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Students are challenged in transitioning from acquiring knowledge and understanding through reading textbooks to their learning to select, read, evaluate, and synthesize the primary literature. A customary approach to teaching this transition to beginning graduate students is for a faculty member to assign “readings” from the recent literature that promise to become key publications; such assignments generally underscore recent, novel scientific content. We advocate here an alternative approach for coaching students very early in their training: first, to read, analyze, and discuss a paper that highlights critically important features of effective and valid experimental design; and, second, to study a paper that can be shown historically to have fundamentally changed the way in which physiological function is understood. We consider as an example of the first goal a study that purports to demonstrate a principle of thermoregulation, but that interaction between students and instructor reveals the study’s lack of an essential control. The second goal requires sufficient time for the publication to concretely validate its contribution(s). The purpose is to identify those essential properties of the selected paper that contributed to its having become a truly exemplary study. We present a 1957 paper by Dr. A. C. Burton ( Am Heart J 54: 801–810, 1957) as an illustration and analyze the study with respect to those attributes that contributed to its lasting importance. These alternative approaches to introduce inexperienced students to the original literature can produce critical insight into the process and can help students inculcate essential practices, guiding them to more productive careers.

Background Rodent models are invaluable for studying biological processes in the context of whole organisms. The reproducibility of such research is based on an assumption of metabolic similarity between experimental animals, controlled for by breeding and housing strategies that minimise genetic and environmental variation. Here, we set out to demonstrate the effect of experimental uraemia on the rat urinary metabolome and gut microbiome but found instead that the effect of vendor shipment batch was larger in both areas than that of uraemia. Results Twenty four Wistar rats obtained from the same commercial supplier in two separate shipment batches underwent either subtotal nephrectomy or sham procedures. All animals undergoing subtotal nephrectomy developed an expected uraemic phenotype. The urinary metabolome was studied using 1H-NMR spectroscopy and found to vary significantly between animals from different batches, with substantial differences in concentrations of a broad range of substances including lactate, acetate, glucose, amino acids, amines and benzoate derivatives. In animals from one batch, there was a complete absence of the microbiome-associated urinary metabolite hippurate, which was present in significant concentrations in animals from the other batch. These differences were so prominent that we would have drawn quite different conclusions about the effect of uraemia on urinary phenotype depending on which batch of animals we had used. Corresponding differences were seen in the gut microbiota between animals in different batches when assessed by the sequencing of 16S rRNA gene amplicons, with higher alpha diversity and different distributions of Proteobacteria subtaxa and short-chain fatty acid producing bacteria in the second batch compared to the first. Whilst we also demonstrated differences in both the urinary metabolome and gut microbiota associated with uraemia, these effects were smaller in size than those associated with shipment batch. Conclusions These results challenge the assumption that experimental animals obtained from the same supplier are metabolically comparable, and provide metabolomic evidence that batch-to-batch variations in the microbiome of experimental animals are significant confounders in an experimental study. We discuss strategies for reducing such variability and the need for transparency in research publications about the supply of experimental animals. Electronic supplementary material The online version of this article (10.1186/s40168-019-0738-y) contains supplementary material, which is available to authorized users.

Pulmonary and cardiovascular dysfunctions are leading causes of morbidity and mortality in patients with chronic Spinal Cord Injury (SCI). Impaired respiratory motor function and decreased Baroreflex Sensitivity (BS) are predictors for the development of cardiopulmonary disease. This observational case-controlled clinical study was undertaken to investigate if respiratory motor control deficits in individuals with SCI affect their ability to perform the Valsalva maneuver, and to determine if a sustained Maximum Expiratory Pressure (MEP) effort can serve as an acceptable maneuver for determination of the BS in the event that the Valsalva maneuver cannot be performed. The BS outcomes (ms/mmHg) were obtained using continuous beat-to-beat arterial blood pressure (BP) and heart rate (HR) recordings during Valsalva or MEP maneuvers in thirty nine individuals with chronic C3-T12 SCI. Twenty one participants (54%) reported signs of intolerance during the Valsalva maneuver and only 15 individuals (39%) were able to complete this task. Cervical level of injury was a significant risk factor (p = .001) for failing to complete the Valsalva maneuver, and motor-complete injury was a significant risk factor for symptoms of intolerance (p = .04). Twenty eight participants (72%) were able to perform the MEP maneuver; the other 11 participants failed to exceed the standard airway pressure threshold of 27 cmH2O. Neither level nor completeness of injury were significant risk factors for failure of MEP maneuver. When the required airway pressure was sustained, there were no significant differences between BS outcomes obtained during Valsalva and MEP maneuvers. The results of this study indicate that individuals with high-level and motor-complete SCI are at increased risk of not completing the Valsalva maneuver and that baroreflex-mediated responses can be evaluated by using sustained MEP maneuver when the Valsalva maneuver cannot be performed.

To evaluate the effects of long term hypoxia exposure on fish spawning, mature common carp, Cyprinus carpio carpio (Linnaeus) were subjected to either normoxia (7.4+/-0.2 mgO(2)mg O(2) L(-1)) or hypoxia (1.0+/-0.2 mgO(2)O(2) L(-1)) for more than two months. Gonadosomatic index (GSI), and concentrations of serum luteinizing hormone (LH), testosterone (T), and estroldiol (E2) were measured and gonad histology examined. Hypoxia inhibits fish spawning even though the gonad and oocytes developed under hypoxia exposure. LH levels of female carp were significantly decreased upon chronic exposure to hypoxia, and the final oocyte maturation in hypoxic females was significantly retarded. The results indicated that hypoxia may inhibit fish spawning through LH-dependent final oocyte maturation. In addition, no courtship was observed in hypoxic males. In conclusion, hypoxia impairs fish ovulation and, therefore, spawning and reproduction. LH levels were reduced leading to a failure of oocyte maturation. This, along with a lack of courtship by males may be the major mechanisms involved in hypoxic inhibition of reproduction in carp.

Human Type 2 diabetes is associated with increased incidence of hypertension and disrupted blood pressure (BP) circadian rhythm. Db/db mice have been used extensively as a model of Type 2 diabetes, but their BP is not well characterized. In this study, we used radiotelemetry to define BP and the circadian rhythm in db/db mice. We found that the systolic, diastolic, and mean arterial pressures were each significantly increased by 11, 8, and 9 mmHg in db/db mice compared with controls. In contrast, no difference was observed in pulse pressure or heart rate. Interestingly, both the length of time db/db mice were active (locomotor) and the intensity of locomotor activity were significantly decreased in db/db mice. In contrast to controls, the 12-h light period average BP in db/db mice did not dip significantly from the 12-h dark period. A partial Fourier analysis of the continuous 72-h BP data revealed that the power and the amplitude of the 24-h period length rhythm were significantly decreased in db/db mice compared with the controls. The acrophase was centered at 0141 in control mice, but became scattered from 1805 to 0236 in db/db mice. In addition to BP, the circadian rhythms of heart rate and locomotor activity were also disrupted in db/db mice. The mean arterial pressure during the light period correlates with plasma glucose, insulin, and body weight. Moreover, the oscillations of the clock genes DBP and Bmal1 but not Per1 were significantly dampened in db/db mouse aorta compared with controls. In summary, our data show that db/db mice are hypertensive with a disrupted BP, heart rate, and locomotor circadian rhythm. Such changes are associated with dampened oscillations of clock genes DBP and Bmal1 in vasculature.

We investigated autonomic control of cardiovascular function in able-bodied (AB), paraplegic (PARA), and tetraplegic (TETRA) subjects in response to head-up tilt following spinal cord injury. We evaluated spectral power of blood pressure (BP), baroreflex sensitivity (BRS), baroreflex effectiveness index (BEI), occurrence of systolic blood pressure (SBP) ramps, baroreflex sequences, and cross-correlation of SBP with heart rate (HR) in low (0.04–0.15 Hz)- and high (0.15–0.4 Hz)-frequency regions. During tilt, AB and PARA effectively regulated BP and HR, but TETRA did not. The numbers of SBP ramps and percentages of heartbeats involved in SBP ramps and baroreflex sequences increased in AB, were unchanged in PARA, and declined in TETRA. BRS was lowest in PARA and declined with tilt in all groups. BEI was greatest in AB and declined with tilt in all groups. Low-frequency power of BP and the peak of the SBP/HR cross-correlation magnitude were greatest in AB, increased during tilt in AB, remained unchanged in PARA, and declined in TETRA. The peak cross-correlation magnitude in HF decreased with tilt in all groups. Our data indicate that spinal cord injury results in decreased stimulation of arterial baroreceptors and less engagement of feedback control as demonstrated by lower 1) spectral power of BP, 2) number (and percentages) of SBP ramps and barosequences, 3) cross-correlation magnitude of SBP/HR, 4) BEI, and 5) changes in delay between SBP/HR. Diminished vasomotion and impaired baroreflex regulation may be major contributors to decreased orthostatic tolerance following injury.

