Ultrastructural changes in the respiratory lamellae of the catfish, Heteropneustes fossilis after sublethal exposure to malathion.

Transmission electron microscopy study of the gills of Heteropneustes fossilis, exposed to 4 mg/liter of malathion (1/3 of LC50) for 24, 48, 72, and 96 h showed significant changes in its ultrastructures. Exposure to the pesticide after 24 h caused a slightly disarrayed condition in the double layered epithelial structure. Lymphatic spaces became more apparent, and a few chloride cells appeared which protruded toward the peripheral margin of the secondary lamellae. Chloride cells were exposed to the exterior by an apical pit. Pinocytosis was observed with marginal folds (MF) originating from the pillar and epithelial cells. Some vascular constrictions were also seen in the capillaries with erythrocytes. After 48 h exposure, the outer epithelial cells were stretched into a thin boundary wall and lymphatic spaces were engorged with plasma exudate. Chloride cells transversed the whole epithelium of the lamella and came into direct contact with lymphoid space and exterior to epithelial lining. Basement membrane of the capillaries became thicker. After 72 h a distorted lamellar epithelium ruptured in a few places allowing many spheroid bodies and some chloride cells come out. Marginal folds of pillar cells migrated into vascular spaces. Basement membrane of capillaries became thicker and blood channels were constricted causing vascular stasis. No erythrocytes were visible. Blood channels were filled with leukocytes and amoebocytes. After 96 h exposure to malathion narrowing of lymphatic spaces, proliferation of epithelial cells and development of pinocytotic vesicles from marginal folds of pillar cell flanges were observed. Only marginal blood channels maintained normal configuration. Vascular stasis due to thickening of the basal lamina were still evident in centrally located blood channels filled with leukocytes. Vascular stasis would likely cause a decrease in respiratory efficiency. This study has revealed that the gills of H. fossilis were affected by a sublethal dose of malathion. The ultrastructural damages to the gills were observed as early as at 24 h exposure, but the most severe damage occurred at 72 h exposure. However, signs of gill structure regeneration were seen in malathion-exposed fish after 96 h.

Microcirculation of gills and accessory respiratory organs of the walking catfish Clarias batrachus.

BACKGROUND: An ability to extract oxygen directly from the atmosphere enables air-breathing fish to survive otherwise debilitating hypoxic environments. Addition of accessory respiratory organs (ARO) necessitates changes in both the general circulatory system and the microcirculation of the respiratory epithelia. Understanding these modifications provides information on the efficiency of gas exchange organs as well as an indication of the evolutionary processes associated with adaptation to terrestrial habitats. METHODS: Vascular organization and structure of gills and ARO of the facultative air-breathing walking catfish Clarias batrachus were examined by scanning electron microscopy of vascular replicas and fixed tissue. RESULTS: Well-developed filaments are present on all four pairs of gill arches and they possess three vascular pathways: respiratory (arterioarterial), nutrient (arteriovenous), and interlamellar (arteriovenous), typical of teleosts. ARO, consisting of gill fans, dendritic organs on the second and fourth gill arch, and the suprabranchial epithelium are derived from gill tissue and retain structural features and arterioarterial vessels similar to gill filaments. Gill and ARO vessels are in parallel with each other, and together they are in series with the systemic circulation. Nutrients and interlamellar vessels are reduced in ARO. CONCLUSIONS: Other than the presence of multiple ventral aortas, and an additional vessel connecting the suprabranchial epithelium to the dorsal aorta, there are no vascular shunts or anatomical modifications that indicate spatial separation of flow through the heart or between gills and ARO. However, a mechanism is proposed that would prevent unsaturation of dorsal aortic blood by local myogenic vasoconstriction of gill vessels when the fish is in hypoxic water. Despite considerable differences in the gross features of ARO in Clarias and Heteropneustes fossilis (Olson et al. 1990 J. Morphol., 203:165), there are striking similarities in vascular organization and respiratory islet structure that suggest these ARO evolved in a common silurid ancestor and were later modified into an everted arborescent organ or inverted air sac, respectively.

Cephalic circulation in the air-breathing snakehead fish, Channa punctata, C. gachua, and C. marulius (Ophiocephalidae, Ophiocephaliformes).

