Central mechanisms of airway sensation and cough ...472018/UQ472018_OA.pdf · Mazzone SB, Central mechanisms of airway sensation and cough hypersensitivity, Pulmonary Pharmacology

Accepted Manuscript

Central mechanisms of airway sensation and cough hypersensitivity

Alexandria K. Driessen, Alice E. McGovern, Monica Narula, Seung-Kwon Yang,Jennifer A. Keller, Michael J. Farrell, Stuart B. Mazzone

PII: S1094-5539(17)30016-0

DOI: 10.1016/j.pupt.2017.01.010

Reference: YPUPT 1584

To appear in: Pulmonary Pharmacology & Therapeutics

Received Date: 12 January 2017

Accepted Date: 25 January 2017

Please cite this article as: Driessen AK, McGovern AE, Narula M, Yang S-K, Keller JA, Farrell MJ,Mazzone SB, Central mechanisms of airway sensation and cough hypersensitivity, PulmonaryPharmacology & Therapeutics (2017), doi: 10.1016/j.pupt.2017.01.010.

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Central mechanisms of airway sensation and

cough hypersensitivity

Alexandria K. Driessen1, Alice E. McGovern1, Monica Narula2, Seung-Kwon

Yang2, Jennifer A. Keller2, Michael J. Farrell3 and Stuart B. Mazzone1*

1Department of Anatomy and Neuroscience, The University of Melbourne, Australia

2 School of Biomedical Sciences, The University of Queensland, Australia

3Biomedicine Discovery Institute and Department of Medical Imaging and Radiation Sciences, Monash

University, Australia

*Author for correspondence: Department of Anatomy and Neuroscience, The University of Melbourne,

Parkville, Melbourne, VIC 3010, Australia

Email: [email protected]

Phone: +61 3 8344 6457

Funding: NHMRC of Australia #1078943

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Abstract

The airway sensory nervous system is composed of two anatomically distinct

processing pathways that allow for the production of respiratory reflexes and voluntary

evoked respiratory behaviours in response to sensing an airway irritation. Disordered

sensory processing is a hallmark feature of many pulmonary disorders and results in

the development of cough hypersensitivity syndrome, characterised by chronic cough

and a persistent urge-to-cough in affected individuals. However, the mechanism

underpinning how the airway sensory circuits become disordered, especially at the

level of the central nervous system, is not well understood. In this mini-review we

present well-defined mechanisms that lead to the development of chronic pain as a

framework to explore the evidence that cough disorders may manifest due to

neuroplasticity and sensitisation of important components of the airway sensory

circuitry in the brain. We highlight recent discoveries of how airway sensory

processing occurs in the brain in health and disease and additionally suggest areas

where gaps exist in our current knowledge on the topic, with the goal of providing a

better understanding of how airway circuits become dysfunctional in disease. This

may in turn help identify novel therapeutic targets for restoring normal airway sensory

processing and alleviating excessive cough.

Keywords: Central sensitization; Vagal afferents; Brainstem; Descending inhibition;

Neuroinflammation; fMRI

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1.0 Introduction

The airways are innervated by heterogeneous populations of jugular and nodose vagal

ganglia sensory neurons that respond to a wide range of chemical and mechanical

stimuli. Activation of these sensory neurons results in the transmission of information

about the airway environment, firstly to the brainstem and then to higher order brain

regions, allowing for the production of both respiratory and autonomic reflexes, as well

as more complex behavioural respiratory responses. Collectively, these reflexes and

respiratory behaviours serve to protect the airways from endogenous or exogenous

stimuli that are potentially damaging and by doing so they contribute to the maintenance

of adequate physiological respiratory function. The characterisation and physiology of

airway vagal afferent neurons has been reviewed in detail, as has the neuroanatomical

organisation of airway vagal afferent pathways in the brain (Widdicombe, 2001, Carr

and Undem, 2003, Canning et al., 2006, Driessen et al., 2016, Mazzone and Undem,

2016), and as such the present review will only briefly cover these topics. Rather, we

will focus on plasticity in the central neural circuitry involved in airway sensory

processing and the current understanding of how this may provide insight into the

airway sensory dysfunctions that underpin two prominent and related clinical

presentations in pulmonary disease; excessive coughing and the urge-to-cough. In doing

so, we will present some original data and draw from the advanced work that has been

conducted in a comparable area of research (chronic pain), with the aim of highlighting

important gaps in our current understanding of central cough neurophysiology.

