Central Chemoreceptors Research Articles

Cardiorespiratory effects of gap junction blockade in the locus coeruleus in unanesthetized adult rats

The locus coeruleus (LC) plays an important role in central chemoreception. In young rats (P9 or younger), 85% of LC neurons increase firing rate in response to hypercapnia vs. only about 45% of neurons from rats P10 or older. Carbenoxolone (CARB – gap junction blocker) does not affect the % of LC neurons responding in young rats but it decreases the % responding by half in older animals. We evaluated the participation of gap junctions in the CO2 ventilatory response in unanesthetized adult rats by bilaterally microinjecting CARB (300μM, 1mM or 3mM/100nL), glycyrrhizic acid (GZA, CARB analog, 3mM) or vehicle (aCSF – artificial cerebrospinal fluid) into the LC of Wistar rats. Bilateral gap junction blockade in LC neurons did not affect resting ventilation; however, the increase in ventilation produced by hypercapnia (7% CO2) was reduced by ∼25% after CARB 1mM or 3mM injection (1939.7±104.8mLkg−1min−1 for the aCSF group and 1468.3±122.2mLkg−1min−1 for 1mM CARB, P<0.05; 1939.7±104.8mLkg−1min−1 for the aCSF group and 1540.9±68.4mLkg−1min−1 for the 3mM CARB group, P<0.05) due largely to a decrease in respiratory frequency. GZA injection or CARB injection outside the LC (peri-LC) had no effect on ventilation under any conditions. The results suggest that gap junctions in the LC modulate the hypercapnic ventilatory response of adult rats.

CrossTalk opposing view: Peripheral and central chemoreceptors have hyperadditive effects on respiratory motor control

CrossTalk opposing view: Peripheral and central chemoreceptors have hyperadditive effects on respiratory motor control

CrossTalk opposing view: Peripheral and central chemoreceptors have hypoadditive effects on respiratory motor output

CrossTalk opposing view: Peripheral and central chemoreceptors have hypoadditive effects on respiratory motor output

Rebuttal from James Duffin and Jason H. Mateika

Our comments in reply to the arguments presented in the CrossTalk articles by Teppema & Smith (2013) and Wilson & Day (2013) are confined to experiments on humans, and in their CrossTalk article Teppema & Smith (2013) cite two studies that found evidence supporting a hyperadditive interaction. First, carotid body denervation or removal in man has consequences for the central chemoreception of CO2. We agree that such denervation removes the tonic drive from the carotid bodies (Dahan et al. 2007), evident from their involvement in sympathetic tone. However, if this drive is indeed tonic, then it does not change with CO2 and so does not function as a hyperadditive factor during hypercapnia. With respect to species differences, it is interesting to note that carotid body denervation in dogs has little effect on the normoxic ventilatory response to CO2 (Mitchell, 1965). The second human study cited by Teppema & Smith (2013) involved acid–base changes induced with acetazolamide and intravenous bicarbonate (Teppema et al. 2010). While arterial tensions and [H+] were determined in these experiments, the stimulus to the central chemoreceptors is unknown, and the effects of acetazolamide on the cerebrovascular sensitivity to CO2 (Fan et al. 2012), and the central and peripheral chemosensitivity to CO2 (Vovk et al. 2000) are confounding factors that make interpretation problematic. There are two common elements in the arguments presented by Wilson & Day (2013) and Teppema and Smith (2013): the recognition that species differences may play a role, and that the various experimental models and scenarios devised to test the interaction between central and peripheral chemoreflexes are complex and subject to limitations. We agree and suggest that a simple test is needed that can be applied across species so that comparisons may be made. The most appropriate experimental preparations therefore need to be identified and employed to move the field forward. For example, vagotomized and decerebrate preparations may have a place in answering specific questions, but it is unlikely that the responses to CO2 and O2 in this type of a preparation would be comparable to a spontaneously breathing intact animal. As we explained in our CrossTalk article (Duffin & Mateika, 2013), the hypoxic, isoxic ventilatory response to hypercapnia can determine the type of interaction between the peripheral and central chemoreflexes. We suggest that this response is the appropriate pan-species test because it is applied to the intact animal and yields the ventilatory regulator characteristics under physiological circumstances.

