Recent Work

In this series of studies, oxygen consumption was measured continuously in young tegu lizards, Tupinambis merianae, exposed to 4 days at 25ºC followed by 7 to 10 days at 17ºC, all in constant dark, at five different times of the year. Under these constant conditions, circadian rhythms of oxygen consumption (higher in the daytime) persisted for anywhere from one day to the entire two weeks in different individuals in all seasons except the winter, at which time the animals had been in constant dark already for over a month. We also saw a progressive decline in standard oxygen consumption (at highly variable rates in different individuals) to a very low rate that was seasonally independent (ranging from 21.1 ± 6.2 to 24.7 ± 0.2 ml.kg-1.hr-1 across seasons). Although this degree of reduction appeared to take longer to invoke when starting from higher metabolic rates, as during the active season, tegu lizards reduced their metabolism to the low rates seen in winter dormancy at all times of the year when given sufficient time in the cold and dark.

Effects of Changes in Season, Temperature and Body Mass on the Standard Metabolic rate of Tegu Lizards (Tupinambis merianae)
Luís F. Toledo, Simone P. Brito, William K. Milsom, Augusto S. Abe and Denis V. Andrade

Abstract

The present study was designed to determine how the standard metabolic rate of tegu lizards, a species that undergoes large ontogenetic changes in body weight with associated changes in life history strategies, is affected by changes in body mass, body temperature, season, and life history traits. We measured oxygen consumption (V·O2) in 156 individuals, ranging in body mass from 10.4 grams to 3.75 kg, at three experimental temperatures (17, 25 and 30ºC) over the four seasons of the year. We found that standard metabolic rate scaled to the power 0.79 at all temperatures in all seasons and that thermal sensitivity of metabolism was relatively low (Q10 ˜ 2.0) over the range from 17 to 25ºC, regardless of body size or season. In winter, however, from 25ºC to 30ºC, Q10 was mass dependent, increasing to values above 4.0 in larger animals (= 3.0 kg). Thus, during the dormancy period, larger tegus were more responsive to changes in the higher temperature range. These mass-dependent changes in Q10 may promote prompt arousal of reproductively active adult lizards during the warmer days at the end of the dormancy period.  

 
The role of branchial and orobranchial O2 chemoreceptors in the control of aquatic surface respiration (ASR) in the neotropical fish tambaqui (Colossoma macropomum): Prolonged and acute responses to hypoxia
Luiz H. Florindo, Cléo A. C. Leite, Ana L. Kalinin, Stephen G. Reid, Milliam K. Milsom and F.Tadeu Rantin

Abstract

The present study examined the role of branchial and orobranchial O2 chemoreceptors in the cardiorespiratory responses, aquatic surface respiration (ASR), and the development of inferior lip swelling in tambaqui during acute and long-term exposure to hypoxia. Intact fish (control) and three groups of denervated fish (bilateral denervation of cranial nerves IX+X (to the gills), of cranial nerves V+VII (to the orobranchial cavity) or of cranial nerves V alone), were exposed to severe hypoxia (PwO2 = 10 mmHg) for 360 min. Respiratory frequency (fR) and heart rate (fH) were recorded simultaneously with ASR. Intact (control) fish increased fR and developed hypoxic bradycardia in the first 60 min of hypoxia. The bradycardia, however, abated progressively and had returned to normoxic levels by the last hour of exposure to hypoxia. The changes in respiratory frequency and the hypoxic bradycardia were eliminated by denervation of cranial nerves IX and X but were not affected by denervation of cranial nerves V or V+VII. The fH in fish with denervation of cranial nerves V or V+VII, however, did not recover to control values as in intact fish. This suggests that orobranchial O2 chemoreceptors innervated by these nerves may mediate the recovery in fH seen during long-term exposure to hypoxia. After 360 min of exposure to hypoxia only the intact and IX+X denervated fish performed ASR. Denervation of cranial nerve V abolished the ASR behavior. However, all (control and denervated (IX+X, V and V+VII)) fish developed inferior lip swelling. These results suggest that ASR is triggered by O2 chemoreceptors innervated by cranial nerve V but that other mechanisms, such as a direct effect of hypoxia on the lip tissue, trigger lip swelling.