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2.0 Neuroanatomy of airway sensation

The central representation of jugular and nodose airway afferents has been investigated

using a variety of physiological and neuroanatomical techniques (McGovern et al.,

2012a, McGovern et al., 2012b, Driessen et al., 2015, McGovern et al., 2015a,

McGovern et al., 2015b). Such studies have described in detail a primary termination

site for airway vagal afferents in the brainstem nucleus of the solitary tract (nTS) and

not surprisingly this nucleus has underpinned a substantial body of work on airway

sensory neurotransmission (Katz and Karten, 1983, Kubin et al., 1991, Panneton, 1991,

Ranson et al., 1995, Mazzone and Canning, 2002, Alheid et al., 2011, Zoccal et al.,

2014). However, our own tracing studies from the airways using a novel Herpes

Simplex Virus 1 strain H129 (HSV1 H129) anterograde transynaptic tracer recently

described a second primary termination site for airway vagal afferents in the brainstem

paratrigeminal nucleus (Pa5) (McGovern et al., 2012a, McGovern et al., 2015a,

McGovern et al., 2015b). The Pa5 is a small obscure region of brainstem neurons

located within the tract of the medullary spinal trigeminal nucleus, previously described

as playing a role in somatic nociception, but not recognised for playing a role in airway

sensation (Phelan and Falls, 1989; Lapa and Watanabe, 2005; Koepp et al., 2006).

Using conventional retrograde neuroanatomical tracing from the nTS and Pa5 we have

further shown that the nodose and jugular vagal ganglia have distinct and specific

projections to these two termination sites (Driessen et al., 2015, McGovern et al.,

2015b). Thus, the nodose vagal ganglia display terminations only in the nTS, while the

jugular vagal ganglia predominately project to the Pa5 (Driessen et al., 2015, McGovern

et al., 2015b). This observation is striking and coincides with the reported distinct

embryological origins of nodose and jugular neurons – the former derived from the

epibranchial placodes and the latter from the somatic neural crest. This anatomical

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segregation of nodose and jugular afferent processing at the level of the brainstem is

conserved in the ascending higher brain pathways, inasmuch as a comparison of airway

specific projections arising from the nTS and Pa5 showed that nodose-nTS afferents

project largely to regions previously identified in autonomic and limbic/paralimbic

pathways, perhaps representing a central viscerosensory processing circuit in the brain,

while the jugular-Pa5 pathway projects largely to regions involved in somatic

nociceptive (somatosensory) processing (McGovern et al., 2015b). This notion of

multiple cortical circuits involved in airway sensations has been supported by studies in

humans using functional brain imaging (Farrell et al., 2012; Ando et al., 2014; Farrell et

al., 2014), however whether this relates to the anatomical segregation of airway afferent

processing observed in animal models remains unclear.

These data clearly argue for two anatomically distinct airway sensory processing

pathways that may form an anatomical framework for functional distinctions between

respiratory reflex production and the conscious perception of airway irritations. For

example, reflexive cough is a brainstem-mediated process that requires sensory input

into the nTS and subsequent alterations to the activity of respiratory cells located in

nuclei of the ventral and pontine respiratory groups (Baekey et al., 2003, Poliacek et al.,

2009, Mazzone et al., 2011, Smith et al., 2013, Dutschmann et al., 2014, Koshiya et al.,

2014, Zoccal et al., 2014, Ferreira et al., 2015, Wang et al., 2015). On the other hand,

the central mechanisms that underpin the conscious perception of airway sensations

leading to voluntary respiratory behaviours are less well understood and could

conceivably be linked to the understudied somatic representation of airway sensation in

the brain that is seemingly carried by the jugular-Pa5 circuitry. Although this

speculation awaits empirical evidence, it provides intriguing possibilities for

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interventional regulation of airway sensations without disturbing fundamental protective

reflexes. It also provides unexplored possibilities by which airway sensory processes

may become disordered in disease.