Rebuttal by Richard J. A. Wilson and Trevor A. Day

Our colleagues skillfully addressed how peripheral chemoreceptors interact with systemic effects of CO2. Unfortunately, systemic CO2 affects oxyhaemoglobin dissociation, sympathetic, endocrine and cardiovascular systems, and afferents from lungs and CO2-sensitive upper airway receptors – all capable of having effects on ventilation independent of central chemoreceptors. Duffin's ingenious rebreathing studies add further concerns, namely psychological and physiological aspects of voluntary hyperventilation (e.g. Steinback et al. 2011). Similarly, carotid body denervation (CBD) studies acclaimed by Teppema & Smith (2013) introduce complications including disruption of baroreflex, abolition of central carotid body regulation of sympathetic and parasympathetic outputs, and unknown contribution of changes associated with long-term plasticity. To mitigate these issues, we developed an artificially perfused rat preparation devoid of intact autonomic and respiratory feedback loops, descending and vagal influences (Day & Wilson, 2009). Demonstrations of hypoadditivity in our preparation are reconcilable with work in humans summarized by Duffin & Mateika (2013). While Duffin & Mateika favour the conclusion of additive interactions, simple addition in ventilation mathematically necessitates hypoadditivity in frequency and/or tidal volume (Mitchell, 1990). Moreover, stimulus and state specific parcelling of responses into frequency and tidal volume suggests hypoadditivity translates to ventilation in certain circumstances. However, our findings are not easily reconciled with a hyperadditive system, as argued by Teppema & Smith (2013). So how do we move forward? Pigeonholed discussions addressing the nature of interaction have illustrated the importance of type, magnitude, order and pattern of stimulus, and species and preparation used, but have yet to reconcile hypoadditive and hyperadditive camps. Therefore, we suggest the need for a new paradigm: a hybrid model incorporating multiple forms of interaction depending on systemic CO2 and/or physiological state. In support of this new paradigm, we note (a) even the hyperadditive O2–CO2 carotid body interaction transitions to hypoadditive at severe levels (Fitzgerald & Parks, 1971), (b) the additive-to-hyperadditive transition with long term facilitation cited by Duffin & Mateika (2013), and (c) Hodges et al. (2005), cited by Teppema & Smith (2013), is only partly consistent with hyperadditivity. Hodges et al. show that, while both mild and severe hypercapnic brainstem aCSF cause increased ventilation, only responses to severe hypercapnic aCSF were reduced by CBD. Thus, while Teppema & Smith considered only the blunted responses to severe hypercapnic aCSF, lack of CBD effect on responses to mild hypercapnic aCSF argues for hypoadditivity. These data are therefore consistent with a hybrid system that endows respiratory chemosensitivity with redundancy and a large dynamic range.

G protein modulation of K2P potassium channel TASK-2

TASK-2 (K2P5.1) is a background K+ channel opened by extra- or intracellular alkalinisation that plays a role in renal bicarbonate handling, central chemoreception and cell volume regulation. Here, we present results that suggest that TASK-2 is also modulated by Gβγ subunits of heterotrimeric G protein. TASK-2 was strongly inhibited when GTP-γ-S was used as a replacement for intracellular GTP. No inhibition was present using GDP-β-S instead. Purified Gβγ introduced intracellularly also inhibited TASK-2 independently of whether GTP or GDP-β-S was present. The effects of GTP-γ-S and Gβγ subunits were abolished by neutralisation of TASK-2 C terminus double lysine residues K257–K258 or K296–K297. Use of membrane yeast two hybrid (MYTH) experiments and immunoprecipitation assays using tagged proteins gave evidence for a physical interaction between Gβ1 and Gβ2 subunits and TASK-2, in agreement with expression of these subunits in proximal tubule cells. Co-immunoprecipitation was impeded by mutating C terminus K257–K258 (but not K296–K297) to alanines. Gating by extra- or intracellular pH was unaltered in GTP-γ-S-insensitive TASK-2-K257A-K258A mutant. Shrinking TASK-2-expressing cells in hypertonic solution decreased the current to 36 % of its initial value. The same manoeuvre had a significantly diminished effect on TASK-2-K257A-K258A- or TASK-2-K296-K297-expressing cells, or in cells containing intracellular GDP-β-S. Our data are compatible with the concept that TASK-2 channels are modulated by Gβγ subunits of heterotrimeric G protein. We propose that this modulation is a novel way in which TASK-2 can be tuned to its physiological functions.

