Static lung compliance and body pressures in Tupinambismerianae with and without post-hepatic septum

Wilfried Klein a,*, Augusto S. Abe b, Steven F. Perry a

a Institut fur Zoologie, Universitat Bonn, Poppelsdorfer Schloß, 53115 Bonn, Germanyb Departamento de Zoologia, Universidade Estadual Paulista-Rio Claro, c.p. 199, 13506-900 Rio Claro, SP, Brazil

Accepted 14 February 2003

Abstract

The surgical removal of the post-hepatic septum (PHS) in the tegu lizard, Tupinambis merianae , significantly reduces

resting lung volume (VLr) and maximal lung volume (VLm) when compared with tegus with intact PHS. Standardised

for body mass (MB), static lung compliance was significantly less in tegus without PHS. Pleural and abdominal

pressures followed, like ventilation, a biphasic pattern. In general, pressures increased during expiration and decreased

during inspiration. However, during expiration pressure changes showed a marked intra- and interindividual variation.

The removal of the PHS resulted in a lower cranio-caudal intracoelomic pressure differential, but had no effect on the

general pattern of pressure changes accompanying ventilation. These results show that a perforated PHS that lacks

striated muscle has significant influence on static breathing mechanics in Tupinambis and by analogy provides valuable

insight into similar processes that led to the evolution of the mammalian diaphragm.

# 2003 Elsevier Science B.V. All rights reserved.

Keywords: Mechanics of breathing; compliance; ventilation; post-hepatic septum; Reptiles; tegu lizard (Tupinambis merianae );

Ventilation; reptile

1. Introduction

The lungs of the tegu lizard, Tupinambis mer-

ianae , are single-chambered and their respiratory

surface is made up of homogeneously distributed

faveoli (Perry, 1983). The lungs and the liver are

enclosed in a pleuro-hepatic cavity and are sepa-

rated from the remaining viscera by a post-hepatic

septum (PHS), a membranous structure containing

bundles of smooth muscle (Broman, 1904; Hoch-

stetter, 1906; Klein et al., in press). The PHS is

highly developed in the teiid genera Tupinambis

and Crocodilurus (Klein et al., 2000). It is trans-

versely oriented in the body cavity and, except for

a persistent dorso-lateral opening on the left side,

resembles the mammalian diaphragm. Recently

(Klein et al., in press), it was shown that the

removal of the PHS has profound effects on the

topology of the viscera: in tegus with a surgically

ruptured PHS, intestine and stomach are displaced

cranially, thereby reducing the space for the lungs.

* Corresponding author. Tel.: �/49-228-735489; fax: �/49-

228-735458.

E-mail address: kleinwilfried@web.de (W. Klein).

Respiratory Physiology & Neurobiology 135 (2003) 73�/86

www.elsevier.com/locate/resphysiol

1569-9048/03/$ - see front matter # 2003 Elsevier Science B.V. All rights reserved.

doi:10.1016/S1569-9048(03)00063-6

The effects of the PHS on the mechanics ofbreathing are unknown.

A measure of the mechanical properties of lungs

is its compliance (distensibility), which is defined

as the change in intrapulmonary volume for a

given intrapulmonary pressure. Surface tension

and elastic properties of the lung tissue are the

main determinants of lung compliance. In the tegu

lung, with its homogeneously distributed anddense parenchyma, a high work of breathing is

necessary to inflate the low-compliance lungs

(Perry and Duncker, 1980). Such a small, par-

enchyma-rich single-chambered lung is considered

an evolutionary dead end with respect to gas

exchange, as the space for elaboration of parench-

ymal gas exchange surfaces is limited to the lung

wall and because the deep faveolar parenchyma isnot accessible to convective air movement (Perry

and Duncker, 1980). Therefore, the PHS may have

evolved as an accessory structure in conjunction

with supporting inflation of the stiff lungs, thereby

partially overcoming the structural limitations set

by the single-chambered lungs.

In mammals, similar functional pressures may

have lead to the evolution of their unique lung�/

diaphragm complex (Perry and Duncker, 1978,

1980). Mammalian lungs show the lowest compli-

ance of all vertebrates and a high work of breath-

ing is necessary to ventilate them (Crosfill and

Widdicombe, 1961; Perry and Duncker, 1980). If

the pleural cavity were opened, the lungs would

collapse completely. In intact mammals an oppos-

ing negative pressure is applied in the pleural spacepreventing the lungs from collapsing (Radford,

1964) while at the same time allowing their free

movement within the closed pleural cavity. There-

fore an undivided pleuro-peritoneal body cavity,

as can be found in lizards, is inappropriate for

mammals and consequently, the evolution of the

mammalian lung seems impossible in the absence

of a complete diaphragm. A functional model forthe stepwise evolution of the diaphragm from an

incomplete septum therefore appears implausible.

The PHS of the tegu could represent such a model.

