Method for estimating absolute at constant inflation pressure B. A. HILLS AND R. E. BARROW Department of Physiology and Biophysics, Galveston, Texas 77550
University
HILLS, B. A., AND R. E. BARROW. Method for estimating absolute lung volumes at constant inflation pressure. J. Appl. Physiol.: Respirat. Environ. Exercise Physiol. 47(4): 907-909, 1979.-A method has been devised for measuring functional residual capacity in the intact killed animal or absolute lung volumes in any excised lung preparation without changing the inflation pressure. This is achieved by titrating the absolute pressure of a chamber in which the preparation is compressed until a known volume of air has entered the lungs. This technique was used to estimate the volumes of five intact rabbit lungs and five rigid containers of known dimensions by means of Boyle’s law. Results were found to agree to within &I% with values determined by alternative methods. In the discussion the advantage of determining absolute lung volumes at almost any stage in a study of lung mechanics without the determination itself changing inflation pressure and, hence, lung volume is emphasized. functional
residual
capacity
IN MANY STUDIES of pulmonary mechanics it is desirable
to measure not only the volume change for a given inflation pressure in the intact lung or isolated lung preparation, but also the absolute volumes involved. For instance, determinations of functional residual capacity (FRC) in a killed animal and the absolute volume when its lungs are subsequently excised and reinflated help in relating measurements of compliance, airway closure, and lung integrity in general to the normal physiological state of the animal. Two principles have been employed in such absolute volume measurements: dilution techniques and various pneumatometric approaches using the compressibility of gasesas expressed by Boyle’s law. Current dilution techniques do not vary appreciably from the original method of Darling et al. (3) but still tend to suffer from one of two problems -absorption of the tracer by blood in the live situation or the difficulty of obtaining uniform distribution in the dead animal. Pneumatometric approaches are of two types depending upon whether the lungs are closed or open to the surrounding atmosphere. The closed type is based on measuring pressure differentials generated by voluntary respiratory effort (5, 9)) 0~~~-75C;7/79/oooO-0$01.25
Copyright
0 1979 the American
lung volumes
Physiological
of Texas Medical
Branch,
which is, therefore, only applicable to live subjects, whereas those classified as “decompression” methods (5) essentially measure the volume exhausted from the lung when ambient pressure is reduced. There are many variations (4, 6, 11) of the latter approach that are good for a lung in the intact thoracic cavity where there is no alveolar-atmospheric pressure differential. These methods are thus well suited to the determination of FRC in the relaxed subject and can easily be adapted to the dead intact animal. However, this advantage does not hold for the isolated lung preparation, because it is particularly undesirable (8,lO) to allow a lung to collapse to minimal volume after excision from the cadaver or sacrificed animal. Even if the lungs are removed with the trachea clamped and compliance studied from the relationship between change in volume and change in inflation pressure, it is highly desirable to be able to redetermine the absolute volume after any cycle in order to relate the findings to the original FRC. Hence a variation of the decompression method has been devised that enables the absolute lung volume to be determined for any inflation pressure. It has the advantage that the same technique can also be used for the particular case of zero inflation pressure in the intact thoracic cavity to determine FRC before the lung is excised. There is a further advantage in that it can be used to determine the absolute volume at any value selected for the constant inflation pressure whether flow is inward (compression) or outward (decompression). MATERIALS
AND
METHODS
The principle used involves measuring the increase in ambient pressure needed to cause a known volume of air to flow into the lung without changing the transmural pressure differential maintaining it at a particular degree of inflation. The intact or isolated lung preparation is intubated and measurements of inflation pressure and volume changes appropriate to the particular study are made via valve X with Y closed (see Fig. 1). Let us consider the stage in a study where we need to know the absolute lung Society
907
Downloaded from www.physiology.org/journal/jappl by ${individualUser.givenNames} ${individualUser.surname} (129.186.138.035) on January 21, 2019.
