Gills of the Snow Trout, Schizothorax curvifrons Heckel: A SEM Study
Using the scanning electron microscope, the gills of Schizothorax curvifrons have been studied after carrying out their primary fixation in 2.5% glutaraldehyde in 0.1 M sodiumcacodylate buffer (pH 7.3) for 24 h at 0-4°C and secondary fixation in 1-4% osmiumtetraoxide in 0.1 M sodiumcacodylate buffer (pH 7.3) for 1-2 h at 0-4°C. The gills are observed to comprise of gill arch, gill rakers having minute projections on their inner sides and gill filaments. The filaments originating from the gill arch are in a linear fashion and equidistant from each other, bearing secondary lamellae on both sides of the filament axis and each lamellae running parallel to the adjacent lamellae. The filament epithelium is found to be equipped with pavement cells, mucous cells and chloride cells. The pavement cells have smooth surface with sparse and irregular microridges defining their cell limits. The mucous cells are mainly found on the leading and trailing edges of the filament and the chloride cells are located close to the onset of secondary lamellae. The possible roles of these structures and cells in relation to respiration, water flow and feeding are discussed, setting thereby a platform for further studies in relation to pathology, pollution and stress conditions in aquaculture, natural and polluted environments.
Regardless of lineage, the majority of fish species uses the gills as the primary
site of aquatic respiration having complex organization that is similar in most
teleosts and are multifunctional in nature. The gills form a highly characteristic
feature of fishes and their presence has a marked effect on the anatomy and
functioning of the rest of the animal. The supporting structure of gills in
teleosts is the branchial arch, the next level of organization is the primary
lamellae that finally supports the terminal secondary lamellae. The most superficial
layer of the gill is made up of pavement cells (90%), chloride cells and mucous
cells. The gross and fine structure of the gills of several teleosts have been
extensively studied (Boyd et al., 1980; Kudo
and Kimura, 1984; Moron and Fernandes, 1996; Carmona
et al., 2004). The teleostean gills are involved in a number of functions
such as: gas transfer, acid-base regulation and ion regulation. Further the
gills are very sensitive to different environmental conditions and can undergo
pathological changes which may be mediated by parasites including microbes and
viruses as well.
The Snow trout, Schizothorax curvifrons locally called as Satter gad is a prized indigenous Schizothorax of Kashmir valley whose population is declining day by day. The fish appears to be the morphometrically and meristically most variable and valuable food species of the Kashmir vale but can always be recognized by the combination of large high scale count, high gill raker number and thin lips. The present investigation has been undertaken as effort to delimit what is known and unknown about this interesting organ, besides providing a base line for comparison with pathological and stress conditions in aquaculture and natural or polluted environment.
MATERIALS AND METHODS
Living specimens of normal and healthy Snow trout, Schizothorax curvifrons
(25-30 cm in length, body weight 300-350 g) were collected from the world famous
Dal Lake of the paradise vale. The fishes were killed on the spot with a blow
to the head. The head was immediately decapitated to remove the gills. The excised
gills were rinsed in physiological saline to remove excess mucin and other adhered
particles. After rinsing the gill samples were fixed in 2.5% glutaraldehyde
in 0.1 M sodiumcacodylate buffer (pH 7.3) for 24 h at 0-4°C (Primary fixation).
Following primary fixation the samples were transferred to 0.1 M sodiumcacodylate
buffer for 24 h and post fixed in 1-4% osmiumtetraoxide in 0.1 M sodiumcacodylate
buffer (pH 7.3) for 1-2 h at 0-4°C (Secondary fixation). The post fixed
samples were then dehydrated in increasing concentrations of ethanol (30-100%)
and critical point dried with carbon dioxide. The gill samples were then mounted
on the metal stubs with a paste of colloidal sliver paste and coated with a
thin conductive film of gold in a sputtering coater and examined under LEO 435
VP scanning electron microscope at All India Institute of Medical Sciences (AIIMS)
New Delhi India.
