INTRODUCTION

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Repetitive episodes of partial or complete upper airway (UA) closure during sleep are a characteristic feature of the obstructive sleep apnea-hypopnea syndrome (OSAHS). The resulting sleep fragmentation, nocturnal oxyhemoglobin desaturation, and periodic increases in sympathetic nerve activity and in systemic arterial pressure can account for the OSAHS- related increase in morbidity and mortality (1, 2). These episodes of UA closure result from an imbalance between the dilating and collapsing forces that apply on UA structures, and also depend on UA compliance, and shape and dimension (3). UA dilating muscles play an unique role in the maintenance of UA patency since they represent the only UA dilating force. They also contribute to determine UA compliance (4). UA dilators activity physiologically precedes that of inspiratory muscles with a steeper rate of rise, thus stabilizing UA during inspiration, decreasing UA resistance, and reducing the work of breathing (5). This preactivation pattern is observed in normals and OSAHS patients during wakefulness (6) and is maintained during sleep in subjects without apnea (5). Sleep-related obstructive breathing disorders are accompanied by a loss of UA muscles preactivation pattern (6). The importance of UA characteristics and of the fine neurophysiological tuning between inspiratory and UA muscles coordination is clearly illustrated by the phenic nerve pacing induced UA collapse during sleep in subjects previously without apnea (7, 8). Indeed, with phrenic pacing, diaphragmatic contractions are by definition independent of any previous UA muscles prestimulation.

We have recently shown that phrenic nerve stimulation can mimic, in normal awake subjects, the conditions associated with UA obstruction during sleep by dissociating UA and inspiratory muscles activation (9). In this experiment, electrical phrenic nerve stimulation and cervical magnetic stimulation (ES and CMS, respectively) performed at end expiration during exclusive nasal breathing induced typical flow-limited breaths. The twitch flow dynamics was different between ES and CMS, and the twitch-induced inspiratory flow limitation pattern was not accounted for by partial nares closure. No genioglossus muscle activity preceded the twitch-related inspiratory flow. From these results, we believe that phrenic nerve stimulation could be used to characterize the dynamics of the passive UA and thus be a useful tool for the management of OSAHS patients. However, our previous study was conducted in normal subjects free of nocturnal breathing abnormalities. The applicability of phrenic nerve stimulation to document flow limitation in patients with OSAHS during wakefulness has not been determined yet. The aim of the present study was to evaluate the feasibility of this method to characterize the twitch flow response in such patients, and to assess different phrenic nerve stimulation techniques in this setting.

	METHODS

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Eight male patients with characteristic clinical features and in whom OSAHS had previously been identified with a conventional polysomnographic sleep recording were included in the study. All of them were treated with continuous positive airway pressure (CPAP) at home. The study was conducted according to the French legislation on human biomedical research ("Comité Consultatif de Protection des Personnes se prêtant à des Recherches Biomédicales, Pitié-Salpêtrière") and each patient signed an informed consent form.

Surface recordings of the right and left costal diaphragmatic electromyogram (EMG) activity were obtained using silver cup electrodes placed on the anterior axillary line in the seventh to eighth right and left intercostal spaces and connected to an electromyograph (Nihon Kohden Neuropack Sigma/Nihon Kohden, Tokyo, Japan). An esophageal balloon was inserted through one nostril following local anesthesia (xylocaine 2% spray) and positioned into the lower third of the esophagus, according to the occlusion test (10). An internal nasal stent (Nozovent; WPM international AB; Göteborg, Sweden) was placed to prevent nasal collapse. A nasal CPAP mask was placed over the nose, its airtightness being assessed during a maximal inspiratory effort against occlusion. The mask was opened to room air through a pneumotachograph (Hans Rudolph, Kansas City, MO). A recording of diaphragm EMG response (M-wave), esophageal pressure referenced to mask pressure, mask pressure, and instantaneous flow was obtained during 1 s following stimulation at 50 kHz sampling rate.

Stimulations

Patients were seated in an armchair with their head maintained in the neutral position by headrests. The investigators paid special attention to avoid any change in body, neck, or head position during the experiment. Stimulations were always performed at end expiration during quiet breathing. All measurements were done with the subject breathing exclusively by the nose.

