Comparative Biochemistry and Physiology Part A: Molecular & Integrative Physiology
This article is part of a Special issue on Physiology from the NeotropicsBuccal jet streaming and dead space determination in the South American lungfish, Lepidosiren paradoxa☆
Introduction
The six species of extant Dipnoi (Neoceratodus forsteri; Lepidosiren paradoxa; Protopterus aethiopicus, P. amphibius, P. annectens, P. dolloi) possess lungs as their principal air-breathing organs (Graham, 1997). The morphological characteristics of dipnoan lungs have been studied for centuries (Owen, 1841; Quekett, 1844; Hyrtl, 1843; Günther, 1871; Parker, 1892; Poll, 1962; Grigg, 1965; Klika and Lelek, 1967; Hughes and Weibel, 1976; Maina and Maloiy, 1985; de Moraes et al., 2005). Several aspects of lung ventilation, gas exchange, blood gas chemistry, or chemoreception have also been of interest for comparative physiologists, especially over the last 50 years (e.g. Johansen and Lenfant, 1967; Johansen and Lenfant, 1968; Lenfant et al., 1970; McMahon, 1970; Fritsche et al., 1993; Amin-Naves et al., 2004; DeLaney et al., 1983; Sanchez and Glass, 2001; Glass, 2010; Sanchez and Glass, 2001; Sanchez et al., 2001b; Bassi et al., 2005; Bassi et al., 2010; Sanchez et al., 2005; Amin-Naves et al., 2007a, Amin-Naves et al., 2007b; da Silva et al., 2008, da Silva et al., 2017; Zena et al., 2017). Such attention given to morphology and physiology of lungfish can be explained by their phylogenetic position close to the origin of the Sarcopterygii (Brinkmann et al., 2004; Yokobori et al., 1994; Zardoya et al., 1998) and their ability to exchange gases bimodally with water (across gills and/or integument) and air (using lungs), representing the remaining living species of a lineage of vertebrates that evolved air breathing in the mid-Devonian (~380 Ma; Clement and Long, 2010).
One aspect of lungfish respiration, however, has not received much attention up to now: the breathing mechanism (Bishop and Foxon, 1968; McMahon, 1969; Brainerd, 1994). While studies of the buccal pump mechanism have been carried out on L. paradoxa (Bishop and Foxon, 1968; Brainerd, 1994) and P. aethiopicus (McMahon, 1969; Brainerd et al., 1993), only superficial descriptions of breathing behavior are available for N. forsteri (Dean, 1906; Grigg, 1965). It seems clear, however, that all lungfish use a two-stroke buccal pump, composed of one expiration and at least one inspiration, to renew the air within the lungs (Brainerd, 1994). The events during one breathing cycle can be described as follows (Brainerd and Ferry-Graham, 2006): 1) Air breathing starts with the animal putting its head out of the water and the mouth being opened; 2) The buccal cavity expands to draw in oxygen-rich air; 3) As the buccal cavity expands, the glottis opens and the lungs empty through elastic recoil of body wall and lungs; 4) Once the buccal cavity has been fully expanded, the mouth closes and the air trapped within the buccal cavity is pushed into the lungs. Since fresh air and gas expired from the lungs enter the buccal cavity during buccal expansion, some degree of mixing of oxygen-rich and oxygen-poor gas occurs during the first phase of the two-stroke buccal pump, reducing the amount of oxygen entering the lungs. McMahon (1969) estimates the gas mixing to be 20–40% of expired air being returned to the lungs in P. aethiopicus.