The macrocirculation in the head of three air-breathing species of Channa was examined with the aid of vascular corrosion replicas and a scanning electron microscopic study was conducted on the pseudobranch, choroid gland and lentiform body. Two facultative air-breathing murrels, C. punctata and C. gachua, and one obligate air-breather, C. marulius were examined. In all three, the air-breathing organs (ABO) and systemic circulations were in-parallel, and both were in-series with the branchial circulation. Efferent branchial arteries from the first and second gill arches formed the arterial supply to the ABO, whereas the third and fourth arch efferents perfused systemic tissues. Postbranchial blood from the second gill arch also entered the systemic circulation directly via a shunt from the efferent branchial artery to the lateral aorta and via hypobranchial arteries. Vascular specialization to prevent mixing of oxygenated ABO venous and deoxygenated systemic venous blood was evident in arterial, but not venous circuits. Pseudobranchs of C. gachua and C. punctata are tri-lobed, in C. marulius they have numerous lobules. Pseudobranch lamellae are wider and shorter along the axis of blood flow than gill lamellae and folded perpendicular to this axis. Pseudobranch lamellae appear to be modified to minimize their epithelial surface while retaining an extensive vascular endothelial-pillar cell surface area, counter-current amplification is also possible. The choroid gland is an extensive planar counter-current capillary rete. The lentiform body of the eye is a globular capillary rete but there is no evidence of a counter-current circulation. The choroid and lentiform rete may have distinct physiological functions.

Microcirculation of gills and accessory respiratory organs from the air-breathing snakehead fish, Channa punctata, C. gachua, and C. marulius.

Snakehead fish of the genus Channa have well-developed air-breathing organs (ABO) yet retain their gill arches for respiratory and non-respiratory functions. Alterations in the macrocirculation accompany inclusion of the ABO and appear to enhance gas exchange efficiency (Munshi et al., 1994. Anat. Rec. 238:77-91). In the present study, the microcirculatory anatomy of gill and ABO from two facultative air-breathing Channa, C. punctata and C. gachua, and one obligate air-breather, C. marulius, were examined in detail using scanning electron microscopy (SEM) of vascular corrosion replicas and fixed whole-sectioned tissue. The results show that the circulation in the filaments from the first, second, and third gill arches is similar to that found in water-breathing teleosts. Fourth gill arch microcirculation of C. punctata is not different from the other three, whereas in C. marulius, it has been greatly modified into a network of low-resistance vascular shunts, although remnants of an intralamellar filamental microcirculation remain. The vascular shunts are formed from extensions of afferent and efferent lamellar arterioles and the complete, or nearly complete, loss of a lamellar sinus. The vasculature of the ABO has been highly modified in all species into a coiled-spiral capillary network with a constricted aperture guarding a dilated capillary dome at the epithelial surface. Microvilli are found congregated on the aperture endothelium of C. punctata but they are virtually absent from C. marulius endothelium. Less than 15% of the ABO capillary surface appears to face the epithelium and thereby contributes directly to gas exchange. These findings suggest that the microvascular modifications observed in Channa entail more than a simple increase in the contact surface between ABO vessels and air and they may serve other unknown physiological functions.

Structure of the respiratory islets of accessory respiratory organs and their relationship with the gills in the climbing perch, Anabas testudineus (Teleostei, Perciformes).

The epithelial and sub-epithelial organization of the accessory respiratory organs of Anabas testudineus has been compared with that of gills by using light and transmission electron microscopy. The details of vascular supply of respiratory islets (RI) and gill filaments and the presence of venous sinusoids in the two systems suggest that the RI have been derived from gill filaments and lamellae. The biserial arrangement of transverse capillaries (TC) in the respiratory islets (RI) is evident under the scanning electron microscope and their homology with the gill filaments and their secondary lamellae has been established. The two sets of transverse capillaries of respiratory islets have been derived either from embryonic transverse or marginal channels of two sets of lamellae of a gill filament. These capillaries with their endothelial septate valves and tongue-like processes offer resistance to blood flow. Gill filaments have two vascular pathways, arterio-arterial and arterio-venous. However, the RI of accessory respiratory organs contain the arterio-venous pathways. This arrangement as well as the septate transverse capillaries may lower the "pulmonary" blood pressure considerably. Two types of mitochondria-rich cells are identified: i) chloride cells with flat microvilli bearing surfaces, devoid of apical pit and (ii) an elongated cell type with sac-like endoplasmic reticulum, with apical pit that comes into close contact with the complex matrix of macrophages, lymphocytes and other loose cells of the epithelium. These cells may be associated with detoxification of the internal media of fish inhabiting foul waters.

Vascular organization of the head and respiratory organs of the air-breathing catfish, Heteropneustes fossilis.