Breathing mechanics was studied in Tupinambis

as part of comparative studies (Perry and

Duncker, 1978, 1980), but these authors examined

mechanical properties only in the closed chest

preparation. These studies, however, reported thelowest values of total compliance (CT) found in a

lizard. A surgical rupture of the PHS not only may

affect lung volumes simply as a result of the

reduced pleuro-hepatic space (Klein et al., in

press), but also could alter the amount and

distribution of elastic fibres in the lung itself,

either stiffening the lungs against the pressure of

the viscera or balancing the lungs at a smallerresting volume. Such a change may be reflected in

decreased lung compliance.

During aspiration breathing, rib movement

generates positive pressure during expiration and

negative pressure during inspiration. In an undi-

vided body cavity, these pressures should apply

equally to all regions. Gans (1970) postulated that

the forces produced by respiratory muscles duringbreathing act directly on the visceral organs and

that these forces are transmitted from one com-

partment to another by the interaction of local

forces between the compartments (linkage) rather

than by hydrostatic forces alone. Therefore the

pressures in the visceral part of an undivided

pleuro-peritoneal cavity could be different from

those found in the pulmonary part (Templeton,1964).

In the present study we test the hypothesis that

the transverse division of the body cavity by the

PHS in Tupinambis is important for lung ventila-

tion and that in spite of the left-side foramen a

large transseptal pressure difference is created,

thereby facilitating breathing.

2. Materials and methods

2.1. Breathing mechanics

2.1.1. Animals and experimental design

Investigations of static breathing mechanics

were performed on neonatal tegus (N�/8; MB:

17.69/5.3 g; SVL: 87.49/7.6 mm), juvenile tegus(N�/5; MB: 120.39/27.6 g; SVL: 160.29/11.0 mm),

sham-operated (SO) adult tegus (N�/6; MB:

707.09/422.5 g; SVL: 267.59/44.3 mm) and adult

tegus maintained with removed PHS (SR-tegus)

for 11 months (N�/8; MB: 1193.89/263.0 g; SVL:

316.49/20.0 mm). The removal of the PHS has

W. Klein et al. / Respiratory Physiology & Neurobiology 135 (2003) 73�/8674

been described in detail elsewhere (Klein et al., inpress). All tegus were killed by intra-peritoneal

injection of 60 mg kg�1 pentobarbital sodium

(SagatalTM), which causes analgesia and cessation

of spontaneous breathing within 15�/20 min, while

the heart continues beating for hours. Experiments

were performed at room temperature (�/28 8C).

In the supine animal the trachea was intubated

to obtain volume�/pressure relationships. Thistube was connected to a three-way stopcock

from which one connection led to a plastic syringe

and the other, to a water-filled tube (PE 60)

connected to the DELTRAN pressure transducer

of a strain gauge. The dead space of all tubing

combined was less than 2 ml. The signal of the

pressure transducer was amplified (1000�/) by a

custom-made pre-amplifier and then sent to ananalogue-to-digital (A�/D) converter and recorded

through a computerised data-acquisition system

(DAC, Sable System). To calibrate the pressure

transducer, the water column was moved in a

stepwise fashion in a range between �/30 and �/30

cm and the regression line derived from this

calibration was used to obtain a regression line

for the V�/P relationships. The experimental pro-tocol followed closely that of Perry and Duncker

(1978). Briefly, resting lung volume (VLr) was

defined as the amount of air that can be removed

from the lungs in a freshly killed animal until a

negative pressure of �/10 cmH2O was reached.

Lung volumes were determined by adding air in

increments to the lungs of an animal with closed

body cavity, starting at a pressure of �/10 cmH2Oand increasing to �/20 cmH2O. The total amount

of air necessary to fill the lungs from the state at

greatest negative pressure until the V�/P diagram

passed its upper inflection point was defined as

maximal lung volume (VLm). To stabilise the V�/P

diagrams (Bernstein, 1957), the lungs were inflated

and deflated several times (usually four) over the

whole pressure range before measurements com-menced. After each determination of VLm, the

system was opened to the atmosphere and lung

volume was allowed to return spontaneously to

VLr. V�/P diagrams were obtained by (1) stepwise

removal of air until a pressure of �/10 cmH2O was

reached, (2) adding of air until pressure reached �/

20 cmH2O, and (3) returning stepwise to VLr.

Pressure was allowed to stabilise for 15�/30 secbefore taking the reading. After each V�/P dia-

gram, air was added until VLm was reached and

the system was opened to return to VLr again. V�/P

diagrams were first made for the animals with

intact body cavity, and then for the in situ lungs

after cutting the mesopneumonia, and widely

opening or resecting the body wall. The lungs

were moistened with saline (0.9% NaCl) through-out. The mean values of three V�/P diagrams were

taken for analysis.

