INTRODUCTION

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When premature infants with established respiratory distress syndrome (RDS) are treated with exogenous surfactant, arterial oxygen tension increases in association with an increase in functional residual capacity (1). However, the immediate improvement in gas exchange and lung volume is usually not accompanied by an increase in dynamic lung compliance measured during mechanical ventilation (2, 4), and some investigators (1, 3) have reported a decrease in dynamic compliance under these circumstances. These effects can be explained by a lung model of RDS which includes three compartments: one compartment of terminal airspaces, which are ventilated and stable, another compartment of terminal airspaces, which are ventilated but unstable, i.e., they collapse during expiration, and a third compartment of terminal airspaces, which are unventilated because they are either collapsed or fluid-filled (8). The immediate increase in functional residual capacity after "rescue" surfactant administration to patients with established RDS can be considered to be the result of two mechanisms: recruitment of new gas exchange units from the previously unventilated compartment, and stabilization of the two ventilated compartments at higher end-expiratory volumes. When end-inspiratory and end-expiratory airway pressures are held constant during mechanical ventilation, stabilization without recruitment will result in a decrease in lung compliance measured between the two pressures. Recruitment of terminal airspaces from a previously unventilated compartment has the opposite effect on compliance. The lack of improvement of compliance immediately following surfactant treatment at a time when gas exchange and lung volume have already improved suggests the possibility that a substantial compartment of unventilated terminal airspaces still exists. If so, mechanical recruitment of newly ventilated terminal airspaces from this compartment at the time of surfactant administration might enhance the immediate physiologic effects of exogenous surfactant treatment.

In order to test this hypothesis, the effect of a mechanical volume recruitment (VR) maneuver on response to surfactant treatment was evaluated in young rabbits made surfactant deficient by airway lavage. The mechanical VR maneuver employed was an increase of the ventilator peak inflating pressure (PIP), which was expected to recruit an additional compartment of terminal airspaces with critical opening pressures greater than those of airspaces which were already being ventilated at the baseline PIP. The effect of this VR maneuver on the response to surfactant treatment was assessed by serial measurements of gas exchange and lung function indices. Gas exchange was assessed by the ratio of arterial to alveolar oxygen tension (a/A ratio) and arterial PCO₂. The lung function indices measured were functional residual capacity (FRC), dynamic compliance of the respiratory system (C_rs), tidal volume (VT), and the alveolar portion of tidal volume (VA).

	METHODS

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Animal Preparation

To approximate lung volumes typical of human premature infants given surfactant treatment, 10 New Zealand white rabbits, 4-6 wk old and approximately 1 kg in weight (range 0.8-1.1 kg) were included in our study. Initially, 0.04 mg/kg atropine, 40-50 mg/kg ketamine, and 8-10 mg/kg xylazine were administered intramuscularly to provide anesthesia for placement of catheters and the tracheostomy. Tracheostomy was performed and the trachea was cannulated with an uncuffed 3.5-mm inner diameter endotracheal tube (ETT) and tightly secured in place by peritracheal ligature to prevent leak. Polyvinyl catheters were inserted into a common carotid artery and into an internal jugular vein. Anesthesia and muscle paralysis were maintained by continuous intravenous infusion of 25 mg/kg pentobarbital and 0.2 mg/kg pancuronium bromide per hour throughout the study period.

After placement of catheters and the tracheostomy, the rabbit was placed in a volume displacement body plethysmograph. All outlets for catheters and the ETT were sealed with silicone putty.

Airway Lavage and Mechanical Ventilation

We used a modification of the original lavage protocol described by Lachmann and coworkers (9). Airway lavage was preceded by an intravenous infusion of 4 µg/kg of isoproterenol over two minutes followed by increasing VT by 33% of baseline values for one minute to enhance surfactant release into small airways (10, 11). Each lavage involved the instillation and removal of 20-30 ml/kg of warmed normal saline via the ETT carried out over a 30 s period. Airway lavage was repeated every 5 min until both the a/A ratio decreased to < 0.12 and a PIP of > 20 cm H₂O was required to maintain VT at 10 ml/kg.