CMS was performed in all patients using a Magstim 200 stimulation unit (Magstim; Whitland, Dyfed, UK) equipped with a standard circular 90-mm coil (maximum output 2.5 Tesla, pulse duration 0.1 ms) used according to the previously reported technique (11). The coil was centered over the spinous process of the seventh cervical vertebra (C7). The adequate positioning of the electrodes and magnetic coil was checked and if necessary readjusted to obtain a maximal M-wave.

In all patients, attempts were made to stimulate the phrenic nerves focally or quasi-focally in the neck using either bilateral ES (four patients) or bilateral anterior magnetic phrenic nerve stimulation (BAMPS, eight patients) according to Mills and coworkers (14). ES was tried out in the first four patients, according to the technique described in the literature (15). The phrenic nerve was looked for at the posterior border of the sternocleidomastoid muscle, at the level of the cricoid cartilage, using two bipolar electrodes with saline-soaked felt tips 5 mm in diameter with 2 cm between electrodes with 0.1 ms square wave pulse. A low-intensity ES stimulation train at 1 Hz was used to localize the right and left phrenic nerves. Then intensity was increased in an attempt to build a recruitment curve, with the aim to achieve a supramaximal stimulation. In no case were we able to obtain adequate M-wave recordings at maximal stimulation intensity despite adequate positioning of the surface electrodes as assessed by inspiratory bursts of EMG activity during tidal inspiration. These technical difficulties were probably related to the morphology of the patients' necks. Therefore, in the subsequent four patients, we resorted to BAMPS only for focal phrenic nerve stimulation. BAMPS was performed using two identical Magstim 200 units similar to the one used for CMS, linked to be simultaneously discharged by one external trigger. One unit was equipped with a figure-of-eight coil similar to the one used by Mills and coworkers (14) (two 43-mm coils mounted on a 90° handle), whereas the other unit was equipped with a single 40-mm coil. The coils were positioned at locations similar to those used for ES and oriented to obtain an optimal M-wave recording.

Ten stimulations were obtained at maximal stimulator output (Imax) with CMS and BAMPS. Then the intensity was progressively decreased using 5% steps, until M-waves disappeared. At least five stimulations were performed at each intermediate level. Focal stimulation and CMS were applied in random sequence.

Five additional stimulations were also obtained with BAMPS at Imax while the subjects had been on CPAP for at least 5 min with the positive pressure level that was prescribed according to a previous titration sleep study.

Data and Statistical Analysis

Resistance of the respiratory system was calculated at peak twitch flow and at peak twitch pressure, as the ratio of driving pressure (esophageal-mask pressure) to twitch flow. Twitch-induced inspirations were defined as flow limited when twitch flow plateaued or decreased in spite of a continuing simultaneous increase of the driving pressure. For tidal breaths and following each stimulation, we collected the following variables: maximal esophageal pressure, peak twitch flow, total respiratory system resistance calculated at peak twitch flow and at peak esophageal pressure, and maximal twitch flow of flow-limited twitches. The influence of the technique of stimulation on each of these variables was studied using analysis of variance for repeated measures. Their change with varying stimulation intensity was analyzed with a regression analysis. In all cases, the threshold for statistical significance was set at 0.05. Values reported in the results section are mean ± SD.

	RESULTS

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The anthropometric characteristics and the results of sleep studies of our patients are described in Table 1. Twitch-related inspiratory flow was produced by both CMS and BAMPS. However, the driving pressure reached at Imax differed in each condition (Figure 1), the peak esophageal pressure at Imax being higher with BAMPS than with CMS in seven of eight patients; for the whole group, the driving pressure measured at Imax was 13.7 ± 11.0 cm H₂O with BAMPS and 6.7 ± 4.3 cm H₂O with CMS (p = 10⁴). No difference was found in peak esophageal pressure values between tidal breathing (12.0 ± 3.9 cm H₂O) and BAMPS, but esophageal pressure was significantly less during CMS than during tidal breathing (p = 10⁴). A positive relationship between the resistance of the respiratory system measured at peak twitch pressure and stimulation intensity was observed in seven of eight patients with BAMPS, but in no patient with CMS (Figure 2). These resistance values measured at Imax were significantly higher with BAMPS (33.1 ± 4.4 cm H₂O/L/s) than with CMS (12.2 ± 4.5 cm H₂O/L/s, p = 0.001). Peak twitch flow measured at Imax was 0.8 ± 0.5 L/s with BAMPS and 0.7 ± 0.4 L/s with CMS (p = 0.1).