2.1.2. Data handling and statistics

Compliance (C) was defined as the slope (b) of

the straight portion of the inflation curve and

obtained by fitting a line (y�/a�/ bx) through the

points given by the V�/P diagram. Compliance ofthe lungs (CL) and total compliance (CT) were

determined from V�/P diagrams where lung vo-

lume was expressed in ml (non-standardised), as

well as after standardisation of lung volume

against MB, VLr and VLm. Body wall compliance

was defined as 1/CB�/1/CT�/1/CL. The index of

hysteresis (H) was defined as the length of a

perpendicular line connecting the midpoints ofthe inflation curve and the deflation curve, and

was calculated by the Pythagorean method (for

details see Perry and Duncker, 1978).

ANOVA was used examine significant differ-

ences between the groups, followed by an all

pairwise multiple comparison procedure (Stu-

dent-Newman�/Keuls Method) to determine which

groups differ significantly from each other. Adifference was considered significant at P B/0.05.

2.2. Pressure measurements

2.2.1. Animals, surgery and experimental protocol

Four tegus without PHS (MB: 1015.259/193.61

g) and four tegus with intact PHS (MB: 1150.09/

298.86 g) were used in this study.

After exposing the animals to CO2 until theydisplayed no reaction to handling or pinching, the

tegus were placed in a prone position on an

operation table and a constant low CO2-flow was

provided to their nostrils to maintain anaesthesia.

The abdomen was disinfected with alcohol and

with an injection needle (¥ 2 mm) a piece of PE

W. Klein et al. / Respiratory Physiology & Neurobiology 135 (2003) 73�/86 75

tubing (Portex, 800/110/240/100) was inserted on

the right dorso-lateral side into the pleural cavity

at the level of the 3th�/5th rib and into the

abdominal cavity 2�/3 cm anterior to the pelvic

girdle. Each tube extended 4�/6 cm into the body

cavity. To prevent blocking of the opening by the

viscera two additional holes were made opposing

each other near the tip of the tube and the tubes

were flushed from time to time with saline (0.9%

NaCl). Pressures were measured using two DEL-

TRAN pressure transducers from the strain gauge

type, each connected to a custom-made pre-

amplifier (amplifying voltage 1000�/).After surgery, which was carried out before

09:00, a mask for ventilation measurements was

put over the tegus head and the animal was placed

into a climatic chamber at 25 or 35 8C to recover.

The mask was connected by tygon tubing to a

pressure transducer (Sable PT-100) for ventilatory

recordings. The signals from pressure transducer

and pre-amplifiers were sent to an A�/D converter

and recorded through a computerised data-acqui-

sition system (DAC, Sable System).

Pressures and spontaneous ventilation data were

measured between 17:00 and 20:00. When these

measurements were completed, the temperature

was increased or decreased and pressures together

with ventilation were measured under the new

conditions the following day between 17:00 and

20:00.

2.2.2. Data handling

To analyse the data, a time was chosen when

tegus appeared to be resting quietly, and trans-

mural pleural and abdominal pressures were

determined: the baseline before each breathing

bout was taken and the change from the baseline

to the maximal or minimal peak was taken as

pressure created during expiration and inspiration,

respectively. Transseptal pressure was obtained bysubtraction of abdominal pressure from pleural

pressure.

3. Results

3.1. Lung volumes

The results for resting (VLr) and maximal lung

volume (VLm) are shown in Fig. 1. VLr of each

group was significant (P B/0.05) different from VLr

of each other group. VLm of SR-tegus was

significantly (P B/0.05) lower compared to neona-

tal, juvenile and adult tegus. In tegus with intact

PHS, the allometric relationships of VLr and VLm

can be described as VLr�/�/0.984�/MB0.84 with

R2�/0.979 and P B/0.0001 and as VLm�/�/

0.679�/MB0.975 with R2�/0.993 and P B/0.0001.

The ratio of non-standardised VLr to VLm was 3.0

for neonatal, 3.8 for juvenile, 5.2 for adult tegus

and 12.7 for tegus with removed PHS.

3.2. Compliance

Sample VLr-standardised V�/P diagrams aregiven in Fig. 2. Non-standardised CT, CB and CL

was significantly lower in neonatal tegus compared

with the other groups and CT, CB and CL of

juvenile tegus were significantly different from the

values of the other groups. No significant differ-

ences were found between adult tegus with and

without PHS (Table 1). Normalised against VLr,

tegus without PHS showed a significantly greaterlung compliance (CL) and total compliance (CT)

compared with neonatal, juvenile and adult tegus

with an intact PHS. Body wall compliance (CB) of

SR-tegus was only significantly different from that

of neonatal and juvenile tegus with intact septum.

VLr-standardised compliance (CT, CL and CB)

Fig. 1. Resting (VLr) and maximal lung volume (VLm) in T.

merianae with removed PHS, and in adult, juvenile and

neonatal T. merianae with intact PHS. Data are presented as

means9/S.E.M. * indicates a value that is significant (P B/0.05)

different from all other values.