High-inspired oxygen concentration further impairs opioid-induced respiratory depression

Hyperoxaemia depresses the output of peripheral and central chemoreceptors. Patients treated with opioids often receive supplemental oxygen to avert possible decreases in oxygen saturation (Sp(O2)).We examined the effect of a single dose of remifentanil in healthy volunteers inhaling room air vs air enriched with 50% oxygen. Twenty healthy volunteers received i.v. 50 mg remifentanil (infused over 60 s) at anormoxic (N) or hyperoxic (FI(O2) 0.5, H) background on separate occasions. Minute ventilation (Vi), respiratory rate (RR), end-tidal PC(O2), and Sp(O2) were collected on a breath to-breath basis. The occurrence of apnoea was recorded. During normoxia, remifentanil decreased Vi from 7.4 (1.3) [mean (SD)] to 2.2 (1.2) litre min 21 (P,0.01), and during hyperoxia from 7.9 (1.0) to 1.2 (1.2) litre min 21 (P,0.01; H vs N: P,0.001). RR decreased from 13.1 (2.9) to 6.1 (2.8) bpm during N (P,0.01) and from 13.2 (3.0) to 3.6 (4.0) bpm during H (P,0.01; H vs N: P,0.01). During normoxia, Sp(O2) decreased from 98.4 (1.5) to 88.6 (6.7)% (P,0.01), while during hyperoxia, Sp(O2) changed from 99.7 (0.7) to 98.7 (1.0)% (P,0.001). Apnoea developed in two subjects during normoxia and 10 during hyperoxia. Respiratory depression from remifentanil is more pronounced in hyperoxia than normoxia as determined from minute ventilation, end-tidal PC(O2), and RR. During hyperoxia, respiratory depression may be masked when measuring Sp(O2) as pulse oximetry remains in normal values during the first minutes of respiratory depression.

Phox2b-Expressing Retrotrapezoid Neurons Are Intrinsically Responsive to H+and CO2

Central respiratory chemoreceptors sense changes in CO2/H(+) and initiate the adjustments to ventilation required to preserve brain and tissue pH. The cellular nature of the sensors (neurons and/or glia) and their CNS location are not conclusively established but the glutamatergic, Phox2b-expressing neurons located in the retrotrapezoid nucleus (RTN) are strong candidates. However, a direct demonstration that RTN neurons are intrinsically sensitive to CO2/H(+), required for designation as a chemosensor, has been lacking. To address this, we tested the pH sensitivity of RTN neurons that were acutely dissociated from two lines of Phox2b-GFP BAC transgenic mice. All GFP-labeled cells assayed by reverse transcriptase-PCR (n = 40) were Phox2b+, VGlut2+, TH-, and ChAT-, the neurochemical phenotype previously defined for chemosensitive RTN neurons in vivo. We found that most dissociated RTN neurons from both lines of mice were CO2/H(+)-sensitive (∼79%), with discharge increasing during acidification and decreasing during alkalization. The pH-sensitive cells could be grouped into two populations characterized by similar pH sensitivity but different basal firing rates, as previously observed in recordings from GFP-labeled RTN neurons in slice preparations. In conclusion, these data indicate that RTN neurons are inherently pH-sensitive, as expected for a respiratory chemoreceptor.

