Respiratory Rates Derived From Arterial Blood Pressure Waveforms in Telemetered Dogs.

The objective of this study was to extract low frequency respiratory "artifacts" from a standard arterial blood pressure (ABP) waveform to simultaneously derive reliable breathing rates (BR). Arterial blood pressure derived BR values were characterized against respiratory rates simultaneously obtained from the Respiratory Inductive Plethysmography (RIP) system (EMKA). Reference compounds were introduced to evaluate responsiveness of the derived measures to respiratory depressants and stimulants. Male beagle dogs (n = 3) were instrumented with minimally invasive telemetry devices for measurements of ABP and heart rate. The RIP system was utilized simultaneously to collect respiratory rate, tidal volume, and minute volume of each animal following pharmacological challenges. Early results revealed the derived BR's from ABP waveforms did not correlate well with those measured from the RIP system. Post study X-ray visualization revealed suboptimal catheter positioning, causing poor concordance of BR tallied from the ABP waveforms. Follow-up evaluations were conducted using additional animals instrumented with the ABP catheter tip placement advanced proximal to the thoracic diaphragm. Preliminary data from this subset of animals significantly improved the correlation of BR derived from ABP and respiratory rates recorded by the RIP. This proof of concept investigation was intended to evaluate an algorithm designed to extract additional data from routine cardiac waveforms. We clearly demonstrated that with optimal blood pressure catheter placement and acquisition algorithm, a reliable breathing rate can also be extracted from safety studies without the need for additional studies/animals to capture those respiratory end points.

Translational assessment of cardiac contractility by echocardiography in the telemetered rat.

INTRODUCTION: Cardiac contractility was evaluated using standard inotropic agents in rats. We compared indices of cardiac contractility, i.e. LV dP/dt max from telemetry while simultaneously collecting EF (ejection fraction) and FS (fractional shortening) measures from echocardiography. METHODS: Male Wistar rats were instrumented with telemetry devices for measurements of blood pressure and left ventricular pressure. Milrinone (PDE III inhibitor) and verapamil (L-type calcium channel blocker) at doses of 0, 3, 10, and 30 mg/kg were administered orally using a 4 × 4 Latin square crossover study design. Telemetry data were recorded at predose and continuously for 24h post-dose. Echocardiographic evaluations were conducted once at predose and at 1 and 2h after milrinone or verapamil administration, respectively. During the recording of echocardiograms, telemetry data were collected simultaneously. Blood samples were also collected to confirm plasma drug exposure. RESULTS: As expected, milrinone increased LV dP/dt max, EF and FS while verapamil decreased LV dP/dt max, EF and FS. Linear regression analysis showed a positive correlation between LV dP/dt max and EF or FS (P<0.001) with both test agents. A change in LV dP/dt max of 1000 mmHg/s was found to correspond with a change in EF and FS of 13 and 16%, respectively, in the telemetered rat. DISCUSSION: The correlation between contractility indices assessed by telemetry and echocardiographic methods in rat models has not received much attention to date. Our results with two reference compounds demonstrate that both methods are sensitive to alterations in contractility induced by inotropic agents administered to rats. The high degree of correlation between changes in LV dP/dt max and EF or FS in the rat enables a translational-element of clinical relevance following changes in contractility indices when measured with telemetry devices in preclinical studies.

Climate change, extinction risks, and reproduction of terrestrial vertebrates.

This review includes a broad, but superficial, summary of our understanding about current and future climate changes, the predictions about how these changes will likely affect the risks of extinction of organisms, and how current climate changes are already affecting reproduction in terrestrial vertebrates. Many organisms have become extinct in the last century, but habitat destruction, disease and man-made factors other than climate change have been implicated as the causal factor in almost all of these. Reproduction is certain to be negatively impacted in all vertebrate groups for a variety of reasons, such as direct thermal and hydric effects on mortality of embryos, mismatches between optimal availability of food supplies, frequently determined by temperature, and reproductive capacities, sometimes determined by rigid factors such as photoperiod, and disappearance of appropriate foraging opportunities, such as melting sea ice. The numbers of studies documenting correlations between climate changes and biological phenomena are rapidly increasing, but more direct information about the consequences of these changes for species survival and ecosystem health is needed than is currently available.

Stability and permanence in gender- and stage-structured models for the boreal toad.

