane (30 µm) and the sections were kept in cryoprotectant awaiting histological procedures.
Tracer injections
Survival surgery was done using aseptic procedures on rats anesthetized with a mixture of ketamine, xylazine, and acepromazine as described previously (Rosin et al. 2006). After surgery and tracer injection, the rats received an antibiotic (ampicillin, 100 mg/kg, administered intramuscularly; American Pharmaceutical Partners, Schaumberg, IL) and an analgesic (ketorolac tromethamine, 0.6 mg/kg, administered subcutaneously; Abbott Laboratories, North Chicago, IL). One week later the rats were deeply anesthetized with pentobarbital (60 mg/kg, administered intraperitoneally; Abbott Laboratories) then killed by intracardiac perfusion with 4% formaldehyde as detailed earlier. Tracer injection was done exactly as described previously (Rosin et al. 2006). The procedure involved iontophoretic delivery of the tracer through glass electrodes drawn from 2-mm glass with an external tip diameter of no more than 5 microns. In four rats, the anterograde tracer biotinylated dextranamide (BDA; lysine fixable, MW 10,000; 10% by weight in 10 mM phosphate buffer, pH 7.4; Molecular Probes, Eugene, OR; 5-µA positive current 5 s ON, 5 s OFF) was injected into the region of the solitary tract nucleus that contains the pump cells (500 µm rostral to the calamus scriptorius, 0.8 mm lateral to the midline, and 0.5 mm below the dorsal surface). Two other rats received an injection of cholera toxin B (CTB, 1% in 0.2 M phosphate buffer, pH 7.35; List Biological, Campbell, CA; 5-µA positive current 5 s ON, 5 s OFF) into the RTN. Injections into the RTN were guided by recording the facial field potential and were placed 200–300 µm below the lower edge of the field, 1.6–1.9 mm lateral to the midline, and 200–400 µm rostral to the caudal end of the field (Rosin et al. 2006). This region contains the largest concentration of RTN chemoreceptors (Stornetta et al. 2006).
Histology
All histochemical procedures were done using 30-µm-thick free-floating sections according to previously described protocols (Rosin et al. 2006; Stornetta et al. 2006; Takakura et al. 2006). Neurons labeled in vivo with biotinamide (juxtacellular recording) or terminal fields labeled anterogradely with BDA were identified by incubating the sections with streptavidin conjuguated with Cy-3 according to previously described methods (Mulkey et al. 2004; Rosin et al. 2006). Glutamic acid decarboxylase 67 (GAD67) mRNA and glycine transporter 2 (GLYT2) mRNA were detected by nonradioactive in situ hybridization using digoxigenin-labeled cRNA probes as described previously (Stornetta and Guyenet 1999; Stornetta et al. 2003b). GAD immunoreactivity was detected by immunofluorescence using a rabbit anti-GAD65/67 antibody (AB 5907, dilution: 1:5,000; Chemicon International, Temecula, CA). Guinea pig anti-GAD65/67 (Chemicon AB1511) was raised against a synthetic peptide (Chemicon AG252) with amino acid sequence CDF LIE EIE RLG QDL from the C-terminal portion of rat glutamate decarboxylase (GAD65; Cys + C-terminal residues 572–585). The manufacturer demonstrated by Western blot of whole brain lysate that the antiserum recognizes bands of the correct molecular weight corresponding to the two isoforms of the enzyme. In our laboratory, preabsorption of the GAD65/67 antibody with a threefold molar excess of the immunogen provided by Chemicon (AG252) eliminated immunolabeling in brain sections.
Cell mapping and imaging
The computer-assisted mapping technique designed to map the location of tracer injection sites, BDA-labeled terminal fields and biotinamide-labeled neurons was described in detail previously (Neurolucida; MicroBrightField, Williston, VT; see Stornetta and Guyenet 1999). Section alignment between brains was done relative to a reference section as follows. Specific segments of the ventral respiratory column were identified by their location caudal to the caudal end of the facial motor neurons in each brain and brain levels were named according to the atlas of Paxinos and Watson (1998). On the basis of our previous work in adult rats of the same age and strain, the pre-Bötzinger region of the ventral respiratory column (for definition see Alheid et al. 2002; Feldman and Del Negro 2006) was judged to lie 900 ± 200 microns caudal to the facial motor nucleus [Bregma –12.5 mm after Paxinos and Watson (1998)] and the rostral ventral respiratory group to reside 1.3 mm or more caudal to facial nucleus (caudal to –12.9 mm in the Paxinos and Watson atlas;Stornetta et al. 2003a).