Serotonergic neurons in the nucleus raphé obscurus are not involved in the ventilatory and thermoregulatory responses to hypoxia in adult rats

The medullary raphé is an important component of the central respiratory network, playing a key role in CO2 central chemoreception. However, its participation in hypoxic ventilatory responses is less understood. In the present study, we assessed the role of nucleus raphé obscurus (ROb), and specifically 5-HT neurons confined in the ROb, on ventilatory and thermoregulatory responses to hypoxia. Chemical lesions of the ROb were performed using either ibotenic acid (non-specific lesion; control animals received PBS) or anti-SERT-SAP (5-HT specific lesion; control animals received IgG-SAP). Ventilation (V˙E; whole body plethysmograph) and body temperature (Tb; data loggers) were measured during normoxia (21% O2, N2 balance) and hypoxia exposure (7% O2, N2 balance, 1h) in conscious adult rats. Ibotenic acid or anti-SERT-SAP-induced lesions did not affect baseline values of V˙E and Tb. Similarly, both lesion procedures did not alter the ventilatory or thermoregulatory responses to hypoxia. Although evidence in the literature suggests a role of the rostral medullary raphé in hypoxic ventilatory responses, under the present experimental conditions our data indicate that caudal medullary raphé (ROb) and its 5-HT neurons neither participate in the tonic maintenance of breathing nor in the ventilatory and thermal responses to hypoxia.

Purinergic signaling in the retrotrapezoid nucleus (RTN) contributes to central and peripheral chemoreflexes by divergent mechansims

Evidence suggests that purinergic signaling in the retrotrapezoid nucleus (RTN) can influence both respiratory and sympathetic activity. Here we evaluate the extent to which purinergic signaling contributes to central and peripheral chemoreception integration at the level of the RTN in vivo. Mean arterial pressure (MAP) and phrenic nerve activity (PNA) were recorded in urethane‐anaesthetized, vagotomized and artificial ventilated male Wistar rats (n = 6–7/group). Bilateral RTN injections of a non‐specific P2‐receptor blocker (PPADS) decreased pressor (8 ± 2 mmHg, vs. saline: 22 ± 4 mmHg) and respiratory (PNA amplitude = 62 ± 3%, vs. saline: 101 ± 5% and PNA frequency = 77 ± 3%, vs. saline: 101 ± 5%) responses to 10% CO2. Conversely, bilateral RTN injections of MRS2179 (P2Y1 receptor antagonist) attenuated cardiorespiratory responses to peripheral chemoreceptor activation, but did not affect CO2‐responsivness. Further, 63% of BDA‐labeled NTS terminals in the RTN are immunoreactive for vesicular glutamate transporter‐2 (VGLUT2) and vesicular nucleotide transporter (VNUT) but only 5% contained glutamic acid decarboxylase‐67 (GAD67). These data indicate that purinergic signaling in the RTN contributes to central and peripheral chemoreception by different, but potentially interactive, mechanisms

Ionotropic but not metabotropic glutamatergic receptors in the Locus Coeruleus modulate the hypercapnic ventilatory response in unanesthetized rats

Central chemoreception is an important mechanism by which changes in tissue CO2/H+ regulate breathing, and the Locus coeruleus (LC) is one of many putative central chemoreceptor sites. Here, we studied the contribution of LC glutamatergic receptors on ventilatory, cardiovascular and thermal responses to hypercapnia. To this end, we measured pulmonary ventilation (VE), body temperatures (Tb), mean arterial pressure (MAP) and heart rate (fH) of male Wistar rats before and after microinjection of Kynurenic Acid (KY, an ionotropic glutamate receptors antagonist, 10 mM) or α‐methyl‐4‐carboxyphenylglycine (MCPG, a metabotropic glutamate receptor antagonist, 10 ηM) unilateraly into the LC, followed by 60 minutes of air or hypercapnia exposure (7% CO2). Ventilatory response to hypercapnia was higher in animals treated with KY intra‐LC (1899 ± 83 mL kg−1 min−1) compared to the control group (1141 ± 64 mL kg−1 min−1; P &lt; 0.01). However, the MCPG treatment within the LC had no effect on the hypercapnia‐induced hyperpnea. The cardiovascular and thermal control were not affected by hypercapnia or by the injection of KY and MCPG in the LC. These data suggest that the glutamate acting on ionotropic but not metabotropic receptors in the LC exerts an inibitory modulation of hypercapnia‐induced hyperpnea.Financial support: FAPESP, CNPq, PROPe.

Effects of leptin in the retrotrapezoid nucleus (RTN) on CO2‐sensitivity and respiration.