View larger version (12K): [in this window] [in a new window]   Figure 1.   Individual values of the maximal esophageal pressure reached at 100% stimulation intensity with CMS and BAMPS. These values represent the maximal subatmospheric pressure value that was reached following twitches. For the whole group, the maximal esophageal pressure was significantly higher with BAMPS than with CMS.

View larger version (28K): [in this window] [in a new window]   Figure 2.   Individual relationship between total respiratory system resistance measured at peak twitch pressure and stimulation intensity with BAMPS and CMS. A significant positive relationship was observed in all but one patient with BAMPS whereas no relationship was noted with CMS. The regression line is indicated when the relationship is significant.

Both resistance of peak flow (RPF) and resistance of peak pressure (RPP) dramatically increased during posterior cervical and bilateral anterior magnetic stimulation; in response to CMS, RPF and RPP were 233 ± 82%, and 443 ± 187% of the values measured during spontaneous breathing, respectively; in response to BAMPS, RPF and RPP were 344 ± 76% and 677 ± 248%, respectively (Figure 1).

Characteristic twitch flow limitation was observed with one magnetic stimulation method or both in seven of eight patients (Figure 3). In these seven patients, 69.0 ± 30.3% of twitches with BAMPS and 32.3 ± 37.1% of twitches with CMS were flow limited (p = 0.06). The minimal stimulation intensity at which flow limited twitches occurred was 73.1 ± 16.7% of Imax with BAMPS and 93.3 ± 6.8% Imax with CMS (p = 0.01). VI_max values of flow-limited twitches obtained at Imax were 0.81 ± 0.5 L/s with BAMPS and 0.87 ± 0.5 L/s with CMS (p = 0.4). Except in one patient, no correlation was found between VI_max of flow-limited twitches and stimulation intensity; in this individual, a positive relationship between VI_max of flow-limited twitches and stimulation intensity was observed with BAMPS only (R = 0.88, p = 10⁴). To assess the influence of stimulation intensity on UA collapsibility, we looked at the influence of stimulation intensity on the esophageal pressure at which flow limitation begun (namely, the pressure at which a plateau or a decrease in twitch flow was observed despite increasing esophageal pressure). A positive relationship was found between these variables in six of eight patients with BAMPS and in two of eight with CMS, suggesting the occurrence of UA closure at a lower (more negative) pressure at low than at high stimulation intensity. No relationship was found between VI_max of flow-limited twitches and the corresponding esophageal pressure with increasing stimulation intensity in any of our patients.

View larger version (19K): [in this window] [in a new window]   Figure 3.   Representative example of the instantaneous flow and esophageal pressure tracings during spontaneous breathing (A) as well with M-wave tracing obtained with BAMPS (B) in the same patient illustrating the presence of a flow-limited twitch. Pes = esophageal pressure; Rdi and Ldi = right and left diaphragm.

CPAP significantly altered the results of BAMPS. As illustrated in Table 2, the percentage of flow-limited twitches, the maximal esophageal pressure level, and peak pressure resistance significantly decreased with CPAP. No difference was found in peak flow resistance with and without CPAP.

	DISCUSSION

TOP ABSTRACT INTRODUCTION METHODS RESULTS DISCUSSION REFERENCES

These results demonstrate that BAMPS can be used in patients with OSAHS during wakefulness to mimic the abnormal flow pattern through passive UA that is characteristic of their disease. Indeed, BAMPS, provoking diaphragm contractions dissociated from UA dilating muscles activity, induced a high proportion of flow-limited breaths, and a dramatic increase in UA resistance. Other phrenic nerve stimulation methods may not all achieve the same result.