908
B. A. HILLS
Mercury
AND
R. E. BARROW
manometer
L
LUI
my
r I cpal
ailul
I
;
\
\ \ \
v Volume:
Impressed-air
Pipette I I \ \
\
c
; I
” Volume:
I \ I LT
; i i ;: II *. p
SUPPlY Pressure FIG.
1. Apparatus
depicting
water
level after
compression
of chamber
volume without further change of inflation pressure. Valve Y is then opened. With the desired inflation pressure still applied to the lung via X, the water reservoir is raised or lowered until the water reaches the mark L in the pipette. The water level in the reservoir must then be above the mark L by a vertical height (H), which now equals the inflation pressure. Valve X is then closed. The whole preparation is placed in the chamber and the pressure raised with compressed air. The absolute pressure (P) is titrated until at an increase (AP) at which the water level in the pipette has moved from the mark L to the mark N (see Fig. 1). In so doing, the water has displaced the accurately known pipette volume of air (v). The whole sequence of events is reversed on decompression after which valve Y can be closed and X opened. This enables the lung study to continue without any change of inflation pressure during the determination of absolute lung volume (V). V is then calculated by applying Boyle’s law to the dry air whose partial pressure is (PO+ H - Pw) before compression and (PO+ AP + H Pw) afterwards (PO+ H - Pw)(V + v + E) = (PO+ P + H - Pw)(V + c) This reduces to give V as V = v(Po + H - Pw)/AP - E
(1)
where PO is atmospheric pressure, Pw is water vapor pressure at ambient temperature, and c is the volume between the lung and the mark N on the pipette. E can be determined directly, while v is selected to be about 25-35s of the anticipated value for V, so that AP is less than one-third atmosphere. This comparatively small pressure change enables a wide range of laboratory vessels to be used as the chamber; any small leaks in it do not affect the result. The pipette needs to be a flat vessel or a horizontal coil of tubing such that the vertical height between marks L and N is negligible with respect to H. The pipette is so arranged to prevent air trapping. The relationship depicted by Eq. 1 is plotted in Fig. 2 for the apparatus used in this study where v = 7.56 ml, c = 3.15 nil, and H = 0 on a day when atmospheric pressure (PO) was 756 Torr.
Chamber
by AP for a lung
maintained
at a constant
inflation
pressure
H.
RESULTS
Volumes of five lengths of rubber tube and five rabbit lungs determined by alternative methods are plotted against the respective value of AP obtained by this method in Fig. 2 in which the resulting points are compared with the Boyle’s law relationship (Eq. 1). Absolute volumes of the rubber tubes with one end closed were determined by weighing them empty and then full of water. Lung volumes were determined by saline immersion after excising them with the trachea clamped. Allowance was made for parenchymal tissue volume, which was estimated from lung weight using a specific gravity of 1.0 gem-” (2) (see Table 1). These data are presented as points in Fig. 2 where it can be seen that the rubber tube and lung volumes are within tl% of the theoretical curve. The use of fixed containers enabled us to test the method well beyond the normal range of lung volumes. Rates of chamber pressurization were very low (94 Torr. min-‘) to avoid any significant intra-airway pressure gradients. DISCUSSION
The foregoing evaluation indicates that this technique of compressing without changing inflation pressure offers an accurate and convenient means of measuring the absolute lung volume of intact lung preparations and should be equally applicable to excised lungs. Moreover, such measurements can be made at any stage in a study of pulmonary mechanics without changing the schedule of inflation pressure to be followed by that lung. This avoids the need to reevaluate the mechanical status of the lung that would have been necessary if the method used had changed the inflation pressure. This is a particular advantage in view of the complex dependence of lung compliance upon its pressure-volume history (1, 7, 12). The authors would like to thank Mrs. Laura Huddleston for her assistance in preparing the manuscript. The research reported here has been supported under the Office of Naval Research Contract NOOO14-75-C-1035 with funds provided by the Naval Medical Research and Development Command. Received
11 October
1978; accepted
in final
form
1 June
1979.
Downloaded from www.physiology.org/journal/jappl by ${individualUser.givenNames} ${individualUser.surname} (129.186.138.035) on January 21, 2019.