Mechanical ventilation was provided by time-cycled pressure limited infant ventilators (Sechrist IV-100B, Anaheim CA). During the sequence of lavage and for a one-hour stabilization period following the last lavage, FI_O2 was adjusted upward from 0.5-0.6 to 1.0 as necessary to keep the oxygen saturation measured by pulse oximetry above 80% if possible. Positive end-expiratory pressure (PEEP) was held constant at approximately 1 cm H₂O. Inspiratory time was held constant at 0.4 and ventilator rate was adjusted to a maximum of 75 bpm as necessary to maintain Pa_CO2 < 7 kPa (54 mm Hg), if possible. PIP was adjusted to keep VT at 10 ml/kg.

Gas exchange and lung function indices were measured prior to the first lavage (pre-lavage measurements) and intermittently during a 1-h stabilization period following the last lavage. The final measurements made during this stabilization period immediately prior to surfactant instillation were regarded as baseline measurements. Rabbits were included for analysis only if there was a decrease in FRC > 4 ml/ kg between the pre-lavage measurement and the baseline measurement in order to decrease the likelihood of including rabbits which did not have significant surfactant deficiency.

Experimental Protocol

Two groups of rabbits were studied. Group S (n = 5) received surfactant given as a 4-min infusion. Group SVR (n = 5) received a 4-min infusion of surfactant accompanied by a mechanical volume recruitment (VR) maneuver. The mechanical VR maneuver consisted of an increase in PIP of approximately 8 to 9 cm H₂O beginning 2 min after beginning the surfactant instillation and lasting for 4 min. The surfactant preparation used was Survanta (provided by Ross Laboratories, Columbus, OH) in a dose of 4 ml/kg instilled manually over 4 min into a sideport of the ETT adapter.

Additional measurements of gas exchange (a/A ratio and arterial PCO₂) and lung function indices (FRC, dynamic C_rs, VT, and VA) were made at 15, 30, 60, 90 and 120 min after the beginning of surfactant instillation. In addition, VT and dynamic C_rs were measured during the final minute of the mechanical VR maneuver in four of the SVR Group rabbits. Except for the 4-min period of mechanical VR, ventilator settings (rate and pressures) were held constant from the time of the baseline measurements immediately before surfactant instillation until the end of the study. After completion of these measurements at 120 min, the rabbit was killed with an overdose of pentobarbital.

Measurement of Gas Exchange and Lung Function Indices

Pa_O2, Pa_CO2 and pH were measured from blood samples taken from the carotid arterial catheter. a/A ratio was calculated as Pa_O2/[(713 · FI_O2)  Pa_CO2].

Airway flow measurements used to calculate FRC, C_rs, VT, and VA were obtained using the whole-body plethysmographic technique described by Sjöqvist and colleagues (12) and Edberg and coworkers (13). The airway flow signal was derived from a differential pressure transducer (Validyne model MP 45-14; Validyne Engineering Corp., Northridge, CA) and a Fleisch No. 1 pneumotachometer in the plethysmograph wall. The flow signal was compensated for the low-pass filtering influence of the plethysmograph. Absence of leak in the system was verified by comparing the flow measured when a flow of oxygen was injected into the plethysmograph and allowed to exit through the pneumotachometer with the flow measured when the same magnitude flow of oxygen was introduced directly through the pneumotachometer.

VT was obtained by integration of the compensated plethysmograph flow signal over a single breath. FRC and VA were measured using a multiple-breath nitrogen washout technique (14) employing a Med Science model 505 Nitralyzer to measure nitrogen concentration in respiratory gas mixtures. In animals requiring > 0.8 FI_O2, the FI_O2, was reduced to 0.8 for 2 min prior to nitrogen washout. Measured gas volumes reflected conditions of body temperature, pressure, and saturation. Dynamic C_rs was estimated by a least square method by fitting airway flow and tidal volume signals to proximal airway pressure using the standard equation of motion (12, 13).

Airway flow, pressure, and nitrogen concentration signals used for the measurement of the lung function indices were sampled at a rate of 200 Hz, digitized, and stored in a personal computer for subsequent analysis. Computer software for data acquisition and analysis was provided by Bengt Arne Sjöqvist and Ants R. Siberberg from Chalmers University of Technology, Gothenburg, Sweden.