In Schizothorax curvifrons the gross structure of the gills are similar
to those found in other teleosts. Each gill is found to consist of gill arch,
gill rakers and gill filaments (Fig. 1). Arising from the
gill arch are a number of gill filaments which are arranged in a single row
and are equidistant from each other giving it a leaf like structure (Fig.
2). The shape of the filaments is of very filamental type but the length
exceeds the breadth. The secondary lamellae are also observed to be projecting
at right angles to the long axis of the filament each lamellae lying parallel
to the adjacent lamellae (Fig. 3). A numbers of rakers (21-28)
are observed to radiate from both sides of the gill arch on the opposite sides
of the gill filament, the inner sides of the rakers are equipped with minute
projections (Fig. 4). The outer most layer of the filament
consists mainly of pavement cells with mucous cells and chloride cells spread
between them (Fig. 5). The chloride cells are found close
to the onset of secondary lamellae and most of the mucous cells are found on
the leading and trailing edges of the filament.
||Scanning electron micrograph of the gill showing gill rakers
(R), gill arch (A) and filaments (F). X200
||Gill filaments (F) with leaf like lamellae on the upper and
lower sides. X200
||SEM of an individual filament showing the parallel arrangement
of the secondary lamellae (Sl) on the primary lamellae (Pl). X500
||Scanning electron micrograph of gill showing distinct rakers
(R) with minute projections ()
on their inner sides. X300
||SEM of filament epithelium showing pavement cells (PVC), chloride
cells (CC), mucous cells (MC) and microridges (→). X959
The pavement cells are polygonal in shape having almost smooth surface with
sparse and irregular microridges and cell limits are well defined by large and
Scanning electron microscopic observations on the surface ultrastructure of
the gills of Schizothorax curvifrons reveal the presence of well developed
and compactly organized primary filaments which radiate from the gill arches.
Both sides of the primary lamellae are noted to be lined up by secondary lamellae
as also reported earlier in other teleosts (Kudo and Kimura,
1984; Ojha et al., 1987; Arellano
et al., 2004). The gill filaments and their secondary lamellae represent
two general types of epithelium (Laurent and Dunel, 1980;
Filaments or the primary lamellae constitute the most prominent respiratory
structures of the gills. Laurent (1984) stated that the
shape of the gill filaments in general varies from being very filamental to
fairly stubby structures. However, only filamental type of gill filaments are
observed in the present study though the length exceeds the breadth. The filamental
type of filaments may probably help in circulating more and more water through
them and may also reduce the diffusion distance between the blood and the respiratory
water. According to Hughes (1966), the number of filaments
do not increase so markedly in adult fishes as during the juvenile growth period,
although each filament exhibits significant increase in the length as the fish
grows. This leads to an increase in the total length of all the filaments, which
is an important morphometric dimension used in calculating the gill area. In
Schizothorax curvifrons the gill filaments observed are long and narrow
projections lateral to the gill arch that taper at their distal ends. The secondary
lamellae are found evenly distributed along the length of the filament and are
perpendicular to the long axis of the filament besides being parallel to each
other. This is in conformity with the findings of Kudo and
Kimura (1984) and Evans et al. (2005). The
lamellae constitute the most important units of the gill system from the point
of view of gas exchange (Laurent, 1984). According to
Evans et al. (2005), the lamellae not only dramatically
increase the surface area of the gill filament epithelium and result in small
diffusion distance between the blood that perfuses each lamellae and the respiratory
water but are also well suited for diffusive loses or gains of ions and water
to or from the environment. From a design point of view, lamellae are required
to have a large surface area where gas exchange can be facilitated without any
excessive exchange of ions and water. A close contact between water and blood
must be achieved so that oxygen uptake can occur in a limited period during
which the water and blood are passing the lamellae.