ESTIMATING
ABSOLUTE
LUNG
909
VOLUMES
320
282
-
CALCULATED MEASURED MEASURED
0 A
VOLUMES (EQ. 1) TUBING VOLUME LUNG VOLUMES
FIG. 2. Volumes of excised lungs (V) estimated by immersion technique and known volumes of rigid containers are compared with values at the same titrated inflation pressure (AP) used to determine V by the Boyle’s law relationship (Eq. I) shown as a solid line.
15
20
25 Absolute
30 Volume
40
35
(V) in ml
1. Comparison of volumes (FRC) determined from AP with values measured by alternative methods
TABLE
Alternative Animal
Rabbit lungs/immersion calibration
wt, kg
Vol
displaced, ml
Method *
Tissue
AP, Method wt, g
3.2 3.6 3.5 4.1
75.0 60.0 65.0
42.9
95.0
3.9
85.3
58.8 51.1
FRC,
functional
residue
capacity.
* Isolated
29.2
lungs.
t Intact
No. of runs
AP, Torr
31.8 36.0 34.2
4 4 4 4 4
157 168 162 142 150
32.2 k 29.9 k 31.2 t 36.0 t, 33.9 t
0.4 0.3 0.3 0.5 0.4
15.3 20.1 25.0 30.0 36.1
4 4 4 4 4
301 243
15.3
0.1
198
25.0 29.2 36.0
32.9 29.8
31.0
Rubber tubing/ weight calibration
FRC,
ml
lungs;
(Eq. I)
values
are means
172 142
Calculated
FRCt,
t
ml
19.8 t 0.2 k 0.2 t 0.3 t 0.4
t SD.
REFERENCES 1. AGOSTONI, E., AND J. MEAD. Statics of the respiratory system. In: Handbook of Physiology. Respiration. Washington, DC: Am. Physiol. Sot., 1969, sect. 3, vol. I, chapt. 13, p. 389-407. 2. CROSFILL, M. L., AND J. G. WIDDICOMBE. Physical characteristics of the chest and lungs and the work of breathing in different mammalian species. J. Physiol. London 158: l-14, 1961. 3. DARLING, R. C., A. COURNAND, AND D. RICHARDS. Studies on the intrapulmonary mixture of gases. III. An open circuit method for measuring (residual) air. J. CZin. Invest. 19: 609, 1940. 4. DEJOURS, P., AND H. RAHN. Residual volume measurements by the gas expansion method and nitrogen dilution method. J. Appl. Physiol. 4: 445-448, 1953. 5. DUBOIS, A. B., S. Y. BOTELHO, G. N. BEDELL, R. MARSHALL, AND J. H. COMROE, JR. A rapid plethysmographic method for measuring thoracic gas volume: a comparison with nitrogen washout method for measuring functional residual capacity in normal subjects. J. CZin. Invest. 35: 322-326, 1956.
6. HARVEY, 7.
8.
9. 10.
11.
12.
R. B., AND J. A. SCHILLING. Measurements of lung volume by rapid decompression. J. AppZ. Physiol. 7: 496, 1955. HILDEBRANDT, J. Dynamic properties of air-filled excised cat lung determined by liquid plethysmograph. J. AppZ. Physiol. 27: 246250, 1969. MEAD, J. Mechanical properties of lungs. Physiol. Rev 4: 281-330, 1961. PFLUEGER, E. Das Pneumonometer. PfZuegers Arch. 7: 496, 1882. RADFORD, E. P., JR. Static mechanical properties of mammalian lungs. In: Handbook of Physiology. Respiration. Washington, DC: Am. Physiol. Sot., 1964, sect. 3, vol. I, chapt. 15, p. 429-449. WILLMON, T. L., AND A. R. BEHNKE. Residual volume determinations by the methods of helium substitution and volume expansion. Am. J. Physiol. 153: 138-142, 1948. WOHL, M. E. B., J. TURNER, AND J. MEAD. Static volume-pressure curves of dog lungsin vivo and in vitro. J. AppZ. Physiol. 24: 348354, 1968.