The Neurolucida files were exported to the Canvas 9 software drawing program (ACD Systems of America, Miami, FL) for final modifications. Photographs were taken with a 12-bit color CCD camera (CoolSnap, Roper Scientific, Tuscon, AZ; resolution 1,392 x 1,042 pixels). IPLab software (Scanalytics, Rockville, MD) was used for merging of color channels in photographs of dual-labeling experiments.
|
|
RESULTS |
|---|
|
NTS pump cells (n = 56) were identified as previously described by others (Ezure and Tanaka 2004; Ezure et al. 2002). These cells were phasically active and their discharge coincided with lung inflation as monitored by the tracheal pressure (Fig. 1, A1–A3). Their activity increased when end-expiratory pressure was raised (Fig. 1, A1, A2, and A5) and the cells became immediately silent when ventilation was interrupted in expiration (Fig. 1A1). Such cells could be found reliably only within a narrow (<300 µm) rostrocaudally oriented cell column centered 0.83 ± 0.04 mm lateral to the midline (n = 13 rats). All recordings were made in the NTS at area postrema level; therefore the full rostrocaudal extent of the cell column was not investigated. Although the pump cells that were recorded had very homogeneous properties with respect to our test criteria, we cannot state whether the sample is representative of the entire population of pump cells present in the NTS.
|
Thirty-six pump cells were tested for antidromic activation from the ipsilateral RTN and 12 could be antidromically activated from this region (latency 6.3 ± 0.18 ms; range: 5.4–7.5 ms). The antidromic nature of the evoked spikes was demonstrated with the collision test (Lipski 1981) (Fig. 2A). There was no statistical difference between the resting discharge rate of pump cells with or without axonal projection to RTN level (Fig. 2B). All the stimulation sites were identified histologically (Fig. 2C). These sites were centered ventromedial to the very caudal end of the facial motor nucleus region [1. 8 mm from the midline; –11.6 mm caudal to Bregma according to the atlas of Paxinos and Watson (1998)] that contains the highest density of chemoreceptors as we have defined them previously (Fig. 2D) (Mulkey et al. 2004; Stornetta et al. 2006).
|
; Patrickson et al. 1991). Seven of the nine recovered pump cells had virtually the same location (Fig. 3B). The labeled pump cell shown in Fig. 3A contained GAD67 mRNA (Fig. 3, C1 and C2). Two other examples of GAD67-positive NTS pump cells that were backfired from RTN are shown in Fig. 3, D1–E2. GAD67mRNA was detected in eight of the nine cells recovered (Fig. 3B).
|
The anterograde transport of biotinamide dextranamide (BDA) was used in an effort to confirm the existence of a GABAergic projection to RTN from the region of the NTS that contains the pump cells. BDA was injected by iontophoresis in four rats at the sites indicated in Fig. 4A. A representative injection site is shown in Fig. 4B. An abundant field of BDA-labeled varicosities was present throughout the ventrolateral medulla (Fig. 4, C and D) extending into the RTN region (Fig. 4, D–F). Synaptic terminals were less numerous in the RTN than in more caudal regions of the ventral respiratory column such as the region of the pre-Bötzinger complex and the rostral ventral respiratory group (rVRG) (Fig. 4G). Simultaneous histological detection of BDA and GAD revealed that a high proportion of the anterogradely labeled varicosities were GABAergic (Fig. 5, A–C). A random sampling of BDA-labeled varicosities located within RTN revealed that approximately 63% of the BDA-labeled terminals were immunoreactive for GAD (total of 55 varicosities sampled in four rats).
