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Fish aquaculture facilities strive to maintain abiotic conditions that optimise somatic growth. Two physiological systems that are critically linked to environmental abiotic factors are ion and acid‐base regulation. Given the detrimental effects of ionic and acid‐base imbalances coupled with the potential energetic costs of ion and acid‐base regulation, it is important to understand how changes in abiotic variables such as salinity, temperature and pH alter ionoregulatory and acid‐base physiology. The gill is the predominant site of ion and acid‐base regulation, two linked processes that are vital to the health of fishes. Most species maintain ion levels in the blood plasma within a narrow range, independent of external ion concentrations. In freshwater (FW), fishes must actively take up ions from their dilute environment to counteract passive ion losses, while in seawater (SW), ion excretion serves to counteract passive salt gain. Ion uptake (FW) and excretion (SW) are coordinated by different subtypes of ion‐transporting cells (ionocytes) expressed in the gill epithelium. In this review, we discuss the function of FW and SW ionocytes in the movement of ions across the gill epithelium and address other features of the gill that are integral to ionoregulation. We also discuss the mechanisms of acid‐base regulation by the gill, which is intimately related to ion regulation because it is coordinated by Na+‐ and Cl−‐linked fluxes of acid (H+) and base (HCO3‐) equivalents. These fundamental physiological principles are integral in understanding how fishes in aquaculture respond to relevant disturbances that may occur under culture conditions. [ABSTRACT FROM AUTHOR]

Clifford et al shrug off these complexities and by using classical approaches, provide direct evidence for a novel Na SP + sp uptake mechanism that serves to sustain Na SP + sp balance in acidic water. Reassessing the contribution of the Na + /H + exchanger Nhe3b to Na + uptake in zebrafish (Danio rerio) using CRISPR/Cas9 gene editing. Over the past century, studies of fish gills have provided much insight into the fundamental mechanisms of ion movements across epithelia. [Extracted from the article]

The present study tested the hypothesis that zebrafish (Danio rerio) aquaporin-1a1 (AQP1a1) serves as a multi-functional channel for the transfer of the small gaseous molecules, CO2 and ammonia, as well as water, across biological membranes. Zebrafish embryos were microinjected with a translation-blocking morpholino oligonucleotide targeted to AQP1a1. Knockdown of AQP1a1 significantly reduced rates of CO2 and ammonia excretion, as well as water fluxes, in larvae at 4 days post fertilization (dpf). Because AQP1a1 is expressed both in ionocytes present on the body surface and in red blood cells, the haemolytic agent phenylhydrazine was used to distinguish between the contributions of AQP1a1 to gas transfer in these two locations. Phenylhydrazine treatment had no effect on AQP1a1-linked excretion of CO2 or ammonia, providing evidence that AQP1a1 localized to the yolk sac epithelium, rather than red blood cell AQP1a1, is the major site of CO2 and ammonia movements. The possibility that AQP1a1 and the rhesus glycoprotein Rhcg1, which also serves as a dual CO2 and ammonia channel, act in concert to facilitate CO2 and ammonia excretion was explored. Although knockdown of each protein did not affect the abundance of mRNA and protein of the other protein under control conditions, impairment of ammonia excretion by chronic exposure to high external ammonia triggered a significant increase in the abundance of AQP1a1 mRNA and protein in 4 dpf larvae experiencing Rhcg1 knockdown. Collectively, these results suggest that AQP1a1 in zebrafish larvae facilitates the movement of CO2 and ammonia, as well as water, in a physiologically relevant fashion. [ABSTRACT FROM AUTHOR]

