Immunohistochemical characterization of nodose cough receptor neurons projecting to the trachea of guinea pigs
© Mazzone and McGovern; licensee BioMed Central Ltd. 2008
Received: 05 September 2008
Accepted: 19 October 2008
Published: 19 October 2008
Cough in guinea pigs is mediated in part by capsaicin-insensitive low threshold mechanoreceptors (cough receptors). Functional studies suggest that cough receptors represent a homogeneous population of nodose ganglia-derived sensory neurons. In the present study we set out to characterize the neurochemical profile of cough receptor neurons in the nodose ganglia.
Nodose neurons projecting to the guinea pig trachea were retrogradely labeled with fluorogold and processed immunohistochemically for the expression of a variety of transporters (Na+/K+/2C1- co-transporter (NKCC1), α1 and α3 Na+/K+ ATPase, vesicular glutamate transporters (vGlut)1 and vGlut2), neurotransmitters (substance P, calcitonin gene-related peptide (CGRP), somatostatin, neuronal nitric oxide synthase (nNOS)) and cytosolic proteins (neurofilament, calretinin, calbindin, parvalbumin).
Fluorogold labeled ~3 per cent of neurons in the nodose ganglia with an average somal perimeter of 137 ± 6.2 μm (range 90–200 μm). All traced neurons (and seemingly all nodose neurons) were immunoreactive for NKCC1. Many (> 90 per cent) were also immunoreactive for vGlut2 and neurofilament and between 50 and 85 per cent expressed α1 ATPase, α3 ATPase or vGlut1. Cough receptor neurons that did not express the above markers could not be differentiated based on somal size, with the exception of neurofilament negative neurons which were significantly smaller (P < 0.05). Less than 10 per cent of fluorogold labeled neurons expressed substance P or CGRP (and these had somal perimeters less than 110 μm) and none expressed somatostatin, calretinin, calbindin or parvalbumin. Two distinct patterns of nNOS labeling was observed in the general population of nodose neurons: most neurons contained cytosolic clusters of moderately intense immunoreactivity whereas less than 10 per cent of neurons displayed uniform intensely fluorescent somal labeling. Less than 3 per cent of the retrogradely traced neurons were intensely fluorescent for nNOS (most showed clusters of nNOS immunoreactivity) and nNOS immunoreactivity was not expressed by cough receptor nerve terminals in the tracheal wall.
These data provide further insights into the neurochemistry of nodose cough receptors and suggest that despite their high degree of functional homogeneity, nodose cough receptors subtypes may eventually be distinguished based on neurochemical profile.
Previous studies have identified a novel vagal sensory nerve subtype that innervates the large airways (larynx, trachea and main bronchi) of guinea pigs and is likely responsible for defensive cough in this species . These sensory neurons (referred to as cough receptors) are derived from the nodose ganglia and are characterized by their insensitivity to capsaicin and their sensitivity to both rapid reductions in pH and punctuate (touch-like) mechanical stimulation [1–3]. However, unlike other classically defined low threshold mechanoreceptors which innervate the airways and lungs, cough receptors display a low sensitivity to mechanical stretch (including inflation/deflation and bronchospasm), conduct action potentials slower (~5 m/sec for cough receptors compared to > 15 m/sec for intrapulmonary stretch receptors) and are unresponsive to the purinergic agonist α,β-methylene ATP . Based on these observations, cough receptors are believed to represent a distinct airway afferent nerve in this species (reviewed in ).
Functional and electrophysiological studies have provided key insights into the role of nodose cough receptors in the cough reflex. In anesthetized guinea pigs, punctuate mechanical stimulation or rapid acidification of the laryngeal or tracheal mucosa evokes coughing, a response that can be abolished by selectively disrupting the afferent pathways from the nodose ganglia [1, 5–7]. Extensive electrophysiological analyses of the activation profiles of nodose neurons projecting to the guinea pig trachea and larynx suggests that the majority (perhaps greater than 95%) of these neurons form a seemingly homogeneous population of neurons that display the functional characteristics of cough receptors [1, 2]. In the guinea pig trachea and larynx, there are very few nodose capsaicin-sensitive nociceptors (tracheal nociceptors are mostly derived from the jugular vagal ganglia) and no classically defined rapidly adapting or slowly adapting stretch receptors [1, 2].
Immunohistochemical studies of other sensory nerve populations have successfully used the expression of proton pump isozymes, vesicular glutamate transporters (vGluts; a marker for glutamatergic neurons), neuropeptides and calcium binding proteins (such as calretinin, calbindin and parvalbumin) as useful markers for characterizing sensory nerve subtypes. Therefore, in the present study we used well characterized antisera raised against these transporters, neurotransmitters and cytosolic proteins to further characterize the guinea pig cough receptor neurons in the nodose ganglia.