The slope of the log of power versus the log of frequency in the arterial blood pressure (BP) power spectrum is classically considered constant over the low-frequency range (i.e., “fractal” behavior), and is quantified by β in the relationship “1/ fβ.” In practice, the fractal range cannot extend to indefinitely low frequencies, but factor(s) that terminate this behavior, and determine β, are unclear. We present 1) data in rats ( n = 8) that reveal an extremely low frequency spectral region (0.083–1 cycle/h), where β approaches 0 (i.e., the “shoulder”); and 2) a model that 1) predicts realistic values of β within that range of the spectrum that conforms to fractal dynamics (∼1–60 cycles/h), 2) offers an explanation for the shoulder, and 3) predicts that the “successive difference” in mean BP (mBP) is an important parameter of circulatory function. We recorded BP for up to 16 days. The absolute difference between successive mBP samples at 0.1 Hz (the successive difference, or Δ) was 1.87 ± 0.21 mmHg (means ± SD). We calculated β for three frequency ranges: 1) 0.083–1; 2) 1–6; and 3) 6–60 cycles/h. The β for all three regions differed ( P < 0.01). For the two higher frequency ranges, β indicated a fractal relationship (β6–60/h = 1.27 ± 0.01; β1–6/h = 1.80 ± 0.16). Conversely, the slope of the lowest frequency region (i.e., the shoulder) was nearly flat (β0.083–1 /h = 0.32 ± 0.28). We simulated the BP time series as a random walk about 100 mmHg with ranges above and below of 10, 30, and 50 mmHg and with Δ from 0.5 to 2.5. The spectrum for the conditions mimicking actual BP time series (i.e., range, 85–115 mmHg; Δ, 2.00) resembled the observed spectra, with β in the lowest frequency range = 0.207 and fractal-like behavior in the two higher frequency ranges (β = 1.707 and 2.057). We suggest that the combined actions of mechanisms limiting the excursion of arterial BP produce the shoulder in the spectrum and that Δ contributes to determining β.

Presenting a 15-s pulsed tone, the conditional stimulus (CS+), followed by 0.5-s tail shock, to a well-trained rat causes a sudden, but transient, pressor response (C1). Blood pressure (BP) then drops before increasing again (C2). A steady tone of the same frequency never followed by a shock (a discriminative stimulus, or CS−) evokes a C1but not a C2response. Experiment 1 tested the hypothesis that this BP response pattern does not depend on the nature of the tone (i.e., pulsed vs. steady) used for CS+and CS−. The tones were reversed from the traditional paradigm, above, in nine rats. The C1BP increase for a steady-tone CS+(+4.8 ± 1.9 mmHg, mean change ± SE) and a pulsed CS−(+2.9 ± 1.3 mmHg) did not differ. Conversely, C2showed a clear discrimination (CS+: +5.1 ± 1.2 mmHg, CS−: +0 .7 ± 0.8 mmHg; P < 0.05). Experiment 2 tested the hypothesis that the C1and C2BP responses first appear at different times during training. On training day 1, five 15-s pulsed tones (CS+) were presented to each of 18 rats; the last tone was followed by a tail shock. Likewise, five steady CS−tones never followed by shock were given. Training continued for 2 more days, with each CS+followed by shock. At the end of day 2, CS+evoked a C1BP response (+3.9 ± 0.9 mmHg) but no C2(+0.6 ± 0.4 mmHg, not significant vs. pretone). By the end of day 3, CS+evoked a significant (vs. baseline) C1(+7.3 ± 1.4 mmHg) and C2(+3.3 ± 0.8 mmHg). Conversely, although CS−evoked a C1response (3.5 ± 1.3 mmHg), there was no C2(+0.7 ± 0.5 mmHg; not significant). We conclude that 1) C1and C2are acquired at different rates, 2) early in training C1is an orienting response evoked by both tones, and 3) C2is only acquired as an animal learns to associate the CS+tone with shock. This suggests that C1and C2are controlled by different processes in the brain.

This study quantified the effect of interrupting the descending input to the sympathetic preganglionic neurons on the dynamic behavior of arterial blood pressure (BP) in the unanesthetized rat. BP was recorded for ∼4-h intervals in six rats in the neurally intact state and in the same animals after complete spinal cord transection (SCT) between T4 and T5. In the intact state, power within the frequency range of 0.35–0.45 Hz was 1.53 ± 0.38 mmHg2/Hz (mean ± SD by fast Fourier transform). One week after SCT, power within this range decreased significantly ( P < 0.05) to 0.43 ± 0.62 mmHg2/Hz. To test for self-similarity before and after SCT, we analyzed data using a wavelet (i.e., functionally, a digital bandpass filter) tuned to be maximally sensitive to fluctuations with periods of ∼2, 4, 8, 16, 32, or 64 s. In the control state, all fluctuations with periods of ≥4 s conformed to a “self-similar” (i.e., fractal) distribution. In marked contrast, the oscillations with a period of ∼2 s (i.e., ∼0.4 Hz) were significantly set apart from those at lower frequencies. One day and seven days after the complete SCT, however, the BP fluctuations at ∼0.4 Hz now also conformed to the same self-similar behavior characteristic of the lower frequencies. We conclude that 1) an intact sympathetic nervous system endows that portion of the power spectrum centered around ∼0.4 Hz with properties (e.g., a periodicity) that differ significantly from the self-similar behavior that characterizes the lower frequencies and 2) even within the relatively high frequency range at 0.4 Hz self-similarity is the “default” condition after sympathetic influences have been eliminated.

68A realistic research experience is beneficial to undergraduate students, but it is often difficult for liberal arts colleges to offer this opportunity. We describe two approaches for developing and maintaining an interdisciplinary research program at small colleges. An active and continuing involvement of an individual with extensive research experience is an essential element in both. One model was developed by the faculty of Taylor University, Upland, IN and a research scientist who had retired from a major university to join the Taylor faculty as their first Research Professor. The school’s Science Research Training Program was initially funded by a modest endowment provided by interested alumni and by extramural grants awarded to the Research Professor and to the institution; the program now enjoys significant funding from diverse sources. Taylor is not located near any large research university and consequently supplies all resources required for the experiments and stipends for students pursuing projects full-time during the summer. The second model was developed by the faculty at Asbury College in Wilmore, KY, working with a scientist having a full-time appointment at the University of Kentucky and a part-time appointment at the college. In this approach, Asbury faculty may place their students for a period of training, often during the summer, in a laboratory of a cooperating host faculty at the University of Kentucky or other institution. The host faculty funds the research and pays a stipend to those students who work full-time during the summer. Relationships established between faculty at the College and at the University of Kentucky have been mutually beneficial. The success of both programs is evidenced by the students’ presenting their data at state and national scientific meetings, by their publishing their results in national journals, and by the undergraduate school faculty developing independent research programs.