Nitric oxide (NO) is a gaseous neurotransmitter, which, in adult mammals, modulates the acute hypoxic ventilatory response; its role in the control of breathing in fish during development is unknown.We addressed the interactive effects of developmental age and NO in the control of piscine breathing by measuring the ventilatory response of zebrafish (Danio rerio) adults and larvae to NO donors and by inhibiting endogenous production of NO. In adults, sodium nitroprusside (SNP), a NO donor, inhibited ventilation; the extent of the ventilatory inhibition was related to the pre-existing ventilatory drive, with the greatest inhibition exhibited during exposure to hypoxia (PO2=5.6 kPa). Inhibition of endogenous NO production using L-NAME suppressed the hypoventilatory response to hyperoxia, supporting an inhibitory role of NO in adult zebrafish. Neuroepithelial cells (NECs), the putative oxygen chemoreceptors of fish, contain neuronal nitric oxide synthase (nNOS). In zebrafish larvae at 4 days post-fertilization, SNP increased ventilation in a concentrationdependent manner. Inhibition of NOS activity with L-NAME or knockdown of nNOS inhibited the hypoxic (PO2=3.5 kPa) ventilatory response. Immunohistochemistry revealed the presence of nNOS in the NECs of larvae. Taken together, these data suggest that NO plays an inhibitory role in the control of ventilation in adult zebrafish, but an excitatory role in larvae. [ABSTRACT FROM AUTHOR]

The ontogeny of carbon dioxide (C02) sensing in zebrafish (Danio rerio) has not been examined. In this study, C02-mediated increases in heart rate were used to gauge the capacity of zebrafish larvae to sense C02. C02 is thought to be detected via neuroepithelial cells (NECs), which are homologous to mammalian carotid body glomus cells. Larvae at 5 days post-fertilization (d.p.f.) exhibited tachycardia when exposed for 30 min to 0.75% C02 (-5.63 mmHg); at 7 d.p.f., tachycardia was elicited by 0.5% C02 (-3.75 mmHg). Based on pharmacological evidence using p-adrenergic receptor (P-AR) antagonists, and confirmed by Pi-AR translational gene knockdown using morpholinos, the reflex tachycardia accompanying hypercapnia was probably mediated by the interaction of catecholamines with cardiac Pi receptors. Because the cardiac response to hypercapnia was abolished by the ganglionic blocker hexamethonium, it is probable that the reflex cardio-acceleration was mediated by catecholamines derived from sympathetic adrenergic neurons. Owing to its likely role in facilitating intracellular acidification during exposure to hypercapnia, it was hypothesized that carbonic anhydrase (CA) is involved in C02 sensing, and that inhibition of CA activity would blunt the downstream responses. Indeed, the cardiac response to hypercapnia (0.75% C02) was reduced in fish at 5 d.p.f. exposed to acetazolamide, a CA inhibitor, and in fish experiencing zCAc (CA2-like a) knockdown. Successful knockdown of zCAc was confirmed by CA activity measurements, western blotting and immunocytochemistry. Co-injection of embryos with zCAc morpholino and mRNA modified at the morpholino binding site restored normal levels of CA activity and protein levels, and restored (rescued) the usual cardiac responses to hypercapnia. These data, combined with the finding that zCAc is expressed in NECs located on the skin, suggest that the afferent limb of the C02-induced cardiac reflex in zebrafish larvae is initiated by coetaneous C02-sensing neuroepithelial cells. [ABSTRACT FROM AUTHOR]