Light and scanning electron microscopy of vascular replicas from the facultative air-breathing fish Heteropneustes fossilis show modifications in the macrocirculation of the respiratory organs and systemic circulation, whereas, gill microcirculation is similar to that found in typical water-breathing fish. Three and sometimes four ventral aortae arise directly from the bulbus. The most ventral vessel supplies the first pair of arches. Dorsal to this another aorta supplies the second gill arches, and a third, dorsal to, and larger than the other two, supplies the third and fourth arches and the air sacs. Occasionally a small vessel that may be the remnant of a primitive aortic arch arises from the first ventral aorta and proceeds directly to the mandibular region without perfusing gill tissue. The air sac is perfused by a large-diameter extension of the afferent branchial artery of the fourth gill arch and its circulation is in parallel with the gill arches. Blood drains from the air sac into the fourth arch epibranchial artery. A number of arteries also provide direct communication between the efferent air sac artery and the dorsal aorta. All four gill arches are well developed and contain respiratory (lamellar) and nonrespiratory (interlamellar and nutrient) networks common to gills of water-breathing fish. Air sac lamellae are reduced in size. The outer 30% of the air sac lamellar sinusoids are organized into thoroughfare channels; the remaining vasculature, normally embedded in the air sac parenchyma, is discontinuous. A gill-type interlamellar vasculature is lacking in the air sac circulation. Despite the elaborate development of the ventral aortae, there is little other anatomical evidence to suggest that gill and air sac outflow are separated and that dorsal aortic oxygen tensions are maintained when the gills are in a hypoxic environment. Physiological adjustments to hypoxic water conditions probably include temporal regulation of gill and air sac perfusion to be effective, if indeed they are so.

Vasculature of the head and respiratory organs in an obligate air-breathing fish, the swamp eel Monopterus (=Amphipnous) cuchia.

Methyl methacrylate vascular corrosion replicas were used to examine the macrocirculation in the head region and the microcirculation of respiratory vessels in the air-breathing swamp eel Monopterus cuchia. Fixed respiratory tissue was also examined by SEM to verify capillary orientation. The respiratory and systemic circulations are only partially separated, presumably resulting in supply of mixed oxygenated and venous blood to the tissues. A long ventral aorta gives rise directly to the coronary and hypobranchial arteries. Two large shunt vessels connect the ventral aorta to the dorsal aorta, whereas the remaining ventral aortic flow goes to the respiratory islets and gills. Only two pairs of vestigial gill arches remain, equivalent to the second and third arches, yet five pairs of aortic arches were identified. Most aortic arches supply the respiratory islets. Respiratory islet capillaries are tightly coiled spirals with only a fraction of their total length in contact with the respiratory epithelium. Valve-like endothelial cells delimit the capillary spirals and are unlike endothelial cells in other vertebrates. The gills are highly modified in that the lamellae are reduced to a single-channel capillary with a characteristic three-dimensional zig-zag pathway. There are no arterio-arterial lamellar shunts, although the afferent branchial artery supplying the gill arches also supplies respiratory islets distally. A modified interlamellar filamental vasculature is present in gill tissue but absent or greatly reduced in the respiratory islets. The macro- and micro-circulatory systems of M. cuchia have been considerably modified presumably to accommodate aerial respiration. Some of these modifications involve retention of primitive vessel types, whereas others, especially in the microcirculation, incorporate new architectural designs some of whose functions are not readily apparent.

Angiotensin-converting enzyme in organs of air-breathing fish.

Angiotensin-converting enzyme (ACE) was measured in tissue homogenates from the African lungfish and six species of air-breathing teleosts (Heteropneustes fossilis, Clarias batrachus, Channa gachua, Anabas testudineus, Notopterus chitala, and Monopterus cuchia) using a standard spectrophotometric assay. In most species, the highest levels of ACE activity were found in the respiratory organs (gills and/or accessory respiratory organs). ACE was also found in heart and kidney tissues from most species and occasionally in liver. Converting enzyme was not found in skin or skeletal muscle from any species and only in blood from H. fossilis and brain from C. batrachus. Captopril, a potent inhibitor of mammalian ACE, inhibited enzymatic activity from all tissues except C. gachua heart and liver and A. testudineus heart. As fish make the transition from aquatic to aerial respiration the gill microcirculation is usually reduced in size and the accessory respiratory organs become elaborated and occupy a more central position in the vascular tree. The presence of ACE in accessory respiratory organs of air-breathing fish appears to greatly enhance the metabolic efficiency of this enzyme on circulating substrates.

Gill microcirculation of the air-breathing climbing perch, Anabas testudineus (Bloch):relationships with the accessory respiratory organs and systemic circulation.