Limits of Phrenic Stimulation Techniques

We have recently shown that both ES and CMS can be used to induce an increase in UA resistance and flow-limited twitches in normal awaked subjects (9). In the patients with OSAHS studied here, we were unable to obtain reliable diaphragmatic stimulation with ES in the four patients in whom it was attempted, despite adequate diaphragmatic EMG signals recorded during natural breathing. We believe that this technical failure is due to the important amount of subcutaneous adipose tissue increasing the skin to phrenic tissular resistance and thus preventing the depolarization of the phrenic nerves. This limitation could theoretically have been bypassed by increasing the duration of the electrical stimulus to increase the amount of current delivered. Nevertheless, this would have precluded any systematic comparison between magnetic and electrical stimulation techniques. In addition, although this has not been precisely evaluated, the tolerability of a prolonged electrical stimulus is likely to be much lower than this of a shorter one. The need to resort to such an alternative to overcome tissue resistance would thus probably further jeopardize ES, already more technically demanding than CMS or BAMPS, as a clinical tool to document flow dynamics in passive UA during wakefulness. This makes magnetic stimulation techniques more adequate for this purpose.

However, it is interesting to note that BAMPS and CMS did not provide similar levels of driving pressure. Furthermore, total respiratory system resistance plateaued with increasing CMS intensity while a progressive increase in resistance was observed with BAMPS, and flow-limited twitches were observed at lower stimulation intensity with BAMPS than with CMS. These differences are possibly due to a lesser degree of diaphragm activation with CMS as compared with BAMPS. It is actually conceivable that in obese patients, the magnetic field produced by CMS is significantly attenuated when it reaches the phrenic nerves due to the loss of energy in neck adipose tissues (16). This should not be the case with BAMPS where the magnetic field is, even in the case of obesity, produced very close to the phrenic nerve. This can account for the fact that minimal stimulation intensity at which flow limitation was observed was less with BAMPS than with CMS and that the esophageal pressure values were similar during tidal breathing and BAMPS but were less with CMS. However, due to the differences in driving pressure obtained with these two magnetic stimulation methods and considering the differences in UA flow dynamics previously observed between ES and CMS, we cannot rule out the possibility of an activation of UA dilator muscles by CMS to explain these findings.

Difference between UA Dynamics Assessed during Sleep and with PNS during Wakefulness

At the highest stimulation intensities phrenic nerve stimulation-induced twitches had a profile characteristic of flow limitation. The VI_max levels obtained in these conditions are significantly higher than those reported in the literature in normal subjects and patients with sleep apnea-hypersomnolence syndrome (SAHS). This can be accounted for by two different interdependent factors. One relates to the magnitude of twitch-induced negative intrathoracic pressures that exceeds by far that of normal inspiratory efforts; this can be expected by the fact that BAMPS and CMS induced supramaximal neuromuscular stimulation. The other concerns the determinants of upper airway collapsibility and particularly the factors influencing the pressure threshold at which flow limitation begins (Pclose). During tidal breathing, the preactivation of UA dilator muscles that precedes the contraction of inspiratory muscles stabilizes the UA and prevents flow limitation (no Pclose value). In sleeping patients with SAHS, the loss of these UA muscles preactivation contributes to destabilize UA leading to obstructive breathing events (hypopneas and apneas) that share the characteristic flow limitation pattern. The importance of UA muscles preactivation on flow regimen is supported by the occurrence of flow-limited twitches in normal awake subjects during phrenic nerve electrical stimulation (9). Therefore, in this condition, Pclose has a subatmospheric value that determines the severity of flow limitation: the more negative Pclose, the higher VI_max. The fact that flow-limited breaths can be induced during wakefulness using phrenic nerve stimulation indicates that the twitch-induced pharyngeal pressure is below Pclose (supramaximal stimulation), whereas high VI_max levels recorded during wakefulness indicate that Pclose is more negative during wakefulness than during sleep. It is known that the sleep-induced decrease in UA muscle EMG tonic activity is accompanied by a dramatic decrease in UA patency independently of any change in their phasic activity (17). Therefore, differences in tonic skeletal muscles activity between wakefulness and sleep may account for the more negative Pclose values observed in awaked subjects. Therefore, it is obvious that the UA dynamics characteristics that we measure using phrenic nerve stimulation in awake subjects is not identical to that of sleeping subjects, but this method mimics during wakefulness neurophysiological abnormalities that are observed only during sleep.