Goldfish acclimated to cold water (e.g. 7°C) experience a marked reduction in functional lamellar surface area owing to the proliferation of an interlamellar cell mass (ILCM), a phenomenon termed gill remodelling. The goal of the present study was to assess the consequences of the reduced functional surface area on the capacity of goldfish to excrete ammonia. Despite the expected impact of ambient temperature on functional surface area, fish acclimated to 7°C and 25°C exhibited similar rates of ammonia excretion (Jnet,amm) the Q10 values for fed and starved fish were 1.07 and 1.20, respectively. To control for possible temperature-related differences in rates of endogenous ammonia production, Jnet,amm was determined at the two acclimation temperatures after loading fish with 1.12μmolg-1 of NH4CI. In the 3h post-injection period, Jnet,amm was elevated to a greater extent in the 25°C fish. To estimate the potential contribution of increased ventilation and cardiac output to ammonia clearance in the warmer fish, the ammonia loading experiment was repeated on the 7°C fish immediately after they were exercised to exhaustion. The rate of excretion of ammonia was significantly increased in the exercised 7°C fish (presumably experiencing increased ventilation and cardiac output for at least some of the measurement period) suggesting that differences in external and internal convection may at least partially explain the enhanced capacity of the 25°C fish to clear the ammonia load. To more specifically assess the contribution of the different functional surface areas on the differing rates of ammonia clearance at the two acclimation temperatures, the 7°C fish were exposed for 7days to hypoxia (PO2=10mmHg=1.33kPa), a treatment known to cause the disappearance of the ILCM. The results demonstrated that the hypoxia-associated loss of the ILCM was accompanied by a significant increase in the rate of ammonia clearance in the 7°C fish when returned to normoxic conditions. To determine whether compensatory changes in the ammonia transporting proteins might be contributing to sustaining Jnet,amm under conditions of reduced functional lamellar surface area, the relative expression and branchial distribution of four Rh proteins were assessed by western blotting and immunocytochemistry. Although the relative expression of the Rh proteins was unaffected by acclimation temperature, there did appear to be a change in the spatial distribution of Rhag, Rhbg and Rhcg1. Specifically, these three Rh proteins (and to a lesser extent Rhcg2) appeared to localize in cells on the outer edge of the ILCM that were enriched with Na+/K+- ATPase. Thus, we suggest that despite the impediment to ammonia excretion imposed by the ILCM, goldfish acclimated to 7°C are able to sustain normal rates of excretion owing to the redistribution of ammonia transporting cells. [ABSTRACT FROM AUTHOR]

Fish exposed to hypoxia develop decreased heart rate, or bradycardia, the physiological significance of which remains unknown. The general muscarinic receptor antagonist atropine abolishes the development of this hypoxic bradycardia, suggesting the involvement of muscarinic receptors. In this study, we tested the hypothesis that the hypoxic bradycardia is mediated specifically by stimulation of the M2 muscarinic receptor, the most abundant subtype in the vertebrate heart. Zebrafish (Danio rerio) were reared at two levels of hypoxia (30 and 40 Torr Po2) from the point of fertilization. in hypoxic fish, the heart rate was significantly lower than in normoxic controls from 2 to 10 days post-fertilization (dpf). At the more severe level of hypoxia (30 Ton Po2), there were significant increases in the relative mRNA expression of M2 and the cardiac type β-adrenergic receptors (β1AR, β2aAR, and β2bAR) at 4 dpf. The hypoxic bradycardia was abolished (at 40 Ton Po2) or significantly attenuated (at 30 Torr Po2) in larvae experiencing M2 receptor knockdown (using morpholino antisense oligonucleotides). Sham-injected larvae exhibited typical hypoxic bradycardia in both hypoxic regimens. The expression of β1AR, β2aAR, β2bAR, and M2 mRNA was altered at various stages between 1 and 4 dpf in larvae experiencing M2 receptor knockdown. Interestingly, M2 receptor knockdown revealed a cardioinhibitory role for the β2-adrenergic receptor. This is the first study to demonstrate a specific role of the M2 muscarinic receptor in the initiation of hypoxic bradycardia in fish. [ABSTRACT FROM AUTHOR]