The objective of this study was to determine how neurons within the right atrial ganglionated plexus (RAGP) and posterior atrial ganglionated plexus (PAGP) interact to modulate right atrial chronotropic, dromotropic, and inotropic function, particularly with respect to their extracardiac vagal and sympathetic efferent neuronal inputs. Surgical ablation of the PAGP (PAGPx) attenuated vagally mediated bradycardia by 26%; it reduced heart rate slowing evoked by vagal stimulation superimposed on sympathetically mediated tachycardia by 36%. RAGP ablation (RAGPx) eliminated vagally mediated bradycardia, while retaining the vagally induced suppression of sympathetic-mediated tachycardia (-83%). After combined RAGPx and PAGPx, vagal stimulation still reduced sympathetic-mediated tachycardia (-47%). After RAGPx alone and after PAGPx alone, stimulation of the vagi still produced negative dromotropic effects, although these changes were attenuated compared with the intact state. Negative dromotropic responses to vagal stimulation were further attenuated after combined ablation, but parasympathetic inhibition of atrioventricular nodal conduction was still demonstrable in most animals. Finally, neither RAGPx nor PAGPx altered autonomic regulation of right atrial inotropic function. These data indicate that multiple aggregates of neurons within the intrinsic cardiac nervous system are involved in sinoatrial nodal regulation. Whereas parasympathetic efferent neurons regulating the right atrium, including the sinoatrial node, are primarily located within the RAGP, prejunctional parasympathetic-sympathetic interactions regulating right atrial function also involve neurons within the PAGP.

We examined the effects of chronic nicotine exposure and dietary salt on the arterial blood pressure (BP) changes learned in response to an acute behavioral stress in the Dahl salt-sensitive rat. Four groups were tested: low salt + vehicle; low salt + nicotine; high salt + vehicle; and high salt + nicotine. Rats were fed a low-salt (0.08% NaCl) or a high-salt (8% NaCl) diet for 4 wk; 2.4 mg · kg−1 · day−1nicotine or vehicle was given via an implanted osmotic minipump for the last 2 wk. All rats were conditioned by following one tone (CS+) with a 0.5-s tail shock; another tone (CS−) was never followed by shock. CS+ in low salt + vehicle and high salt + vehicle-treated rats evoked an initial arterial BP increase (C1), a component of the startle response, and an ensuing, smaller, but more sustained, pressor response (C2), which is acquired with training. In these rats, both C1 and C2 evoked by CS− were significantly smaller than those to CS+, demonstrating that these groups discriminated between the two tests. Conversely, although the low salt + nicotine-treated rats had both the C1 and C2 components of the conditional arterial pressure response, they did not discriminate between CS+ and CS−. Finally, the high salt + nicotine group failed to both discriminate between tones and acquire (i.e., learn) the C2 response. The unconditional response to shock did not differ between groups. We conclude that combined exposure to high salt and to nicotine inhibits the salt-sensitive animal's acquisition of a learned conditional BP response, perhaps because nicotine acts preferentially on those central processes required for associative learning versus those involved in orientating to external stimuli.

This experiment quantified the initial disruption and subsequent adaptation of the blood pressure (BP)-heart rate (HR) relationship after spinal cord transection (SCT). BP and HR were recorded for 4 h via an implanted catheter in neurally intact, unanesthetized rats. The animals were then anesthetized, and their spinal cords were severed at T1–T2 ( n = 5) or T4–T5 ( n = 6) or sham lesioned ( n = 4). BP was recorded for 4 h daily over the ensuing 6 days. The neurally intact rat showed a positive cross correlation, with HR leading BP at the peak by 1.8 ± 0.8 (SD) s. The cross correlation in unanesthetized rats ( n= 2) under neuromuscular blockade was also positive, with HR leading. After SCT at T1–T2, the cross correlation became negative, with BP leading HR, and did not change during the next 6 days. The cross correlation also became negative 1–3 days after SCT at T4–T5, but in four rats by day 6 and thereafter the cross correlation progressively reverted to a positive value. We propose that the positive cross correlation with HR leading BP in the intact rat results from an open-loop control that depends on intact supraspinal input to sympathetic preganglionic neurons in the spinal cord. After descending sympathetic pathways were severed at T1–T2, the intact vagal pathway to the sinoatrial node dominated BP regulation via the baroreflex. We suggest that reestablishment of the positive correlation after SCT at T4–T5 was attributable to the surviving sympathetic outflow to the heart and upper vasculature reasserting some effective function, perhaps in association with decreased spinal sympathetic hyperreflexia. The HR-BP cross correlation may index progression of sympathetic dysfunction in pathological processes.

In marine teleost fishes, the gill mitochondria-rich cells (MRCs) are responsible for NaCl elimination; however, in elasmobranch fishes, the specialized rectal gland is considered to be the most important site for salt secretion. The role of the gills in elasmobranch ion regulation, although clearly shown to be secondary, is not well characterized. In the present study, we investigated some morphological properties of the branchial MRCs and the localization, and activity of the important ionoregulatory enzyme Na(+)/K(+)-ATPase, under control conditions and following rectal gland removal (1 month) in the spiny dogfish, Squalus acanthias. A clear correlation can be made between MRC numbers and the levels of Na(+)/K(+)-ATPase activity in crude gill homogenates (r(2)=-0.69). Strong Na(+)/K(+)-ATPase immunoreactivity is also clearly associated with the basolateral membrane of these MRCs. In addition, the dogfish were able to maintain ionic balance after rectal gland removal. These results all suggest a possible role of the dogfish gill in salt secretion. MRCs were, however, unresponsive to rectal gland removal in terms of changes in number, fine structure and Na(+)/K(+)-ATPase activity, as might be expected if they were compensating for the loss of salt secretion by the rectal gland. Thus, the specific role that these MRCs play in ion regulation in the dogfish remains to be determined

[Figure][1] David Randall discusses George Hughes and Graham Shelton's classic paper ‘Respiratory mechanisms and their nervous control in fish', published in Advances in Comparative Physiology and Biochemistry in 1962. George Hughes and Graham Shelton were members of the Cambridge

Teleost fishes constitute 95% of extant aquatic vertebrates, and we suggest that this is related in part to their unique mode of tissue oxygenation. We propose the following sequence of events in the evolution of their oxygen delivery system. First, loss of plasma-accessible carbonic anhydrase (CA) in the gill and venous circulations slowed the Jacobs–Stewart cycle and the transfer of acid between the plasma and the red blood cells (RBCs). This ameliorated the effects of a generalised acidosis (associated with an increased capacity for burst swimming) on haemoglobin (Hb)–O2 binding. Because RBC pH was uncoupled from plasma pH, the importance of Hb as a buffer was reduced. The decrease in buffering was mediated by a reduction in the number of histidine residues on the Hb molecule and resulted in enhanced coupling of O2 and CO2 transfer through the RBCs. In the absence of plasma CA, nearly all plasma bicarbonate ultimately dehydrated to CO2 occurred via the RBCs, and chloride/bicarbonate exchange was the rate-limiting step in CO2 excretion. This pattern of CO2 excretion across the gills resulted in disequilibrium states for CO2 hydration/dehydration reactions and thus elevated arterial and venous plasma bicarbonate levels. Plasma-accessible CA embedded in arterial endothelia was retained, which eliminated the localized bicarbonate disequilibrium forming CO2 that then moved into the RBCs. Consequently, RBC pH decreased which, in conjunction with pH-sensitive Bohr/Root Hbs, elevated arterial oxygen tensions and thus enhanced tissue oxygenation. Counter-current arrangement of capillaries (retia) at the eye and later the swim bladder evolved along with the gas gland at the swim bladder. Both arrangements enhanced and magnified CO2 and acid production and, therefore, oxygen secretion to those specialised tissues. The evolution of β-adrenergically stimulated RBC Na+/H+ exchange protected gill O2 uptake during stress and further augmented plasma disequilibrium states for CO2 hydration/dehydration. Finally, RBC organophosphates (e.g. NTP) could be reduced during hypoxia to further increase Hb–O2 affinity without compromising tissue O2 delivery because high-affinity Hbs could still adequately deliver O2 to the tissues via Bohr/Root shifts. We suggest that the evolution of this unique mode of tissue O2 transfer evolved in the Triassic/Jurassic Period, when O2 levels were low, ultimately giving rise to the most extensive adaptive radiation of extant vertebrates, the teleost fishes.