Evidence suggests that, in addition to regulate energy homeostasis, leptin also contributes to the CNS control of breathing in response to changes in CO2/H+ (i.e., central chemoreception). However, cellular targets of leptin‐dependent drive to breathe have not been elucidated. Here, we tested the effects of leptin on hypercapnic ventilatory response in leptin deficient mice (ob/ob: 42±2 g) in vivo, and on CO2‐responsiveness of RTN neurons and astrocytes in brain slices isolated from rat pups (P7–12). We found that 3‐ day unilateral RTN injections of leptin (1 μg/100 nl, n=7/group) increased ventilatory responsiveness to 7% CO2 in ob/ob mice (4378±294 μl.min−1.g−1, vs. 2849±319 μl.min−1.g−1in vehicle group). Immunohistochemical analysis using antibodies for leptin receptors, astrocytes (aldehyde dehydrogenase 1L1) and neurons (biocytin) suggest that leptin receptors are expressed by RTN astrocytes but not neurons in ob/ob mice. We also found that acute in vitro exposure to leptin in RTN slices (10 min, 50 nM) elicited no effect on excitability of chemosensitive neurons (cell‐attached current clamp) or conductance of CO2‐sensitive astrocytes (whole‐cell voltage clamp; Ihold −80 mV). These results suggest that although long‐term application of leptin into the RTN increased respiratory drive in ob/ob mice, the cellular and molecular mechanisms remain unclear.

The origin of air breathing in vertebrates: turn (on) your head and cough

Air breathing in tetrapods involves a centrally generated rhythmic activation of respiratory muscles to ventilate a lung, and modulation of this rhythm by central CO2‐sensitive chemoreceptors that match ventilation to metabolic demands. The tetrapod lung arose in primitive air‐breathing fish. The origins of the central rhythm generator and chemoreceptor(s) are unknown. We propose that these critical elements were present in basal vertebrates and were exapted for air breathing with the evolution of the lung. Lamprey represent the basal vertebrate form and as larvae (ammocoetes) are aquatic burrow dwelling filter feeders. We propose that a behavior characterized as “cough” in lamprey is the basis for subsequent evolution of air breathing, and test the hypothesis that the centrally generated cough is stimulated by CO2. Ammocetes in vivo and isolated brainstems in vitro were exposed to hypercapnia. Ammocetes responded with a dose dependant increase in the frequency of spontaneous cough. Brainstems in vitro also produced the cough motor pattern, illustrating that cough results from a centrally CO2‐sensitive rhythm generator. We conclude that a central CO2‐sensitive chemoreceptor and central rhythm generator were present in basal vertebrates. We propose that, with the evolution of the lung, CO2‐sensitive cough is exapted to form the basis for air breathing common to air‐breathing tetrapods. Supported by NSFIOS‐1022442

Ventral respiratory column tonic expiratory neurons: Nodes for convergent afferent modulation of inspiratory drive

The medullary ventral respiratory column (VRC) has an essential role in generating the motor pattern for breathing and inspiratory (I) drive. Contemporary models include a rostral‐to‐caudal excitatory I neuron chain extending to premotor populations that also acts upon inhibitory I neurons with recurrent and feed‐forward connections upon other VRC excitatory and inhibitory I neurons and expiratory (E) neurons. Prior work suggests (i) baroreceptor stimulation inhibits inspiratory drive in part via evoked tonic E (t‐E) neuron activity and (ii) central chemoreceptor stimulation enhances I drive via disinhibition through reduced t‐E neuron activity. A recent computational neuromechanical model (O'Connor et al. 2012) predicted that inhibition of t‐E neurons amplifies I drive during coughing ‐ a result supported by in vivo data (Segers et al. 2012). We now report recording sites of pericolumnar t‐E neurons (n=140) and show that selective stimulation of carotid chemoreceptors via close injection of CO2‐saturated saline reduces the I‐phase activity of t‐E cells coincident with increased I drive in vagotomized decerebrate cats. Spike train cross correlation and gravity clustering methods revealed short‐time scale impulse synchrony in 24 t‐E neuron pairs. We conclude that multiple afferent systems tune inspiratory drive via actions of coordinated groups of VRC t‐E neurons. Support: NIH grant NS19814.