A two-stroke buccal pump ventilation has also been identified in anuran amphibians. De Jongh and Gans (1969) and Gans et al. (1969) performed classical experiments using the bullfrog (Lithobates catesbeianus) to describe the complex functioning of the buccal pump. De Jongh and Gans (1969) identified three different types of cyclical phenomena associated to buccal movements. 1) Oscillations of the buccal floor to ventilate the buccal cavity through the open nares. 2) Ventilatory cycles to renew the air within the lungs through coordinated opening and closing of nares and glottis. 3) Inspiratory movements with pulmonary ventilation interspaced by non-ventilatory periods. One fundamental proposal of de Jongh and Gans (1969) was the presence of an expiratory airflow, known as jet stream, postulated to result in minimal mixing of oxygen rich air in the buccal cavity with the oxygen poor air leaving the lungs during expiration, and passing rapidly along the dorsal part of the buccal cavity. For Lithobates pipiens a similar ventilatory pattern as in the bullfrog has been described, but no evidence for the presence of an expiratory jet stream has been found (Vitalis and Shelton, 1990), since the air being pumped into the lungs during inspiration represents a mixture of air, containing 30–50% of air from the previous expiration (West and Jones, 1975). Brett and Shelton (1979), studying Xenopus laevis, found in this species a ventilatory cycle starting with an expiration that is followed by an inspiration, thereby suggesting no mixing of fresh and used air within the buccal cavity. Fernandes et al. (2005), investigating the ventilatory pattern and jet stream hypothesis in the toad Rhinella schneideri, found a buccal cavity dead-space representing 30–40% of tidal volume, suggesting some degree of inspiratory and expiratory flow separation, with limited mixing within the buccal cavity.
Regarding the ventilatory cycle of L. paradoxa, it has to be assumed that this species possibly expires almost all of the air contained within the lungs at the beginning of a ventilatory cycle. This was demonstrated by Bishop and Foxon (1968) using X-ray imaging of a breathing cycle, where the lungs were no longer visible during expiration due to a near-complete loss of air, also has been suggested by da Silva et al. (2017), based on measurements of expiratory tidal volume. Since pulmonary dead space seems to be very small, a significant amount of gas mixing may only be possible during the expansion phase of the two-stroke buccal pump, when air is being expired from the lungs and fresh air is being drawn into the buccal cavity. To determine the degree of mixing of fresh air and expired gas during the breathing cycle of L. paradoxa and to verify the possible presence of a jet stream during expiration in this species, we performed measurements of buccal pressure, buccal and pulmonary PCO2 and PO2, pulmonary ventilation and calculated dead space and effective ventilation by using the Bohr dead space equation (see Eq. 3).
Section snippets
Animals
The lungfish specimens of Lepidosiren paradoxa were collected in the region of Cuiabá – MT (Brazil) and transported to the University of São Paulo, campus Ribeirão Preto (FMRP/USP) (authorized by IBAMA # 02027. 002172/2005–68). Upon arrival, the animals were placed in tanks of 1000 L with aerated and temperature-controlled water (25 °C) and were fed three times a week with chicken liver. Before the experiments, each animal remained 24 h without access to food. The experimental procedures were
Equations used and statistics
The lung O2 extraction coefficient (EO2) (which expresses the percentage of O2 absorbed by the pulmonary capillaries) and the respiratory exchange ratio (RER) were calculated following (Dejours (1981):where: PIO2 gives the oxygen partial pressure of inspired air (in mmHg), PLO2 the oxygen partial pressure in the lungs (in mmHg), and PLCO2 the lung carbon dioxide partial pressure (mmHg). Eq. 2 is a simple rearrangement of the alveolar gas equation,
Results
Fig. 1 depicts a representative buccal pressure profile along with airflow measurements during a breathing event in the lungfish. A typical ventilatory cycle in L. paradoxa was characterized by a slight increase in buccal pressure (‘a’ in Fig. 1) followed by a decrease in buccal pressure to sub-atmospheric values. Once negative pressure stopped decreasing, expiratory flow initiated (‘b’) and continued for about 4 s. With expiratory flow still occurring, buccal pressure increased sharply to
Discussion
Pulmonary airflow measurements in the current study confirm the presence of a two-stroke breathing mechanism in L. paradoxa, composed of a single expiration followed by 2–4 buccal inspirations. Our recordings of buccal cavity pressure, however, were different from previous recordings, since a negative buccal pressure was found at the beginning of each ventilatory cycle, always coinciding with expiratory flow. While McMahon (1969) and Brainerd et al. (1993) did record in P. aethiopicus small
Conflict of interests
The authors confirm that there is no conflict of interest
Acknowledgements
Funding: This study was financially supported by the Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) and Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), through the National Institute of Science and Technology on Comparative Physiology (INCT em Fisiologia Comparada, proceeding nos. 2008/57712-4 and 573921/2008-3). G.S.F. da Silva was supported by a Young Investigator Award (FAPESP no 2013/17606-9 and 2014/12190-1) and is currently supported by CNPq
References (53)
- et al.