 
Gill chemoreceptors and cardio-respiratory reflexes in the neotropical teleost pacu, Piaractus mesopotamicus
Leite, C. A. C., Florindo, L. H.,Kalinin, A. L.,Milsom, W. K. and F. T. Rantin

Abstract

This study examined the location and distribution of O2 chemoreceptors involved in cardio-respiratory responses to hypoxia in the neotropical teleost pacu, Piaractus mesopotamicus. We measured heart rate, ventilation rate and ventilation amplitude in fish exposed to gradual hypoxia (PWO2 of water gradually reduced from 140 to 10 mmHg) and submitted to intrabuccal and intravenous injections of NaCN (1.0 mL and 0.5 mL of 1 mg/mL NaCN, respectively). The protocol was performed on intact fish and on fish following progressive gill denervation by selective transection of cranial nerves IX and X. We found that the chemoreceptors producing reflex bradycardia were confined to, but distributed along, all gill arches and were sensitive to O2 levels in the water and the blood. We also found receptors initiating changes in ventilation frequency and amplitude distributed along all gill arches sensitive to O2 levels in the water and the blood. Ventilatory responses to all stimuli, though modified, continued following gill denervation, however, indicated the presence of internally and externally oriented receptors either in the pseudobranch or at extra-branchial sites. Chemoreceptors located on the first pair of gill arches and innervated by the glossopharyngeal (IXth) nerve appeared to attenuate the cardiac and respiratory responses to hypoxia..

 
Control of breathing in anuran amphibians
Luciane H. Gargaglioni and William K. Milsom

 Abstract

One of the defining characteristics of many amphibians is their free-living aquatic larval stage and semi-aquatic/terrestrial adult stage. This gave rise to their name that derives from the Greek for "double life" (amphibios). In many ways the larval stage reflects their ancestral origin from fish, while their adult stage resembles that of the more derived tetrapods. The three living groups (caecilians, salamanders and frogs), however, include almost 4,000 species that display a wide range of life histories and that are evolutionarily distant from their ancestral past. Thus, while amphibians stand as an intermediate stage in the evolution of the tetrapods, all modern amphibians are highly specialized and represent a significant departure in morphology, ecology and behaviour from the stem group that gave rise to the later tetrapods.

Despite their high degree of specialization, and the tremendous differences that exist between the three major lineages, there are also many features that they share; including up to three respiratory surfaces (skin, gills and lungs). In most amphibians at least two of these are functional at any given time during development. Aquatic amphibians (including all larvae) primarily rely on gills for gas exchange while terrestrial amphibians primarily rely on lungs. In both, however, the skin may serve as a major surface for gas exchange. Indeed, some terrestrial species have reduced or lost their lungs and some aquatic forms have lost their gills; both groups now relying solely on cutaneous gas exchange. Given this, study of the control of breathing in amphibians may be regarded as a regulatory physiologists dream (or nightmare). These animals can 'ventilate' up to three different exchange surfaces (skin, gills and lungs) with different respiratory media (water and air) and can independently perfuse these surfaces in different proportions due to the existence of highly regulated intra and extra-cardiac shunts.

A review of all exchange processes occurring at all surfaces in all groups is beyond the scope of this article. The focus of this review will be solely on the control of gill and lung ventilation in anuran amphibians (primarily of the genera Rana and Bufo). These are the groups that have attracted the most attention from respiratory physiologists and ventilatory control is an area that has attracted much recent research and is ripe for review.