The New Mode to Neurohumoral Regulation in Respiratory and Circulatory Systems

We defined the sophisticated organic whole by placing oxygen (along with CO2 and [H+] Trinity) as the core. The regulation through integrative nervous system is still unclear. In order for us to easily comprehend the working mode of respiratory and circulatory regulations, we use an analogy of ¡°sound system regulation mode¡±: 1. The wave signal generated by the circulating blood in the circulatory system due to pulmonary ventilation is compared to the speech acoustic signals produced by person. 2. Multiple peripheral chemoreceptors and baroreceptors are compared to the sonic microphones that sense sound waves; they collect and upload distinct signals. 3. The central nervous system is equivalent to the volume amplifier control system. 4. The central chemoreceptors act as a volume‐adjusting knob; it balances the central incoming/outgoing integration. 5. The respiratory muscle and thorax lung jointly play the role of a speaker.This concept explains why we can stably maintain our respiration by breath‐by‐breath pattern via respiratory muscles contraction/diastole conversion. Similarly, our circulation can be maintain stable by beat‐by‐beat pattern.

New Theory of Respiratory Control and Regulation

We recently finished ¡°Holistic Regulation of LifeCIntegrative Physiology¡± framework as a whole. The part of respiratory control and regulation is decribed as below: The key control of singles should be considered O2, while CO2 and [H+] act as subordinates to the core integration of breathing, circulation, metabolism, and other multi‐functional system control. The trigger to inhale during the termination of exhale is when the level of PO2 gradually decreases (PCO2 and [H+], opposite). The trigger to exhale during the termination of inhale is when the level of PO2 gradually increases (PCO2 and [H+], opposite), creating the cycle of breathing. Cardiovascular function is the key factor in determining a normal breathing rhythm and frequency. Regulation mode of breathing amplitude (depth): the next breath is induced as above to achieve a fast response through peripheral chemoreceptors. The breathing rate is exaggerated by the last breath, for example, a larger breath induces a larger breath, a smaller breath induces a smaller breath, and so on. Regulatory control the steady breathing pattern must be integration with slow response of central chemoreceptors.

Ionotropic but not metabotropic glutamatergic receptors in the locus coeruleus modulate the hypercapnic ventilatory response in unanaesthetized rats

Central chemoreceptors are important to detect changes of CO2/H(+), and the Locus coeruleus (LC) is one of the many putative central chemoreceptor sites. Here, we studied the contribution of LC glutamatergic receptors on ventilatory, cardiovascular and thermal responses to hypercapnia. To this end, we determined pulmonary ventilation (V(E)), body temperatures (T(b)), mean arterial pressure (MAP) and heart rate (HR) of male Wistar rats before and after unilateral microinjection of kynurenic acid (KY, an ionotropic glutamate receptor antagonist, 10 nmol/0.1 μL) or α-methyl-4-carboxyphenylglycine (MCPG, a metabotropic glutamate receptor antagonist, 10 nmol/0.1 μL) into the LC, followed by 60 min of air breathing or hypercapnia exposure (7% CO2). Ventilatory response to hypercapnia was higher in animals treated with KY intra-LC (1918.7 ± 275.4) compared with the control group (1057.8 ± 213.9, P < 0.01). However, the MCPG treatment within the LC had no effect on the hypercapnia-induced hyperpnea. The cardiovascular and thermal controls were not affected by hypercapnia or by the injection of KY and MCPG in the LC. These data suggest that glutamate acting on ionotropic, but not metabotropic, receptors in the LC exerts an inhibitory modulation of hypercapnia-induced hyperpnea.