3.0 Can airway afferent circuits in the brain be altered in disease?

Neuronal sensitization. Disordered airway sensory neural circuit activity, precipitated

(for example) by the inflammatory processes induced by acute viral or allergen

exposure or that associated with chronic illness in asthma or other respiratory diseases,

contribute to excessive coughing and the increased perception of airway irritation

common to many pulmonary disorders (Chung et al., 2013, Ando et al., 2014, Hilton et

al., 2015, Ando et al., 2016, Zaccone et al., 2016). As such, it has become increasingly

important to understand how airway sensory circuits are altered in these disease states, a

topic that has been investigated in some detail with respect to the primary afferent

neurons themselves (peripheral sensitisation) but largely unexplored in terms of the

central neural circuits in receipt of airway sensory inputs (central sensitisation).

Surprisingly, the notion of central sensitisation as a mechanism of airway sensory

hypersensitivity has been widely presented in the literature without substantive evidence

for central circuit plasticity or even a well-presented framework for what could be

expected if airway sensory circuits in the brain undergo disease-induced changes. Such

a framework can be laid down by looking at other nociceptive systems where the

mechanisms of central neural plasticity have been extensively investigated. Perhaps the

best example of this is in the chronic pain field where the inflammatory and neuropathic

mechanisms leading to changes in central circuit connectivity, neuronal excitability and

gene expression are well described (Latremoliere and Woolf, 2009, Undem et al., 2015,

Bettini and Moore, 2016).

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The similarities between, somatic and airway afferent neurophysiology supports the

notion that common mechanisms may underlie the development of pain and cough

hypersensitivities (Mazzone et al., 2009, Ji, 2015, Undem et al., 2015, Ando et al.,

2016). Indeed, chronic pain and cough are characterised by common phenotypes that

relate to underlying inflammatory and/or neuropathic processes (Latremoliere and

Woolf, 2009, Chung et al., 2013, Ji, 2015). These include hyperalgesia/hypertussia

(increased pain/cough sensitivity to noxious stimuli), allodynia/allotussia (increased

pain/cough responsivity to innocuous stimuli) and paraesthesia (abnormal sensations

such as numbness or tingling in the affected tissue) (Latremoliere and Woolf, 2009,

Chung et al., 2013, Ji, 2015). In chronic pain, these phenotypes occur due to peripheral

and/or central sensitisation processes (Latremoliere and Woolf, 2009). Peripheral

sensitisation occurs following tissue damage, whereby the release of inflammatory

mediators alters sensory neuron excitability and gene expression within the sensory

neuron cell bodies and at their peripheral terminals (Latremoliere and Woolf, 2009,

Bettini and Moore, 2016). This sensitisation is restricted to a localised area of damage

and drives hypersensitivities at the injury site (Latremoliere and Woolf, 2009, Bettini

and Moore, 2016). The evidence for peripheral sensitisation in chronic cough has been

reviewed elsewhere and won’t be repeated here (Undem et al., 2015; Zaccone and

Undem, 2016; Mazzone and Undem, 2016).

Second order neuron plasticity. Persistent sensory nerve input to the central nervous

system can induce alterations in central neural circuits by changing second order

sensory neuron excitability and their projection profiles, although in the case of nerve

injury central sensitisation can be induced without prior peripheral sensitisation

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(Latremoliere and Woolf, 2009, Liang et al., 2016). This allows for an uncoupling of

responses to any direct noxious stimulation and thus these processes are important for

the development of secondary hyperalgesia (hypersensitivity outside of the injury zone)

and allodynia (Latremoliere and Woolf, 2009). The induction and maintenance of

centrally sensitised pain states depends on both acute and chronic processes, initially at

the level of the primary afferent synapse in the spinal dorsal horn (Bettini and Moore,