The branchial epithelium of the mudskipper Periophthalmodon schlosseri is densely packed with mitochondria-rich (MR) cells. This species of mudskipper is also able to eliminate ammonia against large inward gradients and to tolerate extremely high environmental ammonia concentrations. To test whether these branchial MR cells are the sites of active ammonia elimination, we used an immunological approach to localize ion-transport proteins that have been shown pharmacologically to be involved in the elimination of NH(4)(+) (Na(+)/NH(4)(+) exchanger and Na(+)/NH(4)(+)-ATPase). We also investigated the role of carbonic anhydrase and boundary-layer pH effects in ammonia elimination by using the carbonic anhydrase inhibitor acetazolamide and by buffering the bath water with Hepes, respectively. In the branchial epithelium, Na(+)/H(+) exchangers (both NHE2- and NHE3-like isoforms), a cystic fibrosis transmembrane regulator (CFTR)-like anion channel, a vacuolar-type H(+)-ATPase (V-ATPase) and carbonic anhydrase immunoreactivity are associated with the apical crypt region of MR cells. Associated with the MR cell basolateral membrane and tubular system are the Na(+)/K(+)-ATPase and a Na(+)/K(+)/2Cl(−) cotransporter. A proportion of the ammonia eliminated by P. schlosseri involves carbonic anhydrase activity and is not dependent on boundary-layer pH effects. The apical CFTR-like anion channel may be serving as a HCO(3)(−) channel accounting for the acid-base neutral effects observed with net ammonia efflux inhibition.

A 43-year-old woman was admitted with progressive leg pains and weakness and was found to have rhabdomyolysis. Prior to this admission the patient had hypercalcemia, but this returned to normal following treatment with calcitonin. During the hospitalization, she developed the syndrome of calciphylaxis consisting of necrotic skin and muscle associated with vascular calcification. This is the first case report of rhabdomyolysis caused by calciphylaxis in a patient without chronic renal failure.

A quantitative analysis of O2 and CO2 transport was conducted in resting and exercising rainbow trout, and these data were used to quantify the magnitude of coupling between O2 and CO2 exchange, in vivo. The release of Bohr protons during haemoglobin-oxygenation was non-linear over the Hb-O2 equilibrium curve used in trout subjected to different levels of sustained exercise. At low swimming speeds, when venous blood O2 content (CvO2) was high, there was a small acidosis as blood passed through the gills, indicating more protons were released during oxygenation of Hb than were consumed during HCO3- dehydration. At higher swimming speeds, when CvO2 was low, there was a significant alkalosis in arterial relative to venous blood, indicating that fewer protons were released upon oxygenation than HCO3- ions were dehydrated to CO2. Haldane coefficients (moles of protons released per mole of O2 which binds to Hb), calculated from steady state arterial and mixed-venous parameters, revealed that under resting conditions all blood CO2 removed from the blood during gill transit was stoichiometrically related to O2 uptake through the release of Bohr protons during Hb oxygenation. The magnitude of coupling between CO2 excretion and O2 uptake decreased from 100% to less than 40% at the maximal swimming velocity when the largest region of the Hb-O2 equilibrium curve was used for gas exchange. The non-linear release of Bohr protons over the range of Hb-O2 saturation in the blood reduces HCO3- dehydration at the gills during greater work loads elevating arterial P(CO2) levels, leading to an increase in HCO3- buffer capacity of the blood and tissues.

Exposure of fish to alkaline conditions clearly inhibits the rate of ammonia excretion (JAMM) leading to problems of ammonia accumulation and toxicity. There is some evidence that higher environmental calcium levels may ameliorate the negative effects of alkaline exposure. In the present study, we investigated the effects of elevated environmental calcium on the recovery of ammonia excretion during acute (6 h) and chronic (15 day) exposures to high pH (pH 10), and the levels of nitrogenous products (ammonia, urea, and glutamine) in tissues and plasma (1–15 days). Acclimation to alkaline conditions results in a chronically depressed ammonia excretion rate. Glutamine as well as urea do not appear to be important in ammonia detoxification and storage. Calculations of the production rates from storage and excretion data indicate a reduction in the rate of ammonia production. Elevating environmental calcium during acute exposure to alkaline conditions does not affect the recovery of JAMM but may reduce stress. We conclude that rainbow trout survive alkaline exposure by reducing ammonia production and that the role of calcium may be in the amelioration of the stress of alkaline exposure.

The role of the posterior atrial ganglionated plexus (PAGP) in heart rate (HR) control was tested in unanesthetized dogs ( n = 8). Resting HR was unchanged before (85 ± 20 beats/min, mean ± SD) versus after (87 ± 18 beats/min) surgical ablation of these intrinsic cardiac ganglia (PAGPX). However, the peak tachycardia to a 30-s stressful stimulus was significantly increased ( P < 0.05) from +53 ± 22 beats/min before the denervation to +77 ± 13 beats/min after PAGPX. Conversely, the peak HR increase during the stress after β-adrenergic blockade was the same before (36 ± 24 beats/min) versus after (38 ± 14 beats/min) PAGPX. Moreover, the HR response to a neutral behavioral stimulus, which is mediated primarily by withdrawal of parasympathetic inhibition of the sinoatrial (SA) node, was unaltered by PAGPX. Thus the augmented tachycardia subsequent to PAGPX was attributable primarily to increased sympathetic action at the SA node. These findings indicate that a major role of PAGP parasympathetic neurons is to inhibit sympathoexcitatory effects on HR, probably either via interactions between neurons comprising the intrinsic plexus(es) or perhaps via presynaptic inhibition of sympathetic neurotransmitter release. This organization would allow parasympathetic ganglia within the PAGP to selectively modify sympathetic input to the SA node independent of direct vagal inhibition of pacemaker activity.

We have described a 0.4-Hz rhythm in renal sympathetic nerve activity (SNA) that is tightly coupled to 0.4-Hz oscillations in blood pressure in the unanesthetized rat. In previous work, the relationship between SNA and fluctuations in mean arterial blood pressure (MAP) was described by a set of two first-order differential equations. We have now modified our earlier model to test the feasibility that the 0.4-Hz rhythm can be explained by the baroreflex without requiring a neural oscillator. In this baroreflex model, a linear feedback term replaces the sympathetic drive to the cardiovascular system. The time delay in the feedback loop is set equal to the time delay on the efferent side, approximately 0.5 s (as determined in the initial model), plus a time delay of 0.2 s on the afferent side for a total time delay of approximately 0.7 s. A stability analysis of this new model yields feedback resonant frequencies close to 0.4 Hz. Because of the time delay in the feedback loop, the proportional gain may not exceed a value on the order of 10 to maintain stability. The addition of a derivative feedback term increases the system's stability for a positive range of derivative gains. We conclude that the known physiological time delay for the sympathetic portion of the baroreflex can account for the observed 0.4-Hz rhythm in rat MAP and that the sensitivity of the baroreceptors to the rate of change in blood pressure, as well as average blood pressure, would enhance the natural stability of the baroreflex.

Effects of adrenaline on the equilibrium distributions of Na(+) , K(+) , H(+) , Cl(-) , and H2O across the cell membrane of rainbow trout (Salmo gairdneri) erythrocytes were determinedin vitro, as a function of P CO2 (1.76-7.77 torr). CO2-carrying capacity of the blood was also examined. Plasma catecholamine concentrations inunanaesthetized, unrestrained trout were 3.1 nM adrenaline and 1.2 nM noradrenaline. Elevation of the plasma adrenaline concentrationin vitro to 4.6 × 10(3) nM resulted in net gains of Na(+) , Cl(-) and H2O by red cells, a net loss of H(+) from red cells, and a pronounced red cell swelling. Adrenaline also reduced the CO2-carrying capacity of trout bloodin vitro. The magnitudes of these effects increased with PCO2 and, thus, were sensitive to blood HCO3 (-) concentrations. The distribution of K(+) between red cells and plasma was unaffected by adrenaline. Adrenergic-mediated ion movements and red cell swelling were sensitive to both propranolol and SITS. These results are consistent with the symport NaCl uptake model for adrenergic-mediated swelling of Baroinet al. (1984). The adrenergic response of fish erythrocytes may function to ameliorate the effects of blood acidoses on O2-carrying capacity by maintaining red cell pH in the face of a decrease in plasma pH.