Clinical Care

A radiant warmer was used to maintain a constant core temperature between 37.0 and 39.0° C, monitored by a rectal probe. Oxygen saturation was monitored continuously by pulse oximetry with the probe fixed to the rabbit's left hind leg. Heart rate and arterial blood pressure were monitored online by a pressure transducer and displayed on an 8-track chart recorder. In addition to these vital signs, VT, airway flow, and proximal airway pressure were displayed continuously on the chart recorder.

The protocol was approved by the Vanderbilt University Animal Care Committee in accordance with guidelines of the American Physiologic Society.

Statistical Methods

In order to establish comparability of the two experimental groups before surfactant instillation, pre-lavage and baseline measurements were compared between the two groups using a standard nonparametric method (Wilcoxon rank sum test). The average response of each of the gas exchange and lung function indices following surfactant instillation was assessed using a trapezoidal approximation for the area under the curve of the observations at 15, 30, 60, 90, and 120 min. This calculation was time-averaged in order to weight the observation at 15 min appropriately. Because there were only five animals in each group, which precludes achieving statistical significance for a paired analysis with a nonparametric test, we used a paired t test to determine whether a given time-averaged response was significantly different from baseline. We also used a paired t test to compare PIP, VT, and C_rs measured during the mechanical VR maneuver with baseline measurements of these variables. Repeated measures analysis of variance was used to assess differences between groups in change over time following surfactant instillation for the gas exchange and lung function indices.

Mean values are expressed ± 1 SD unless specified otherwise.

	RESULTS

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Comparability of Study Groups

The two groups were similar in regard to body weight (S: 0.98 ± 0.08 kg; SVR: 0.92 ± 0.08 kg) and number of lavages required (S: 3.6 ± 0.55 lavages; SVR: 3.0 ± 0.71 lavages). Ventilator settings, gas exchange, and pulmonary function indices were similar between the two groups prior to airway lavage and at baseline (Tables 1 and 2).

Mechanical Volume Recruitment (VR) Maneuver

PIP, VT, and C_rs measured during the final minute of the mechanical VR maneuver in the SVR group were all significantly increased over baseline (Table 3).

Response Following Surfactant Instillation

The changes from baseline in gas exchange and pulmonary function over the two hours following surfactant instillation are shown in Figure 1 for the two study groups. In the S group, the only significant response was an increase in a/A ratio. In contrast, there were significant responses observed in the SVR group for all indices: a decrease in Pa_CO2 and an increase in a/ A ratio, FRC, dynamic C_rs, VT, and VA. There were also significant differences between the S and SVR subgroups in the change over time of FRC, C_rs, and VT following baseline. These results suggest an enhancement of response when surfactant was administered in conjunction with a VR maneuver when compared with surfactant administration without a VR maneuver.

View larger version (18K): [in this window] [in a new window]   Figure 1.   Responses in gas exchange and pulmonary function indices following intervention are shown according to study group: S = surfactant alone; SVR = surfactant + volume recruitment maneuver. The mean and standard error of the change from baseline of each index is shown at 15, 30, 60, 90, and 120 min into the response period for each of the study groups. The statistical significance of a difference between the time-averaged area under the curve across the five observations and the baseline value for a variable is shown by asterisks over the corresponding study group. The significance of a difference between the S and SVR groups in the change over time of a variable is shown by the bracket and asterisk beneath the study groups (*p < 0.05, **p < 0.01, ***p < 0.001).

	DISCUSSION

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This study was designed to test the hypothesis that the immediate effect of surfactant treatment on gas exchange and lung function can be enhanced if a mechanical volume recruitment maneuver is used in conjunction with surfactant administration.

A significant improvement was observed in all gas exchange and lung function indices following surfactant when administered in conjunction with a mechanical VR maneuver (Figure 1). When surfactant was administered without a VR maneuver, the only significant response was an increase in a/A ratio. There were also significant differences detected between these two study groups in the change over time of FRC, C_rs, and VT following baseline. These findings are consistent with the hypothesis that the immediate physiologic effects of exogenous surfactant treatment can be enhanced by mechanical volume recruitment provided at the time of surfactant instillation.