Ojha et al. (1987) observed two types of gill
rakers in the gills of freshwater mullet, Rhinomugil corsula and single
type of gill rakers in Sicamugil cascasia. However, during the present
study, only single type of gill rakers are noticeable which may reflect the
variety of food passing through them and the selection of a particular type
of food. The gill rakers are long and slender bearing minute projections on
the inner side. These projections were also observed by Ojha
et al. (1987) on the first type of gill rakers of the gills of Rhinomugil
corsula. The well developed gill rakers and their specific orientation may
possibly be related to the herbivorous nature of the fish studied. The densely
packed rakers aid in the detection and capturing of food flowing through water.
The presence of microridges on the gill rakers can be associated with the holding
of mucous secreted by the goblet cells. The mucous serve to increase the efficiency
of gill rakers by cleaning the surface and exposing the chemoreceptors for the
detection of food and chemical characteristics of ambient water. Moreover, the
arrangement and density of gill rakers are specific for different fishes and
may be a guiding factor for determining food and feeding habits and the taxonomic
status of the fish.
The filament epithelium of teleosts consists of pavement cells, mucous cells
and chloride cells (Morgan and Tovell, 1973; Kendall
and Dale, 1979; Fernandes and Martins, 2001) as
is also evident from the present investigation. The major surface area of the
gill filament is covered by pavement cells, which are largely considered to
play a passive role in gill physiology and are significant or gaseous exchange.
The microridges are noted to be present in between the pavement cells defining
their cell boundaries. The presence of microridges on the apical membrane of
pavement cells seem to increase the functional surface area of the epithelium
and may also play a role in anchoring mucous to the surface (Hughes
and Wright, 1970; Lewis, 1979; Fernandes
and Martins, 2001; Evans et al., 2005). According
to Olson and Fromm (1973), the nature of the microridges
varies in fish species but no correlation between microridge structure and the
physiology and ecology has yet been established.
The mucous cells are more or less similar to those found in the gills of other
teleosts (Laurent, 1984; Fernandes
and Martins, 2001). The mucous secreted by the mucous cells have different
roles such as control of infection, prevention of dehydration, ion regulation
and ion diffusion (Handy et al., 1989; Diaz
et al., 2001). In addition, the mucous of the gill epithelia is also
involved in swimming and defense against pollutants and parasites (Fletcher,
1978). Schizithorax curvifrons is a freshwater fish living generally
in ion poor waters. The released mucin plays a significant role in the prevention
of loss of ions.
The chloride cells in the gills of the fish studied are found close to the
onset of secondary lamellae. Similar observations have been made by Fernandes
and Martins (2001) with respect to the gills of Hypostomus plecostomus.
Franklin and Davison (1989) while working on the scanning
electron microscopy of the gills of freshwater adopted sockeye salmon, Oncorhynchus
nerka, have reported two morphologically different chloride cells on the
afferent surface of the gill filament. However, during the present study, only
single type of chloride cells are noted to be present on the gill epithelium.
The occurrence of single type of chloride cells is possibly due to the fact
that the Schizothorax curvifrons is generally living in cold and ion
poor waters Teleostean chloride cells seem to play an important role in ionic
and osmotic regulation (Karnaky, 1986). According to
Evans et al. (2005) chloride cells occupy a much
smaller fraction of the branchial epithelia, but they are considered to be the
primary active sites of physiological processes in the gills.
The authors not only appreciate the excellent assistance of Dr. T.C. Nag (Associate Prof. Department. of Anatomy AIIMS) but also his helpful suggestions and sharing of knowledge on the gills throughout this study.
1: Arellano, J.M., V. Storch and C. Sarasquete, 2004. Ultrastructural and histochemical study on gills and skin of the Senegal sole Solea senegalensis. J. Appl. Ichthyol., 20: 452-460.
2: Boyd, R.B., A.L. DeVries, J.T. Eastman and G.G. Pietra, 1980. The secondary lamellae of the gills of cold water (high latitude) teleosts: A comparative light and electron microscopic study. Cell. Tissue Res., 213: 361-367.
