Design and setup of intermittent-flow respirometry system for aquatic organisms.

Intermittent-flow respirometry is an experimental protocol for measuring oxygen consumption in aquatic organisms that utilizes the best features of closed (stop-flow) and flow-through respirometry while eliminating (or at least reducing) some of their inherent problems. By interspersing short periods of closed-chamber oxygen consumption measurements with regular flush periods, accurate oxygen uptake rate measurements can be made without the accumulation of waste products, particularly carbon dioxide, which may confound results. Automating the procedure with easily available hardware and software further reduces error by allowing many measurements to be made over long periods thereby minimizing animal stress due to acclimation issues. This paper describes some of the fundamental principles that need to be considered when designing and carrying out automated intermittent-flow respirometry (e.g. chamber size, flush rate, flush time, chamber mixing, measurement periods and temperature control). Finally, recent advances in oxygen probe technology and open source automation software will be discussed in the context of assembling relatively low cost and reliable measurement systems.

Sources of variation in oxygen consumption of aquatic animals demonstrated by simulated constant oxygen consumption and respirometers of different sizes.

As intermittent-flow respirometry has become a common method for the determination of resting metabolism or standard metabolic rate (SMR), this study investigated how much of the variability seen in the experiments was due to measurement error. Experiments simulated different constant oxygen consumption rates (M˙O2 ) of a fish, by continuously injecting anoxic water into a respirometer, altering the injection rate to correct for the washout error. The effect of respirometer-to-fish volume ratio (RFR) on SMR measurement and variability was also investigated, using the simulated constant M˙O2 and the M˙O2 of seven roach Rutilus rutilus in respirometers of two different sizes. The results show that higher RFR increases measurement variability but does not change the mean SMR established using a double Gaussian fit. Further, the study demonstrates that the variation observed when determining oxygen consumption rates of fishes in systems with reasonable RFRs mainly comes from the animal, not from the measuring equipment.

Metabolic and cardiorespiratory responses of summer flounder Paralichthys dentatus to hypoxia at two temperatures.

To quantify the tolerance of summer flounder Paralichthys dentatus to episodic hypoxia, resting metabolic rate, oxygen extraction, gill ventilation and heart rate were measured during acute progressive hypoxia at the fish's acclimation temperature (22° C) and after an acute temperature increase (to 30° C). Mean ±s.e. critical oxygen levels (i.e. the oxygen levels below which fish could not maintain aerobic metabolism) increased significantly from 27 ± 2% saturation (2·0 ± 0·1 mg O(2) l(-1)) at 22° C to 39 ± 2% saturation (2·4 ± 0·1 mg O(2) l(-1)) at 30° C. Gill ventilation and oxygen extraction changed immediately with the onset of hypoxia at both temperatures. The fractional increase in gill ventilation (from normoxia to the lowest oxygen level tested) was much larger at 22° C (6·4-fold) than at 30° C (2·7-fold). In contrast, the fractional decrease in oxygen extraction (from normoxia to the lowest oxygen levels tested) was similar at 22° C (1·7-fold) and 30° C (1·5-fold), and clearly smaller than the fractional changes in gill ventilation. In contrast to the almost immediate effects of hypoxia on respiration, bradycardia was not observed until 20 and 30% oxygen saturation at 22 and 30° C, respectively. Bradycardia was, therefore, not observed until below critical oxygen levels. The critical oxygen levels at both temperatures were near or immediately below the accepted 2·3 mg O(2) l(-1) hypoxia threshold for survival, but the increase in the critical oxygen level at 30° C suggests a lower tolerance to hypoxia after an acute increase in temperature.

Comparative metabolic rates of common western North Atlantic Ocean sciaenid fishes.

The resting metabolic rates (R(R)) of western North Atlantic Ocean sciaenids, such as Atlantic croaker Micropogonias undulatus, spot Leiostomus xanthurus and kingfishes Menticirrhus spp., as well as the active metabolic rates (R(A)) of M. undulatus and L. xanthurus were investigated to facilitate inter and intraspecific comparisons of their energetic ecology. The R(R) of M. undulatus and L. xanthurus were typical for fishes with similar lifestyles. The R(R) of Menticirrhus spp. were elevated relative to those of M. undulatus and L. xanthurus, but below those of high-energy-demand species such as tunas Thunnus spp. and dolphinfish Coryphaena hippurus. Repeated-measures non-linear mixed-effects models were applied to account for within-individual autocorrelation and corrected for non-constant variance typical of noisy R(A) data sets. Repeated-measures models incorporating autoregressive first-order [AR(1)] and autoregressive moving average (ARMA) covariances provided significantly superior fits, more precise parameter estimates (i.e. reduced s.e.) and y-intercept estimates that more closely approximated measured R(R) for M. undulatus and L. xanthurus than standard least-squares regression procedures.

CARDIO--a Lotus 1-2-3 based computer program for rapid calculation of cardiac output from dye or thermal dilution curves.

We have developed a menu-driven computer program (CARDIO), based on a Lotus 1-2-3 template and a series of macrocommands, that rapidly and semiautomatically calculates cardiac output from dye or thermal dilution curves. CARDIO works with any dye or thermal dilution recorder with an analog output, any analog to digital (A-to-D) conversion system, and any computer capable of running Lotus 1-2-3 version 2. No prior experience with Lotus 1-2-3 is needed to operate CARDIO, but experienced users can take full advantage of Lotus 1-2-3's graphics, data manipulation, and data retrieval capabilities.

Oxygen transport and cardiovascular responses in skipjack tuna (Katsuwonus pelamis) and yellowfin tuna (Thunnus albacares) exposed to acute hypoxia.

