New developments on gill innervation: insights from a model vertebrate

Jonz, Michael G.; Nurse, Colin A.

doi:10.1242/jeb.010587

Cited by 30 publications

(19 citation statements)

References 70 publications

(131 reference statements)

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“…Previous studies on ASICs in fishes have suggested that they are primarily associated with neuronal tissue and sensory cells (27,35,42,46). In adult and developing zebrafish, morphological studies using confocal immunofluorescence techniques demonstrated that gill MRCs come into direct contact with nerve fibers in the basolateral region (21,22). However, in our study, ASIC4 protein was clearly localized to the apical region of the MRCs, as demonstrated by three-dimensional reconstruction of con- focal images.…”

Section: Discussioncontrasting

confidence: 57%

“…For all flux experiments, fish were transferred to individual darkened 60-ml flux chambers, supplied with a constant flow of aerated low-ionic-strength, low-pH water, and acclimated to the chambers for 24 h prior to experimentation. Unidirectional Na ϩ influx was measured using radiolabeled 22 Na, as described previously (12). Briefly, flow of water to chambers was turned off, 0.1 Ci/l 22 Na was added to each chamber and allowed to mix for 5 min, and either the DMSO control (0.05% DMSO) or pharmacological agents dissolved in DMSO were added to the chambers.…”

Section: Methodsmentioning

confidence: 99%

“…At the completion of the experiment, fish were terminally anesthetized (MS-222, 1 g/l) and weighed. Water samples (3 ml) were analyzed for 22 Na radioactivity using a gamma counter (Packard Cobra II, Auto Gamma, model 5010, Perkin Elmer, Waltham, MA), and total concentration of Na ϩ was measured using atomic absorption spectrophotometry (model 3300, Perkin Elmer, Shelton, CT). Unidirectional 22 Na ϩ influx (mol·kg Ϫ1 ·h Ϫ1 ) was calculated for each flux period according to the following equation: JNain ϭ ⌬cpm ϫ V/(SA ϫ t ϫ M), where ⌬cpm is the difference between the initial and final radioactivities in the water (cpm/ml), V is the water flux volume (ml), SA is the average specific activity (cpm/mol Na ϩ ) as measured at the beginning and end of the flux period, t is the time elapsed (h), and M is the mass of the fish (kg).…”

Section: Methodsmentioning

confidence: 99%

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Acid-sensing ion channels are involved in epithelial Na⁺ uptake in the rainbow trout Oncorhynchus mykiss