Summary Teleost fishes and mammalian lineages diverged 400 million years ago, and environmental requirements (water vs. air) have resulted in marked differences in cardiovascular function between fish and mammals. Suggestions that the fish secondary vascular system (SVS) could be used as a model for the mammalian lymphatic system should be taken with caution. Despite molecular markers indicating similar genetic origin, functions of the SVS in teleost fish are probably different from those of the mammalian lymphatic system. We determined that, in resting glass catfish, (Kryptopterus bicirrhis), plasma moves from the primary vascular system (PVS) to the SVS through small connecting vessels less than 10 μm in diameter, smaller than the red blood cells (RBCs). During and following hypoxia or exercise, flow increases, and RBCs enter the SVS, possibly via β-adrenoreceptor-mediated dilation of the connecting vessels. The volume of the SVS can be large and, as RBCs flow into the SVS, the haematocrit of the PVS falls by as much as 50% of the resting value. Possible functions of the SVS, including skin respiration, ionic and osmotic buffering, and reductions in heart work and RBC turnover, are discussed.

The purpose of this study was to examine the role of adenosine receptors (ARs) in (1) the regulation of catecholamine secretion and (2) the modulation of blood oxygen capacitance by catecholamines. To this end, we assessed the response of rainbow trout and Pacific hagfish treated with either an AR blocker, theophylline, or saline under hypoxic and normoxic conditions. Compared with the control hypoxic rainbow trout, AR blockade resulted in a smaller increase in haematocrit and haemoglobin (Hb) concentration of the blood, smaller red blood cell transmembrane pH differences and mean cellular [Hb] (MCHC), as well as a 16-fold higher plasma adrenaline concentration after only 10 min of acute hypoxic exposure. In hypoxic hagfish, AR blockade had no effect on the [Hb] of the blood, and there was no regulation of red blood cell pH or changes in MCHC. However, whereas plasma [adrenaline] did not change following exposure to a of 1.33 kPa in the hypoxic sham group, the concentration increased 3.8-fold within 10 min in the theophylline-injected group. These results suggest that adenosine modulates the circulating level of catecholamines in both hypoxic rainbow trout and hypoxic Pacific hagfish.

An extracorporeal blood circulation and a stopflow technique were used to examine the acid-base status of arterial blood in the rainbow trout, Oncorhynchus mykiss. Arterial blood was routed from the coeliac artery through an external circuit in which pH (pHa), partial pressure of oxygen (PaO2) and partial pressure of carbon dioxide (PaCO2) were monitored continuously. The stopflow condition was imposed by turning off the pump which drove the external loop. A radioisotopic CO2 excretion assay was performed on blood samples collected periodically to evaluate plasma carbonic anhydrase (CA) activity and hence red blood cell (rbc) lysis. An acid-base disequilibrium was found in the post-branchial blood; pHa increased by 0.04-0.06 units, and PaCO2 by 0.03-0.10 Torr, during the stopflow period. The disequilibrium appeared to arise primarily from the slow (uncatalyzed) rate of plasma H2CO3 dehydration. This was confirmed by the intra-arterial injection of bovine CA (22 mg kg-1) prior to the stopflow; the disequilibrium was abolished. When the CA inhibitor acetazolamide (30 mg kg-1) was injected, a negative pH disequilibrium of 0.04 units, accompanied by a rise in PaCO2 of 0.57 Torr, was observed during the stopflow. These results can be explained by the acetazolamide-induced inhibition of rbc CA, which leads to continuing rbc CO2 "excretion" in the post-branchial blood.

The effects of neuropeptide Y (NPY) alone and in combination with noradrenaline (NA) on cardiac performance were determined in isolated perfused dogfish (Squalus acanthias) hearts. Since inositol 1, 4,5-trisphosphate (Ins(1,4,5)P₃) is known to be an important second messenger in the regulation of cardiac function, Ins(1,4,5)P₃ levels in these hearts were measured with an Ins(1,4,5)P₃ protein binding assay kit. Neuropeptide Y increased cardiac output by increasing heart rate, as stroke volume was not changed, whereas NA increased cardiac output by increasing stroke volume. Neuropeptide Y or NA alone did not have significant effect on pressure development. Neuropeptide Y plus NA, however, increased pressure development. Inositol 1,4,5-trisphosphate level was elevated by NA alone and was further increased by NPY plus NA. The present study indicates that NPY could potentiate the effect of NA in the dogfish heart. The changes in Ins(1,4,5)P₃ may be implicated in the cardiac effects of NA and NPY in these hearts.

We recorded via telemetry the arterial blood pressure (BP) and heart rate (HR) response to classical conditioning following the spontaneous onset of autoimmune diabetes in BBDP/Wor rats versus age-matched, diabetes resistant control (BBDR/Wor) rats. Our purpose was to evaluate the autonomic regulatory responses to an acute stress in a diabetic state of up to 12 months duration. The stress was a 15 sec. pulsed tone (CS+) followed by a 0.5 sec. tail shock. The initial, transient increase in BP (i.e., the ‘first component’, or C1), known to be derived from an orienting response and produced by a sympathetic increase in peripheral resistance, was similar in diabetic and control rats through ~9 months of diabetes; it was smaller in diabetic rats 10 months after diabetes onset. Weakening of the C1 BP increase in rats that were diabetic for > 10 months is consistent with the effects of sympathetic neuropathy. A longer-latency, smaller, but sustained ‘second component’ (C2) conditional increase in BP, that is acquired as a rat learns the association between CS+ and the shock, and which results from an increase in cardiac output, was smaller in the diabetic vs. control rats starting from the first month of diabetes. A concomitant HR slowing was also smaller in diabetic rats. The difference in the C2 BP increase, as observed already during the first month of diabetes, is probably secondary to the effects of hyperglycemia upon myocardial metabolism and contractile function, but it may also result from effects on cognition. The small HR slowing concomitant with the C2 pressor event is probably secondary to differences in baroreflex activation or function, though parasympathetic dysfunction may contribute later in the duration of diabetes. The nearly immediate deficit after disease onset in the C2 response indicates that diabetes alters BP and HR responses to external challenges prior to the development of structural changes in the vasculature or autonomic nerves.

Nitrite oxidizes haemoglobin (Hb) to methaemoglobin (MetHb), which is unable to bind oxygen. Nitrite exposure can therefore be used as a tool to manipulate the oxygen-carrying capacity of the blood without changing haematocrit. The objective of this study is to examine the relationship between the critical swimming velocity (Ucrit) and the functional haemoglobin concentration ([Hb]) of the blood in adult chinook salmon. Functional [Hb] was reduced by increasing MetHb levels through intraperitoneal administration of a mass-dependent volume of sodium nitrite. In resting fish, MetHb levels were found to stabilize at 25% of total [Hb] 3h after the injection of 30mgkg−1 sodium nitrite. Methaemoglobin levels increased in proportion to the amount of sodium nitrite injected and reached a maximum (following the injection of 90mgkg−1 sodium nitrite) of 51.8% in resting fish and 72% in fish forced to swim to Ucrit. At 60 and 90mgkg−1 sodium nitrite, MetHb formation was greater in exercised than in resting fish. A second-order regression revealed that Ucrit was virtually independent of functional [Hb] between 51 and 100% of control functional [Hb], but was positively correlated with functional [Hb] below 51% of total [Hb] (4.5 g dl−1). The insensitivity of Ucrit to a functional [Hb] greater than 51% may be partly due to the exponential increase in aerobic metabolism required to provide the power to overcome hydrodynamic drag at higher water velocities. There were no significant changes in intraerythrocytic organic phosphate (adenylates and guanylates) concentrations standardized to [Hb] in swimming or resting fish over the range of MetHb levels induced in this study. Fish may encounter nitrite naturally; if MetHb levels become severely elevated as a result, swimming ability will be significantly impaired.