Mitrovic D, Dymowska A, Nilsson GE, Perry SF. Physiological consequences of gill remodeling in goldfish (Carassius auratus) during exposure to long-term hypoxia. Am J Physiol Regul Integr Conip Physiol 297: R224-R234, 2009. First published May 20, 2009; doi: 10.1152/ajpregu.00l89.2009.-Goldfish (Carassius auratus) acclimated to 7°C and exposed to hypoxia (-10 mmHg) for 7 days exhibited a pronounced remodeling of the gill consisting of the removal of an interlamellar cell mass (ILCM). Subsequent experiments were designed to assess the impact of gill remodeling and the associated increase in functional lamellar surface area on the distribution of branchial ionocytes and Cl[sup-] flux across the gill. Despite the increased functional lamellar surface area during hypoxia, there was no corresponding increase in Cl[sup-] loss or efflux of the extracellular marker polyethylene glycol (PEG 4000). However, when hypoxic fish were returned to normoxic water for 12 h, rates of Cl[sup-1] and PEG efflux were markedly stimulated in keeping with an increased surface area for solute movement. Similarly, the rate of branchial Cl uptake was reduced (105 ± 22 vs. 45 ± 8 μmol∙kg[sup-1]∙h[sup-1]) in normoxic and hypoxic fish, respectively, bUt then stimulated (345 μmol∙kg[sup-1]∙h[sup-1]) upon reestablishment of norinoxic conditions. Hypoxia (7 days) was accompanied by a significant decrease in the total cross-sectional area of branchial ionocytes owing to a decrease in their numbers and individual sizes. Thus, despite experiencing an increase in functional lamellar surface area, hypoxic goldfish limit branchial Cl[sup-] loss likely by a hypoxia-mediated decrease in paracellular permeability. In normoxic fish, the ionocytes were largely confined to the outer edges of the ILCM. During hypoxia, preexisting ionocytes migrated with the shrinking ILCM, while a smaller proportion of newly differentiated cells appeared below the surface of the ILCM. The capacity to maintain a population of ionocytes in contact with the water is an appropriate strategy to retam ionoregulatory capabilities regardless of whether the lamellae are uncovered or covered. [ABSTRACT FROM AUTHOR]

The goal of the present study was to investigate the role of circulating cortisol and urea in the transcriptional regulation of branchial glutamine synthetase (GS), which incorporates NH3 into glutamate to form glutamine, and the toadfish urea transporter, tUT, which is involved in urea excretion across the gill of the gulf toadfish. GS (of which there are two isoforms, LGS and GGS) and tUT mRNA expression and activity were measured in toadfish exposed to treatments that would induce variable stress responses. In addition, the role of circulating urea in tUT regulation was investigated by infusing toadfish with urea alone or in combination with intraperitoneal injection of RU486, a corticosteroid type II receptor antagonist. There was a 4.8-fold upregulation in the mRNA expression of the gill-specific GS isoform (GGS) in response to cortisol infusion and a similar upregulation in the more ubiquitous isoform (LGS). Furthermore, there was a significant 1.9-fold and 3.3-fold upregulation in the mRNA expression of the toadfish urea transporter, tUT, in response to stress through crowding or exogenous cortisol loading through infusion, respectively. In addition, tUT was found to have a urea-sensitive component to transcriptional regulation that was independent of circulating cortisol concentrations. However, the changes measured in mRNA expression of GGS, LGS and tUT did not correspond with changes in protein activity. To determine the cell type(s) involved in glutamine production and urea excretion, we attempted to localize GGS, LGS and tUT using in situ hybridization. This study is the first to show that GGS and tUT expression appear to occur in gill mitochondria-rich cells of toadfish, suggesting that these cells play a combined glutamine production and urea excretion role, which may have implications for predator avoidance. [ABSTRACT FROM AUTHOR]

Tests the hypothesis that efficiency of carbon dioxide (CO[sub2]) excretion is sensitive to changes in blood flow owing to chemical equilibrium limitations in trout. Discussion on CO[sub2] excretion; Relationship of arterial partial pressure of CO[sub2] to cardiac output; Influence of carbonic anhydrase on the relationship of CO[sub2] pressure and cardiac output.

Assesses the role of renin-angiotensin system in catecholamine release during hypoxia periods in rainbow trout. Increase of plasma catecholamine levels upon nicotinic receptor desensitization; Activation of the renin-angiotensin system during hypoxia; Role of angiotensin II in facilitating catecholamine release.

Focuses on a study which assessed the potential direct and modulating effects of acute hypoxia on catecholamine secretion in rainbow trout, Oncorhynchus mykiss using a perfused cardinal vein preparation and chronically cannulated fish. Materials and methods; Effects of hypoxic perfusate on in situ basal catecholamine secretion rates; Influence of acute moderate hypoxia on nicotine-evoked catecholamine release in vivo.

Discusses a study which assessed the relative involvement of nicotinic and muscarinic cholinergic receptors in the neuronal control of catecholamine secretion from the chromaffin tissue of rainbow trout. Background information on the mechanism of catecholamine secretion in teleosts; Materials and methods; Results; Discussion.

