Morphology Of The Respiratory System In Ichthyophis Bannanicus

Background:China is located in eastern Asia,with a vast territory,across the Palaearctic and Oriental zoogeographical regions,and the natural environment is extremely complex and diverse,containing rich amphibians resources.However,Gymnophiona in China only has a species,a genus and a family.Its Chinese Name is????,and scientific name is Ichthyophis bannanicus Yang,which was discovered by Yang Datong et al.,the Chinese Amphibious Herpetologists in 1974 near Mengla County,Xishuangbanna in Yunnan.So far,it is confirmed to be the only representative of amphibians Gymnophiona in China.It's called Taro Plant Snake,Double-headed Snake,Gilt-edged Snake or Double Gold Striped Snake by the local people in the habitat,i.e.the Bone Melt Snake in Chinese legend.The population of this species is very rare and mainly distributed in Yunnan,Guangxi,and Guangdong of China,and it has been proved that this species also is distributed in Vietnam and Thailand,which is included in the list of China Main Terrestrial Wildlife and the"Three-haves(beneficial value,economic value and scientific research value)" Animal.I.bannanicus is oviparous;the larva is aquatic and the adult is terrestrial.Significance:It is widely accepted that the Gymnophiona is the most primitive group in the three existing amphibians,so theoretically it should retain more plesiomorphies that provide more evidences for the research on the evolution history of terrestrial vertebrates from aquatic fish ancestors and the disclosing of the evolutional process of the animals from the aquatic to the terrestrial.The primary problem to be solved for landing of ancestral terrestrial vertebrates is breathing,rather than food;therefore the best content for the study on the landing process of ancestral vertebrates is the evolutional history of respiratory system.Certainly,in this study,in addition to this evolutionary significance,as I.bannanicus is the only species resource in the biodiversity resources repository in China,the study and understanding on it is a symbol of the sovereign resources and is conducive to ensure the Chinese property related to the knowledge of the species as well as the protection and development of the species.Objectives:To obtain the structure characteristics of organs composition of I.bannanicus respiratory system in there different living environments-oviparous,aquatic and terrestrial environments,so as to master the evolvement rules of I.bannanicus respiratory system formed in oviparous,aquatic and terrestrial environments;and to discuss the relationship between the morphology and function of the organs composition of I.bannanicus respiratory system;the phylogenetic relationship among the three orders existing in amphibians;the evolution process of respiratory system of vertebrates from aquatic to terrestrial evolution and biological evolution history and phylogenic history presented in the evolution process of respiratory system.Materials:Three developmental levels of I.bannanicus corresponding to the oviparous,aquatic and terrestrial environment conditions:embryo,larva and adult.Methods:Mainly use light microscopy and electron microscopy to fully observe the morphological characteristics of I.bannanicus respiratory system presented in there different living environments(oviparous,aquatic and terrestrial)and build the morphological evolution sequence of I.bannanicus respiratory system.Results:1.Structure characteristics of organs composition of I.bannanicus respiratory system in there different living environments-oviparous,aquatic and terrestrial environments(1)In Oviparous EnvironmentSkin is thin,and the layer of epidermal cells developed from one to three.At Stage 34,the epidermis consists of two layers of pavement cells;in the epidermis of head,middle body and tail,the thickness of trunk and tail has no significant difference(n=135,P>0.05),but the thickness between head and middle body as well as tail has significant difference(n=135,P<0.05).The variation in epidermal thickness among each part is very great;the thinnest place in the whole body is about 5pm,and the thickest place is about 30?m,with the average thickness being about 17?m.The thin Leydig cells exist between two layers of epidermal cells.Basilar membrane is under the epidermis,distributing the rich capillaries and mixing with a lot of pigment cells.The thickness of water-blood barrier is about 17?m.The immature gland cells in the dermis are gathered in groups,and the glandular cavity begins forming.At Stage 35,the epidermis consists of three layers of cells,which are mostly flat with other shapes;the large Leydig cells are distributed intensively in the intermediate layer,which is the densest in the head,with 44800 pcs/mm2,and is relatively sparse in the body and tail;the size of Leydig cells is generally 4-6 times of the adjacent epithelial cells.There is a significant difference between the epidermal thickness of the middle body and the head as well as tail(n=135,P<0.05),but the thickness between head and middle body and tail has no significant difference(n=135,P>0.05).The basilar membrane under epidermis is thicker than that at Stage 34,distributing rich blood vessels and mixing with a lot of melanophore.The thickness of water-blood barrier is about 23?m.The glandular cavity with duct in the dermis have formed,and the glandular cavity is filled with secreta.At Stage 34,the gill is well-developed and there are 6 gill ramus,i.e the left front gill ramus,the right front gill ramus,the left middle gill ramus,the right middle gill ramus,the left rear gill ramus,and the right rear gill ramus;each three gills on the left and right are located on both side of the neck.Gill ramus consists of gill axis and gill filament inserted in it.A pair of gill rami in the middle is the longest,which is almost twice as much as the front and rear pair of gill rami,and its left and right gill ramus have no significant difference(n=6,P>0.05);the front and rear pair of gill rami are relatively short,and the length between them has no significant difference(n=6,P>0.05).The length between left and right gill ramus in the same pair has no significant difference(n=6,P>0.05).The base width of the front and middle gill axis has no significant difference(n=6,P>0.05);but there is a significant difference between them and the base width of the rear gill axis(n=6,P<0.05).There is no significant difference among the tail width of the front,middle and rear gill axis(n=6,P>0.05).The number of gill filaments in different pairs of gill rami is different,and the number of gill filaments in middle pair has significant