2016). Alterations that occur in the acute phase are remarkably similar to those seen in

peripheral sensitisation, except that they affect second order sensory neurons and their

projections throughout the central nervous system (Latremoliere and Woolf, 2009,

Cornelison et al., 2016, Liang et al., 2016). That is the release of inflammatory

cytokines, neurotrophins and neuropeptides drives hyperexcitability in second order

sensory neurons (Tao et al., 2014, Bettini and Moore, 2016). Collectively these

processes increase the excitability of the post-synaptic neurons in the spinal dorsal horn

and therefore lead to increased activation of neurons elsewhere in key pain processing

regions such as the raphe nuclei in the brainstem and the anterior cingulate and

somatosensory cortices in the higher brain (Li et al., 2014, Tao et al., 2014, Liang et al.,

2016, Potter et al., 2016). This enhanced excitability is largely driven by increased

synaptic delivery of ion channels, transporters, neuropeptides and vesicle release

machinery that are necessary to facilitate and strengthen glutamatergic transmission

(Latremoliere and Woolf, 2009, Tao et al., 2014, Radwani et al., 2016).

With respect to cough, studies have shown that airway afferent second order neurons in

the brainstem can similarly become hyperexcitable (Mutoh et al., 2000, Mazzone and

Canning, 2002, Mazzone et al., 2005, Kline et al., 2007, Sekizawa et al., 2008, Undem

et al., 2015). More specifically, exposure of the airways to environmental pollutants,

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such as cigarette smoke, is known to increase excitability of nTS neurons with the

neuropeptide substance P playing an essential role in driving increased synaptic

neurotransmission (Mutoh et al., 2000, Sekizawa et al., 2008). Consistent with this, the

activation of pulmonary C-fibre afferents (many of which express neuropeptides)

acutely enhances mechanoreceptor (glutamatergic) reflexes via a brainstem

neuropeptide-dependent mechanism (Mazzone and Canning, 2002, Mazzone et al.,

2005). While glutamatergic and ionic changes may also be directly altered, it is apparent

that dysregulation of neuropeptide expression and transmission is a key component in

the development of central sensitisation of airway sensory circuits at the level of the

sensory integration nuclei in the brainstem. Whether jugular afferent pathways similarly

drive neuropeptide-dependent sensitisation of second order Pa5 neurons has not yet

been studied. However, pulmonary infections induced by pneumovirus exposure in mice

results in elevated neuronal activity within the Pa5 (Mazzone et al., unpublished data),

suggesting that there is persistent sensory input to this region as a consequence of

airway inflammation.

Glia and neuroinflammation. The transition from acute to chronic central sensitisation

includes activation of glia that induce a state of spinal neuroinflammation, which may

reflect the persistent or altered primary sensory neuron input to the central nervous

system that promotes the development of central neuroinflammation (Gwak et al., 2012,

Ikeda et al., 2012, Trang et al., 2012, Chen et al., 2014, Gu et al., 2016, Lei et al., 2016,

Liu et al., 2016). Therefore, both peripheral nerve injury and inflammation can

upregulate the expression of glia, as marked by increased expression of CD11b (OX-42)

or ionized calcium binding adaptor molecule 1 (Iba1) for microglia activation and Glial

Fibrillary Acidic Protein (GFAP) for astrocyte activation, in both sensory ganglia

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themselves and in the central nervous system (Ikeda et al., 2012, Romero et al., 2013,

M’Dahoma et al., 2015, Liu et al., 2016, Lei et al., 2016). More importantly,

pharmacological inhibition of glia has been shown to largely abolish enhanced neuronal

excitation in models of chronic pain, suggesting that glial cells are the primary drivers

of the neuronal hyperexcitability underpinning painful phenotypes (Ikeda et al., 2012).