The boreal toad Bufo boreas boreas, once common in the western USA, is listed as an endangered species in Colorado and New Mexico, and protected in Wyoming. Populations have dramatically declined due to the presence of the fungal pathogen Batrachochytrium dendrobatidis (Bd). A gender- and stage-structured model for the boreal toad is formulated which depends on its life cycle and breeding strategies. In addition, an epizootic model for the spread of Bd is formulated. Analysis of these models provides two thresholds. The first threshold, the basic reproduction number for the population, ℛ(0), determines whether the population persists and the second threshold, the basic reproduction number for the fungal disease, ℛ(F), determines whether the disease persists. If ℛ(0)>1 and ℛ(F)<1, then the population becomes disease-free. However, if both thresholds are greater than one, the population levels are severely reduced by the fungal pathogen.

Amphibian chytridiomycosis.

Amphibian chytridiomycosis is a fungal disease caused by the chytrid fungus Batrachochytrium dendrobatidis. It is arguably the most significant recorded infectious disease of any vertebrate class. The disease is reducing amphibian biodiversity across most continents and regions of the world, affecting the resilience of surviving populations and driving multiple species to extinction. It is now recognised by the World Organisation for Animal Health (OIE) as an internationally notifiable disease. Collaborative research in areas including the development of diagnostic assays, distribution and impact of the disease, and management (treatment and policy) has assisted in leading a paradigm shift in accepting infectious disease as a major factor influencing wildlife population stability and biodiversity.

Effects of temperature and hydric environment on survival of the Panamanian Golden Frog infected with a pathogenic chytrid fungus.

Considerable controversy exists concerning whether or not climate changes (particularly global warming) are causing outbreaks of a lethal amphibian pathogen, the chytrid fungus Batrachochytrium dendrobatidis (Longcore, Pessier & D.K. Nichols 1999). In the present study, groups of Panamanian golden frogs (Atelopus zeteki Dunn, 1993), a critically endangered amphibian thought to be nearly extinct in Panama, were exposed to varying dosages of zoospores of Batrachochytrium dendrobatidis, temperatures and hydric environments in order to learn whether this species is susceptible to this pathogen and, if so, how environmental factors affect survival. This pathogen proved to be highly lethal for A. zeteki. Frogs exposed to a dosage of 100 Bd zoospores survived significantly (P<0.0001) longer than those that had been exposed to 10(4) or 10(6) zoospores. Exposed frogs housed at 23 °C survived significantly (P<0.0001) longer than those that were housed at 17 °C. Exposed frogs held in dry conditions survived significantly longer than those in wet conditions (P<0.0001). As a laboratory study, these results do not directly test hypotheses about the relation between climate change and the decline of these frogs in the field, but they inform the discussion about how environmental conditions can have an impact on the interaction between a susceptible amphibian and this pathogen. These data do not support the contention that rising global temperatures are necessary to cause the death of amphibians infected with this pathogen because the pathogen was equally lethal at 17 as at 23 °C, and frogs at the warmer temperature lived significantly longer than those at the cooler one.

The impacts of climate change on the annual cycles of birds.

Organisms living today are descended from ancestors that experienced considerable climate change in the past. However, they are currently presented with many new, man-made challenges, including rapid climate change. Migration and reproduction of many avian species are controlled by endogenous mechanisms that have been under intense selection over time to ensure that arrival to and departure from breeding grounds is synchronized with moderate temperatures, peak food availability and availability of nesting sites. The timing of egg laying is determined, usually by both endogenous clocks and local factors, so that food availability is near optimal for raising young. Climate change is causing mismatches in food supplies, snow cover and other factors that could severely impact successful migration and reproduction of avian populations unless they are able to adjust to new conditions. Resident (non-migratory) birds also face challenges if precipitation and/or temperature patterns vary in ways that result in mismatches of food and breeding. Predictions that many existing climates will disappear and novel climates will appear in the future suggest that communities will be dramatically restructured by extinctions and changes in range distributions. Species that persist into future climates may be able to do so in part owing to the genetic heritage passed down from ancestors who survived climate changes in the past.

Immune defenses of Xenopus laevis against Batrachochytrium dendrobatidis.