W. Klein et al. / Respiratory Physiology & Neurobiology 135 (2003) 73�/8676

showed no significant differences between neona-

tal, juvenile and adult tegus (Table 1). When CT,

CL and CB were standardised against MB, CT and

CB were significantly lower in SR-tegus compared

with the other groups with intact septum and CL

was significantly less than in adults. In neonatal

tegus CT was significantly greater compared withadult and juvenile tegus. CT in the VLm-standar-

dised V�/P diagram was highest in neonatal tegus

and significantly different from all other groups,

and CB was significantly greater compared with

SR- and adult tegus (Table 1).

3.3. Hysteresis

The index of hysteresis (Table 2) determined for

lungs and body wall (HT) and for the lungs alone

(HL) on a VLr-standardised V�/P diagram showed

no significant differences between the different

groups of tegus. HT and HL in SR-tegus were

significantly greater (P B/0.05) compared with

neonatal, juvenile and adult tegus on a non-

standardised V�/P diagram. HL of neonatal tegus

was also significantly less (P B/0.05) in compar-

ison with adult and juvenile tegus. When normal-ised against MB and VLm the index of hysteresis

showed no significant differences among the

groups.

3.4. Pressures

Samples of pressure recordings in spontaneous

breathing tegus can be seen in Fig. 3 and the

pressures measured in each individual are shown in

Table 3. Several different pictures of pressure

changes were found. (1) Expiration could be

Fig. 2. VLr-standardised sample V�/P diagrams of a neonatal (A), juvenile (B), an adult T. merianae (C) with intact PHS and an adult

T. merianae without PHS (D). Note the different y -axis in (D). Circles indicate V�/P diagram of lungs and body wall together and

triangles indicate V�/P diagrams of lungs alone.

W. Klein et al. / Respiratory Physiology & Neurobiology 135 (2003) 73�/86 77

accompanied by a large increase in pressure in the

abdominal and pleural part of the body cavity, (2)

only an increase in pressure in the pleural cavity or

(3) no marked change in pressure at all. (4) In

some cases the pleural pressure decreased at the

beginning of an expiration, followed by a slight

increase at the end of the expiration. During

inspiration, on the other hand, a large decrease

in pressures was always found in the pleural as well

as in the abdominal part of the body cavity.The recorded pressures were similar at 25 and

35 8C, and only SO-tegus showed a larger decrease

in pressure during inspiration at 35 8C compared

with 25 8C. Tegus with a surgically ruptured PHS

showed some differences compared with those

having an intact PHS. Those differences were

Table 1

VLr-, VLm-, VMB- and non-standardised compliance of body wall and lungs combined (CT), and for body wall (CB) and lungs (CL)

alone in T. merianae without PHS (SR), and adult (SO), juvenile and neonatal T. merianae with intact PHS

Without PHS/SR Adult/SO Juvenile Neonatal

CL/VLr 6.409/2.30a 1.39/0.16 1.69/0.34 1.49/0.18

CB/VLr 1.929/0.47b 1.099/0.26 0.789/0.14 0.809/0.16

CT/VLr 1.449/0.39a 0.559/0.06 0.499/0.06 0.519/0.06

CL/Vol 69.69/16.0 48.19/5.0 14.49/6.7a 1.69/0.16a

CB/Vol 24.09/3.0 25.39/5.4 4.09/0.5a 1.29/0.20a

CT/Vol 17.59/2.6 15.89/3.2 2.89/0.4a 0.69/0.06a

CL/VLm 0.449/0.05 0.579/0.10 0.509/0.13 0.489/0.07

CB/VLm 0.169/0.01 0.199/0.02 0.259/0.06 0.339/0.04c

CT/VLm 0.129/0.01 0.139/0.02 0.149/0.01 0.199/0.01a

CL/VMB 49.59/6.0d 90.79/12.6 62.09/11.2 68.49/4.7

CB/VMB 21.29/1.3a 39.79/5.6 42.39/4.6 57.29/7.5

CT/VMB 1.479/1.0a 23.19/4.3 24.79/3.0 34.49/3.4

Data are means9/S.E.M. Units are as follows: C/VLr and C/VLm: ml cmH2O�1 ml�1; C/Vol: ml cmH2O�1; C/VMB:

ml cmH2O�1 kg�1.a Indicates a value that is significant (P B/0.05) different from all other values.b Indicates a value that is significant (P B/0.05) different from neonatal and juvenile tegus.c Indicates a value that is significant (P B/0.05) different from adult SO- and SR-tegus.d Indicates a value that is significant (P B/0.05) different from adult SO-tegus.