Downloaded from www.physiology.org/journal/jappl by ${individualUser.givenNames} ${individualUser.surname} (129.186.138.035) on January 21, 2019.
University
HILLS, B. A., AND R. E. BARROW. Method for estimating absolute lung volumes at constant inflation pressure. J. Appl. Physiol.: Respirat. Environ. Exercise Physiol. 47(4): 907-909, 1979.-A method has been devised for measuring functional residual capacity in the intact killed animal or absolute lung volumes in any excised lung preparation without changing the inflation pressure. This is achieved by titrating the absolute pressure of a chamber in which the preparation is compressed until a known volume of air has entered the lungs. This technique was used to estimate the volumes of five intact rabbit lungs and five rigid containers of known dimensions by means of Boyle’s law. Results were found to agree to within &I% with values determined by alternative methods. In the discussion the advantage of determining absolute lung volumes at almost any stage in a study of lung mechanics without the determination itself changing inflation pressure and, hence, lung volume is emphasized. functional
residual
capacity
IN MANY STUDIES of pulmonary mechanics it is desirable
to measure not only the volume change for a given inflation pressure in the intact lung or isolated lung preparation, but also the absolute volumes involved. For instance, determinations of functional residual capacity (FRC) in a killed animal and the absolute volume when its lungs are subsequently excised and reinflated help in relating measurements of compliance, airway closure, and lung integrity in general to the normal physiological state of the animal. Two principles have been employed in such absolute volume measurements: dilution techniques and various pneumatometric approaches using the compressibility of gasesas expressed by Boyle’s law. Current dilution techniques do not vary appreciably from the original method of Darling et al. (3) but still tend to suffer from one of two problems -absorption of the tracer by blood in the live situation or the difficulty of obtaining uniform distribution in the dead animal. Pneumatometric approaches are of two types depending upon whether the lungs are closed or open to the surrounding atmosphere. The closed type is based on measuring pressure differentials generated by voluntary respiratory effort (5, 9)) 0~~~-75C;7/79/oooO-0$01.25
Copyright
0 1979 the American
lung volumes
Physiological
of Texas Medical
Branch,
which is, therefore, only applicable to live subjects, whereas those classified as “decompression” methods (5) essentially measure the volume exhausted from the lung when ambient pressure is reduced. There are many variations (4, 6, 11) of the latter approach that are good for a lung in the intact thoracic cavity where there is no alveolar-atmospheric pressure differential. These methods are thus well suited to the determination of FRC in the relaxed subject and can easily be adapted to the dead intact animal. However, this advantage does not hold for the isolated lung preparation, because it is particularly undesirable (8,lO) to allow a lung to collapse to minimal volume after excision from the cadaver or sacrificed animal. Even if the lungs are removed with the trachea clamped and compliance studied from the relationship between change in volume and change in inflation pressure, it is highly desirable to be able to redetermine the absolute volume after any cycle in order to relate the findings to the original FRC. Hence a variation of the decompression method has been devised that enables the absolute lung volume to be determined for any inflation pressure. It has the advantage that the same technique can also be used for the particular case of zero inflation pressure in the intact thoracic cavity to determine FRC before the lung is excised. There is a further advantage in that it can be used to determine the absolute volume at any value selected for the constant inflation pressure whether flow is inward (compression) or outward (decompression). MATERIALS
AND
METHODS
The principle used involves measuring the increase in ambient pressure needed to cause a known volume of air to flow into the lung without changing the transmural pressure differential maintaining it at a particular degree of inflation. The intact or isolated lung preparation is intubated and measurements of inflation pressure and volume changes appropriate to the particular study are made via valve X with Y closed (see Fig. 1). Let us consider the stage in a study where we need to know the absolute lung Society
907
Downloaded from www.physiology.org/journal/jappl by ${individualUser.givenNames} ${individualUser.surname} (129.186.138.035) on January 21, 2019.