Determines the contribution of angiotensin II (Ang-II)-elicited increases in humoral catecholamines levels to rainbow trout cardiovascular control. Effect of homologous rainbow trout Ang-II in plasma catecholamine levels; Cardiovascular effects of Ang-II with and without alpha-adrenoreceptor blockade; Relationship between plasma concentrations and their cardiovascular effects.

Investigates the relationships between angiotensins and catecholamine release from the chromaffin tissue of rainbow trout. Evidence that angiotensins can elicit catecholamine release; Method in which angiotensins effects this release.

Characterizes the acid-base disequilibrium in the arterial blood of rainbow trout Oncorhynchus mykiss during environmental hypoxia, hyperoxia or hypercapnia. Extracorporeal circulation; Measurement of pH, partial pressure of oxygen and carbon dioxide; Ventilation.

Examines the response of cannulated rainbow trout to acute hypoxia in fish acclimated to two temperatures (5 and 15 degrees Celsius). Effects of acclimation on catecholamine release; Effects of acclimation temperature on hemoglobin oxygen-affinity; Other potential causes/correlates of catecholamine release.

Zebrafish ( Danio rerio ) is an emerging model for integrative physiological research. In this mini-review, we discuss recent advances in the neuroendocrine control of ionic balance in this species, and identify current knowledge gaps and issues that would benefit from further investigation. Zebrafish inhabit a hypo-ionic environment and therefore are challenged by a continual loss of ions to the water. To maintain ionic homeostasis, they must actively take up ions from the water and reduce passive ion loss. The adult gill or the skin of larvae are the primary sites of ionic regulation. Current models for the uptake of major ions in zebrafish incorporate at least three types of ion transporting cells (also called ionocytes); H + -ATPase-rich cells for Na + uptake, Na + /K + -ATPase-rich cells for Ca 2+ uptake, and Na + /Cl − -cotransporter expressing cells for both Na + and Cl − uptake. The precise molecular mechanisms regulating the paracellular loss of ions remain largely unknown. However, epithelial tight junction proteins, including claudins, are thought to play a critical role in reducing ion losses to the surrounding water. Using the zebrafish model, several key neuroendocrine factors were identified as regulators of epithelial ion movement, including the catecholamines (adrenaline and noradrenaline), cortisol, the renin-angiotensin system, parathyroid hormone and prolactin. Increasing evidence also suggests that gasotransmitters, such as H 2 S, are involved in regulating ion uptake. [ABSTRACT FROM AUTHOR]

Objective-To characterize physiologic responses of ball pythons (Python regius) following a minor surgical procedure and investigate the effects of 2 commonly used analgesics on this response. Animals-15 healthy ball pythons. Procedures-Snakes were randomly assigned to receive 1 of 3 treatments: meloxicam (0.3 mg/kg [0.14 mg/lb]; n = 5), butorphanol (5 mg/kg [2.3 mg/lb]; 5), or saline (0.9% NaCl) solution (5) before catheterization of the vertebral artery. Plasma concentrations of catecholamines and cortisol, blood pressure, heart rate, and blood gas values were measured at various times for 72.5 hours after catheterization. The 72.5-hour point was defined as baseline. Results-Heart rate of ball pythons increased significantly during the first hour following surgery. Mean plasma epinephrine concentration increased slightly at 2.5 hours after surgery, whereas mean plasma cortisol concentration increased beginning at 1.5 hours, reaching a maximum at 6.5 hours. Mean blood pressure increased within the first hour but returned to the baseline value at 2.5 hours after surgery. After 24.5 hours, blood pressure, heart rate, and plasma hormone concentrations remained stable at baseline values. There were no significant differences in values for physiologic variables between snakes that received saline solution and those that received meloxicam or butorphanol. Conclusions and Clinical Relevance-Measurement of physiologic variables provides a means of assessing postoperative pain in snakes. Meloxicam and butorphanol at the dosages used did not decrease the physiologic stress response and did not appear to provide analgesic effects in ball pythons. [ABSTRACT FROM AUTHOR]