The VR maneuver employed in this study was designed to expose a compartment of collapsed or liquid-filled terminal airspaces to intermittent peak pressures which exceeded their critical opening pressures. By applying the increased PIP during and immediately following surfactant instillation, we anticipated that those terminal airspaces newly recruited and ventilated at the higher PIP, having become stabilized by exposure to the instilled surfactant, would remain recruited and ventilated after return of PIP to baseline level. The effectiveness of the mechanical VR maneuver in actually increasing the number of ventilated terminal airspaces was assessed by comparing C_rs during the final minute of the VR maneuver with baseline C_rs (Table 3). If the increase in VT accompanying the VR maneuver had been only the result of further inspiratory distention of terminal airways already being ventilated, C_rs would have remained the same or decreased. The increase in C_rs observed implies that additional parallel terminal airspaces were being ventilated during the VR maneuver.

Alveolar instability refers to the tendency for an alveolus to switch abruptly between the inflated state and the collapsed state (15). Inflation of an unstable alveolus occurs suddenly when transalveolar pressure exceeds the critical opening pressure. During deflation, an unstable alveolus collapses abruptly when transalveolar pressure decreases below the critical closing pressure. When unstable alveoli have been stabilized by surfactant repletion, they deflate progressively without collapse so that they retain gas volume at end-expiratory pressures below their critical closing pressures (15, 16).

The lung model on which our hypothesis is based assumes the presence of three compartments which differ in ventilation and in stability: (1) a compartment made up of terminal airspaces, which are not ventilated; (2) a compartment made of terminal airspaces that are ventilated, but are unstable, i.e., they collapse during expiration; and (3) a compartment made up of terminal airspaces that are ventilated and do not collapse during expiration. The inhomogeneity of ventilation implied by this multi-compartmental model has been previously demonstrated by analysis of passive exhalation lung mechanics of the respiratory system in preterm lambs (17) and by analysis of multiple-breath nitrogen washout curves obtained from newborn human infants with severe RDS (18). In addition, inhomogeneity of ventilation demonstrated by indicator gas washout analysis in premature lambs (19) and in premature infants with RDS (20) has been shown to be normalized by surfactant treatment.

Our hypothesis is also based on the assumption that the immediate effect of instilled surfactant is exhibited mainly by those terminal airspaces which are already ventilated at the time of surfactant administration. Previously reported studies (21) have demonstrated that under conditions in which inhomogeneity of distribution of instilled surfactant is exaggerated, surfactant distribution is correlated with distribution of ventilation. That distribution of ventilation can determine the distribution of instilled surfactant was demonstrated in adult sheep with lung injury induced by airway lavage with and without the right upper lobe protected from lavage (23). In animals which had the right upper lobe spared from injury there was preferential deposition of exogenous surfactant into the right upper lobe. Based on these observations, it is reasonable to expect that mechanical recruitment of terminal airspaces from a previously unventilated compartment at the time of surfactant administration would facilitate the distribution of surfactant to this compartment.

This study was designed to assess the effect of a mechanical VR maneuver employed at the time of surfactant administration. Other investigators have reported the effects of different methods and strategies of mechanical ventilation initiated and continued after surfactant instillation (24). A mechanical VR maneuver is commonly employed at the time of surfactant instillation, both in experimental studies and in clinical practice. The Survanta package insert recommends manual ventilation with sufficient positive pressure to provide adequate air exchange and chest wall excursion. The use of a mechanical VR maneuver in connection with surfactant instillation apparently has been empirically based. We are not aware of any previously published study in which the rationale and effectiveness of a VR maneuver in this context have been examined.

The end-expiratory pressure used in this study was lower than the level commonly used when premature infants with RDS are mechanically ventilated. However, the purpose of this study was to test an hypothesis regarding a proposed model of surfactant treatment effect rather than to evaluate a clinical intervention. Nevertheless, the results of this study do support the concept that some form of volume recruitment maneuver such as high frequency ventilation or sustained inflations (29) might be effective in a clinical context of surfactant replacement therapy.

Rabbits which received surfactant treatment in conjunction with a mechanical VR maneuver showed a significant improvement in all measured gas exchange and lung function indices. Rabbits in the group which received surfactant without the maneuver did not show improvement in any of these indices except a/A ratio. A beneficial effect of the VR maneuver was also seen when changes over time in FRC, C_rs, and VT following surfactant replacement were compared between these two study groups. These results support the hypothesis that mechanical recruitment of terminal airspaces from a previously unventilated compartment will enhance the effectiveness of surfactant replacement therapy by facilitating the distribution of instilled surfactant to this compartment.