Experiments were approved by the Howard Florey Institute Animal Ethics Committee and conducted on male albino Hartley guinea pigs (200–350 g, n = 36, IVMS, South Australia) at the Howard Florey Institute (The University of Melbourne, Australia).
Guinea pigs (n = 32) were anesthetized with 1.8–2.2 per cent isoflurane in oxygen. The extrathoracic trachea was exposed via a ventral incision in the animal's neck. Using a 10 μl Hamilton glass microsyringe fitted with a 32 gauge needle, 10 μl of 4 per cent fluorogold (Fluorochrome LLC, Colorado, USA) was injected into the rostral extrathoracic tracheal lumen (on to the mucosal surface). Following injection, the wound was sutured and the animals were allowed to recover for 7 days at which time they were anesthetized with sodium pentobarbital (100 mg/kg i.p.) and transcardially perfused with 10 mM phosphate buffered saline (PBS) followed by 4% paraformaldehyde in PBS. The nodose ganglia was removed and placed in 4% paraformaldehyde at 4°C for 2 hours, then cyroprotected in 20% sucrose solution at 4°C overnight prior to immunohistochemical processing (see below).
Preparation of tracheal wholemounts
Wholemount preparations of guinea pig (n = 4) tracheal segments were prepared using a modification of previously described methods [6, 8]. Briefly, animals were deeply anesthetized with sodium pentobarbital (80 mg/kg i.p) and transcardially perfused with 500 mL of 10 mM phosphate buffered saline (PBS). The entire trachea was removed, cleaned of excess connective tissue, and opened longitudinally via a midline incision along the ventral surface. The epithelium was gently rubbed off the trachea with a cotton swab and tracheal segments (8–10 rings in length) were pinned flat onto a piece of cork board and placed in fixative (4% paraformaldehyde) for 2–3 hours at 4°C, and then transferred to blocking solution (10 mM PBS and 10% horse serum) for one hour prior to immunohistochemical staining (see below). Epithelial removal is necessary to visualize cough receptor nerve terminals in the guinea pig trachea which are confined to the extracellular matrix below the epithelium. This procedure would be expected to remove some tracheal nociceptors  but does not disrupt cough receptors [7, 9].
Immunohistochemistry and microscopy
Details of the primary and secondary antibodies used for immunohistochemical staining.
Primary Antibodies – Transporters
α1 Na+/K+ ATPase (clone 05–369)
α3 Na+/K+ ATPase (clone XVIF9-G10)
Biomol, PA, USA.
Gift Dr RJ Turner, National Institute of Dental and Craniofacial Research, USA.
vGLUT1 (catalogue# 135 302)
Synaptic Systems Goettingen, Germany.
vGLUT2 (catalogue# 135 402)
Synaptic Systems Goettingen, Germany.
Primary Antibodies – Neurotransmitters
CGRP (catalogue# RPN 1842)
Neuronal nitric oxide synthase (nNOS)
Gift Dr Colin Anderson, University of Melbourne, Australia.
Somatostatin (catalogue# AB5494)
Substance P (clone NC1)
Primary Antibodies – Cytosolic Proteins
Calbindin D28k (number CB-38A)
Swant Bellinzona, Switzerland.
Calretinin (number 7699/4)
Swant Bellinzona, Switzerland.
Neurofilament 160KD (clone NN18)
Parvalbumin (number 235)
Swant Bellinzona, Switzerland.
Secondary Antibodies (IgG, H+L, 2 mg/ml)
AlexaFluor 488 or 594 anti-goat
Molecular Probes Eugene, OR, USA
AlexaFluor 488 or 594 anti-mouse
Molecular Probes Eugene, OR, US
AlexaFluor 488 or 594 anti-rabbit
Molecular Probes Eugene, OR, USA
AlexaFluor 488 anti-rat
Molecular Probes Eugene, OR, USA
Immunohistochemical processing of tracheal wholemounts was performed using a modification of the methods described above for nodose sections. Tissues were first pinned flat to a sylgard-filled tissue culture dish and incubated for 1 hour in blocking solution (10% normal horse serum in 10 mM PBS) and then overnight (at 37°C) in 10 mM PBS/0.3% Triton X-100/2% horse serum containing the primary antisera of interest (refer Table 1). After washing thoroughly with 10 mM PBS (for at least 3 hours), wholemounts were then incubated for 1 hour at room temperature in the appropriate AlexaFluor-conjugated secondary antibody (refer Table 1).
Labeling of wholemounts and slide mounted sections was visualized using an Olympus BX51 fluorescent microscope equipped with appropriate filters and an Optronics digital camera. Low and high magnification images were captured and stored digitally for subsequent off-line analysis of somal size (see below) and preparation of representative photomicrographs. Negative control experiments, in which the primary antisera were excluded, were carried out where necessary.