The Lake Magadi tilapia, Oreochromis alcalicus grahami, is remarkable among teleosts in that it flourishes under extremely well-buffered alkaline water conditions (pH10, 180mmol l−1) at temperatures of 30–40°C (Wood et al. 1989). As expected from current models in teleosts, ammonia excretion into such water would be difficult at best (Wood, 1993). Part of the survival strategy of the Lake Magadi tilapia is that it has a complete ornithine–urea cycle (O–UC) in the liver and excretes virtually all of its waste nitrogen as urea (Randall et al. 1989). Ammonia toxicity in ammoniotelic teleosts has been studied extensively, and typical values for unionized ammonia (NH3) 96h LC 50 (the concentration at which half of test subjects die after 96h) are well below 100 μmol l−1 (Haywood, 1983; Thurston et al. 1983a,b; Campbell, 1991). Surprisingly, no ammonia LC50 values are available for ureogenic teleost fish, and one would predict that fish synthesizing and excreting urea for whatever purpose would have higher LC50 values than their ammoniotelic counterparts. Additionally, since ammonia exposure has been implicated in the functional response of urea excretion in the Lake Magadi tilapia (Wood et al. 1989) and another ureogenic teleost (the gulf toadfish Opsanus beta) (Walsh et al. 1990), we reasoned that ammonia exposure in the Lake Magadi tilapia might reveal insights into the biochemical regulation of the O–UC in this species; in particular that it might induce enzyme activity. We report here that the Lake Magadi tilapia has a rather high ammonia LC50 compared to values for other teleosts, but that short-term ammonia exposure has very limited effects on the activities of the enzymes of nitrogen metabolism and on swimming performance.

N-ethymaleimide-sensitive ATPase activity was measured in crude homogenates of gill tissue from rainbow trout using a coupled-enzyme ATPase assay in the presence of EGTA, ouabain and azide. This NEM-sensitive ATPase activity, determined to be about 1.5 μmolmg−1 protein h−1 at 15°C for freshwater trout, is also inhibited by other H+-ATPase blockers such as DCCD, DES, PCMBS and bafilomycin. It is concluded, therefore, that the NEM-sensitive ATPase activity was generated by a proton-translocating ATPase. Since this NEM-sensitive ATPase was also sensitive to the plasma membrane ATPase inhibitor vanadate, we conclude that the H+-ATPase in fish gill is of the plasma membrane type. The major role of the H+-ATPase in the gill epithelium is to facilitate Na+ uptake from fresh water. Sodium concentration in the external medium was the primary regulator of the H+-ATPase in fish gills, with low sodium levels being associated with high H+-ATPase activity. High external calcium concentration had a marked stimulatory effect on H+-ATPase activity in fish gills when the sodium level was low. Environmental hypercapnia induced a 70% increase in the H+-ATPase activity in fish gills. H+-ATPase activity was also elevated in freshwater fish after chronic cortisol infusion.

We investigated ventillatory responses to a plasma alkaloids and hypocapnia,a nd the basis for the ventilatory response to sodium bicarbonate (NaHCO3) infusion in rainbow trout. Plasma alkalosis and hypocapnia created by infusion of sodium hydroxide (NaOH) did not cause hypoventilation, whereas infusion of hydrochloric acid (HCl) caused vigorous hyperventilation, associated with an acidosis, a reduction in blood O2 content (CaO 2) and a release of circulating catecholamines. Infusion of NaHCO3 stimulated ventilation and caused an increase in plasma pH, total carbon dioxide content (CaCO 2) and catecholamine levels, and a reduction in oxygen tension (PaO 2). Infusion of ammonium bicarbonate (NH 4HCO3) caused hyperventilation and was associated with an increase in CaCO 2 and plasma total ammonia (Camm) and ammonia gas (NH3) concentration. Infusion of sodium chloride (NaClI) and Cortland's saline had no effect on ventilation. The results indicate that trout do not exhibit the ventilatory sensitivity to pH seen in terrestrial vertebrates. Ventilatory responses to NaHCO3 appear to have been a result of reductions in PaO 2, a release of catecholamines and an increase in CaCO 2 whereas responses to NH4HCO3 appear to have been a result of increases in CaCO 2 and Camm.

This study investigated the possible role of catecholamines in the ventilatory response of rainbow trout (Oncorhynchus mykiss) to acute external hypercapnia. The ventilatory response to hypercapnia [partial pressure of CO2 in water of fish pre-treated with the selective β-adrenoceptor antagonist, D,L-propranolol, was compared with that of D-propranolol (an isomer with minimal β-antagonistic activity) and saline pre-treated fish (sham). A sustained 3.6-fold increase in gill ventilation volume was observed in the sham and D-propranolol-treated groups during the 30min interval of hypercapnia. Fish pre-treated with D,L-propranolol displayed a blunted hyperventilatory response to hypercapnia (1.9-fold increase at 30min). These results indicate that the β-component of an adrenergic response is involved in the usual hyperventilatory response to external hypercapnia. It is suggested that the impaired hyperventilatory response of the D,L-propranolol-treated group reflects an inhibition of central adrenergic mechanism(s) involved in the hyperventilatory reflex to respiratory acidosis.

We investigated the effects of a diet enriched in omega-3 (ω3) polyunsaturated fatty acids (PUFA) and vitamin E on responses of sturgeon (Acipenser naccarii) to hypoxia. After 3 months of feeding, there were significant increases in ω3 PUFA in liver and muscle, and of vitamin E in muscle, of fish fed the enriched diet (ED) compared with fish on a standard diet (SD), indicating that tissue composition is influenced by diet. Acute exposure to hypoxia (10 min at 10.8 kPa water O2 tension, ) had no effect on oxygen consumption , increased gill ventilation frequency (fg) and reduced arterial blood O2 content in both dietary groups, but ED sturgeon exhibited a significantly smaller decrease in than did SD animals. Progressive hypoxic exposure ( decreasing gradually from 20.5 to 3.6kPa within 45–60min) led to a significant increase in at intermediate levels of in SD sturgeon that was not seen in ED animals. Furthermore, ED sturgeon showed no significant reduction in arterial plasma pH (pHa) and at levels that caused significant reductions in these variables in SD sturgeon. ED sturgeon exhibited a smaller increase in plasma lactate level than did SD fish. We suggest that PUFA and/or vitamin E contribute significantly to regulation of metabolism in hypoxia.

The carbon dioxide content of the intracellular compartment of fish muscle was determined by direct measurements of CO2 and pH in tissue homogenates of rainbow trout, Oncorhynchus mykiss. The results agree with the concept that compartmental distribution of CO2 is pH-dependent and that muscle membranes are not very permeable to bicarbonate. The interaction between CO2 and ammonia excreted from fish muscle was also investigated by altering plasma CO2 content using bicarbonate infusion following exhaustive exercise. Removal of the acid boundary layer in white muscle by bicarbonate infusion resulted in retention of ammonia in the muscle, indicating that ammonia excretion across the muscle membrane might be enhanced by the hydration of excreted CO2 in the extracellular fluid. Passive diffusion of NH3, rather than NH4+ transfer, is probably the dominant pathway of ammonia excretion through fish muscle membranes.

Ion transport inhibitors, amiloride, SITS, vanadate and acetazolamide, were added to the water to determine the effect of ion transfer mechanisms on the acidification of water passing over the gills. In neutral water, proton excretion causes a marked reduction in gill water pH. If water pH is 2.5 units lower than blood pH, however, then this proton excretion is inhibited and all water pH changes can be accounted for by CO2 hydration and ammonia protonation. Proton excretion across the gills is insensitive to 0.1 mmol l−1 amiloride and SITS but sensitive to vanadate, acetazolamide and water pH; thus, we conclude that proton excretion is mediated by an active proton pump on the apical membrane of the gill epithelium similar to that reported for the frog skin. Higher concentrations of amiloride (0.5 and 1 mmol l−1) reduced both ammonia and acid excretion, presumably because of inhibition of Na+/K+-ATPase on the basolateral border of the gill epithelium.

The role of sensory afferent information from the gills of Amia calva in cardiovascular and ventilatory control was investigated by bilateral branchial denervation and pseudobranch ablation. Aquatic hypoxia or 1 mg of sodium cyanide (NaCN) in the water flowing over the gills stimulated bradycardia, and gill and air ventilation in sham-operated fish. Sodium cyanide, noradrenaline (NA) and adrenaline (A) infusion into the dorsal aorta increased gill ventilation, and NA and A infusion also stimulated tachycardia and an increase in blood pressure. Following denervation and pseudobranch ablation, O2 consumption (V˙O2) air-breathing frequency (fAB) and arterial O2 tension (PaO2) declined, and circulating NA levels increased, as compared with sham-operated fish. Cardiovascular and air-breathing responses to hypoxia were abolished and gill ventilatory responses attenuated. All ventilatory and cardiovascular responses to NaCN were abolished and gill ventilatory responses to NA and A were attenuated in animals following denervation and pseudobranch ablation. These results demonstrate that O2-sensitive chemoreceptors in the gills and pseudobranch control reflex bradycardia and air-breathing responses in Amia, but that gill ventilatory responses to hypoxia, NA and A are partially mediated by extrabranchial mechanisms. Plasma NA levels increased during hypoxia in sham-operated and denervated animals, indicating that circulating NA may have mediated gill ventilatory responses in denervated animals.

