First published online March 28, 2008
Journal of Experimental Biology 211, 1231-1242 (2008)
Published by The Company of Biologists 2008
doi: 10.1242/jeb.015248
A comparative analysis of putative oxygen-sensing cells in the fish gill
Emily H. Coolidge,
Cosima S. Ciuhandu* and
William K. Milsom
Department of Zoology, University of British Columbia, 6270 University
Blvd, Vancouver, BC, V6T 1Z4, Canada
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Fig. 1. Distribution of serotonergic immunoreactive cells (darkly stained cells)
along the filament (F) and lamellae (L) in (A) O. mykiss, (B) C.
auratus, (C) H. lacerdae, and (D) H. malabaricus. Scale
bars, (A) 200 µm, (B–D) 100 µm. Images were colour-inverted to
emphasize bright immunofluorescent structures.
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Fig. 2. Distribution of 5-HT-IR cells expressed as the average staining intensity
(pixels) from lamellar tips (0) to the central axis of the filament (100) in
(A) O. mykiss, (B) H. lacerdae, (C) H. malabaricus
and (D) C. auratus. Shaded area represents the 95% confidence
interval associated with the mean distribution (solid line) for each fish
species. (E) Comparison of the mean distribution of 5-HT-IR cells among the
four species. Broken line represents the division between lamellae and
filament.
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Fig. 3. Number of 5-HT-IR cells on the filament and lamellae in rainbow trout
(O. mykiss), trairão (H. lacerdae), traira (H.
malabaricus) and goldfish (C. auratus). Values are expressed as
means ± s.e.m. At least 50 cells per individual and six individuals per
species were measured. Letters (a–d) indicate cell counts significantly
different from columns labelled with different letters.
P50 values are given in the key as good indicators of
relative hypoxia tolerance.
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Fig. 4. Double immunolabelling of serotonin (5-HT, green) and synaptic vesicles
(SV2, red) along a single gill filament with multiple lamellae in (A) O.
mykiss, (B) C. auratus, (C) H. lacerdae and (D) H.
malabaricus. Scale bars, 100 µm. Colocalization appears yellow.
Arrowheads indicate non-5-HT-IR but SV2-IR cells and arrows indicate
non-SV2-IR but 5-HT-IR cells.
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Fig. 5. Double immunolabelling of serotonin (5-HT; green) and nerves (using a
neuronal marker zn12; red) in longitudinal sections (30 µm thick) of the
gill filament and lamellae of (A) O. mykiss, (B) C. auratus,
(C) H. lacerdae and (D) H. malabaricus. Images were captured
using a confocal scanning system. Inset in B shows close association of nerves
(red) and 5-HT-IR cell at the tip of the lamellae of C. auratus.
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Fig. 6. TEM images of neuroepithelial cells (NECs) in (A) the filament of O.
mykiss and (B) the lamella of C. auratus. Lower left-hand insets
(areas indicated by broken red boxes) show the presence of dense-cored
vesicles (arrowheads). Note the proximity of NECs in both A and B to a red
blood cell (RBC), as well as a nerve (N) and the basal lamina (BL) in A. The
NEC of the goldfish lamella (B) is also juxtaposed to a pillar cell (pc) and
near the ambient environment (H2O). Scale bars, 500 nm.
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Fig. 7. Cross sections of gill filaments from (A) O. mykiss, (B) C.
auratus, (C) H. lacerdae and (D) H. malabaricus showing
double immunolabelling of 5-HT (green) and SV2 (red; Ai–Di) and 5-HT
(green) and zn12 (red; Aii–Dii). (E) Schematic of a filament cross
section showing the location of the efferent filament artery (eFA), central
venous sinus (cvs), afferent filament artery (aFA), lamellar pillar cells (pc)
and companion vessels (cv) [adapted from Farrell
(Farrell, 1979 )]. Note that
lamellae are rarely sectioned in a straight plane. Scale bars, 100 µm.
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Fig. 8. Comparison of the tip of the filaments in all four species using antibody
immunolabelling of serotonin (5-HT) and synaptic vesicle marker (SV2): (A)
O. mykiss, (B) C. auratus, (C) H. lacerdae and (D)
H. malabaricus. Colocalization appears yellow. Scale bars, 100
µm.
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Fig. 9. Innervated serotonergic cells located at the filament tips in (A) O.
mykiss, (B) C. auratus, (C) H. lacerdae and (D) H.
malabaricus immunolabelled for serotonin (5-HT, green) and a neuronal
marker (zn12, red). Scale bars, 25 µm.
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Fig. 10. Innervated serotonergic cells located in the gill rakers in (A) O.
mykiss and (B) C. auratus immunolabelled for serotonin (5-HT,
green) and a synaptic vesicle marker (sv2, red). Insets show magnification of
individual cells. Colocalization appears yellow. Arrowheads indicate
non-5-HT-IR but SV2-IR cells. Scale bars, 50 µm.
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Fig. 11. Innervated serotonergic cells located in the gill rakers in (A) O.
mykiss and (B) C. auratus immunolabelled for serotonin (5-HT,
green) and a neuronal marker (zn12, red). Insets show magnification of
innervated 5-HT-IR cells in both trout and goldfish. Scale bars, 50 µm.
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Fig. 12. Schematic of proposed locations of putative O2 receptor cells in
fish gills: gill rakers (R), lamellae (L), tips of filaments (FT) and centre
of filament (F) (efferent filament artery). Note that all fish species contain
putative O2 receptor cells in at least two of these proposed
locations and variation between species seems to be related to hypoxia
tolerance.
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© The Company of Biologists Ltd 2008