908
B. A. HILLS
Mercury
AND
R. E. BARROW
manometer
L
LUI
my
r I cpal
ailul
I
;
\
\ \ \
v Volume:
Impressed-air
Pipette I I \ \
\
c
; I
” Volume:
I \ I LT
; i i ;: II *. p
SUPPlY Pressure FIG.
1. Apparatus
depicting
water
level after
compression
of chamber
volume without further change of inflation pressure. Valve Y is then opened. With the desired inflation pressure still applied to the lung via X, the water reservoir is raised or lowered until the water reaches the mark L in the pipette. The water level in the reservoir must then be above the mark L by a vertical height (H), which now equals the inflation pressure. Valve X is then closed. The whole preparation is placed in the chamber and the pressure raised with compressed air. The absolute pressure (P) is titrated until at an increase (AP) at which the water level in the pipette has moved from the mark L to the mark N (see Fig. 1). In so doing, the water has displaced the accurately known pipette volume of air (v). The whole sequence of events is reversed on decompression after which valve Y can be closed and X opened. This enables the lung study to continue without any change of inflation pressure during the determination of absolute lung volume (V). V is then calculated by applying Boyle’s law to the dry air whose partial pressure is (PO+ H - Pw) before compression and (PO+ AP + H Pw) afterwards (PO+ H - Pw)(V + v + E) = (PO+ P + H - Pw)(V + c) This reduces to give V as V = v(Po + H - Pw)/AP - E
(1)
where PO is atmospheric pressure, Pw is water vapor pressure at ambient temperature, and c is the volume between the lung and the mark N on the pipette. E can be determined directly, while v is selected to be about 25-35s of the anticipated value for V, so that AP is less than one-third atmosphere. This comparatively small pressure change enables a wide range of laboratory vessels to be used as the chamber; any small leaks in it do not affect the result. The pipette needs to be a flat vessel or a horizontal coil of tubing such that the vertical height between marks L and N is negligible with respect to H. The pipette is so arranged to prevent air trapping. The relationship depicted by Eq. 1 is plotted in Fig. 2 for the apparatus used in this study where v = 7.56 ml, c = 3.15 nil, and H = 0 on a day when atmospheric pressure (PO) was 756 Torr.
Chamber
by AP for a lung
maintained
at a constant
inflation
pressure
H.
RESULTS
Volumes of five lengths of rubber tube and five rabbit lungs determined by alternative methods are plotted against the respective value of AP obtained by this method in Fig. 2 in which the resulting points are compared with the Boyle’s law relationship (Eq. 1). Absolute volumes of the rubber tubes with one end closed were determined by weighing them empty and then full of water. Lung volumes were determined by saline immersion after excising them with the trachea clamped. Allowance was made for parenchymal tissue volume, which was estimated from lung weight using a specific gravity of 1.0 gem-” (2) (see Table 1). These data are presented as points in Fig. 2 where it can be seen that the rubber tube and lung volumes are within tl% of the theoretical curve. The use of fixed containers enabled us to test the method well beyond the normal range of lung volumes. Rates of chamber pressurization were very low (94 Torr. min-‘) to avoid any significant intra-airway pressure gradients. DISCUSSION
The foregoing evaluation indicates that this technique of compressing without changing inflation pressure offers an accurate and convenient means of measuring the absolute lung volume of intact lung preparations and should be equally applicable to excised lungs. Moreover, such measurements can be made at any stage in a study of pulmonary mechanics without changing the schedule of inflation pressure to be followed by that lung. This avoids the need to reevaluate the mechanical status of the lung that would have been necessary if the method used had changed the inflation pressure. This is a particular advantage in view of the complex dependence of lung compliance upon its pressure-volume history (1, 7, 12). The authors would like to thank Mrs. Laura Huddleston for her assistance in preparing the manuscript. The research reported here has been supported under the Office of Naval Research Contract NOOO14-75-C-1035 with funds provided by the Naval Medical Research and Development Command. Received
11 October
1978; accepted
in final
form
1 June
1979.
Downloaded from www.physiology.org/journal/jappl by ${individualUser.givenNames} ${individualUser.surname} (129.186.138.035) on January 21, 2019.