Interestingly, in a model of nerve injury, spinal dorsal horn neurons become

hyperexcitable and this coincides with an increase in connexion-43 expression in

astrocytes (Chen et al., 2014). Connexin-43 is a gap junction protein that facilitates glia-

neuron cross talk, and when inhibited leads to suppression of neuronal excitation,

suggesting that more complex interactions between activated glia and neurons after

injury may further enhance the neuroinflammatory state (Chen et al., 2014). Since glia

are not electrically active they interact with neurons via neurotransmitter release, such

as cytokines or other mediators, and in line with this there is a growing body of

evidence that suggests adenosine tri-phosphate (ATP), chemokine CXCL12, interleukin

1 beta (IL-1β), tumor necrosis factor alpha (TNFα) and/or Brain Derived Neurotrophic

Factor (BDNF) may be the primary mediators released to maintain a state of central

sensitisation after peripheral injury and inflammation (Henn, 1976, Guthrie et al., 1999,

Fields and Burnstock, 2006, Ulmann et al., 2008, Trang et al., 2011, Trang et al., 2014,

M’Dahoma et al., 2015, Bai et al., 2016, Gu et al., 2016, Lalo et al., 2016). Furthermore,

the neuroinflammatory state in chronic pain is known to persist due to epigenetic

changes to second order sensory neurons, including histone modifications on important

genes including BDNF (Crow et al., 2013, Tao et al., 2014).

There is a paucity of data to indicate whether vagal ganglia or the brainstem processing

sites of airway afferents display glial activation in models of airway inflammation.

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However, recent studies may support this notion (Chounlamountry et al., 2015,

Spaziano et al., 2015). Allergen sensitisation as well as ozone exposure cause gliosis

within the nTS, and it has been speculated that this upregulation plays a role in airway

hyper-responsiveness (Chounlamountry et al., 2015, Spaziano et al., 2015). Gliosis at

glutamatergic synapses may indicate that the activation of glia could strengthen

glutamatergic transmission and in turn drive neuronal hyper-excitability in second order

neurons (Chounlamountry et al., 2015). In our own studies, neonatal mice infected with

murine pneumovirus demonstrate a modest increase in Iba1 expression (microglial

cells) and a robust increase in GFAP expression (astrocytes), along with increased

cytokine expression, in the region of the nTS (Figure 1). This gliosis may reflect both

migration and proliferation of glia cells given that we also noted an increase in the cell

proliferation marker PCNA at this site (Figure 1). Correspondingly, influenza infection

alters nTS neuron resting membrane potential and baseline action potential firing,

consistent with a centrally sensitised state (Figure 1). Nevertheless, more extensive

studies still need to be conducted to better understand the role of activated glia in

altering airway sensory neurons but data to date suggests that it is likely an avenue

worth investigating to better understand the role of central neural plasticity in

respiratory disease.

FIGURE 1 HERE

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Figure 1. Brainstem neuroinflammation following respiratory tract viral infection in mice. Central sensitisation in chronic pain is typically underpinned by glial cell activation and central inflammation, which contributes to synaptic plasticity and neuronal hyperresponsiveness. (A) 7 day old neonatal mice infected with an intranasal inoculation of 5 plaque forming units of pneumovirus show increased immunostaining density for the glial markers GFAP (astrocytes) and IBA1 (microglia) 3-10 days post-inoculation (DPI) in the nucleus of the solitary tract. Virus induced increase in the number of cell nuclei expressing the proliferation marker PCNA is consistent with glial cell expansion at this site (data represent the mean ± SEM of 5-7 animals per group). (B) Consistent with this, pneumovirus infected mice have elevated transcripts for the inflammatory mediators Tumor Necrosis Factor alpha (TNFα) and Interleukin 1β in brainstem homogenates (quantitative PCR, mean ± SEM fold change expression over β-actin, n=3 per group). (C) Representative patch clamp electrophysiological recording of two neurons in nucleus of the solitary tract in brainstem slices of two mice, one inoculated intranasally 7 days earlier with vehicle (VEH, lower trace) and the other inoculated with 103 plaque forming units of influenza Pr8 strain (FLU, upper trace). Of note is the more depolarised resting membrane potential and induction of spontaneous action potential discharge after viral infection. Mean data show an average 40mV shift in the resting membrane potential of nucleus of the solitary tract neurons after infection (mean ± SEM resting membrane potential of 10 neurons per group recorded from 3-4 separate preparations per treatment). *, P<0.05 significantly different to vehicle inoculated (VEH) controls.