Cell counts in a given field of view were performed either online (during microscopy) or offline (using high resolution digital images) at 100–200× magnification. 3–10 representative replicate sections were assessed per animal and a minimum of 4 animals were analyzed per group. For somal size analysis, stored images were imported into ImageJ software (NIH, USA http://rsb.info.nih.gov/ij/) and cell edges were traced on screen using a calibrated scale tool. Only cells with a distinct nuclear region were measured in order to increase the likelihood that perimeters were measured close to the middle of the neuron and therefore accurately reflected the true somal size. A minimum of 100 labeled cells, taken from at least 3 different animals, were used to estimate somal sizes for each marker. Data are expressed as a mean ± SEM. Differences between group data are compared using a Student's t-test and significance was set at P < 0.05.
Fluorogold retrograde labeling
Immunohistochemical expression of transporter proteins in nodose ganglia
Mean cell sizes of guinea pig nodose neurons.
Average Cell Perimeter (μm)
Average Cell Perimeter (μm)
113.9 ± 3.1*
99.1 ± 3.7*
108.3 ± 5.9*
α1 Na+/K+ ATPase
109.3 ± 4.8*
131.3 ± 8.4
90.0 ± 2.6*
α3 Na+/K+ ATPase
139.1 ± 1.8
128.5 ± 5.1
80.4 ± 2.2*
135.7 ± 2.6
126.8 ± 4.7
98.3 ± 2.6*
109.5 ± 8.9*
109.6 ± 3.0*
138.6 ± 1.8
177.9 ± 2.6*
142.1 ± 5.7
105.3 ± 2.4*
173.3 ± 2.1*
112.6 ± 2.9*
126.8 ± 5.9
Immunohistochemical expression of neurotransmitters in nodose ganglia
Immunohistochemical expression of cytosolic proteins in nodose ganglia
Approximately 90 per cent of the neurons retrogradely labeled with fluorogold expressed neurofilament (Fig 3 and Fig 9A). By contrast there were no fluorogold-positive neurons that exhibited either calretinin or calbindin (or parvalbumin) immunoreactivity (Fig 3 and Fig 9B–D). The population (approximately 10 per cent) of fluorogold-positive neurofilament negative neurons were significantly (P < 0.05) smaller in size compared to the traced neurons that were neurofilament-positive (Table 2).
In the present study we investigated the expression of a variety of neurochemical markers in cough receptor neurons in the nodose ganglia. Retrograde neuronal tracing from the airways confirmed previous studies showing that the majority of nodose neurons projecting to the trachea have medium somal sizes and express neurofilament, a marker for myelinated neurons [2, 11, 12]. The minor population of small sized neurons that were retrogradely labeled did not express neurofilament, but rather stained positively for neuropeptides such as substance P or CGRP. All traced neurons in the nodose ganglia expressed the Na+/K+/2Cl- co-transporter, NKCC1. By contrast, although many medium sized traced neurons (cough receptor neurons) expressed α1 or α3 Na+/K+ ATPase, vGlut1 or vGlut2, none of these markers were universally expressed by all cough receptor cells. Most neurons in the nodose ganglia displayed detectable levels of nNOS immunoreactivity. However, intense immunolabeling for nNOS was not characteristic of cough receptor neurons and nNOS was not observed in cough receptor nerve terminals in the tracheal wall. Furthermore, cough receptors did not express somatostatin, calretinin, calbindin or parvalbumin. These data provide a detailed immunohistochemical characterization of guinea pig cough receptor neurons in the nodose ganglia. Furthermore our data suggest that, despite the evidence suggesting homogeneity in their peripheral physiology, it is likely that variations exist in the neurochemical profile of some cough receptors.
Characterization of cough receptors in guinea pigs
Previous studies have characterized a novel airway afferent nerve subtype in guinea pigs that appears to be essential for defensive cough in this species [[1, 7], reviewed in ]. These cough receptors represent a subset of mechanically sensitive afferent nerves innervating the extrapulmonary airways. This distribution (at least in guinea pigs) is in contrast to the terminal location of the classically defined rapidly and slowly adapting receptors (RARs and SARs) which are mainly confined to the intrapulmonary airways and lungs. Cough receptors also display very distinct activation profiles and electrophysiological properties compared to RARs and SARs . Cough receptors are readily differentiated from bronchopulmonary C-fibers by their lack of sensitivity to capsaicin and bradykinin, faster conduction velocity and lack of expression of substance P and TRPV1 [1, 2, 13], and from the vagal afferents that innervate neuroepithelial bodies (NEBs) by their terminal locations (sub-epithelial, rather than associated with specialized epithelial cells, and exclusively extrapulmonary) . Guinea pigs also have reportedly very few NEBs .