ESTIMATING
ABSOLUTE
LUNG
909
VOLUMES
320
282
-
CALCULATED MEASURED MEASURED
0 A
VOLUMES (EQ. 1) TUBING VOLUME LUNG VOLUMES
FIG. 2. Volumes of excised lungs (V) estimated by immersion technique and known volumes of rigid containers are compared with values at the same titrated inflation pressure (AP) used to determine V by the Boyle’s law relationship (Eq. I) shown as a solid line.
15
20
25 Absolute
30 Volume
40
35
(V) in ml
1. Comparison of volumes (FRC) determined from AP with values measured by alternative methods
TABLE
Alternative Animal
Rabbit lungs/immersion calibration
wt, kg
Vol
displaced, ml
Method *
Tissue
AP, Method wt, g
3.2 3.6 3.5 4.1
75.0 60.0 65.0
42.9
95.0
3.9
85.3
58.8 51.1
FRC,
functional
residue
capacity.
* Isolated
29.2
lungs.
t Intact
No. of runs
AP, Torr
31.8 36.0 34.2
4 4 4 4 4
157 168 162 142 150
32.2 k 29.9 k 31.2 t 36.0 t, 33.9 t
0.4 0.3 0.3 0.5 0.4
15.3 20.1 25.0 30.0 36.1
4 4 4 4 4
301 243
15.3
0.1
198
25.0 29.2 36.0
32.9 29.8
31.0
Rubber tubing/ weight calibration
FRC,
ml
lungs;
(Eq. I)
values
are means
172 142
Calculated
FRCt,
t
ml
19.8 t 0.2 k 0.2 t 0.3 t 0.4
t SD.
REFERENCES 1. AGOSTONI, E., AND J. MEAD. Statics of the respiratory system. In: Handbook of Physiology. Respiration. Washington, DC: Am. Physiol. Sot., 1969, sect. 3, vol. I, chapt. 13, p. 389-407. 2. CROSFILL, M. L., AND J. G. WIDDICOMBE. Physical characteristics of the chest and lungs and the work of breathing in different mammalian species. J. Physiol. London 158: l-14, 1961. 3. DARLING, R. C., A. COURNAND, AND D. RICHARDS. Studies on the intrapulmonary mixture of gases. III. An open circuit method for measuring (residual) air. J. CZin. Invest. 19: 609, 1940. 4. DEJOURS, P., AND H. RAHN. Residual volume measurements by the gas expansion method and nitrogen dilution method. J. Appl. Physiol. 4: 445-448, 1953. 5. DUBOIS, A. B., S. Y. BOTELHO, G. N. BEDELL, R. MARSHALL, AND J. H. COMROE, JR. A rapid plethysmographic method for measuring thoracic gas volume: a comparison with nitrogen washout method for measuring functional residual capacity in normal subjects. J. CZin. Invest. 35: 322-326, 1956.
6. HARVEY, 7.
8.
9. 10.
11.
12.
R. B., AND J. A. SCHILLING. Measurements of lung volume by rapid decompression. J. AppZ. Physiol. 7: 496, 1955. HILDEBRANDT, J. Dynamic properties of air-filled excised cat lung determined by liquid plethysmograph. J. AppZ. Physiol. 27: 246250, 1969. MEAD, J. Mechanical properties of lungs. Physiol. Rev 4: 281-330, 1961. PFLUEGER, E. Das Pneumonometer. PfZuegers Arch. 7: 496, 1882. RADFORD, E. P., JR. Static mechanical properties of mammalian lungs. In: Handbook of Physiology. Respiration. Washington, DC: Am. Physiol. Sot., 1964, sect. 3, vol. I, chapt. 15, p. 429-449. WILLMON, T. L., AND A. R. BEHNKE. Residual volume determinations by the methods of helium substitution and volume expansion. Am. J. Physiol. 153: 138-142, 1948. WOHL, M. E. B., J. TURNER, AND J. MEAD. Static volume-pressure curves of dog lungsin vivo and in vitro. J. AppZ. Physiol. 24: 348354, 1968.
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