Dymowska

Schultz

Blair

et al. 2014

American Journal of Physiology-Cell Physiology

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Dymowska AK, Schultz AG, Blair SD, Chamot D, Goss GG. Acidsensing ion channels are involved in epithelial Na ϩ uptake in the rainbow trout Oncorhynchus mykiss. Am J Physiol Cell Physiol 307: C255-C265, 2014. First published June 4, 2014 doi:10.1152/ajpcell.00398.2013.-A role for acid-sensing ion channels (ASICs) to serve as epithelial channels for Na ϩ uptake by the gill of freshwater rainbow trout was investigated. We found that the ASIC inhibitors 4=,6-diamidino-2-phenylindole and diminazene decreased Na ϩ uptake in adult rainbow trout in a dose-dependent manner, with IC 50 values of 0.12 and 0.96 M, respectively. Furthermore, we cloned the trout ASIC1 and ASIC4 homologs and demonstrated that they are expressed differentially in the tissues of the rainbow trout, including gills and isolated mitochondrion-rich cells. Immunohistochemical analysis using custom-made anti-zASIC4.2 antibody and the Na ϩ -K ϩ -ATPase (␣5-subunit) antibody demonstrated that the trout ASIC localizes to Na ϩ /K ϩ -ATPase-rich cells in the gill. Moreover, three-dimensional rendering of confocal micrographs demonstrated that ASIC is found in the apical region of mitochondrion-rich cells. We present a revised model whereby ASIC4 is proposed as one mechanism for Na ϩ uptake from dilute freshwater in the gill of rainbow trout. sodium uptake; gill; acid-sensing ion channels; mitochondrion-rich cells; fish; ionoregulation FISHES LIVING IN FRESHWATER (FW) must actively take up Na ϩ against a steep concentration gradient. Na ϩ uptake occurs via specialized mitochondrion-rich cells (MRCs), located on the fish gill epithelium (26, 29), and was initially proposed to be linked to NH 4 ϩ /H ϩ excretion (26). Subsequent studies have described two models of Na ϩ transport (for review see Refs. 8 and 18). In the first proposed model, Na ϩ is exchanged for H ϩ via an electroneutral Na ϩ /H ϩ exchanger (NHE) located on the apical membrane of MRCs in fish gill epithelia. The identification of multiple NHEs in the gills of zebrafish (Danio rerio) (51), Osorezan dace (Tribolodon hakonensis) (15), rainbow trout (Oncorhynchus mykiss) (7,19), and tilapia (Oreochromis mossambicus) (48) by immunocytochemistry, Western blot analysis, and RT-PCR, supports this model. However, significant thermodynamic constraints associated with the function of NHEs at very low ion concentrations (Na ϩ Ͻ0.1 mM) and low environmental pH (pH Ͻ5) (1, 34) suggest that fishes living in very soft and poorly buffered water would not be able to rely on a NHE-based mechanism for sufficient Na ϩ uptake. Recently, the NHE model was revised, whereby the ammonia transporter [rhesus (Rh) protein] present on the apical membrane of MRCs (30, 31) forms a functional metabolon with NHE2/3 (50). The revised model does alleviate the thermodynamic constraints associated with a low-pH environment, but not those imposed by low Na ϩ concentrations in the FW aquatic environment (6). Therefore, it is unlikely that this mechanism is the sole contributor to Na ϩ uptake by FW fishes living in low-ionic-strengt...

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Section: Discussioncontrasting

confidence: 57%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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Acid-sensing ion channels are involved in epithelial Na⁺ uptake in the rainbow trout Oncorhynchus mykiss

Dymowska

Schultz

Blair

et al. 2014

American Journal of Physiology-Cell Physiology

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“…Because the distally located NECs are innervated, they are presumed to be fully functional O 2 chemoreceptors. The nerve fibres innervating the lamellar NECs are protrusions of the branchial nerves which branch from bundles within the filament to innervate the lamellae (Jonz and Nurse, 2008). Interestingly, there appeared to be little, if any, nerve fibres present within the ILCM, itself, a finding which is consistent with the apparent absence of NECs on the outer edge of the ILCM.…”

Section: Branchial Nec Distribution In Fish With or Without An Ilcmmentioning

confidence: 51%

The control of breathing in goldfish (Carassius auratus) experiencing thermally induced gill remodelling

Tzaneva

Perry

2010

Journal of Experimental Biology

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SUMMARY At temperatures below 15°C the gill lamellae of goldfish (Carassius auratus) are largely covered by an interlamellar cell mass (ILCM) which decreases the functional surface area of the gill. The presence of the ILCM in goldfish acclimated to cold water conceivably could lead to a covering of the neuroepithelial cells (NECs), which are believed to be important for sensing ambient O2 and CO2 levels. In this study we tested the hypothesis that goldfish with covered lamellae (and presumably fewer NECs exposed to the water) exhibit a decreased capacity to hyperventilate in response to hypoxic stimuli. Measurements of ventilation amplitude and frequency were performed during exposure to acute hypoxia (PwO2=30 mmHg) or following injections of the O2 chemoreceptor stimulant NaCN into the buccal cavity or caudal vein of fish acclimated to 25°C (uncovered lamellae) or 7°C (covered lamellae) to stimulate predominantly the externally or internally oriented NECs, respectively. The results demonstrated no significant differences in the response to hypoxia, with each group exhibiting similar percentage increases in ventilation amplitude (90–91%) and frequency (34–43%). Similarly, with the exception of a rightward shift of the ventilation frequency dose–response in the fish acclimated to 7°C, there were no significant differences between the two groups of fish in the ED50 values. These findings suggest that goldfish with covered lamellae retain the capacity to sense external hypoxic stimuli. Using immunohistochemistry to identify serotonin-enriched NECs, it was demonstrated that the presence of the ILCM results in the NECs being redistributed towards the distal regions of the lamellae. In 25°C-acclimated fish, the NECs were distributed evenly along the length of the lamellae with 53±3% of them in the distal half, whereas in fish acclimated to 7°C, 83±5% of the NECs were confined to the distal half. Using the neuronal marker antibody ZN-12, it was demonstrated that the NECs at the distal edges of the lamellae are innervated by nerve fibres. Thus, it is hypothesised that the capacity to sense external hypoxic stimuli in goldfish acclimated to cold water is maintained despite the increasing coverage of the gill epithelial surfaces because of a redistribution of innervated NECs to the exposed distal regions of the lamellae.