The available electrophysiological data suggests that almost all of the nodose neurons projecting to the guinea pig trachea display activation profiles that classify them as cough receptors [1–3, 13, 16–18]. Few capsaicin-sensitive airway afferents arising from the nodose ganglia innervate the guinea pig trachea (most originate from the jugular ganglia) and in guinea pigs the mechanically-sensitive nodose nerve endings in the trachea don't display the characteristics of RARs or SARs (although other species such as dogs and rabbits possess RARs and/or SARs in the trachea) [1, 2, 19, 20]. There is also no evidence to suggest that individual cough receptors vary significantly in their basic physiology. The available anatomical data would also support the assertion that nodose neurons innervating the trachea are relatively homogenous. These observations make guinea pigs an ideal species for characterizing tracheal cough receptors. Thus, retrograde labeling of tracheal afferent nerves in the guinea pig nodose ganglia reveals a major population (95–99%) of medium to large sized neurons that do not express substance P or TRPV1 (markers used to define capsaicin-sensitive nociceptors) but do express neurofilament proteins (a marker for myelinated axons) [2, 11, 12]. These neurons are presumably the cough receptor neurons that have been identified functionally. The minor population of nodose neurons that project to the trachea (~5%) show the characteristics of small, unmyelinated nociceptors. Our data are consistent with these observations.
The results of the present study, however, provide some evidence that not all cough receptor neurons are identical. Not all neurons in the nodose ganglia that were retrogradely labeled from the trachea expressed α1 or α3 Na+/K+ ATPase, vGlut1 or vGlut2. Unlike the neurofilament negative traced neurons which displayed a significantly smaller somal size compared to the neurofilament positive traced neurons (suggesting that they are small diameter nociceptors), the average somal sizes of the retrogradely labeled neurons that were negative for α1 and α3 Na+/K+ ATPase, vGlut1 and vGlut2 immunoreactivity were not significantly different to the size of cough receptor neurons. This would suggest that some cough receptors likely differ in their expression of certain neuronal markers. A similar observation has been made with respect to the myelinated vagal afferent nerves that innervate pulmonary NEBs in rats (two myelinated vagal afferent types can be differentiated by the expression of α3 Na+/K+ ATPase, and P2X3 receptors) . Whether clearly definable and distinct subsets of cough receptor neurons can be differentiated based on these, or other neuronal markers, awaits additional analyses.
NKCC1 and Na+/K+ ATPase expression by cough receptors
The results from the present study confirm our previous data which showed that nodose neurons and cough receptor nerve terminals in the airways express NKCC1 , although NKCC1 is not a specific marker for cough receptor neurons as seemingly all neurons in the nodose ganglia showed NKCC1 labeling. NKCC1 functions to accumulate intracellular chloride ions above the electrochemical equilibrium in somatic (and likely vagal) sensory neurons [6, 22, 23], allowing a depolarizing chloride current to contribute to the regulation of afferent activity. Depolarizing chloride currents in sensory neurons are particularly critical to GABA-evoked primary afferent depolarization and inhibition of neurotransmitter release from the central projections of somatic and vagal sensory terminals in the spinal cord and brainstem (reviewed in ). More recently, NKCC1-mediated chloride uptake has also been suggested to be an important mechanism regulating the peripheral excitability of sensory neurons [6, 23, 25] and may represent a useful peripheral target for suppressing afferent nerve excitability [6, 23, 26].
Unlike NKCC1 expression, populations of sensory neurons may be differentiated based on the sodium pump isozyme which they express. For example, previous studies have revealed that in the dorsal root ganglia (DRG) α1 Na+/K+ ATPase is commonly expressed across many DRG neurons of varying somal sizes whilst α3 Na+/K+ ATPase is apparently restricted to medium and large sized neurons that presumably give rise to mechanically sensitive afferent fibers [27, 28]. This is in agreement with our data in the nodose ganglia showing that α3 Na+/K+ ATPase, but not α1 Na+/K+ ATPase, immunolabeling is restricted to medium and large sized neurons. In the airways, α3 Na+/K+ ATPase is not expressed in substance P-containing C-fibers (Fig 1 of present study; ) but is present in the peripheral terminals of SARs in rabbits and rats [21, 29], myelinated afferent fibers associated with NEBs in rats  and cough receptor terminals in guinea pigs (Figs 1 and 7 of present study; [6, 30]). Moreover, few neurons in the jugular ganglia that project to the trachea (exclusively C- and Aδ-fiber nociceptors; [1, 2]) express α3 Na+/K+ ATPase (S.B. Mazzone and A.E. McGovern, unpublished data). There have been no previous reports of α1 Na+/K+ ATPase expression in vagal afferent nerves, but our data suggest that more than half of the cough receptors in the nodose ganglia express detectable levels of α1 Na+/K+ ATPase immunoreactivity. It is not known what proportion of cough receptors express one versus both isozymes of Na+/K+ ATPase.