Effects of acute temperature changes on aerial and aquatic gas exchange, pulmonary ventilation and blood gas status in the south American lungfish, Lepidosiren paradoxa
Comp. Biochem. Physiol A
(2004) - et al.
Components to the acid–base related ventilatory drives in the south American lungfish Lepidosiren paradoxa
Respir. Physiol. Neurobiol.
(2007) - et al.
Blood gases and cardiovascular shunt in the south American lungfish (Lepidosiren paradoxa) during normoxia and hyperoxia
Respir. Physiol. Neurobiol.
(2010) - et al.
Lung ventilation in salamanders and the evolution of vertebrate air-breathing mechanisms
Biol. J. Linn. Soc.
(1993) - et al.
Aestivation in the south American lungfish, Lepidosiren paradoxa: effects on cardiovascular function, blood gases, osmolality and leptin levels
Respir. Physiol. Neurobiol.
(2008) - et al.
Combined ventilatory responses to aerial hypoxia and temperature in the south American lungfish Lepidosiren paradoxa
J. Therm. Biol.
(2011) - et al.
Effects of aerial hypoxia and temperature on pulmonary breathing pattern and gas exchange in the south American lungfish, Lepidosiren paradoxa
Comp. Biochem. Physiol. A.
(2017) - et al.
An assessment of dead space in pulmonary ventilation of the toad Bufo schneideri
Comp. Biochem. Physiol. A
(2005) - et al.
Respiratory and cardiovascular responses hypoxia in the Australian lungfish
Respir. Physiol.
(1993) - et al.
The differential cardio-respiratory responses to ambient hypoxia and systemic hypoxaemia in the south American lungfish, Lepidosiren paradoxa
Comp. Biochem. Physiol.
(2001)
Temperature effects on lung and blood gases in Bufoparacnemis: consequences of bimodal gas exchange
Respir. Physiol.
Acute effects of temperature and hypercarbia on cutaneous and branchial gas exchange in the south American lungfish, Lepidosiren paradoxa
J. Therm. Biol.
Central ventilatory control in the south American lungfish, Lepidosiren paradoxa: distinct responses to pH and CO2
J. Comp. Physiol. B.
Pulmonary oxygen diffusing capacity of the south American lungfish Lepidosiren paradoxa: physiological values by the Bohr integration method
Physiol. Biochem. Zool.
The mechanism of breathing in the south American lungfish, Lepidosiren paradoxa; a radiological study
J. Zool. Lond.
The evolution of lung gill bimodal breathing and the homology of vertebrate respiratory pumps
Amer. Zool.
Mechanics of respiratory pumps
Ventilatory mechanisms of the amphibian, Xernopus laevis; the role of the buccal force pump
J. Exp. Biol.
Nuclear protein-coding genes support lungfish and not the coelacanth as the closest living relatives of land vertebrates
Proc. Natl. Acad. Sci.
Air-breathing adaptation in a marine Devonian lungfish
Biol. Lett.
On the mechanism of respiration in the bullfrog Rana catesbeiana. A reassessment
J. Morphol.
Morphometric comparison of the respiratory organs in the south American lungfish Lepidosiren paradoxa (Dipnoi)
Physiol. Biochem. Zool.
Notes on the living specimens of the Australian lungfish, ceratodus forsteri, in the zoological Society's collection
Proc. Zool. Soc. Lond.
Principles of Comparative Respiratory Physiology
Pulmonary mechanoreceptors in the Dipnoi lungfish Protopterus and Lepidosiren
Am. J. Phys.
Bullfrog (Rana catesbeiana) ventilation: how does the frog breathe?
Science.
Cited by (3)
Editorial on physiology from the neotropics
2020, Comparative Biochemistry and Physiology -Part A : Molecular and Integrative PhysiologyCircumventing surface tension: Tadpoles suck bubbles to breathe air
2020, Proceedings of the Royal Society B: Biological SciencesThe mechanics of air breathing in gray tree frog tadpoles, hyla versicolor (Anura: Hylidae)
2020, Journal of Experimental Biology
- ☆
This article is part of a special issue entitled: Physiology from the Neotropics, edited by: Dr. Kenia Bicego, Dr. Luciane Gargaglioni and Dr. Mike Hedrick
- 1
In memorian, Deceased October 4th, 2018.