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“…The gills, and presumably the branchial ICs, are innervated by the IX and X cranial nerves 7,[23][24][25] . These nerves carry both efferent and afferent inputs to and from the gill, respectively.…”

Section: Discussionmentioning

confidence: 99%

A Time Differential Staining Technique Coupled with Full Bilateral Gill Denervation to Study Ionocytes in Fish

Tzaneva

Perry

2015

JoVE

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Branchial ionocytes (ICs) are the functional units for ionic regulation in fish. In adults, they are found on the filamental and lamellar epithelia of the gill where they transport ions such as Na + , Cl -and Ca 2+ via a variety of ion channels, pumps and exchangers. The teleost gill is extrinsically innervated by the facial (VI), glossopharyngeal (IX) and vagus (X) nerves. The IX and X nerves are also the extrinsic source of branchial IC innervation. Here, two techniques used to study the innervation, proliferation and distribution of ICs are described: a time differential staining technique and a full bilateral gill denervation technique. Briefly, goldfish are exposed to a vital mitochondrion-specific dye (e.g., MitoTracker Red) which labels (red fluorescence) pre-existing ICs. Fish were either allowed to recover for 3 -5 days or immediately underwent a full bilateral gill denervation. After 3 -5 days of recovery, the gills are harvested and fixed for immunohistochemistry. The tissue is then stained with an α-5 primary antibody (targets Na + /K + ATPase containing cells) in conjunction with a secondary antibody that labels all (both new and pre-existing)ICs green. Using confocal imaging, it was demonstrated that pre-existing ICs appear yellow (labelled with both a viable mitochondrion-specific dye and α-5) and new ICs appear green (labelled with α-5 only). Both techniques used in tandem can be applied to study the innervation, proliferation and distribution of ICs on the gill filament when fish are exposed to environmental challenges.

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New developments on gill innervation: insights from a model vertebrate

Cited by 30 publications

References 70 publications

Acid-sensing ion channels are involved in epithelial Na⁺ uptake in the rainbow trout Oncorhynchus mykiss

Acid-sensing ion channels are involved in epithelial Na⁺ uptake in the rainbow trout Oncorhynchus mykiss

The control of breathing in goldfish (Carassius auratus) experiencing thermally induced gill remodelling

A Time Differential Staining Technique Coupled with Full Bilateral Gill Denervation to Study Ionocytes in Fish

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New developments on gill innervation: insights from a model vertebrate

Cited by 30 publications

References 70 publications

Acid-sensing ion channels are involved in epithelial Na+ uptake in the rainbow trout Oncorhynchus mykiss

Acid-sensing ion channels are involved in epithelial Na+ uptake in the rainbow trout Oncorhynchus mykiss

The control of breathing in goldfish (Carassius auratus) experiencing thermally induced gill remodelling

A Time Differential Staining Technique Coupled with Full Bilateral Gill Denervation to Study Ionocytes in Fish

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Product

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Acid-sensing ion channels are involved in epithelial Na⁺ uptake in the rainbow trout Oncorhynchus mykiss

Acid-sensing ion channels are involved in epithelial Na⁺ uptake in the rainbow trout Oncorhynchus mykiss