The apparent selective expression of α1 or α3 containing isozymes of Na+/K+ ATPase in different sensory neurons raises the question of what specific contribution the various sodium pump isozymes have on sensory neuron function. The more ubiquitous expression of α1 Na+/K+ ATPase across neurons of all sizes may suggest that pump isozymes containing this subunit play more of a housekeeping role in regulating sensory neuron Na+ and K+ gradients . By contrast, some evidence suggests that α3 Na+/K+ ATPase may be specialized to mechanoreceptors. Indeed, the Na+/K+ ATPase inhibitor ouabain, at doses that are reportedly selective for inhibiting the α3 subunit, inhibits cough receptor activation and coughing evoked by citric acid, mechanical stimulation or electrical stimulation of the tracheal mucosa in anesthetized guinea pigs while having no effect on C-fiber dependent reflexes evoked from the trachea . This effect may be due to unique kinetic properties of the α3 Na+/K+ ATPase isozyme that facilitate high frequency action potential conduction along the cough receptor axons . Experiments investigating the high frequency firing properties of rat calyx of Held nerve terminals also support this assertion . However, it is not known if the population of nodose mechanoreceptors that lack α3 Na+/K+ ATPase (present study) display different physiological attributes. It is also worth adding that Na+/K+ ATPase activity may be intrinsically linked to NKCC1 in cough receptors. NKCC1 would presumably elevate intracellular sodium ion concentrations (in addition to chloride) thereby facilitating Na+/K+ ATPase activity.
Cough receptor neurotransmitters
Although a rigorous analysis of the neurotransmitters expressed by cough receptors was not conducted, several points are worthy of note. The expression of vGlut1 and/or vGlut2 by cough receptor neurons would suggest that cough receptors utilize glutamate as a key neurotransmitter. The vGluts are a family of transporters that are responsible for packaging glutamate into synaptic vesicles, and a substantial body of evidence shows that these proteins are reliable markers for glutamatergic neurons (e.g., ). Consistent with this, cough receptor evoked coughing in rabbits and guinea pigs is inhibited by selective ionotropic glutamate receptor antagonists injected in to the nucleus of the solitary tract [34, 35].
In line with previous studies, our experiments have also failed to identify any neuropeptides in healthy cough receptor neurons. Studies to date have shown convincingly that cough receptors do not normally express substance P, CGRP, somatostatin or dynorphin [present study; [1, 11, 12, 36]]. However, this is not to say that cough receptors don't contain a yet to be identified neuropeptide, as many neurons utilize neuropeptides as co-transmitters in the mammalian nervous system. The present data also suggest that cough receptor neurons may not use nitric oxide as a neurotransmitter. However, the results from these experiments are more difficult to interpret. In our hands, nNOS immunoreactivity in nodose ganglia neurons showed two distinct patterns: many cells possessed dense nNOS immunoreactive clusters, whereas substantially fewer cells displayed uniform intense somal labeling. Importantly, very few nodose neurons that projected to the airways showed intense uniform immunoreactivity for nNOS (those that did tended to be smaller cells, probably nodose nociceptors). Similar results have been previously reported in guinea pigs . Moreover, nNOS was not present in cough receptor terminals, but was expressed by fine varicose fibers, in the guinea pig trachea. Nevertheless, given that nNOS expression was not assessed in the central terminals of cough receptors we cannot conclude definitively that NOS is not a neurotransmitter of cough receptors. The exact nature of the clustered nNOS immunoreactivity is also unclear at present. It may represent labeling of non-neuronal structures (e.g., glia associated with nodose neurons), labeling of nNOS in specific cellular compartments that have no relation to neurotransmission, or non-specific labeling produced by the antisera. It is, however, interesting that nitric oxide has been shown to mediate inter-somal transmission in the guinea pig nodose ganglia, suggesting the existence of a specific source of NOS for generating releasable nitric oxide .
Absence of calcium binding proteins in cough receptors
A final interesting observation to arise from the present studies was the distinct lack of expression of calretinin, calbindin or parvalbumin in nodose cough receptor neurons. This is in contrast to studies of other vagal mechanosensitive and/or myelinated afferent nerve populations. In rats, the vagal afferents that innervate NEBs are defined in part by their expression of calbindin . Furthermore, pulmonary smooth muscle associated receptors (likely SARs) and laryngeal mechanosensors express calretinin in rats [21, 40, 41], and parvalbumin and calbindin have been shown to be reliable markers for a variety of other vagal (or other visceral) mechanosensory neurons [42–44]. In our studies, calretinin and calbindin only labeled a subset of very large neurons in the nodose, whereas parvalbumin failed to label any structures in the nodose ganglia (despite labeling neurons and fibers in the guinea pig brainstem, not shown). Whether the large neurons immunoreactive for calbindin and calretinin consist in part of mechanoreceptive neurons that project to the intrapulmonary airways (i.e., RARs and/or SARs) is unknown, but may reveal an alternative method for differentiating cough receptors from other airway mechanosensors.