|
|
|
. This larger cell group contained scant GAD67 mRNA (Fig. 6C) and no detectable GLYT2 mRNA (not illustrated). To quantify the proportion of retrogradely labeled GAD67-positive neurons present in the lateral and medial clusters, the NTS was divided in two regions by a vertical line drawn 500 µm lateral to the midline. In the region of the NTS situated 0–500 µm from the midline only 10% of the CTB-labeled neurons contained GAD67 mRNA (five of 50 cells counted in three sections/rat in two rats). This proportion was 40% in the lateral portion of the NTS extending 500–1,000 µm from the midline (10 of 24 CTB-labeled cells counted). |
|
DISCUSSION |
|---|
|
). We also show that a subset of these pump cells innervate the RTN region and we conclude that this projection could be responsible for the previously described inhibitory effect of lung inflation on RTN chemoreceptors (Moreira et al. 2007). Inhibitory pump cells innervate RTN
The NTS neurons that we identified as pump cells displayed the same characteristics as the cells previously recorded in both cats and rats and their location generally conformed to prior descriptions (Bonham and McCrimmon 1990; Ezure and Tanaka 1996; Ezure et al. 2002). In the present study, evidence that these cells receive excitatory inputs from slowly adapting pulmonary afferents (SARs) was based on the following criteria: a discharge pattern that is strictly synchronized with lung inflation, a slowly adapting excitatory response to lung inflation, and the absence of a central respiratory activity. In addition, others have shown that these cells are driven at very short latencies by low-intensity vagus nerve stimulation, which suggests that their vagal input is from myelinated afferents and is probably monosynaptic (Ezure and Tanaka 1996; Ezure et al. 2002; Miyazaki et al. 1999). Finally, because our rats were paralyzed and artificially ventilated through a tracheal cannula, the pump cells that we recorded could not have been driven by laryngeal or pharyngeal mechanoreceptor inputs sensitive to air flow or to changes in upper airway diameter.
A minor debate persists as to the exact location of the NTS pump cells (Kubin et al. 2006). Depending on the study, these neurons have been described as residing predominantly lateral to the tractus solitarius (Ezure et al. 2002), within the interstitial subnucleus (Ezure and Tanaka 2004), or medial to the tractus solitarius (Bonham and McCrimmon 1990). These apparent discrepancies may be explained by the fact that the tractus solitarius is not a compact fiber tract but a collection of bundles of differing size and geometry with an imprecise ventrolateral border made of smaller fiber tracts. This fact can be readily appreciated when nondelipidated brain tissue is observed under dark-field illumination (see Fig. 3A). The interstitial subnucleus of the NTS is usually defined as a sizable and roughly circular myelin-poor region located amid the dorsomedial fascicles of the tractus solitarius (Altschuler et al. 1989) (Fig. 3A). This zone and the immediately overlying neuropil have been described as receiving a very dense input from laryngeal and pharyngeal afferents in rats (Altschuler et al. 1989; Furusawa et al. 1996; Patrickson et al. 1991). The pump cells that we labeled (eight of nine) were located at the periphery of this circular region (in all directions except dorsal) but never in it. If one ignores the smaller outer bundles of the tractus solitarius, one could easily conclude that some of the pump cells reside ventrolateral to the solitary tract, as suggested by Ezure and colleagues in their earlier work (Ezure and Tanaka 1996; Miyazaki et al. 1998). Our electrophysiological sampling of 56 pump cells suggests that these cells reside within a longitudinal column that is about 300 microns wide in the rat. This lateral spread clearly exceeds the width of the interstitial nucleus narrowly defined as the small myelin-poor circular zone alluded to earlier. Based on this observation and the anatomical location of the biotinamide labeled neurons, our results suggest that the cell bodies of the pump cells reside at the immediate periphery of the interstitial nucleus. Some of these cells are located medial to the tractus solitarius as suggested by Bonham and McCrimmon (1990). In their latest work, Ezure and Tanaka (2004) describe the pump cells as residing in "the interstitial subnucleus of the solitary tract nucleus and its immediate vicinity." This definition is in reasonable agreement with the present finding if one defines the interstitial nucleus more broadly as the neuropil that is interspersed with the myelinated bundles of the tractus solitarius.