Amphibian populations are declining at an unprecedented rate worldwide. A number of declines have been linked to a pathogenic skin fungus, Batrachochytrium dendrobatidis. Although amphibians have robust immune defenses, many species seem to be very susceptible to infection by this fungus and to development of the lethal disease called chytridiomycosis. One species that is relatively resistant to B. dendrobatidis is Xenopus laevis. Because X. laevis has been used as a model for studies of immunity in amphibians and because it is relatively resistant to chytridiomycosis, it is a good model to examine immune defenses against B. dendrobatidis. Although much less is known about immune defenses in Bufo boreas, it serves as a second model species because it is very susceptible to B. dendrobatidis. Here we review what is known about innate antimicrobial peptide defenses in the skin and the development of immune responses following experimental immunization with heat-killed fungal cells. Development of an immunization protocol in X. laevis that induces effective defenses may suggest better strategies for protecting vulnerable species such as B. boreas.

Predicted disease susceptibility in a Panamanian amphibian assemblage based on skin peptide defenses.

Chytridiomycosis is an emerging infectious disease of amphibians caused by a chytrid fungus, Batrachochytrium dendrobatidis. This panzootic does not equally affect all amphibian species within an assemblage; some populations decline, others persist. Little is known about the factors that affect disease resistance. Differences in behavior, life history, biogeography, or immune function may impact survival. We found that an innate immune defense, antimicrobial skin peptides, varied significantly among species within a rainforest stream amphibian assemblage that has not been exposed to B. dendrobatidis. If exposed, all amphibian species at this central Panamanian site are at risk of population declines. In vitro pathogen growth inhibition by peptides from Panamanian species compared with species with known resistance (Rana pipiens and Xenopus laevis) or susceptibility (Bufo boreas) suggests that of the nine species examined, two species (Centrolene prosoblepon and Phyllomedusa lemur) may demonstrate strong resistance, and the other species will have a higher risk of disease-associated population declines. We found little variation among geographically distinct B. dendrobatidis isolates in sensitivity to an amphibian skin peptide mixture. This supports the hypothesis that B. dendrobatidis is a generalist pathogen and that species possessing an innate immunologic defense at the time of disease emergence are more likely to survive.

Emerging infectious disease and the loss of biodiversity in a Neotropical amphibian community.

Pathogens rarely cause extinctions of host species, and there are few examples of a pathogen changing species richness and diversity of an ecological community by causing local extinctions across a wide range of species. We report the link between the rapid appearance of a pathogenic chytrid fungus Batrachochytrium dendrobatidis in an amphibian community at El Copé, Panama, and subsequent mass mortality and loss of amphibian biodiversity across eight families of frogs and salamanders. We describe an outbreak of chytridiomycosis in Panama and argue that this infectious disease has played an important role in amphibian population declines. The high virulence and large number of potential hosts of this emerging infectious disease threaten global amphibian diversity.

Population trends associated with skin peptide defenses against chytridiomycosis in Australian frogs.

Many species of amphibians in the wet tropics of Australia have experienced population declines linked with the emergence of a skin-invasive chytrid fungus, Batrachochytrium dendrobatidis. An innate defense, antimicrobial peptides produced by granular glands in the skin, may protect some species from disease. Here we present evidence that supports this hypothesis. We tested ten synthesized peptides produced by Australian species, and natural peptide mixtures from five Queensland rainforest species. Natural mixtures and most peptides tested in isolation inhibited growth of B. dendrobatidis in vitro. The three most active peptides (caerin 1.9, maculatin 1.1, and caerin 1.1) were found in the secretions of non-declining species (Litoria chloris, L. caerulea, and L. genimaculata). Although the possession of a potent isolated antimicrobial peptide does not guarantee protection from infection, non-declining species (L. lesueuri and L. genimaculata) inhabiting the rainforest of Queensland possess mixtures of peptides that may be more protective than those of the species occurring in the same habitat that have recently experienced population declines associated with chytridiomycosis (L. nannotis, L. rheocola, and Nyctimystes dayi). This study demonstrates that in vitro effectiveness of skin peptides correlates with the degree of decline in the face of an emerging pathogen. Further research is needed to assess whether this non-specific immune defense may be useful in predicting disease susceptibility in other species.

How physiological methods and concepts can be useful in conservation biology.