In summary, the present data provide further characterization of the nodose neurons that innervate the guinea pig trachea and suggest that immunohistochemically distinct subtypes of presumed cough receptors likely exist.
List of abbreviations
Calcitonin Gene-Related Peptide
Dorsal Root Ganglia
Intensely Fluorescent Cells
Na+/K+/2Cl- Co-transporter 1
neuronal Nitric Oxide Synthase
Rapidly Adapting Receptor
Slowly Adapting Receptor
Transient Receptor Potential Vanilloid 1
vesicular Glutamate transporter
This research was funded by the National Health and Medical Research Council (NH&MRC) of Australia (grant numbers 350333, 454776).
- Canning BJ, Mazzone SB, Meeker SN, Mori N, Reynolds SM, Undem BJ: Identification of the tracheal and laryngeal afferent neurones mediating cough in anaesthetized guinea-pigs. J Physiol. 2004, 557 (Pt 2): 543-58. 10.1113/jphysiol.2003.057885.PubMed CentralView ArticlePubMedGoogle Scholar
- Riccio MM, Kummer W, Biglari B, Myers AC, Undem BJ: Interganglionic segregation of distinct vagal afferent fibre phenotypes in guinea-pig airways. J Physiol. 1996, 496 (Pt 2): 521-30.View ArticleGoogle Scholar
- Kollarik M, Undem BJ: Mechanisms of acid-induced activation of airway afferent nerve fibres in guinea-pig. J Physiol. 2002, 543 (Pt 2): 591-600. 10.1113/jphysiol.2002.022848.PubMed CentralView ArticlePubMedGoogle Scholar
- Mazzone SB: An overview of the sensory receptors regulating cough. Cough. 2005, 1: 2-10.1186/1745-9974-1-2.PubMed CentralView ArticlePubMedGoogle Scholar
- Mazzone SB, Mori N, Canning BJ: Synergistic interactions between airway afferent nerve subtypes regulating the cough reflex in guinea-pigs. J Physiol. 2005, 569 (Pt 2): 559-73. 10.1113/jphysiol.2005.093153.PubMed CentralView ArticlePubMedGoogle Scholar
- Mazzone SB, McGovern AE: Na+-K+-2Cl- cotransporters and Cl- channels regulate citric acid cough in guinea pigs. J Appl Physiol. 2006, 101 (2): 635-43. 10.1152/japplphysiol.00106.2006.View ArticlePubMedGoogle Scholar
- Canning BJ, Farmer DG, Mori N: Mechanistic studies of acid-evoked coughing in anesthetized guinea pigs. Am J Physiol Regul Integr Comp Physiol. 2006, 291 (2): R454-63.View ArticlePubMedGoogle Scholar
- Baluk P, Gabella G: Afferent nerve endings in the tracheal muscle of guinea-pigs and rats. Anat Embryol (Berl). 1991, 183 (1): 81-7.View ArticleGoogle Scholar
- Mazzone SB, Mori N, Burman M, Palovich M, Belmonte KE, Canning BJ: Fluorescent styryl dyes FM1-43 and FM2-10 are muscarinic receptor antagonists: intravital visualization of receptor occupancy. J Physiol. 2006, 575 (Pt 1): 23-35. 10.1113/jphysiol.2006.106351.PubMed CentralView ArticlePubMedGoogle Scholar
- Lawson SN, Perry MJ, Prabhakar E, McCarthy PW: Primary sensory neurones: neurofilament, neuropeptides, and conduction velocity. Brain Res Bull. 1993, 30 (3–4): 239-43. 10.1016/0361-9230(93)90250-F.View ArticlePubMedGoogle Scholar
- Mazzone SB, Canning BJ: Synergistic interactions between airway afferent nerve subtypes mediating reflex bronchospasm in guinea pigs. Am J Physiol Regul Integr Comp Physiol. 2002, 283 (1): R86-98.View ArticlePubMedGoogle Scholar
- Kummer W, Fischer A, Kurkowski R, Heym C: The sensory and sympathetic innervation of guinea-pig lung and trachea as studied by retrograde neuronal tracing and double-labelling immunohistochemistry. Neuroscience. 1992, 49 (3): 715-37. 10.1016/0306-4522(92)90239-X.View ArticlePubMedGoogle Scholar
- McAlexander MA, Myers AC, Undem BJ: Adaptation of guinea-pig vagal airway afferent neurones to mechanical stimulation. J Physiol. 1999, 521 (Pt 1): 239-47. 10.1111/j.1469-7793.1999.00239.x.PubMed CentralView ArticlePubMedGoogle Scholar