Collectively and individually, NTS pump cells innervate large portions of the ventral respiratory column as well as selected dorsal pontine regions that are also involved in respiratory pattern generation (Ezure and Tanaka 1996; Ezure et al. 2002). The present study suggests that RTN is among the structures that receive a direct input from these pump cells. This conclusion derives first and foremost from the fact that we could antidromically activate about 30% of the pump cells by microstimulation within RTN. The relatively long latency of the antidromic spikes (5.4–7.5 ms) suggests that this projection probably consists of unmyelinated or lightly myelinated axons as is the case of other pump cells according to Ezure and Tanaka (1996). The long antidromic latencies that we observed may also denote the fact that we stimulated fine axonal collaterals rather than the main axon of the pump cells. This interpretation is supported by the fact that the minimum latency of activation of these cells from the dorsal pons is 1.9 to 4.5 ms (Ezure et al. 2002), which is slightly less than from RTN. Nonetheless, we were unable to elicit antidromic spikes at multiple discrete latencies from the RTN stimulation sites; therefore our electrophysiological approach alone did not provide definite evidence that the pump cells that we recorded had a terminal field in RTN rather than just an axon of passage. Accordingly, we sought additional anatomical evidence of pump cell projection to RTN. The anterograde and retrograde tracing experiments provided supportive evidence by showing that GABAergic neurons located in the region of the NTS that contains the pump cells do innervate RTN.
The present study confirms and extends prior evidence that a large majority of NTS pump cells in the rat are GABAergic and that a small fraction of these cells may also release glycine (Ezure and Tanaka 2004). In our case, eight of nine pump cells were clearly positive for GAD67 (89%) versus 10/14 (71%) in Ezure and Tanaka's report (2004). The difference could be explained by slight variations in histological protocol or by the fact that we sampled only pump cells with axonal projection to RTN, whereas Ezure and Tanaka labeled generic pump cells without regard to projection pattern.
If the NTS pump cells are inhibitory, how does the Breuer–Hering reflex work?
NTS pump cells mediate the Breuer–Hering reflex prolongation of expiration (Bonham and McCrimmon 1990; Bonham et al. 1993; Hayashi et al. 1996). The exact neuronal targets of the NTS pump cells have been difficult to determine because most ventral respiratory column (VRC) neurons are influenced by lung stretch afferent stimulation arising from the complex interactions that exist within the CPG network (Krolo et al. 2005). Based on the assumption that the most likely targets of the pump cells are neurons in which single-shock stimulation of the vagus nerve elicits the shortest-latency postsynaptic currents (PSCs), this reflex is currently thought to be mediated by the disynaptic activation of expiratory decremental (E-DEC, also known as postinspiratory) medullary interneurons (Hayashi et al. 1996; Kubin et al. 2006). The theory is compatible with the projection pattern of NTS pump cells and is consistent with the fact that E-DEC interneurons inhibit inspiratory and other neurons (Richter et al. 1987) by releasing glycine (Ezure et al. 2003) and/or 
-aminobutyric acid (GABA) (Okazaki et al. 2001). Also in accordance with the theory, the latency of the excitatory PSCs recorded in E-DEC neurons is among the shortest of any VRC neuron and this latency is shorter than that of the inhibitory postsynaptic currents recorded in other VRC neurons (Hayashi et al. 1996; Kubin et al. 2006). The weakest link of this theory is that it postulates the existence of excitatory pump cells for which no anatomical evidence exists yet. However, this possibility is not ruled out by the present study nor by that of Ezure and Tanaka (2004) because neither of these studies found that 100% of the pump cells are GABAergic. Furthermore, the hypothetical excitatory pump cells may reside at some distance from the region that was preferentially explored by Ezure and Tanaka (2004) and by our research group.
Physiological role of the GABAergic pump cell input to RTN
The existence of a direct projection from GABAergic NTS pump cells to RTN chemoreceptors accounts for the fact that many of these cells are inhibited by lung inflation (Moreira et al. 2007). Indeed, the inhibitory effect of lung inflation on RTN neurons is eliminated by silencing neurons located in the NTS region that contains the GABAergic pump cells (Moreira et al. 2007) and many RTN neurons are phasically inhibited during each lung inflation, as would be expected if they received a periodic inhibitory input from the type of pump cells identified in the present study (Moreira et al. 2007). Finally, the inhibitory effect of lung inflation on RTN neurons is unaffected by silencing neurons within the rVRG on both sides using the GABAA agonist muscimol (Moreira et al. 2007). This procedure is likely to silence the CPG altogether and, at the very least, its ineffectiveness reduces the plausibility that the pathway between pump cells and RTN neurons involves an interneuron located within the ventrolateral medulla.