Table 2

VLr-, VLm-, VMB- and non-standardised index of hysteresis of body wall and lungs combined (HT), and lungs (HL) alone in T. merianae

without PHS (SR), and adult (SO), juvenile and neonatal T. merianae with intact PHS

Without PHS/SR Adult/SO Juvenile Neonatal

HT/VLr 1.59/0.10 1.29/0.30 1.89/0.47 1.39/0.21

HL/VLr 0.99/0.17 0.79/0.14 1.79/0.86 1.29/0.13

HT/Vol 8.09/0.86* 3.69/1.13 2.69/0.40 1.39/0.22

HL/Vol 8.59/1.99* 2.99/0.44 2.79/0.96 0.89/0.19*

HT/VLm 1.29/0.18 1.29/0.32 2.09/0.40 1.39/0.23

HL/VLm 0.69/0.14 0.79/0.15 1.69/0.84 1.19/0.13

HT/VMB 9.29/2.4 10.49/3.6 12.19/2.8 6.29/0.9

HL/VMB 8.89/2.4 6.39/1.3 8.39/3.3 4.49/0.4

Data are means (in cm)9/S.E.M.

* Indicates a value that is significant (P B/0.05) different from all other values.

W. Klein et al. / Respiratory Physiology & Neurobiology 135 (2003) 73�/8678

Fig. 3. Sample recordings of ventilation (upper trace), abdominal pressure (middle trace) and pleural pressure (lower trace) at 25 8C(A) and 35 8C (B�/C) of T. merianae with intact PHS. B and C show the variation in the pressure changes accompanying expiration.

W. Klein et al. / Respiratory Physiology & Neurobiology 135 (2003) 73�/86 79

more pronounced at 35 than at 25 8C. During

expiration at 35 8C, for example, the increases in

pressures in the pleural and the abdominal part of

the body cavity were greater in SR- than in SO-

tegus. During inspiration at 35 8C, however,

pressure changes in the pleural cavity were smaller

and pressure changes in the abdominal cavity were

larger in SR-tegus than in SO-tegus. The resulting

intracoelomic pressure gradient was about 2

cmH2O greater in SO- than in SR-tegus (Table 3).

4. Discussion

4.1. Lung volumes

Perry and Duncker (1978) reported for similar

sized Tupinambis teguixin (earlier T . nigropuncta-

tus , Avila-Pires, 1995) a resting lung volume of 43

ml kg�1 and a maximum lung volume of 151

ml kg�1. The slightly lower values found in this

study on T . merianae may be due to differences in

Table 3

Pressures measured in the pleural and abdominal body cavity of individual adult T. merianae with (SO) and without PHS (SR) at 25

and 35 8C

25 8C Pleural Abdominal Transseptal

Expiration Inspiration Expiration Inspiration Expiration Inspiration

SR

T1 0.55 �/0.62 �/0.03 �/1.74 0.59 1.12

T2 2.11 �/2.46 1.69 �/1.55 0.42 �/0.91

T3 3.92 �/4.83 1.39 �/2.94 2.54 �/1.88

T5 �/0.17 �/0.91 0.98 �/2.01 �/1.15 1.10

Mean 1.60 �/2.20 1.01 �/2.06 0.59 �/0.14

S.D. 1.82 1.93 0.75 0.62 1.51 1.50

SO

T6 1.58 �/3.74 �/0.25 �/1.67 1.34 �/2.06

T8 1.66 �/0.76 0.82 �/1.52 0.85 0.76

T11 4.37 �/2.16

T13 1.25 �/0.31 0.33 �/0.31 0.92 0.003

Mean 2.22 �/1.74 0.46 �/1.17 1.03 �/0.43

S.D. 1.45 1.55 0.30 0.75 0.26 1.46

35 8C

SR

T1 0.05 �/0.97 1.19 �/2.16 �/1.14 1.20

T2 2.85 �/5.12 1.61 �/3.50 1.22 �/1.61

T3 2.80 �/1.60 0.22 �/1.90 2.58 0.30

T5 0.28 �/0.92 1.03 �/1.37 �/0.76 0.45

Mean 1.49 �/2.15 1.02 �/2.23 0.48 0.08

S.D. 1.54 2 0.58 0.91 1.74 1.46

SO

T6 1.55 �/1.62 0.55 �/0.83 1.00 �/0.79

T8 4.97 �/7.83 �/0.35 �/2.93 5.31 �/4.90

T11 2.31 �/2.94 0.21 0.02 2.10 �/2.96

T13 0.55 �/2.44 0.02 �/1.07 0.53 �/1.37

Mean 2.34 �/3.71 0.11 �/1.20 2.24 �/2.51

S.D. 1.89 2.80 0.37 1.24 2.16 1.84

Pressures are given in cmH2O and represent the maximal deviations from the baseline pressure during expiration and inspiratio. The

two columns on the right show calculated (transseptal) pressure differences between pleural and abdominal cavities.

W. Klein et al. / Respiratory Physiology & Neurobiology 135 (2003) 73�/8680

the species or to better maintenance conditionsand the resulting greater body mass in the

Brazilian animals. With increasing body mass,

VLr increased less than VLm, increasing the ratio

VLm to VLr from 3.0 in neonatals to 5.1 in adults.