The single and synergistic effects of man-made changes to the environment, such as habitat destruction, climate change, introduction of novel, long-lived chemicals into the environment, transport of exotic species and pathogens into new geographical areas, and other factors are predicted to cause widespread population declines and species extinctions of plants and animals in this century. From its inception, physiology has dealt with organismal capacities to deal with environmental change. This essay argues that physiologists, their methods and concepts can make more substantial contributions to Conservation Biology than they have to date. A few of the many ways in which physiologists can participate in Conservation Biology include formulating standards for proof of cause-and-effect relations and providing information about how environmental change could affect organismal energetics, host-pathogen relations, immune defenses, and others.

Effects of ultraviolet radiation on toad early life stages.

BACKGROUND: Exposure to harmful levels of ultraviolet-B radiation (UVB), a component of solar radiation, has been suggested as a potential cause of amphibian declines. METHODS: We measured solar radiation (UVB, ultraviolet-A, and visible) wavebands in breeding ponds of Bufo boreas (boreal toad, a montane species that has undergone severe population declines) and Bufo woodhousii (Woodhouse's toad, a plains toad that has not experienced declines)and examined tolerances of these species to simulated solar UVB exposures in the laboratory. RESULTS: We found larvae of both species to be tolerant of simulated solar UVB in excess of solar UVB levels observed in their breeding ponds. B. boreas tadpoles were more tolerant of simulated solar UVB exposure than B. woodhousii tadpoles, possibly because of greater amounts of photoprotective melanin in B. boreas skin. CONCLUSIONS: UVB levels observed in B. boreas habitats do not currently appear to constitute a threat to the survival of these animals; however, long-term (> 1 month) exposure to UVB levels comparable to levels associated with the water interface appears to reduce survival in B. woodhousii tadpoles. Therefore, future increases in surface and water column UVB radiation in bufonid habitats might pose significant survival risks to B. boreas or B. woodhousii populations.

Activities of temporin family peptides against the chytrid fungus (Batrachochytrium dendrobatidis) associated with global amphibian declines.

Temporin A and structurally related peptides produced in amphibian dermal granular glands and in wasp venom were tested for growth inhibition of Batrachochytrium dendrobatidis, a pathogen associated with global amphibian declines. Two natural amphibian temporins, a wasp temporin, and six synthetic analogs effectively inhibited growth. Differences in potency due to amino acid substitution suggest that ability to penetrate membranes and form an alpha-helical structure is important for their effectiveness against this pathogen.

Activity of antimicrobial skin peptides from ranid frogs against Batrachochytrium dendrobatidis, the chytrid fungus associated with global amphibian declines.

Accumulating evidence suggests that a chytrid fungus, Batrachochytrium dendrobatidis, is responsible for recent declines in amphibian populations in Australia, Central America, Europe, and North America. Because the chytrid infects the keratinized epithelium of the skin, we investigated the possible role of antimicrobial peptides produced in the skin as inhibitors of infection and growth. We show here that 10 peptides representing eight families of peptides derived from North American ranid frogs can effectively inhibit growth of this chytrid. The peptides are members of the ranatuerin-1, ranatuerin-2, esculentin-1, esculentin-2, brevinin-2, temporin, palustrin-3, and ranalexin families. All the tested peptides inhibit growth of mature fungal cells at concentrations above 25 microM, and some of them inhibit at concentrations as low as 2 microM. A comparison of the sensitivity of infectious zoospores with that of mature cells showed that the zoospores are inhibited at significantly lower concentrations of peptides. To determine whether cold temperature interferes with the inhibitory effects of these peptides, we tested their effectiveness at both 22 and 10 degrees C. Although the peptides inhibit at both temperatures, they appear to be more effective against zoospores at the lower temperature. These results suggest that the ranid frogs have, within their repertoire of antimicrobial substances, a number of skin peptides that should be a deterrent to chytrid infection. This may provide some natural resistance to infection, but if environmental factors inhibit the synthesis and release of the skin peptides, the pathogen could gain the advantage.

Antimicrobial peptide defenses against pathogens associated with global amphibian declines.