Higher level plasticity. Plasticity and sensitisation following peripheral injuries and

inflammation are not confined to second order neurons in primary afferent processing

sites, but rather can occur at all levels of the central sensory processing circuit. In this

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regard, it is interesting to note that unlike the data described above, which is almost

exclusively obtained from animal studies, higher brain plasticity in sensory

hypersensitivities has been studied in both animals and humans. Indeed, functional

magnetic resonance imaging (fMRI) has uniquely allowed higher brain physiological

sensory circuits to be studied in humans in both healthy and disease states (Farrell et al.,

2005, Mazzone et al., 2009, Wager et al., 2013, Farrell and Mazzone, 2014, Hodkinson

et al., 2015, Ando et al., 2016, Flodin et al., 2016). The available evidence would

suggest that sensory hypersensitivity can result from both an enhanced activity of the

brain regions encoding sensation as well as dysfunctional responses in the brain circuits

that ordinarily provide descending control over primary afferent processing (the

descending analgesia system).

In patients with chronic pain, fMRI studies have reported that there is an increased

functional connectivity at most levels of the higher order pain circuit (Zambreanu et al.,

2005, Hodkinson et al., 2015, Flodin et al., 2016). A region of particular importance in

this circuit is the somatosensory cortex where the conscious perception of sensory

stimulation for each body part is mapped onto a restricted cortical region (Endo et al.,

2007). fMRI studies in rodents have shown that in models of chronic pain the integrity

of the primary somatosensory cortex organisation is lost and instead activity extends

beyond the boundaries of the affected region resulting in an increased receptive field

and hyperalgesia (Endo et al., 2007, Potter et al., 2016). Although these changes may

result due to increased afferent input, local mechanisms within the primary

somatosensory cortex have also been identified in rodent models of chronic pain. These

include upregulation of excitatory synapses by significantly increased vesicular

glutamate transporter 1 (VGlut1) synaptic density as well as activation of local

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astrocytes (Kim et al., 2016, Potter et al., 2016). These profound alterations drive an

excitatory-inhibitory imbalance allowing for dysfunctional pain processing to persist.

Similarly, imaging studies in humans with painful trigeminal neuropathy show signs of

decreased grey matter volume and blood flow to the primary somatosensory cortex as

well as the thalamus (Henderson et al., 2013). These changes represent functional

cortical plasticty as they are correlated with reported increased ongoing pain (Henderson

et al., 2013). In line with human imaging studies, the thalamus and subthalamus have

also been implicated in rodent models of chronic pain whereby changes in the gating

and processing of nociceptive information have been observed (Whitt et al., 2013, Masri

et al., 2009). For example, thalamic sensory regulation is in part dependent on tonic

inhbitory signals from the zona incerta (a subthalamic nucleus) and after spinal cord

injury, neurons of the zona incerta have a decreased firing rate leading to increased

thalmic neuron discharge in response to noxious stimulation (Whitt et al., 2013, Masri et

al., 2009). Thus, the balance between excitatory and inhibitory control within the pain

circuitry is lost and pain signals are processed without appropriate gating allowing for

both hyperalgesia and allodynia to result.