Direct Link |
3: Carmona, R., M. Garcia-Gallego, A. Sanz, A. Domezain and M.V. Ostos-Garrido, 2004. Chloride cells and pavement cells in gill epithelia of Acipenser naccarii: Ultrastructural modifications in seawater-acclimated specimens. J. Fish. Biol., 64: 553-566.
CrossRef | Direct Link |
4: Diaz, A.O., A.M. Garcia, C.V. Devincenti and A.L. Goldemberg, 2001. Mucous cells in-Micropogonias furnieri gills: Histochemistry and ultrastructure. Anat. Histol. Embryol., 30: 135-139.
Direct Link |
5: Evans, D.H., P.M. Piermarini and K.P. Choe, 2005. The multifunctional fish gill: Dominant site of gas exchange, osmoregulation, acid-base regulation and excretion of nitrogenous waste. Physiol. Rev., 85: 97-177.
CrossRef | Direct Link |
6: Fernandes, M.N. and S.A.P. Martins, 2001. Epithelial gill cells in the armored catfish, Hypostomus plecostomus (Loricariidae). Rev. Braz. Biol., 61: 69-78.
7: Fletcher, T.C., 1978. Defense mechanism in fish. J. Histochem. Cytochem., 32: 681-689.
8: Franklin, C.E. and W. Davison, 1989. SEM observations of morphologically different chloride cells in freshwater adopted sockeye salmon, Oncorhynchus nerka. J. Fish Biol., 34: 803-804.
9: Handy, R.D., F.B. Eddy and G. Romain, 1989. In vitro evidence for the iono regulation- role of rainbow trout mucus in acid/aluminum and zinc toxicity. J. Fish Biol., 35: 737-747.
10: Hughes, G.M., 1966. The dimensions of fish gills in relation to their function. J. Expt.-Biol., 45: 177-195.
Direct Link |
11: Hughes, G.M. and D.E. Wright, 1970. A comparative study of the ultrastructure of the water-blood pathway in the secondary lamellae of teleost and elasmobranch fishes-benthic forms. Z. Zelloforsch. Mikrosk. Anat., 104: 478-493.
12: Karnaky, K.J., 1986. Structure and function of the chloride cells of Fundulus heteroclitusand other teleosts. Am. Zool., 26: 209-244.
13: Kendall, M.W. and J.E. Dale, 1979. Scanning and transmission electron microscopic-observations of rainbow trout (Salmo gairdneri) gill. J. Fish. Res. Bd. Can., 36: 1072-1079.
14: Kudo, S. and N. Kimura, 1984. Scanning electron microscopic studies on bacterial gill-disease in rainbow trout fingerlings. Japan. J. Ichthyol., 30: 393-403.
15: Laurent, P. and S. Dunel, 1980. Morphology of gill epithelia in fish. Am. J. Physiol., 238: 147-159.
16: Laurent, P., 1984. Gill Internal Morphology. In: Fish Physiology, Hoar, W.S. and D.J. Randall (Eds.). Academic Press, New York, pp: 73-183.
17: Lewis, S.V., 1979. A scanning electron microscopic study of the gills of the air breathing catfish Clarias batrachus L. J. Fish Biol., 15: 381-384.
18: Morgan, M. and P.W. Tovell, 1973. The structure of the gill of the trout Salmo gairdneri (Rich.). Z. Zellforsch Mikrosk Anat., 142: 147-162.
19: Moron, S.E. and M.N. Fernandes, 1996. Pavement cell ultrstructural differences on Hoplias malabaricus gill epithelia. J. Fish Biol., 49: 357-362.
Direct Link |
20: Ojha, J., A.K. Mishra and J.S.D. Munshi, 1987. Interspecific variations in the surface ultrastructure of the gills of freshwater mullets. Ichthyol. Res., 33: 388-393.
21: Olson, K.R. and P.O. Fromm, 1973. A scanning electron microscopic study of secondary lamellae and chloride cells of rainbow trout (Salmo gairdneri). Z. Zellforsch. Mikrosk. Anat., 143: 439-449.