- Van Genechten J, Brouns I, Burnstock G, Timmermans JP, Adriaensen D: Quantification of neuroepithelial bodies and their innervation in fawn-hooded and Wistar rat lungs. Am J Respir Cell Mol Biol. 2004, 30 (1): 20-30. 10.1165/rcmb.2003-0097OC.View ArticlePubMedGoogle Scholar
- Van Lommel A, Lauweryns JM: Postnatal development of the pulmonary neuroepithelial bodies in various animal species. J Auton Nerv Syst. 1997, 65 (1): 17-24. 10.1016/S0165-1838(97)00030-1.View ArticlePubMedGoogle Scholar
- Pedersen KE, Meeker SN, Riccio MM, Undem BJ: Selective stimulation of jugular ganglion afferent neurons in guinea pig airways by hypertonic saline. J Appl Physiol. 1998, 84 (2): 499-506.PubMedGoogle Scholar
- Kajekar R, Proud D, Myers AC, Meeker SN, Undem BJ: Characterization of vagal afferent subtypes stimulated by bradykinin in guinea pig trachea. J Pharmacol Exp Ther. 1999, 289 (2): 682-7.PubMedGoogle Scholar
- Undem BJ, Oh EJ, Lancaster E, Weinreich D: Effect of extracellular calcium on excitability of guinea pig airway vagal afferent nerves. J Neurophysiol. 2003, 89 (3): 1196-204. 10.1152/jn.00553.2002.View ArticlePubMedGoogle Scholar
- Bergren DR, Sampson SR: Characterization of intrapulmonary, rapidly adapting receptors of guinea pigs. Respir Physiol. 1982, 47 (1): 83-95. 10.1016/0034-5687(82)90094-9.View ArticlePubMedGoogle Scholar
- Keller E, Kohl J, Koller EA: Location of pulmonary stretch receptors in the guinea-pig. Respir Physiol. 1989, 76 (2): 149-57. 10.1016/0034-5687(89)90093-5.View ArticlePubMedGoogle Scholar
- Brouns I, De Proost I, Pintelon I, Timmermans JP, Adriaensen D: Sensory receptors in the airways: neurochemical coding of smooth muscle-associated airway receptors and pulmonary neuroepithelial body innervation. Auton Neurosci. 2006, 126–127: 307-19. 10.1016/j.autneu.2006.02.006.View ArticlePubMedGoogle Scholar
- Sung KW, Kirby M, McDonald MP, Lovinger DM, Delpire E: Abnormal GABAA receptor-mediated currents in dorsal root ganglion neurons isolated from Na-K-2Cl cotransporter null mice. J Neurosci. 2000, 20 (20): 7531-8.PubMedGoogle Scholar
- Lee MG, Macglashan DW, Undem BJ: Role of chloride channels in bradykinin-induced guinea pig airway vagal C-fibre activation. J Physiol. 2005, 566 (Pt 1): 205-12. 10.1113/jphysiol.2005.087577.PubMed CentralView ArticlePubMedGoogle Scholar
- Rudomin P, Schmidt RF: Presynaptic inhibition in the vertebrate spinal cord revisited. Exp Brain Res. 1999, 129 (1): 1-37. 10.1007/s002210050933.View ArticlePubMedGoogle Scholar
- Granados-Soto V, Arguelles CF, Alvarez-Leefmans FJ: Peripheral and central antinociceptive action of Na+-K+-2Cl- cotransporter blockers on formalin-induced nociception in rats. Pain. 2005, 114 (1–2): 231-8. 10.1016/j.pain.2004.12.023.View ArticlePubMedGoogle Scholar
- Mazzone SB, McGovern AE: Sensory neural targets for the treatment of cough. Clin Exp Pharmacol Physiol. 2007, 34 (10): 955-62. 10.1111/j.1440-1681.2007.04702.x.View ArticlePubMedGoogle Scholar
- Dobretsov M, Hastings SL, Stimers JR: Non-uniform expression of alpha subunit isoforms of the Na+/K+ pump in rat dorsal root ganglia neurons. Brain Res. 1999, 821 (1): 212-7. 10.1016/S0006-8993(98)01361-4.View ArticlePubMedGoogle Scholar
- Dobretsov M, Hastings SL, Sims TJ, Stimers JR, Romanovsky D: Stretch receptor-associated expression of alpha 3 isoform of the Na+, K+-ATPase in rat peripheral nervous system. Neuroscience. 2003, 116 (4): 1069-80. 10.1016/S0306-4522(02)00922-3.View ArticlePubMedGoogle Scholar
- Yu J, Wang YF, Zhang JW: Structure of slowly adapting pulmonary stretch receptors in the lung periphery. J Appl Physiol. 2003, 95 (1): 385-93.View ArticlePubMedGoogle Scholar