 
Age, temperature and the pons: effects on the respiratory rhythm of rat brainstem-spinal cord preparations
M. Beth Zimmer and W.K. Milsom

 Abstract

To determine the role of age (0-6 days after birth) and pontine inputs on the respiratory response to hypothermia, respiratory-like motor output was examined in neonatal, rat brainstem-spinal cord preparations during step-wise cooling and re-warming. Fictive breathing continued at colder temperatures in preparations with higher starting frequencies, and the duration and area of each burst increased during cooling. Inputs from the pons reduced this increase. Young preparations tended to have a bell-shaped burst profile with the pons intact, but when the pons was removed, the shape was rapid in onset and decrementing. Older preparations only produced the decrementing profile, indicating that inputs from the pons shape the motor discharge pattern in an age-dependent fashion. Finally, fictive breathing became episodic at lower temperatures (= 23ºC). Interactions between age and temperature were important in generating and modulating this episodic pattern, and may reflect an intrinsic feature of central respiratory control in all vertebrate species under conditions of reduced drive.

Effects of Cortical Activation State on Respiration and Respiratory Chemoreflexes: Role of the Parabrachial/Kölliker Fuse Complex
Joyce A. Boon and W.K. Milsom

 Abstract

Microinjection of MK-801, a non-competitive inhibitor of NMDA type glutamate receptors (NMDAr), into the Parabrachial/Kölliker Fuse (PBrKF) complex of the pons in vagally intact urethane anaesthetized Sprague Dawley rats caused a reduction in the frequency of respiration, due to increases in both inspiratory and expiratory times, and an increase in tidal volume. MK-801 also reduced the increase in ventilation that was usually seen on cortical activation (i.e. the change from State III (the slow wave sleep-like state) to State I (the wake-like state)) under urethane anesthesia, indicating that the "wakefulness" stimulus for breathing involves glutamate activation of NMDAr on PBrKF neurons. In addition, blockade of NMDAr in this region also caused the rats to spend more time in the SWS-like state (State III) possibly due to diffusion of the MK-801 into the adjacent pontine reticular formation. Nevertheless, there was a reduction in ventilation in both State I and State III, indicating that this reduction was not simply a result of the change in state. Our results also indicate that PBrKF neurons with NMDAr are not involved in the response to hypoxia or hypercapnia, nor, interestingly, in the change in chemosensitivity associated with cortical activation.   

The present study was designed to explore systematically the midbrain of unanesthetized, decerebrate anuran amphibians (cane toads and bullfrogs), using electrical stimulation and midbrain transections, to identify sites capable of exciting and inhibiting breathing. Ventilation was measured as fictive motor output from the mandibular branch of the trigeminal nerve and the laryngeal branch of the vagus nerve. The results of our transection studies suggest that, under resting conditions, the net effect of inputs from sites within the rostral half of the midbrain is to increase breathing frequency, while inputs from sites within the caudal half of the midbrain have no net effect on breathing frequency but appear to act on the medullary central rhythm generator to produce episodic breathing. The results of our stimulation experiments indicate that the principal sites in the midbrain that are capable of exciting or inhibiting the frequency of buccal oscillations and lung ventilations, and potentially clustering breaths into episodes, appear to be those primarily involved in visual and auditory integration, motor functions and attentional state.

The role of persistent sodium currents in respiratory rhythm generation in vitro
Lieneke H. Marshall and William K. Milsom

 Abstract

Persistent sodium currents (INaP) contribute to both pacemaker and network properties involved in the generation of breathing. We tested the hypothesis that INaP is essential for rhythm generation and autoresuscitation in neonatal mammals using the in vitro brainstem-spinal cord preparation. We applied riluzole (RIL), a drug known to block INaP, to preparations from neonatal rats and hamsters (P0-4), both at 27ºC and at low temperatures, while recording fictive breathing from cervical rootlets. Preparations from rats responded to RIL at 27ºC with decreased motor burst amplitude with no change in frequency whereas those from hamsters responded with decreased frequency with no change in amplitude. Upon re-warming from low temperatures, RIL had no effect on the ability of rat preparations to autoresuscitate (re-start fictive breathing) but blocked autoresuscitation completely in 70% of hamster preparations. The data suggest that RIL acts primarily to block network bursting properties in neonatal rats but pacemaker properties in neonatal hamsters.