A well described descending circuit provides top down control over ascending

nociceptive processing, and recent studies suggest that this circuit may also undergo

changes in patients with chronic pain. The most extensively studied of these descending

circuits is that involved in pain inhibition, also known as the descending analgesia

system, largely controlled by a neuronal circuit that originates in the frontal cortex and

descends upon the spinal dorsal horn via connections in the midbrain periaqueductal

gray (PAG) and rostral ventromedial medulla (RVM) (Boadas-Vaello et al., 2016). The

RVM contains both ON and OFF cells, which enhance or inhibit pain respectively

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(Boadas-Vaello et al., 2016). ON cells receive inhibitory control from the PAG in order

to allow for the firing of OFF cells and it is this balance that ultimately provides

endogenous anti-nociceptive benefits (Boadas-Vaello et al., 2016). However, in chronic

pain conditions alterations in the PAG change the balance of excitatory to inhibitory

control, leading to the development of primary and secondary hyperalgesia (Liu et al.,

2010, Hahm et al., 2011, Schwedt et al., 2014, Ho et al., 2015, Boadas-Vaello et al.,

2016). For example, spinal nerve ligations in the rat resulted in both altered

glutamatergic and GABAergic transmission in the PAG and both the PAG and RVM

display signs of neuroinflammation following peripheral injuries, which is presumably

involved in the maintenance of a centrally sensitised state (Ho et al., 2015, Zhuang et

al., 2016). In particular, models of Parkinson’s disease, where pain is a commonly

reported symptom, the dorsal lateral PAG has increased activation of pro-inflammatory

cytokines, while microglia have been shown to be activated in the RVM under

conditions of peripheral inflammatory pain (Roberts et al., 2009, Zhuang et al., 2016).

In human functional brain imaging studies, experimentally induced hyperalgesia

following subcutaneous injections of capsaicin results in a rapid induction of PAG

neuronal activity, suggestive that alterations in the descending analgesia pathway plays

an essential role in the development of hyperalgesia (Zambreanu et al., 2005).

Consistent with this, it has also been shown that these descending pain circuits have

altered connectivity in chronic pain conditions that present with allodynia (Schwedt et

al., 2014).

Patients with chronic cough may similarly display alterations in their higher brain

processing of airway irritant stimuli (Ando et al., 2016). We recently reported that

patients with chronic cough demonstrated enhanced urge-to-cough sensitivity during

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inhaled capsaicin challenges and that this was associated with elevated activity in

midbrain regions known to be involved in descending nociceptive control. Thus, in

fMRI studies the PAG and neighbouring cuneiform nucleus showed elevated activity in

response to inhaled capsaicin, regardless of whether capsaicin concentrations were the

same for healthy and cough participants or if they were tailored to produce equivalent

behavioural outcomes in the two groups (i.e. to match the urge-to-cough intensities)

(Ando et al., 2016). Remarkably, the midbrain areas activated in our study overlap with

the areas active during experimental induction of pain hypersensitivity (Figure 2),

strongly suggesting that common central mechanisms are indeed involved in the

development or maintenance of cough and pain hypersensitivity. The midbrain,

however, was not the only higher brain region demonstrating changes in activity. Cough

hypersensitivity patients showed diminished activity compared to healthy controls in

parts of a brain network previously implicated in the voluntary suppression of coughing

(Mazzone et al., 2011, Farrell et al., 2012; Farrell et al., 2014, Ando et al., 2016).

Collectively these findings suggest that altered descending control and/or altered

voluntary cough suppression may contribute to excessive coughing and the urge-to-

cough in airways disease. However, more research is needed in this area as we don’t

know if cough hypersensitivity coincides with any changes in sensory representations in

the brain, nor do we understand the mechanisms that precipitate these changes seen with

fMRI. Furthermore, as our study described above was conducted by investigating brain

responses during inhaled irritant challenges, it will be important to know whether there

are any changes in brain activity, connectivity and/or brain morphometry in cough

patients at baseline in the absence of exogenous stimuli. In addition, whether current or

future antitussive therapies can correct altered brain activity is also unknown.

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FIGURE 2 HERE

Figure 2. A common mechanism of central sensitisation in pain and cough: functional

brain imaging studies in humans. Left panel, Subjects were treated unilaterally on the

right lower eg with a combination of heat and capsaicin to induce hyperalgesia.