Perry and Duncker (1978) found a ratio of 3.4 in

T . teguixin , similar to those of neonatals and

juveniles in the present study. The ratios reported

for man (3.0; Rahn et al., 1946), mouse (4.8;Crosfill and Widdicombe, 1961) and some lizards

(Lacerta 4.0; Gecko 4.7; Chamaeleon 5.0; Varanus

4.2; Perry and Duncker, 1978) are, despite lung

types varying from single- to multichambered

lungs, all in the same range, whereas SR-tegus

(VLm/VLr�/12.7) lie outside this range.

Maximum lung volume was found to increase as

MB0.975 in tegus, which is similar to the slopes

reported by Schmidt-Nielsen (1984) for mammals

and birds (MB0.94 and MB

1.06, respectively) and

found by Tenney and Tenney (1970) for amphi-

bians (MB1.05). Thus, maximum lung volume seems

to be proportional to body mass in tetrapods in

general.

VLr as well as VLm were significantly less in SR-

tegus than in other groups. VLr reaches 37.8% andVLm 73.4% of the respective volumes shown by

adults with intact PHS. The large difference in VLr

can be explained by the cranial movement of the

viscera in PHS-ectomotised tegus as described by

Klein et al. (in press). The smaller difference

between tegus with intact and surgically ruptured

PHS in VLm could be explained by the high

pressure (�/20 cmH2O) which was used to inflatethe lungs. The maximal change in lung volume

occurred in tegus at pressures between 0 and �/6

cmH2O. An excursion to �/20 cmH2O only reflects

the maximal possible distension of the lung tissues

and this may not change significantly in one year

without PHS. The decrease in lung volume seen in

tegus without a PHS does not necessarily lead to a

significant decrease in respiratory surface areabecause of the way in which faveolar parenchyma

responds to volume changes. When lung volume

decreases, the walls of the faveoli form accordion

pleats until parts of the epithelia come into contact

with each other (Daniels et al., 1994, 1995). If the

faveolar walls in tegus without PHS do not

collapse completely, the area for gas exchange

remains unaffected and oxygen uptake may re-main unchanged.

4.2. Compliance

The value for total compliance from the

non-standardised V�/P diagram found in this

study is similar to CT reported by Perry and

Duncker (1978) for T . teguixin (15.8 vs. 13.65

ml cmH2O�1, respectively). Perry and Duncker(1978) reported 0.47 ml cmH2O�1 for VLr-

standardised CT, which is similar to the value

found in this study (CT: 0.55 ml cmH2O�1). In

comparison with other lizards studied by Perry

and Duncker (1978) CT and CB of tegus are most

similar to those of the emerald lizard Lacerta

viridis , the tokay Gekko gecko , and the monitor

Varanus exanthematicus , but the values for CL

found in the present study are the lowest ones for

lizards reported so far. Only the emerald lizard has

a CL as low as that of the tegu. Both species belong

to the superfamily of lacertiform lizards (Estes et

al., 1988), which characteristically possess single-

chambered lungs with dense, homogeneously dis-

tributed parenchyma (Duncker, 1978). No differ-

ences were found in compliance between tegus ofdifferent sizes, which indicates that not only lung

volume but also static mechanical properties are

proportional to MB throughout development.

Further evidence for this hypothesis comes from

the values of compliance obtained from non-

standardised V�/P diagrams. Here, neonatal tegus

showed the lowest values of CT, CB and CL which

are comparable with those reported by Perry andDuncker (1978) for similar sized emerald lizards

(MB: 28 g; CT: 0.51 ml cmH2O�1; CB: 1.39

ml cmH2O�1; CL: 1.74 ml cmH2O�1). The allo-

metric relationships for CT, CL and CB of tegus

with intact PHS are shown in Table 4. With a mass

exponent close to 1.0, CL shows a similar propor-

tionality to MB as VLm, whereas CB and CT show

the same allometric relationship as VLr. VLr is aresult of forces interacting between lungs and

thorax. Due to surface tension phenomena and

elastic recoil, lungs tend to collapse, whereas the

rib cage shows a tendency to spring outwards. The

resulting balance of forces determines VLr (Agos-

tini et al., 1959). Apparently, neonatal tegus have a

W. Klein et al. / Respiratory Physiology & Neurobiology 135 (2003) 73�/86 81

stiffer body wall than adults, which leads to a

relatively greater VLr and thus to a smaller VLm/

VLr ratio than in adult tegus with intact PHS.

Tegus with ruptured PHS showed, on a VLr-

standardised V�/P diagram, significantly greater

CT, CB and CL than tegus with intact PHS and

especially CL is very high, exceeding CL of the

chameleon Chamaeleo chamaeleon (6.4 vs. 5.7

ml cmH2O�1, respectively; Perry and Duncker,

1978). This apparently high compliance in SR-

tegus was the result of the significantly lower VLr,

which leads to a steeper slope in the V�/P diagram

and thus to a greater VLr-standardised compliance.