The critical swimming velocity of fish is affected by environmental conditions and the highest swimming speed is obtained only under specific circumstances. The mechanisms causing a reduction in exercise capacity depend on the type of environmental change. Acid waters exert an effect by reducing oxygen transport capacity, whereas reduced temperatures act largely on muscle contractility. Alkaline conditions and salinity change may affect both gas transport and muscle contractility. A fish must operate over a wide range of internal and external conditions and must possess both short- and long-term mechanisms to maintain function under a wide variety of conditions. These mechanisms may be part of the immediate reflex arsenal available to the fish (for example, catecholamine release) or they may be mechanisms induced during acclimation. The nature of the acclimation process is an indication of what may be limiting exercise under a particular set of circumstances. The concept of symmorphosis, especially when applied to ectotherms, needs to be viewed in a broader context. A symmetry of component parts may exist such that structures are designed to satisfy functional requirements for operation over a wide range of conditions, rather than optimally for a given set of conditions. This must involve compromises in design, especially for ectotherms such as fish. Thus, the more variable the conditions under which a system must operate, the less apparent it will be that symmorphosis exists between the component parts.

Pharmacological blockade of the renin-angiotensin converting enzyme reportedly alters the heart rate (HR) power spectrum in conscious dogs, suggesting that these hormones contribute to the short-term regulation of arterial blood pressure. We tested this possibility using four independent procedures. First, HR power spectrum was determined in seven awake dogs before and after administration of enalaprilat (300 ng/kg), a converting-enzyme inhibitor. There were no significant changes in the average amplitude for the spectral peak between 0.003 and 0.1 Hz (i.e., the "low-frequency peak"). Second, the HR power spectrum was measured in 11 awake rabbits before and after treatment with deoxycorticosterone acetate (1 mg.kg-1.day-1) and salt (0.9% saline ad libitum) for 7 days to depress plasma renin levels. There were no significant changes in the amplitude of the HR power spectrum, although mean HR decreased from 206 +/- 3 to 184 +/- 4 beats/min after treatment. In the third experiment, another group of rabbits (n = 8) was tested after 2 wk on a low-salt diet to elevate plasma angiotensin levels and then after 2 wk on a normal salt diet. Once again there were no significant effects on the HR power spectrum. Finally, tranquilized dogs (n = 9) were subjected to sinusoidally varying lower body negative pressure at selected frequencies of 0.008-0.12 Hz. Tests were conducted in the control state and after administration of an angiotensin receptor antagonist (saralasin, 1 microgram.kg-1.min-1). Lower body negative pressure-induced fluctuations in arterial blood pressure were similar in both states. We find no evidence for the role of the renin-angiotensin system in the moment-to-moment regulation of arterial pressure and HR.

Hydrochloric acid (HCl) infusion during aquatic normoxia increased blood pressure, gill ventilation, and air breathing in bowfin. These responses were associated with a reduction in arterial blood pH ($pH_{a}$) and O₂, content ($Ca_{O_{2}}$) and an increase in arterial plasma CO₂ tension ($Pa_{CO_{2}}$). Ammonium bicarbonate (NH₄HCO₃) infusion did not change $pH_{a}$ or $Ca_{O_{2}}$ but increased $Pa_{CO_{2}}$ and had no effects on blood pressure or ventilation. In aquatic hyperoxia, HCl infusion lowered $pH_{a}$ and $Ca_{O_{2}}$. Arterial blood O₂ content, however, was maintained above normoxic levels, and there were no cardiovascular or ventilatory responses. These results indicate that cardiovascular and ventilatory responses to acid-base disturbances in bowfin are correlated with reductions in $Ca_{O_{2}}$ below normoxic levels and not with decreases in $pH_{a}$ or increases in $Pa_{CO_{2}}$. Hydrochloric acid infusion in normoxia stimulated a release of endogenous catecholamines, whereas NH₄HCO₃ infu...

The purpose of this study was to quantify the relative roles of the canine cardiac parasympathetic and sympathetic nerves in controlling the distribution of power within the heart rate (HR) power spectrum using a highly selective surgical technique to parasympathectomize the SA node. DAta were recorded in awake dogs (n = 6) before and after the selective denervation; the animals were isolated from human contact and their behavior carefully monitored during the measurements. The average amplitude in the high-frequency (approximately 0.32 Hz) peak in the HR power spectrum decreased from a predenervation control of 2.68 +/- 1.54 (mean +/- SD, arbitrary units) to 0.07 +/- 0.06 (P less than 0.05). Corresponding resting HR increased from 80 +/- 9 to 106 +/- 16 beats/min (P less than 0.05). The low-frequency peak (approximately 0.02 Hz) also decreased from a control of 2.45 +/- 1.18 to a postparasympathectomy value of 1.25 +/- 0.92 (P less than 0.05). beta-Adrenergic blockade (propranolol, 1 mg/kg) further decreased the latter peak to 0.59 +/- 0.52 (P less than 0.05). These data directly demonstrate that the high-frequency peak of the HR power spectrum 1) results from parasympathetic control of SA nodal automaticity, while 2) the low-frequency peak reflects activity in both divisions of the autonomic nervous system.

Plasma catecholamine levels and gill ventilation were measured in rainbow trout (Oncorhynchus mykiss) during acidosis and hypoxia. There was an increase in both plasma catecholamines and ventilation correlated with the acidosis. Fish exposed to hyperoxia prior to acid infusion did not show significant changes in catecholamines or ventilation. Those treated with the β-adrenergic antagonist propranolol before acidosis showed increases in catecholamines but not ventilation. Hypoxia was also associated with increases in endogenous catecholamines and ventilation, and the increase in ventilation could be partially blocked with propranolol. This increase in ventilation during hypoxia was not inhibited by a saline injection alone. It is proposed that catecholamines act to modulate ventilatory responses in fish under both acidotic and hypoxic conditions. If a central H+ chemoreceptor exists in fish to control breathing, it can be inhibited by hyperoxia or by β-receptor blockade.

Intrinsic heart rate is defined as the rate at which the heart beats when all cardiac neural and hormonal inputs are removed. We determined the effect of prevailing autonomic innervation of the heart on the intrinsic heart rate in chronically maintained, sedated, normally innervated dogs (n = 14), and in 14 other dogs that had previously (greater than 12 day) undergone complete surgical cardiac denervation. Intrinsic rate was determined in both groups using the following two procedures: 1) pharmacological effector blockade; and 2) pharmacological ganglionic blockade. The intrinsic rate determined by effector blockade was 142.9 +/- 7.2 (SE) beats/min in the dogs with intact cardiac innervation. When the same treatment was given after total surgical cardiac denervation, intrinsic rate was 97.9 +/- 4.8 beats/min. Intrinsic heart rate was significantly (P less than 0.05) lower in surgically denervated dogs. Ganglionic blockade in surgically denervated animals yielded an intrinsic rate of 90.0 +/- 8.5 beats/min, which was again significantly lower than the corresponding value of 128.4 +/- 5.5 beats/min in normal dogs. There was no difference in the intrinsic heart rate as determined by effector vs. ganglionic blockade in either group of dogs. An additional six dogs were subjected to selective surgical sinoatrial nodal parasympathectomy; their intrinsic rate (effector blockade) in the conscious state was 115.8 +/- 4.3 beats/min; this was significantly lower than the corresponding value for normal dogs and significantly greater than in dogs subject to total surgical cardiac denervation. The lower rate observed in the totally denervated and selectively denervated dogs after effector and/or ganglionic blockades implies that intrinsic heart rate depends on the level or nature of prevailing autonomic activity.