Twitch Flow Dynamics in Patients with OSAHS

The potential interest of phrenic nerve stimulation is to allow the study of passive upper airway during wakefulness. We have recently found that the genioglossus muscle EMG activity effectively follows the airway pressure drop with electric twitch stimulation in normal subjects (9). It is interesting to note that in the present study even the esophageal pressure levels that were reached following both magnetic stimulation methods were identical or less than tidal inspiratory efforts, these twitches were characterized by a significant increase in respiratory resistance and the occurrence of flow limitation. We believe that this demonstrates that phrenic nerve stimulation-induced airflow obstruction developed because upper airway properties differ during unstimulated and stimulated breaths, probably as a consequence of the occurrence of inspiratory efforts without previous activation of upper airway dilating muscles. We found that the closing pressure defined as the esophageal pressure at which flow limitation occurs decreased with increasing BAMPS intensity in most of our OSAHS patients, and that this was not accompanied by a rise in VI_max. According to previous studies conducted in collapsible conduits (18), the slope of the flow/pressure relationship preceding flow limitation is inversely proportional to the sum of the resistance of the segments upstream and downstream to the point of UA collapse, and the pressure at which flow limitation occurs is inversely proportional to downstream resistance. Therefore, the decrease in closing pressure with increasing stimulation intensity can be accounted for by an increase in downstream resistance. Since VI_max of flow-limited events was inversely proportional to upstream resistance and did not change with changing stimulation intensity, it can be assumed that upstream resistance did not change in these conditions. This indicates that the increase in total respiratory resistance that we observed with increasing stimulation intensity can be attributed to a simultaneous increase in resistance at the collapsing site but also downstream to this level.

Comparison with Normal Subjects

Since the aim of the present study was to evaluate the feasibility of different phrenic stimulation methods in OSAHS, we did not use a control group. Nevertheless, although the normal subjects included in our previous study were younger and did not have focal phrenic stimulation with BAMPS but with ES only, it is interesting to compare the results of these two studies. The total resistance measured at peak pressure obtained in the present OSAHS study population was 33.1 ± 4.4 cm H₂O/L/s. This figure is dramatically higher than that obtained with ES at the same anatomical site in normal subjects (11.4 ± 3.8 cm H₂O/L/s). Furthermore, even if the percentage of flow-limited twitches does not seem to differ between ES and BAMPS in normal subjects and in patients with OSAHS, respectively (71.5 ± 27.2 and 69.0 ± 30.3%), the VImax value of flow-limited twitches appears to be less in patients than in normal subjects(1.2 ± 0.3 L/s). This justifies the need for a controlled study comparing flow dynamics with increasing stimulation intensity with BAMPS in patients with OSAHS and in normal subjects. Such a study would be useful to determine the normal physiological characteristics of twitch-induced flow and to analyze the relationship between these characteristics and sleep-related obstructive breathing disorders.

Effects of CPAP and Perspectives

Interesting results were obtained with CPAP using BAMPS at maximal stimulation intensity. The reduced percentage of flow-limited twitches, the decrease in twitch flow esophageal pressure, and the decrease in resistance observed at the measured effective pressure level can be accounted for by the pneumatic splinting effect of CPAP (21). It can therefore be speculated that phrenic nerve stimulation in general, and BAMPS in particular, could theoretically be used to determine the effective pressure level during the day. However, it is not currently possible to determine what parameters should be used and what values of these parameters should be reached to accurately determine the effective therapeutic pressure value using BAMPS.

Naturally, this reasoning could be expanded to other therapeutic strategies such as mandibular anterior prosthesis, to determine the protrusion distance that suppresses sleep-related obstructive events. Further prospective studies have to be conducted to specifically examine this issue, which may open the field of mechanical devices titration studies realized during wakefulness.

We conclude that BAMPS allows the characterization of the dynamics of flow through the passive UA in awaked patients with OSAHS. This technique could provide very useful information on UA dynamics during wakefulness with several clinical (determination of effective pressure setting for home CPAP therapy) and physiological (measurement of UA critical pressure) applications.