Responses to acute hypoxia were measured in skipjack tuna (Katsuwonus pelamis) and yellowfin tuna (Thunnus albacares) (approximately 1-3 kg body weight). Fish were prevented from making swimming movements by a spinal injection of lidocaine and were placed in front of a seawater delivery pipe to provide ram ventilation of the gills. Fish could set their own ventilation volumes by adjusting mouth gape. Heart rate, dorsal and ventral aortic blood pressures, and cardiac output were continuously monitored during normoxia (inhalant water (PO2 greater than 150 mmHg) and three levels of hypoxia (inhalant water PO2 approximately 130, 90, and 50 mmHg). Water and blood samples were taken for oxygen measurements in fluids afferent and efferent to the gills. From these data, various measures of the effectiveness of oxygen transfer, and branchial and systemic vascular resistance were calculated. Despite high ventilation volumes (4-7 l.min-1.kg-1), tunas extract approximately 50% of the oxygen from the inhalant water, in part because high cardiac outputs (115-132 ml.min-1.kg-1) result in ventilation/perfusion conductance ratios (0.75-1.1) close to the theoretically ideal value of 1.0. Therefore, tunas have oxygen transfer factors (ml O2.min-1.mmHg-1.kg-1) that are 10-50 times greater than those of other fishes. The efficiency of oxygen transfer from water in tunas (approximately 65%) matches that measured in teleosts with ventilation volumes an order of magnitude lower. The high oxygen transfer factors of tunas are made possible, in part, by a large gill surface area; however, this appears to carry a considerable osmoregulatory cost as the metabolic rate of gills may account for up 70% of the total metabolism in spinally blocked (i.e., non-swimming) fish. During hypoxia, skipjack and yellowfin tunas show a decrease in heart rate and increase in ventilation volume, as do other teleosts. However, in tunas hypoxic bradycardia is not accompanied by equivalent increases in stroke volume, and cardiac output falls as HR decreases. In both tuna species, oxygen consumption eventually must be maintained by drawing on substantial venous oxygen reserves. This occurs at a higher inhalant water PO2 (between 130 and 90 mmHg) in skipjack tuna than in yellowfin tuna (between 90 and 50 mmHg). The need to draw on venous oxygen reserves would make it difficult to meet the oxygen demand of increasing swimming speed, which is a common response to hypoxia in both species.(ABSTRACT TRUNCATED AT 400 WORDS)

Metabolic effects of dehydroabietic acid on rainbow trout erythrocytes.

Oxygen consumption and ATP concentration were measured in rainbow trout erythrocytes incubated in a physiological saline containing 0, 5, 15, 30 or 60 mg/l dehydroabietic acid. DHAA caused a decrease in cellular ATP level and oxygen consumption at concentrations above 15 mg/l. Haemolysis increased markedly, when the cellular ATP concentration decreased below 1 mM. These data suggest that increased breakdown of red cells may be the primary reason for jaundice which is observed in resin acid-toxicated fish.

The metabolic rate of an active, tropical elasmobranch, the lemon shark (Negaprion brevirostris).

The oxygen consumption of an active tropical elasmobranch, the lemon shark (Negaprion brevirostris), was measured at different swimming speeds in an annular respirometer. The resulting power-performance curve had a slope of 0.36 which is similar to that of moderately active teleosts. The standard metabolic rate of this elasmobranch (95 ml O2.kg-1.h-1 was higher than that of other sharks, whereas the routine metabolic rate (110 ml O2.kg-1.h-1) was less than that of other elasmobranchs previously studied. Active metabolic rate (226 ml O2.kg-1.h-1), measured in one individual, was the highest for any elasmobranch measured to date. The cost of living of the tropical water lemon shark was calculated to be approximately 2.5 times greater than that of a temperate water shark, but not as high as a salmonid. It is possible that previous generalizations on the poor aerobic performance of sharks are based on data from the less active, temperate water sharks, and are misleading since the metabolic rate of this more active species compares well with active teleosts.

Spontaneous contractions in elasmobranch vessels in vitro.

Isolated vessels from four elasmobranchs, yellow stingray (Urolophus jamaicensis), clearnose skate (Raja eglanteria), ghost shark (Hydrolagus novaezelandiae), and spiny dogfish (Squalus acanthias), were examined for the presence of spontaneous contractions (SC). SC were observed in otherwise unstimulated dorsal aortas (DA) from stingray and ghost shark, but not in skate DA. Unstimulated ventral aortas (VA) did not exhibit SC. After treatment of VA with a contractile agonist, SC appeared in stingray and skate but not ghost shark or dogfish. SC in stingray VA were subsequently inhibited by either epinephrine (10(-5) M) or indomethacin (10(-4) M). Agonist contraction also elicited strong SC in ductus Cuvier from stingray, but not from ghost shark or dogfish. SC in dogfish hepatic portal veins (HPV) produced a rhythmical oscillation in tension. The frequency of HPV SC was highest (approximately 1 min(-1)) in intact veins and lower (approximately 3 min(-1)) in vein segments, indicative of a dominant pacemaker in the intact vessel. SC in HPV were depressed during the first 30 min of hypoxia, but there was substantial recovery over an additional 30 min of hypoxia and complete recovery upon return to normoxia. Addition of 80 mM KCl completely inhibited HPV SC and lowered resting tone. These results show that SC are a common feature of elasmobranch vessels and there appears to be a correlation between swimming behavior and the propensity for SC. KCl inhibition of SC and tonus in HPV is highly unusual for vascular smooth muscle.