- Canning BJ, Kollarik M, Undem BJ, Mazzone SB: Demonstration of the essential role of the alpha3-expressing isozyme of the Na+-K+-ATPase in regulating cough receptor activation in guinea pigs. Am J Respir Crit Care Med. 2004, 169 (7): A799-Google Scholar
- Dobretsov M, Hastings SL, Stimers JR: Functional Na+/K+ pump in rat dorsal root ganglia neurons. Neuroscience. 1999, 93 (2): 723-9. 10.1016/S0306-4522(99)00122-0.View ArticlePubMedGoogle Scholar
- Kim JH, Sizov I, Dobretsov M, von Gersdorff H: Presynaptic Ca2+ buffers control the strength of a fast post-tetanic hyperpolarization mediated by the alpha3 Na(+)/K(+)-ATPase. Nat Neurosci. 2007, 10 (2): 196-205. 10.1038/nn1839.View ArticlePubMedGoogle Scholar
- Takamori S, Rhee JS, Rosenmund C, Jahn R: Identification of a vesicular glutamate transporter that defines a glutamatergic phenotype in neurons. Nature. 2000, 407 (6801): 189-94. 10.1038/35025070.View ArticlePubMedGoogle Scholar
- Mutolo D, Bongianni F, Fontana GA, Pantaleo T: The role of excitatory amino acids and substance P in the mediation of the cough reflex within the nucleus tractus solitarii of the rabbit. Brain Res Bull. 2007, 74 (4): 284-93. 10.1016/j.brainresbull.2007.07.001.View ArticlePubMedGoogle Scholar
- Canning BJ, Mori N, Mazzone SB: Vagal afferent nerves regulating the cough reflex. Respir Physiol Neurobiol. 2006, 152 (3): 223-42. 10.1016/j.resp.2006.03.001.View ArticlePubMedGoogle Scholar
- Myers AC, Kajekar R, Undem BJ: Allergic inflammation-induced neuropeptide production in rapidly adapting afferent nerves in guinea pig airways. Am J Physiol Lung Cell Mol Physiol. 2002, 282 (4): L775-81.View ArticlePubMedGoogle Scholar
- Fischer A, Mayer B, Kummer W: Nitric oxide synthase in vagal sensory and sympathetic neurons innervating the guinea-pig trachea. J Auton Nerv Syst. 1996, 56 (3): 157-60. 10.1016/0165-1838(95)00085-2.View ArticlePubMedGoogle Scholar
- Moore KA, Taylor GE, Weinreich D: Serotonin unmasks functional NK-2 receptors in vagal sensory neurones of the guinea-pig. J Physiol. 1999, 514 (Pt 1): 111-24. 10.1111/j.1469-7793.1999.111af.x.PubMed CentralView ArticlePubMedGoogle Scholar
- Brouns I, Adriaensen D, Burnstock G, Timmermans JP: Intraepithelial vagal sensory nerve terminals in rat pulmonary neuroepithelial bodies express P2X(3) receptors. Am J Respir Cell Mol Biol. 2000, 23 (1): 52-61.View ArticlePubMedGoogle Scholar
- Yamamoto Y, Atoji Y, Kuramoto H, Suzuki Y: Calretinin-immunoreactive laminar nerve endings in the laryngeal mucosa of the rat. Cell Tissue Res. 1998, 292 (3): 613-7. 10.1007/s004410051091.View ArticlePubMedGoogle Scholar
- Yamamoto Y, Atoji Y, Suzuki Y: Calretinin immunoreactive nerve endings in the trachea and bronchi of the rat. J Vet Med Sci. 1999, 61 (3): 267-9. 10.1292/jvms.61.267.View ArticlePubMedGoogle Scholar
- Ichikawa H, Helke CJ: Parvalbumin and calbindin D-28k in vagal and glossopharyngeal sensory neurons of the rat. Brain Res. 1995, 675 (1–2): 337-41. 10.1016/0006-8993(95)00071-W.View ArticlePubMedGoogle Scholar
- Ichikawa H, Jin HW, Terayama R, Yamaai T, Jacobowitz DM, Sugimoto T: Calretinin-containing neurons which co-express parvalbumin and calbindin D-28k in the rat spinal and cranial sensory ganglia; triple immunofluorescence study. Brain Res. 2005, 1061 (2): 118-23. 10.1016/j.brainres.2005.08.014.View ArticlePubMedGoogle Scholar
- Yamamoto Y, Atoji Y, Suzuki Y: Calbindin D28k-immunoreactive afferent nerve endings in the laryngeal mucosa. Anat Rec. 2000, 259 (3): 237-47. 10.1002/1097-0185(20000701)259:3<237::AID-AR20>3.0.CO;2-P.View ArticlePubMedGoogle Scholar
This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.