The general macrocirculation and branchial microcirculation of the air-breathing climbing perch, Anabas testudineus, was examined by light and scanning electron microscopy of vascular corrosion replicas. The ventral aorta arises from the heart as a short vessel that immediately bifurcates into a dorsal and a ventral branch. The ventral branch distributes blood to gill arches 1 and 2, the dorsal branch to arches 3 and 4. The vascular organization of arches 1 and 2 is similar to that described for aquatic breathing teleosts. The respiratory lamellae are well developed but lack a continuous inner marginal channel. The filaments contain an extensive nutritive and interlamellar network; the latter traverses the filament between, but in register with, the inner lamellar margins. Numerous small, tortuous vessels arise from the efferent filamental and branchial arteries and anastomose with each other to form the nutrient supply for the filament, adductor muscles, and arch supportive tissues. The efferent branchial arteries of arches 1 and 2 supply the accessory air-breathing organs. Arches 3 and 4 are modified to serve primarily as large-bore shunts between the dorsal branch of the ventral aorta and the dorsal aorta. In many filaments from arches 3 and 4, the respiratory lamellae are condensed and have only 1-3 large channels. In some instances in arch 4, shunt vessels arise from the afferent branchial artery and connect directly with the efferent filamental artery. The filamental nutrient and interlamellar systems are poorly developed or absent. The respiratory and systemic pathways in Anabas are arranged in parallel. Blood flows from the ventral branch of the ventral aorta, through gill arches 1 and 2, into the accessory respiratory organs, and then returns to the heart. Blood, after entering the dorsal branch of the ventral aorta, passes through gill arches 3 and 4 and proceeds to the systemic circulation. This arrangement optimizes oxygen delivery to the tissues and minimizes intravascular pressure in the branchial and air-breathing organs. The efficiency of this system is limited by the mixing of respiratory and systemic venous blood at the heart.

Morphology and vascular anatomy of the accessory respiratory organs of the air-breathing climbing perch, Anabas testudineus (Bloch).

The vascular organization and endothelial cell specialization of the air-breathing organs of Anabas testudineus were examined by light and scanning electron microscopy of fixed tissue and vascular corrosion replicas. The vessels supplying blood to the lining of paired suprabranchial chambers and the plicated labyrinthine organs within the chambers are tripartite, having a median artery and paired, lateral veins. Hundreds of respiratory islets, the functional units of gas exchange, cover the surfaces of both the chamber and labyrinthine organ. A median islet artery supplies the central aspect of each islet and gives rise to numerous short arterioles from which the transverse channels are formed. Transverse channels are parallel capillary-sized vessels that extend in two rows away from the medial arterioles and drain laterally into one of two lateral islet veins. Basally situated single rows of endothelial cells lining the transverse channels form thick, evaginated, tongue-like cytoplasmic processes that project freely into the lumen from the tissue side of the channel. Other thin, septate, cytoplasmic extensions of the same cells form valve-like septa that extend across the channel. Both the septa and tongue-like processes appear to direct the red blood cells to the epithelial side of the channel and thus decrease the diffusion distance between the air and red cell. A large sinusoidal space lies under the transverse channels and may support the channels and even elevate them during increased oxygen demand. The epithelium covering the transverse channels is smooth, which enhances air convection and minimizes unstirred layer effects. The epithelium between the channels contains microvilli that may serve to trap bacteria or particulates and to humidify the air chambers.

Blood of two siluroid fishes during different respiratory conditions.

In the present study an attempt has been made to investigate the changes in different blood parameters in two siluroid fishes, Heteropneustes (H.) fossilis. Bloch and Clarias (C.) batrachus. Linn, both air breathing fishes, during different respiratory conditions, Viz, 1. normal, 2. submerged water condition and 3. exclusive air breathing condition. An increase in different blood parameters [Viz, red blood corpuscles (RBC) counts, haemoglobin (Hb) content and packed cell volume (PCV) etc.] was observed under submerged water and exclusive air breathing conditions. Possible reasons and the mechanism of such changes in different blood parameters have been discussed.

Cytochemical differentiation of tonic (slow) and phasic (fast) muscle units in the ocular muscles of a fresh-water carp, Catla catla (Ham.).

1. Functional morphology of the ocular muscles with special reference to their topography, innervation and action have been studied in a fresh-water carp Catla catla. An attempt has also been made to differentiate tonic (slow) and phasic (fast) muscle units in the various ocular muscles by their SDH activities and their lipid contents. 2. Catla catla has large eyes (2 cm in diameter) in a fish measuring 30 cms. There are four recti (inferior, superior, anterior and posterior) and two obliquii (superior and inferior) muscles for the synchronized rotation of the eye ball within the orbit. Anterior and posterior myodomes are well developed. The former accommodates two obliquii muscles, while the latter gives space for the posterior and anterior recti. 3. In this fish, all the ocular muscles are composite in nature being made up of tonic (slow) and phasic (fast) muscle units. The former is made up of red fibers whereas the latter have only white fibers. In the tonic muscle units lipid is more concentrated. 4. The superior and the inferior obliquii muscles rotate the eye ball on its antero-posterior axis. The upward and downward movements of the eye balls are achieved by the superior and the inferior recti. The anterior and posterior recti move the eye ball in the antero-lateral and postero-lateral directions. 5. The superior obliquii contains about 41% of the tonic and 59% of phasic muscle units. The inferior oblique is made up of 25% of tonic and 75% of the phasic muscle units. 6. The Anterior rectus contains greater percentage of tonic muscle fibers (48%) than the superior rectus (40%), the Interior rectus (34%) and the posterior rectus (35%).