Therefore, to compare the effect of a removed

PHS on static breathing mechanics, compliance

was also determined on V�/P diagrams standar-

dised with MB and VLm. On a body mass basis, CT,

CB and CL were significantly less in tegus without

PHS than in neonatal, juvenile and adult tegus

with intact PHS. In particular, CB was only half as

great as in the other groups. Not only the

abdominal wall but also the viscera belong to the

category ‘body wall’. There is no reason to believe

that in tegus without PHS the elastic properties of

the body wall changed due to the removal of the

PHS, as dissection did not reveal a marked

hypertrophy of the intercostal and abdominal

musculature. Therefore, the significantly lower

CB is solely due to the lack of a PHS. Tegus

without PHS must inflate the lungs not only

against the resistance of the abdominal wall but

also must displace the viscera which occupy a large

part of the former pleuro-hepatic cavity.

The VLm-standardised V�/P diagram gave no

significant differences in the compliance, which

may be expected from the proportionality of VLm

with MB and the small difference in VLm betweenoperated and intact tegus.

4.3. Hysteresis

The index of hysteresis (H) for the total

respiratory system of intact adult tegus found in

the present study is similar to HT reported by

Perry and Duncker (1978; 1.2 vs. 1.6 cm, respec-tively) and in the same range like HT of Lacerta .

HL in tegus, however, is less than HL of the

emerald lizard, but somewhat greater than HL of

the tokay and the chameleon (Perry and Duncker,

1978). The removal of the PHS had no significant

effect in H, based on VLr-, VLm- or MB-standar-

dised V�/P diagrams. Perry and Duncker (1978)

postulated that HL depends upon the degree ofpartitioning of the lungs, being greater in lungs

with uniformly distributed, fine parenchyma as

seen in Lacerta and less in lungs with ventral and

caudal dilatations as seen in Chamaeleo or Gecko .

The present results are therefore surprising, be-

cause the surface-to volume ratio for Lacerta and

Tupinambis are nearly identical (Perry, 1983).

Tegus in this study showed no difference inhysteresis over a body mass ranging from 17 to 700

g. The non-standardised H, however, showed a

markedly greater HT and HL in tegus without PHS

compared to the other groups.

During the determination of open-body V�/P

diagrams some lungs showed alternating contrac-

tions at mid-inflation causing a slight increase in

pressure. Such peristaltic waves are a long knowncharacteristics for turtles (Francois-Franck, 1906)

and lizards (Saalfeld, 1934) and have been attrib-

uted to spontaneous contractions of trabecular

smooth muscle (Perry and Duncker, 1978). They

may be of significant importance in the dense

parenchyma of tegus and could promote mixing of

air in the faveoli under apneic conditions and

allow more effective use oxygen stores in thecentral lumen of the lungs.

4.4. Pressures

In human or veterinary medicine, internal body

pressures are measured by inserting balloon cathe-

ters into the esophagus and stomach to measure

Table 4

Allometric relationships for non-standardised CT, CL and CB of

T. merianae with intact PHS expressed as y�/aMBb with a as

proportionally coefficient, b as body-mass exponent and y as

dependent variable

y a b P 9/S.E.M. R2

CT �/1.27 0.851 B/0.0001 0.05 0.954

CL �/1.06 0.996 B/0.0001 0.1 0.89

CB �/1.13 0.884 B/0.0001 0.08 0.916

W. Klein et al. / Respiratory Physiology & Neurobiology 135 (2003) 73�/8682

e.g. transdiaphragmatic pressures (e.g. Gilbert etal., 1979; Desmecht et al., 1992). Some problems

that arise with this technique are like different

pressures depending on whether or not the gastric

balloon lies in a fluid-filled part of the stomach, or

pressure decreases when the balloon is moved

caudally (Desmecht et al., 1992). Therefore, pres-

sures were measured with small, open tubes which

could be implanted quickly with minor surgery,and tegus seemed to be unaffected by this proce-

dure. Although the exact position of the tip of the

tubes could not be determined, their position in

the middle of the pleural or abdominal cavity,

respectively, was sufficient to reflect pressures in

these regions of the body cavity. It is possible, that

the lack of large pressure changes during expira-

tion may be caused by blocking of the tube by theviscera. In this case, there would be no signal at all

during expiration or inspiration. It is unlikely that

both the abdominal and pleural tubes were

blocked at the same time throughout the entire 3

h of recording and despite several flushing events.