Subsequent mechanical stimulation of the area evoked increased pain sensations

which were associated with unilateral increased neural activity in the midbrain nucleus

cuneiformis (reproduced with permission from Zambreanu et al., 2005). Right Panel,

Comparable activations in the midbrain nucleus cuneiformis extending into the

periaqueductal grey (PAG) in subjects with chronic cough hypersensitivity exposed to

inhaled capsaicin challenges (data adapted from Ando et al., 2016). Not shown is an

absence of midbrain activity in control subjects, consistent with the midbrain playing a

specific role in the development or maintenance of hypersensitivity.

3.0 Concluding remarks

Cough and the urge-to-cough contribute significantly to morbidity in pulmonary

diseases and as such understanding the neuronal processes that underpin the

development and maintenance of cough hypersensitivity is essential. We now have a

reasonably advanced knowledge of the central processing circuits involved in airway

sensation that will allow for hypothesis driven investigations of how the brain may

contribute to chronic coughing in disease. Although this area of research is still in its

infancy, the information that we have presented in this review highlights the likely

importance of central plasticity in the development of cough in respiratory disease.

Indeed, there is evidence that sensory circuits are subject to disease induced acute

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changes in neuronal excitability through alterations in neurotransmission and ion

channel activity in the brainstem, and that this enhanced excitability may be maintained

and transformed to a chronic neuroinflammatory state by glial cell activation. The net

outcome is altered activity of cough regulatory circuits in the brains of patients with

chronic cough. However, there are clearly still many gaps in our understanding of the

processes and it will be important to define the mechanisms underpinning central

sensitisation and plasticity in the airway sensory circuitry as this may help to identify

novel therapeutic targets to better treat patients with sensory hypersensitivities

associated with respiratory diseases.

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Figure Legends

Figure 1. Brainstem neuroinflammation following respiratory tract viral infection in mice.

Central sensitisation in chronic pain is typically underpinned by glial cell activation and

central inflammation, which contributes to synaptic plasticity and neuronal

hyperresponsiveness. (A) 7 day old neonatal mice infected with an intranasal inoculation

of 5 plaque forming units of pneumovirus show increased immunostaining density for the

glial markers GFAP (astrocytes) and IBA1 (microglia) 3-10 days post-inoculation (DPI) in the

nucleus of the solitary tract. Virus induced increase in the number of cell nuclei expressing

the proliferation marker PCNA is consistent with glial cell expansion at this site (data

represent the mean ± SEM of 5-7 animals per group). (B) Consistent with this, pneumovirus

infected mice have elevated transcripts for the inflammatory mediators Tumor Necrosis

Factor alpha (TNFα) and Interleukin 1β in brainstem homogenates (quantitative PCR, mean

± SEM fold change expression over β-actin, n=3 per group). (C) Representative patch clamp

electrophysiological recording of two neurons in nucleus of the solitary tract in brainstem

slices of two mice, one inoculated intranasally 7 days earlier with vehicle (VEH, lower trace)

and the other inoculated with 103 plaque forming units of influenza Pr8 strain (FLU, upper

trace). Of note is the more depolarised resting membrane potential and induction of

spontaneous action potential discharge after viral infection. Mean data show an average

40mV shift in the resting membrane potential of nucleus of the solitary tract neurons after

infection (mean ± SEM resting membrane potential of 10 neurons per group recorded from

3-4 separate preparations per treatment). *, P<0.05 significantly different to vehicle

inoculated (VEH) controls.

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Figure 2. A common mechanism of central sensitisation in pain and cough: functional brain

imaging studies in humans. Left panel, Subjects were treated unilaterally on the right lower

eg with a combination of heat and capsaicin to induce hyperalgesia. Subsequent

mechanical stimulation of the area evoked increased pain sensations which were associated

with unilateral increased neural activity in the midbrain nucleus cuneiformis (reproduced

with permission from Zambreanu et al., 2005). Right Panel, Comparable activations in the

midbrain nucleus cuneiformis extending into the periaqueductal grey (PAG) in subjects with

chronic cough hypersensitivity exposed to inhaled capsaicin challenges (data adapted from

Ando et al., 2016). Not shown is an absence of midbrain activity in control subjects,

consistent with the midbrain playing a specific role in the development or maintenance of

hypersensitivity.