Global declines of amphibian populations are a source of great concern. Several pathogens that can infect the skin have been implicated in the declines. The pathogen most frequently associated with recent die-offs is a chytrid fungus, Batrachochytrium dendrobatidis. A second fungus, Basidiobolus ranarum, was isolated from declining populations of Wyoming toads. A third pathogen, Aeromonas hydrophila, is an opportunistic bacterium found in healthy frogs, but capable of inducing disease. Among the immune defense mechanisms used by amphibians is the production of antimicrobial peptides in granular glands in the skin. These packets of natural antibiotics can be emptied onto the skin when the amphibian is injured. To determine whether antimicrobial skin peptides defend against these amphibian pathogens, six peptides (magainin I, magainin II, PGLa, CPF, ranalexin, and dermaseptin), from three species, and representing three structurally different families of peptides, were tested in growth inhibition assays. We show here that the peptides can kill or inhibit growth of both fungi but not Aeromonas. Although each peptide varied in its effectiveness, at least one from each species was effective against both fungi at a concentration of about 10-20 microM. This is the first direct evidence that antimicrobial peptides in the skin can operate as a first line of defense against the organisms associated with global amphibian declines. It suggests that this innate defense mechanism may play a role in preventing or limiting infection by these organisms.

Effect of constant and fluctuating temperatures on resting and active oxygen consumption of toads, Bufo boreas.

The relations of standard and active rates of oxygen consumption to body temperature (Tb) were tested in montane Bufo b. boreas and lowland Bufo boreas halophilus acclimated to constant T b of 10, 20, or 30° C or to a fluctuating cycle of 5-30° C. Standard metabolic rates (SMR) of boreas acclimated to 30° C and halophilus acclimated to 10° C show pronounced regions of thermal independence but all other standard and active metabolic rates of groups acclimated to other thermal regimes are thermally sensitive. The SMR of both subspecies acclimated to the 5-30° C cycle are more thermally sensitive than those of similar individuals acclimated to constant T b. In cases where the relation between SMR and T b is linear for both halophilus and boreas at the same acclimation temperature, the slope and Q10 of the relation for boreas are significantly higher than those of halophilus. Acclimation had little or no effect on the active metabolic rates of either subspecies. The relation between SMR and T b of boreas maintained under field conditions (Carey, 1979) is matched only by those of individuals from the same population acclimated to 20° C.

Aerobic and anaerobic energy expenditure during rest and activity in montane Bufo b. boreas and Rana pipiens.

The relations of standard and active aerobic and anaerobic metabolism and heart rate to body temperature (T b) were measured in montane groups of Bufo b. boreas and Rana pipiens maintained under field conditions. These amphibians experience daily variation of T b over 30°C and 23°C, respectively (Carey, 1978). Standard and active aerobic and anaerobic metabolism, heart rate, aerobic and anaerobic scope are markedly temperature-dependent with no broad plateaus of thermal independence. Heart rate increments provide little augmentation of oxygen transport during activity; increased extraction of oxygen from the blood probably contributes importantly to oxygen supply during activity. Development of extensive aerobic capacities in Bufo may be related to aggressive behavior of males during breeding. Standard metabolic rates of both species are more thermally dependent than comparable values for lowland relatives. Thermal sensitivity of physiological functions may have distinct advantages over thermally compensated rates in the short growing season and daily thermal fluctuations of the montane environment.

Factors affecting body temperatures of toads.

Factors influencing levels and rates of variation of body temperature (T b) in montane Bufo boreas boreas and in lowland Bufo boreas halophilus were investigated as an initial step toward understanding the role of natural thermal variation in the physiology and energetics of these ectothermic animals. Body temperatures of boreas can vary 25-30° C over 24-h periods. Such variation is primarily due to both nocturnal and diurnal activity and the physical characteristics of the montane environment. Bufo boreas halophilus are primarily nocturnal except during breeding and are voluntarily active at body temperatures ranging between 10 and 25° C. Despite variation in T b encountered in the field, boreas select a narrow range of T b in a thermal gradient, averaging 23.5 and 26.2° C for fasted individuals maintained under field conditions or acclimated to 20° C, respectively. In a thermal gradient the mean T b of fasted halophilus acclimated to 20° C is 23.9° C. Skin color of boreas varies in the field from very dark to light. The dark skins absorb approximately 4% more radiation than the light ones. Light colored boreas should absorb approximately 5% more radiation than similarly colored halophilus. Evaporative water losses increase directly with skin temperatures and vapor pressure deficit in both subspecies. Larger individuals heat and cool more slowly than smaller ones. Calculation of an enery budget for boreal toads suggests that they could sit in direct sunlight for long periods without fatally overheating, providing the skin was continually moist.