In general, expiration in tegus was accompanied

by an increase and inspiration by a decrease in

pressure compared with the breath-hold period.The pressure changes accompanying expiration,

however, were not continuous throughout a mea-

suring period in an individual tegu, and when the

animals were moving, the signals could change

(Fig. 4A). Other tegus showed a continuous

decrease in expiratory pressures after an activity

bout (Fig. 4B). During rest, expiration was mainly

an active process and the increase in pressure wascaused by a contraction of the body wall muscles,

thus moving the ribs caudad and mediad and

reducing the volume of the body cavity. The small

pressures recorded in some animals during expira-

tion may be explained by the elastic recoil or

surface tension of the lungs. A decrease in lung

volume together with an unchanged volume in rib

cage position could cause a small negative pressurein the pleural cavity at the beginning of expiration.

Afterwards, the chest wall also moved inward,

either by its elastic recoil or by active contraction,

and caused a small increase in pressure. Whenever

the expiratory air flow stopped and inspiration

began, pressures fell dramatically, causing a nega-

tive pressure in the pleural cavity, and air flowed

into the lungs. Termination of inspiration wascoincident with abolition of the transthoracic

pressure differential.

The changes in pressures in the abdominal

cavity were, in general, smaller than those in the

pleural cavity. This held true even for tegus

without PHS, which agrees with the hypothesis

of Gans (1970), that pressures on the viscera are

transmitted by local interaction of forces betweenthe different compartments rather than by the

whole body cavity acting as a single fluid filled

space. The residual transseptal pressure difference

in SR-tegus could be induced by the liver, which

partially protects the right lung from the viscera.

At the greater temperature, the transseptal pres-

sure was much greater in SO- than in SR-tegus:

i.e., the pressures in the body cavity were moreevenly distributed in animals that lacked a PHS.

Farmer and Hicks (2000) measured pressures in

the abdominal part of the body cavity in Iguana

iguana . They recorded an increase in pressure in

the body cavity during exercise caused by the

activation of the trunk muscles during locomotion,

and further that elevated pressures provoke col-

lapsing of abdominal veins and thereby reduce thevenous return from the caudal body.

In tegus with intact PHS, the greatest part of the

pressure changes necessary for ventilation may be

restricted by the PHS to the thoracic region,

particularly under conditions of elevated meta-

bolic demand. This situation would maintain low

pressures in the abdominal part of the body cavity,

and thus would not reduce venous return evenduring light exercise. The fact that the role of the

PHS in increasing the intracoelomic pressure

gradient at rest is equivocal, however, suggests

that this is not the primary function of the septum.

Rather, its main function appears to be preventing

the craniad migration of the viscera during in-

spiration (Klein et al., in press).

The respiratory system of T. merianae givesvaluable insights in the breathing mechanics in

lizards and allows us to construct a functional

morphological scenario about the evolution of the

mammalian lung/diaphragm complex. In lizards,

costal aspiration breathing may be efficient when

the lungs are highly compliant. With reduced lung

compliance some mechanisms appears to be ne-

W. Klein et al. / Respiratory Physiology & Neurobiology 135 (2003) 73�/86 83

cessary, which restrict visceral kinesis. One such

mechanism is the use of the liver as a barrier to

impede the cranial movement of the gut during

inspiration. The role of liver retraction in inspira-

Fig. 4. Sample recordings of ventilation (upper trace), abdominal pressure (middle trace) and pleural pressure (lower trace) at 25 8C(A) and 35 8C (B) of T. merianae showing the variation in the pleural pressures related to expiration. (A): Only a small positive

pressure change accompanies expiration before movement of the animal but a large change after movement of animal. (B): Decreasing

pressures accompanying expiration after movement.

W. Klein et al. / Respiratory Physiology & Neurobiology 135 (2003) 73�/8684

tion in crocodiles and testudines is well known(Gans and Clark, 1976; Gaunt and Gans, 1969),

and ‘abdominal breathing’ in mammals also in-

volves liver displacement. The present studies

suggest that dynamic fixation of the liver during

inspiration may precede hepatic kinesis as an

accessory respiratory mechanism in general.

The superficial similarity of the teiid PHS and

the embryonic mammalian diaphragm as a trans-verse perforated barrier, which fixes the liver in the

tegu and divides the body cavity between lungs

and liver in the mammal, is striking. Later in

ontogeny the mammalian diaphragm closes its

perforations whereas in the tegu an opening

persists, suggesting that the basic function of

PHS and diaphragm may have been similar during

early phylogenetic stages in mammalian ancestors.In both cases the presence of this transverse barrier

allows the efficient inflation of low-compliance

lungs. In the mammal striated muscle may have

replaced smooth muscle (Brink, 1956), or may

have been present from the beginning (Keith,

1905; Graper, 1928) allowing breath-by-breath

participation of the diaphragm, whereas the

smooth muscle of the teiid PHS is expected toallow only tonal increase of the efficiency of costal

breathing.

Acknowledgements

Financial support was provided by a PRO-

BRAL (CAPES/DAAD) grant to ASA and SFP

and a Graduiertenstipendium des Landes NRW to

WK. We also thank two anonymous reviewers for

helpful comments on the manuscript.

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