As mentioned earlier, SAR activation reduces the duration of the inspiratory phase and prolongs expiration by activating E-DEC inhibitory neurons (Hayashi et al. 1996). In theory, the central apnea caused by higher levels of lung inflation could simply result from a more intense activation of the E-DEC neurons by the hypothetical excitatory pump cells located in the NTS. However, present and prior results suggest that SAR activation may also reduce the frequency and intensity of the respiratory pattern generator by suppressing the excitatory drive that originates from RTN chemoreceptors (Moreira et al. 2007). This second mechanism is almost certainly mediated by NTS pump cells that are inhibitory.
Chemoreceptor stimulation and lung inflation usually exert opposing effects on the activity of the respiratory network (Coleridge and Coleridge 2001; Hayashi et al. 1996; Kubin et al. 2006; Vatner and Uemura 2001). The effects of chemoreceptors and lung inflation are described as additive or as interacting in more complex ways depending on the species, the anesthetic, and the type of chemoreceptors (central, peripheral, or both) that were stimulated (Bajic et al. 1994; Kubin et al. 2006; Mitchell and Selby 1988; Mitchell et al. 1982). The present study suggests that the interaction between the chemical drive of respiration and its control by lung stretch afferent occurs, at least in part, at the RTN level. Because RTN neurons are central chemoreceptors that also receive strong excitatory inputs from the carotid body (Takakura et al. 2006), RTN should also be viewed as a site of interaction between lung stretch afferent input and both central and peripheral chemoreception.
|
|
GRANTS |
|---|
|
|
|
FOOTNOTES |
|---|
The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
Address for reprint requests and other correspondence: P. G. Guyenet, Department of Pharmacology, University of Virginia Health System, P.O. Box 800735, 1300 Jefferson Park Avenue, Charlottesville, VA 22908-0735 (E-mail: pgg{at}virginia.edu)
|
|
REFERENCES |
|---|
|
Alheid GF, Gray PA, Jiang MC, Feldman JL, McCrimmon DR. Parvalbumin in respiratory neurons of the ventrolateral medulla of the adult rat. J Neurocytol 31: 693–717, 2002.[CrossRef][Web of Science][Medline]
Altschuler SM, Bao X, Bieger D, Hopkins DA, Miselis RR. Viscerotopic representation of the upper alimentary tract in the rat: sensory ganglia and nuclei of the solitary and spinal trigeminal tracts. J Comp Neurol 283: 248–268, 1989.[CrossRef][Web of Science][Medline]
Bajic J, Zuperku EJ, Tonkovic-Capin M, Hopp FA. Interaction between chemoreceptor and stretch receptor inputs at medullary respiratory neurons. Am J Physiol Regul Integr Comp Physiol 266: R1951–R1961, 1994.
Bonham AC, Coles SK, McCrimmon DR. Pulmonary stretch receptor afferents activate excitatory amino acid receptors in the nucleus-tractus-solitarii in rats. J Physiol 464: 725–746, 1993.
Bonham AC, McCrimmon DR. Neurones in a discrete region of the nucleus tractus solitarius are required for the Breuer–Hering reflex in rat. J Physiol 427: 261–280, 1990.
Coleridge HM, Coleridge JC. Afferent innervation of lungs, airways, and pulmonary artery. In: Reflex Control of the Circulation, edited by Zucker IH, Gilmore JP. Boca Raton, FL: CRC Press, 2001, p. 579–607.