The essential role of peripheral respiratory chemoreceptor inputs in maintaining breathing revealed when CO2 stimulation of central chemoreceptors is diminished

Central sleep apnoea is a condition characterized by oscillations between apnoea and hyperpnoea during sleep. Studies in sleeping dogs suggest that withdrawal of peripheral chemoreceptor (carotid body) activation following transient ventilatory overshoots plays an essential role in causing apnoea, raising the possibility that sustaining carotid body activity during ventilatory overshoots may prevent apnoea. To test whether sustained peripheral chemoreceptor activation is sufficient to drive breathing, even in the absence of central chemoreceptor stimulation and vagal feedback, we used a vagotomized, decerebrate dual-perfused in situ rat preparation in which the central and peripheral chemoreceptors are independently and artificially perfused with gas-equilibrated medium. At varying levels of carotid body stimulation (CB PO2/PCO2: 40/60, 100/40, 200/15, 500/15 Torr), we decreased the brainstem perfusate PCO2 in 5 Torr steps while recording phrenic nerve activity to determine the central apnoeic thresholds. The central apnoeic thresholds decreased with increased carotid body stimulation. When the carotid bodies were strongly stimulated (CB 40/60), the apnoeic threshold was 3.6 ± 1.4 Torr PCO2 (mean ± SEM, n = 7). Stimulating carotid body afferent activity with either hypercapnia (60 Torr PCO2) or the neuropeptide pituitary adenylate cyclase-activating peptide restored phrenic activity during central apnoea. We conclude that peripheral stimulation shifts the central apnoeic threshold to very hypocapnic levels that would likely increase the CO2 reserve and have a protective effect on breathing. These data demonstrate that peripheral respiratory chemoreceptors are sufficient to stave off central apnoeas when the brainstem is perfused with low to no CO2.

Arterial chemoreceptor activation reduces the activity of parapyramidal serotonergic neurons in rats

The parapyramidal (ppy) region targets primarily the intermediolateral cell column and is probably involved in breathing and thermoregulation. In the present study, we tested whether ppy serotonergic neurons respond to activation of central and peripheral chemoreceptors. Bulbospinal ppy neurons (n=30) were recorded extracellularly along with the phrenic nerve activity in urethane/α-chloralose-anesthetized, paralyzed, intact (n=7) or carotid body denervated (n=6) male Wistar rats. In intact animals, most of the ppy neurons were inhibited by hypoxia (n=14 of 19) (8% O2, 30s) (1.5±0.03 vs. control: 2.4±0.2Hz) or hypercapnia (n=15 of 19) (10% CO2) (1.7±0.1 vs. control: 2.2±0.2Hz), although some neurons were insensitive to hypoxia (n=3 of 19) or hypercapnia (n=4 of 19). Very few neurons (n=2 of 19) were activated after hypoxia, but not after hypercapnia. In carotid body denervated rats, all the 5HT-ppy neurons (n=11) were insensitive to hypercapnia (2.1±0.1 vs. control: 2.3±0.09Hz). Biotinamide-labeled cells that were recovered after histochemistry were located in the ppy region. Most labeled cells (90%) showed strong tryptophan hydroxylase immunocytochemical reactivity, indicating that they were serotonergic. The present data reveal that peripheral chemoreceptors reduce the activity of the serotonergic premotor neurons located in the ppy region. It is plausible that the serotonergic neurons of the ppy region could conceivably regulate breathing automaticity and be involved in autonomic regulation.

A computerized integrated system for the assessment of central and peripheral chemoreflex sensitivity

The assessment of chemoreflex sensitivity (CRS) is of major importance in studies investigating the adaptation of ventilation to the needs of human body. Increased sensitivity of chemoreceptors to both hypoxia and hypercapnia has recently been shown to be a powerful and independent prognosticator in heart failure (HF) patients, thus highlighting the importance of the assessment of CRS also in the clinical setting. In spite of this, the measurement of CRS is currently limited to the research setting. One possible reason might be the lack of suitable commercial equipments.On the basis of these considerations, we designed a system to carry out a comprehensive assessment of CRS, including both central and peripheral chemoreceptors. The system is based on the integration of different commercial devices and is entirely managed by a custom software written in Matlab language. The main features of our system are: (1) the implementation of standard methods (the Read's rebreathing test, the CO2 single breath test and the transient hypoxia test) suitable for both pathological and healthy subjects, (2) data quality assurance and reduction of subjective judgment in the analysis through advanced analysis procedures and statistical outliers rejection, and (3) full interactive control of every step of the recording and analysis procedures.The system is currently used in our Institution in the assessment of CRS in HF patients, chronic obstructive pulmonary disease patients and healthy subjects. It has proven to be very effective and easy to use even by clinical personnel without a specific background in respiratory function assessment.