During gradual air exposure, Amia calva show no reduction in oxygen consumption, no increase in plasma urea levels or in urea excretion. Blood pH remains constant, and plasma total CO2, PCO 2, HCO3 -. total ammonia and NH3 concentrations all rise significantly. Exposure to 923 μmol/l NH4Cl does not elicit an increase in urea production or airbreathing. Aquatic hypoxia without access to air does not cause a reduction in aerobic metabolism, and moderate levels result in death. These results suggest that Amia are incapable of aestivation, due to an inability to detoxify ammonia to urea and reduce metabolism, and die following three to five days of air exposure.

Pavlovian conditioning was accomplished in two groups of dogs by following a 30-s tone (the CS+) with a short (less than or equal to 1 s) electric shock). An inflatable occluder was implanted around the left circumflex coronary artery in all animals. The CS+ was presented to Group I dogs (n = 6) during control (i.e., no coronary occlusion) and at 30 s and 3 min after the onset of a 4-min left circumflex coronary occlusion. The CS+ evoked a robust increase in heart rate and blood pressure in the control state. A conditional cardiovascular response was still evoked during the acute coronary occlusion despite the evolving myocardial ischemia. No increase in ventricular ectopic beats occurred during the CS+. An anterior myocardial infarction (16.8 +/- 1.7% of left ventricular mass, mean +/- SEM) was created in Group II dogs (n = 8) at the time of surgery. A CS+ was presented, as above, both alone and 1 min after the beginning of a 2-min left circumflex coronary occlusion. There were significant differences in the conditional response in heart rate, left ventricular (systolic) pressure (LVP) and d(LVP)/dt for trials given during the coronary occlusion compared with no occlusion. These dogs were then classified as 'resistant' or 'susceptible' to ventricular arrhythmias approx. 4 weeks later using an exercise and coronary occlusion test. Three of five susceptible dogs, but none of three resistant dogs, evidenced increased severity of arrhythmias during the CS+. We conclude that the nature of the cardiovascular response to behavioral stress during myocardial ischemia depends in part upon the timing of the CS+, the presence or absence of a resident myocardial infarction and the status of the autonomic reflexes controlling the heart.

Acidification of expired water was studied in rainbow trout exposed to water of pH9.91, 6.80 (control) and 3.88. For the high-pH and the control exposures, the water flowing over the gills was acidified because of the dominant effect of CO2 hydration. During the low-pH treatment, the water was alkalized because of ammonium ion formation and perhaps bicarbonate dehydration. Carbon dioxide excretion was not significantly affected by the high-pH and low-pH treatments but ammonia accumulated in the plasma in both cases.

▪ Abstract The ability of animals to survive food deprivation is clearly of considerable survival value. Unsurprisingly, therefore, all animals exhibit adaptive biochemical and physiological responses to the lack of food. Many animals inhabit environments in which food availability fluctuates or encounters with appropriate food items are rare and unpredictable; these species offer interesting opportunities to study physiological adaptations to fasting and starvation. When deprived of food, animals employ various behavioral, physiological, and structural responses to reduce metabolism, which prolongs the period in which energy reserves can cover metabolism. Such behavioral responses can include a reduction in spontaneous activity and a lowering in body temperature, although in later stages of food deprivation in which starvation commences, activity may increase as food-searching is activated. In most animals, the gastrointestinal tract undergoes marked atrophy when digestive processes are curtailed; this structural response and others seem particularly pronounced in species that normally feed at intermittent intervals. Such animals, however, must be able to restore digestive functions soon after feeding, and these transitions appear to occur at low metabolic costs.

Previous studies in our laboratory demonstrated that rats exhibiting obesity in response to a moderately high-fat (MHF) diet developed hypertension associated with activation of the local and systemic renin-angiotensin system. In this study, we examined the effect of the angiotensin type 1 (AT1)-receptor antagonist, losartan, on blood pressure in obesity-prone (OP) and obesity-resistant (OR) rats fed a MHF diet. Using telemetry monitoring, we characterized the evolution of blood pressure elevations during the development of obesity. Male Sprague-Dawley rats were implanted with telemetry transducers for chronic monitoring of blood pressure, and baseline measurements were obtained. Rats were then switched to the MHF diet (32% kcal as fat) and were segregated into OP and OR groups at week 5. At week 9 on the MHF diet, OP rats exhibited significantly greater 24-h mean arterial blood pressure compared with OR rats (OP: 105 ± 4 mmHg, OR: 96 ± 2 mmHg; P < 0.05). Elevations in blood pressure in OP rats were manifest as an increase in systolic pressure. Administration of losartan to all rats at week 9 resulted in a reduction in blood pressure; however, losartan had the greatest effect in OP rats (percent decrease in mean arterial pressure by losartan; OP: 19 ± 4, OR: 10 ± 2%; P < 0.05). These results demonstrate that elevations in blood pressure occur subsequent to established obesity in rats fed a high-fat diet. Moreover, these results demonstrate the ability of losartan to reverse the blood pressure increase from diet-induced obesity, supporting a primary role for the renin-angiotensin system in obesity-associated hypertension.

In obesity-related hypertension, activation of the renin-angiotensin system (RAS) has been reported despite marked fluid volume expansion. Adipose tissue expresses components of the RAS and is markedly expanded in obesity. This study evaluated changes in components of the adipose and systemic RAS in diet-induced obese hypertensive rats. RAS was quantified in adipose tissue and compared with primary sources for the circulating RAS. Male Sprague-Dawley rats were fed either a low-fat (LF; 11% kcal as fat) or moderately high-fat (32% kcal as fat) diet for 11 wk. After 8 wk, rats fed the moderately high-fat diet segregated into obesity-prone (OP) and obesity-resistant (OR) groups based on their body weight gain (body weight: OR, 566 ± 10; OP, 702 ± 20 g; P < 0.05). Mean arterial blood pressure was increased in OP rats (LF: 97 ± 2; OR: 97 ± 2; OP: 105 ± 1 mmHg; P < 0.05). Quantification of mRNA expression by real-time PCR demonstrated a selective increase (2-fold) in angiotensinogen gene expression in retroperitoneal adipose tissue from OP vs. OR and LF rats. Similarly, plasma angiotensinogen concentration was increased in OP rats (LF: 390 ± 48; OR: 355 ± 24; OP: 530 ± 22 ng/ml; P < 0.05). In contrast, other components of the RAS were not altered in OP rats. Marked increases in the plasma concentrations of angiotensin peptides were observed in OP rats (angiotensin II: LF: 95 ± 31; OR: 59 ± 20; OP: 295 ± 118 pg/ml; P < 0.05). These results demonstrate increased activity of the adipose and systemic RAS in obesity-related hypertension.

The possible interactions between inhibitory vagal control of the heart and circulating levels of catecholamines in dogfish (Squalus acanthias) were studied using an in situ preparation of the heart, which retained intact its innervation from centrally cut vagus nerves. The response to peripheral vagal stimulation typically consisted of an initial cardiac arrest, followed by an escape beat, leading to renewed beating at a mean heart rate lower than the prestimulation rate (partial recovery). Cessation of vagal stimulation led to a transient increase in heart rate, above the prestimulation rate. This whole response was completely abolished by 10(-4) M atropine (a muscarinic cholinergic antagonist). The degree of vagal inhibition was evaluated in terms of both the initial, maximal cardiac interval and the mean heart rate during partial recovery, both expressed as a percentage of the prestimulation heart rate. The mean prestimulation heart rate of this preparation (36+/-4 beats min(-1)) was not affected by noradrenaline but was significantly reduced by 10(-4) M nadolol (a beta-adrenergic receptor antagonist), suggesting the existence of a resting adrenergic tone arising from endogenous catecholamines. The degree of vagal inhibition of heart rate varied with the rate of stimulation and was increased by the presence of 10(-8) M noradrenaline (the normal in vivo level in routinely active fish), while 10(-7) M noradrenaline (the in vivo level measured in disturbed or deeply hypoxic fish) reduced the cardiac response to vagal stimulation. In the presence of 10(-7) M noradrenaline, 10(-4) M nadolol further reduced the vagal response, while 10(-4) M nadolol + 10(-4) M phentolamine had no effect, indicating a complex interaction between adrenoreceptors, possibly involving presynaptic modulation of vagal inhibition.