Cream C, Li A, Nattie E. The retrotrapezoid nucleus (RTN): local cytoarchitecture and afferent connections. Respir Physiol Neurobiol 130: 121–137, 2002.[CrossRef][Web of Science][Medline]
Ezure K, Tanaka I. Pump neurons of the nucleus of the solitary tract project widely to the medulla. Neurosci Lett 215: 123–126, 1996.[CrossRef][Web of Science][Medline]
Ezure K, Tanaka I. GABA, in some cases together with glycine, is used as the inhibitory transmitter by pump cells in the Hering–Breuer reflex pathway of the rat. Neuroscience 127: 409–417, 2004.[CrossRef][Web of Science][Medline]
Ezure K, Tanaka I, Kondo M. Glycine is used as a transmitter by decrementing expiratory neurons of the ventrolateral medulla in the rat. J Neurosci 23: 8941–8948, 2003.
Ezure K, Tanaka I, Saito Y, Otake K. Axonal projections of pulmonary slowly adapting receptor relay neurons in the rat. J Comp Neurol 446: 81–94, 2002.[CrossRef][Web of Science][Medline]
Feldman JL, Del Negro CA. Looking for inspiration: new perspectives on respiratory rhythm. Nat Rev Neurosci 7: 232–242, 2006.[CrossRef][Web of Science][Medline]
Feldman JL, Mitchell GS, Nattie EE. Breathing: rhythmicity, plasticity, chemosensitivity. Annu Rev Neurosci 26: 239–266, 2003.[CrossRef][Web of Science][Medline]
Furusawa K, Yasuda K, Okuda D, Tanaka M, Yamaoka M. Central distribution and peripheral functional properties of afferent and efferent components of the superior laryngeal nerve: morphological and electrophysiological studies in the rat. J Comp Neurol 375: 147–156, 1996.[CrossRef][Web of Science][Medline]
Guyenet PG, Mulkey DK, Stornetta RL, Bayliss DA. Regulation of ventral surface chemoreceptors by the central respiratory pattern generator. J Neurosci 25: 8938–8947, 2005.
Hayashi F, Coles SK, McCrimmon DR. Respiratory neurons mediating the Breuer–Hering reflex prolongation of expiration in rat. J Neurosci 16: 6526–6536, 1996.
Krolo M, Tonkovic-Capin V, Stucke AG, Stuth EA, Hopp FA, Dean C, Zuperku EJ. Subtype composition and responses of respiratory neurons in the pre-Bötzinger region to pulmonary afferent inputs in dogs. J Neurophysiol 93: 2674–2687, 2005.
Kubin L, Alheid GF, Zuperku EJ, McCrimmon DR. Central pathways of pulmonary and lower airway vagal afferents. J Appl Physiol 101: 618–627, 2006.
Li A, Nattie E. CO2 dialysis in one chemoreceptor site, the RTN: stimulus intensity and sensitivity in the awake rat. Respir Physiol Neurobiol 133: 11–22, 2002.[CrossRef][Web of Science][Medline]
Lipski J. Antidromic activation of neurones as an analytical tool in the study of the central nervous system. J Neurosci Methods 4: 1–32, 1981.[CrossRef][Web of Science][Medline]
Mitchell GS, Cross BA, Hiramoto T, Scheid P. Interactions between lung stretch and PaCO2 in modulating ventilatory activity in dogs. J Appl Physiol 53: 185–191, 1982.
Mitchell GS, Selby BD. Ventilatory responses to lung inflation and arterial CO2 in halothane-anesthetized dogs. J Appl Physiol 64: 1433–1438, 1988.
Miyazaki M, Arata A, Tanaka I, Ezure K. Activity of rat pump neurons is modulated with central respiratory rhythm. Neurosci Lett 249: 61–64, 1998.[CrossRef][Web of Science][Medline]
Miyazaki M, Tanaka I, Ezure K. Excitatory and inhibitory synaptic inputs shape the discharge pattern of pump neurons of the nucleus tractus solitarii in the rat. Exp Brain Res 129: 191–200, 1999.[CrossRef][Web of Science][Medline]
Moreira TS, Takakura AC, Colombari E, West GH, Guyenet PG. Inhibitory input from slowly-adapting lung stretch receptors to retrotrapezoid nucleus (RTN) chemoreceptors. J Physiol 580: 285–300, 2007.
Mulkey DK, Stornetta RL, Weston MC, Simmons JR, Parker A, Bayliss DA, Guyenet PG. Respiratory control by ventral surface chemoreceptor neurons in rats. Nat Neurosci 7: 1360–1369, 2004.[CrossRef][Web of Science][Medline]
Nattie E, Li A. Muscimol dialysis in the retrotrapezoid nucleus region inhibits breathing in the awake rat. J Appl Physiol 89: 153–162, 2000.
Nattie E, Shi J, Li A. Bicuculline dialysis in the retrotrapezoid nucleus (RTN) region stimulates breathing in the awake rat. Respir Physiol 124: 179–193, 2001.[CrossRef][Web of Science][Medline]
Okada Y, Chen Z, Jiang W, Kuwana S, Eldridge FL. Anatomical arrangement of hypercapnia-activated cells in the superficial ventral medulla of rats. J Appl Physiol 93: 427–439, 2002.
Okazaki M, Takeda R, Haji A, Yamazaki H. Glutamic acid decarboxylase-immunoreactivity of bulbar respiratory neurons identified by intracellular recording and labeling in rats. Brain Res 914: 34–47, 2001.[CrossRef][Web of Science][Medline]
Patrickson JW, Smith TE, Zhou SS. Afferent projections of the superior and recurrent laryngeal nerves. Brain Res 539: 169–174, 1991.[CrossRef][Web of Science][Medline]
Paxinos G, Watson C. The Rat Brain in Stereotaxic Coordinates. San Diego, CA: Academic Press, 1998.
Putnam RW, Filosa JA, Ritucci NA. Cellular mechanisms involved in CO(2) and acid signaling in chemosensitive neurons. Am J Physiol Cell Physiol 287: C1493–C1526, 2004.
Richter DW, Ballantyne D, Remmers JE. The differential organization of medullary post-inspiratory activities. Pfluegers Arch 410: 420–427, 1987.[CrossRef][Web of Science][Medline]
Rosin DL, Chang DA, Guyenet PG. Afferent and efferent connections of the rat retrotrapezoid nucleus. J Comp Neurol 499: 64–89, 2006.[CrossRef][Medline]
Stornetta RL, Guyenet PG. Distribution of glutamic acid decarboxylase mRNA-containing neurons in rat medulla projecting to thoracic spinal cord in relation to monoaminergic brainstem neurons. J Comp Neurol 407: 367–380, 1999.[CrossRef][Web of Science][Medline]
Stornetta RL, Moreira TS, Takakura AC, Kang BJ, Chang DA, West GH, Brunet JF, Mulkey DK, Bayliss DA, Guyenet PG. Selective expression of Phox2b by brainstem neurons involved in chemosensory integration in the adult rat. J Neurosci 26: 10305–10314, 2006.
Stornetta RL, Rosin DL, Wang H, Sevigny CP, Weston MC, Guyenet PG. A group of glutamatergic interneurons expressing high levels of both neurokinin-1 receptors and somatostatin identifies the region of the pre-Bötzinger complex. J Comp Neurol 455: 499–512, 2003a.[CrossRef][Web of Science][Medline]
Stornetta RL, Sevigny CP, Guyenet PG. Inspiratory augmenting bulbospinal neurons express both glutamatergic and enkephalinergic phenotypes. J Comp Neurol 455: 113–124, 2003b.[CrossRef][Web of Science][Medline]
Takakura AC, Moreira TS, Colombari E, West GH, Stornetta RL, Guyenet PG. Peripheral chemoreceptor inputs to retrotrapezoid nucleus (RTN) CO2-sensitive neurons in rats. J Physiol 572: 503–523, 2006.
Vatner SF, Uemura N. Integrative cardiovascular control by pulmonary inflation reflexes. In: Reflex Control of the Circulation, edited by Zucker IH, Gilmore JP. Boca Raton, FL: CRC Press, 2001, p. 609–626.
This article has been cited by other articles:
|
|
 |
|
  A. C. Takakura, T. S. Moreira, R. L. Stornetta, G. H. West, J. M. Gwilt, and P. G. Guyenet Selective lesion of retrotrapezoid Phox2b-expressing neurons raises the apnoeic threshold in rats J. Physiol., June 15, 2008; 586(12): 2975 - 2991. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
| Visit Other APS Journals Online |
TOP
ABSTRACT
INTRODUCTION