difference with that in front and rear pairs after comparison(n=6,P<0.05),and there is no significant difference between the number of gill filaments in front and rear pairs(n=6,P>0.05).There is no significant difference between the number of gill filament on the left and right gill rami in the same pair(n=6,P>0.05),but the case with equal number is rare.In an individual,the longest filament is commonly located in the middle axis of the mid-pair of the gills;by contrast,the shortest filaments are found in the tails of the three pairs.The length of the filaments at the base of the axis and in the middle part of the mid-pair of the gills did not significantly differ from that of the corresponding filaments in the front pair(n=36,P>0.05);conversely,the length of the filaments at the base of the axis and in the middle part of the mid-pair of the gills significantly differed from that of the corresponding filaments in the rear pair(n=36,P<0.05).The lengths of the gill filaments in the gill axis tail of three pairs of gill rami have no significant difference(n=36,P>0.05).in the same pair of gill rami,the length of the gill filamentsin the corresponding part on the gill axis of left and right gill rami has no significant difference(n=36,P>0.05).In the same pair of gill rami,the gill filaments in the middle of gill axis is longer than that in the base and tail,but has no significant difference from the gill filaments in the base(n=36,P>0.05)and has a significant difference from the gill filaments in the tail(n=36,P<0.05).Two major blood vessels are parallel throughout the gill axis and filament.The gill epithelium is thin,which consists of two layers of cells,and the thickness of the thinnest place is less than 2?n.At Stage 35,the gill becomes degenerated,and the gill rami are shorten than at Stage 34,and the statistics(n=48)significances of various comparisons are the same as Stage 34;the degenerative process of gill has obvious degradation characteristics,and the random detachment will occur at Stage 36.Six different forms of epithelial cells are distributed in the gill epithelium with two layers of cells:pavement cells with microridges in the surface layer,the pavement cells with mucus and cilia in the surface layer,the pea shaped Langerhans cells with mucus in surface layer,the pavement cells in the lower layer,the rich granular cells in the lower layer,and the round Merkel cells in the lower layer.Lung has been formed in embryonic period and consists of trachea,left lung and right lung;the right lung is about three times as long as the left lung,and the left lung is basically as long as trachea.At Stage 34,the lung meshes(lung sac)are apparent,with about 2000 in the entire lung.The lung development has obvious time sequence,and the septa surrounding the meshes gradually get shorter from front to back,and the septa in the section turn to nodositas backwards from shape[of trachea segment.At Stage 35,the lung sacs further expand outward,and the septa become high,from shape ? of trachea segment to shape U of front and middle segment of left and right lungs to shape[of the tail from front to back.Lung cell with only one layer has various shapes,and most of cells are protuberant towards the cavity,and the surface is covered with microvillus;the electron-dense granules,lamellar bodies,vesicles and other organelles appear within the cells;non-respiratory blood-air barriers appear,and the average of the thinnest place is 5.23?m and the span is among 0.16-10.05?m;the thinnest place is only 0.16?m.(2)In Aquatic EnvironmentThere are four layers of skin epidermal cells,and each layer of cells has various shapes,and the outer superficial cells have keratinization integration and obscission.Each part of epidermal thickness was significantly different(n = 180,P<0.05).Leydig cells are distributed in all cell layers and degenerated and ruptured on the outer surface layer,with the number being significantly reduced;Nevertheless,there are still the most cells on the head,which reaches 100 pcs/mm2.The average thickness of water-blood barrier is 61?m,and the span is among 35-100?m.Dermal glands are mostly distributed on the head and tail and less in middle body.Right lung is about four times as long as left lung,and trachea is about twice as long as left lung.Trachea segment has cartilaginous bars,and tracheal lung is obvious.Lung sac further expand outward and is shaped basically.Lung epithelial cell has various shapes,with lamellar bodies and microvillus.The electron-dense granules or lamellar bodies in the cytoplasmic process of lung cells are less than that in the part of cell body;a large number of electron-dense granules or vesicles or lamellar bodies may arrive to extracellular space through the exocytosis via the top of lung cells.The average in the thinnest place of blood-air barrier is 1.13?m,and the span is among 0.77-1.62?m;the thinnest place is only 0.77?m.(3)In Terrestrial EnvironmentEpidermis consists of 5-6 layers of cells,and cell polarization is obvious.The outward two to three layers of cells are flat,and the superficial cells are fully integrated and keratinized.The skin epithelial is irregularly thickening,which is outward raised to form skin fold;the sinking between skin folds is skin groove.There is no significant difference between the thickness of middle body epidermis and head epidermis(n=180,P>0.05),but there is a significant difference between the thickness of the middle body epidermis and tail epidermis(n= 180,P<0.05);and there is a significant difference between the thickness of head and tail epidermis(n=180,P<0.05).There are Merkel cells in the epidermis.The basilar membrane of epidermis is quite wide,which is full of melanophore and blood capillaries under it.The blood capillary is quite thick however,with a diameter of 20-36?m.The number of mucous gland in the head is the largest,whereas the number of granular glands in the tail is the largest.Lung can be clearly divided into three sections,i.e.tracheal lung,left lung,and right lung.Tracheal lung is the lung tissue growing between the incomplete cartilage ring gap and upper and lower cartilage rings in the trachea,and is an important part of the lung as closing to the glottis and directly attaching to the trachea.Right lung is about three times as long as left lung,and trachea is about 1.3 times as long as left lung.The lungs are fully vascularized;there is something opacified at the end of the left and right lungs,whose degree of vascularization is low and is filled with air.Lung sac has radial arrangement,and individual lung sac which grows independently protrudes out of the lung.The front of the trachea segment have incomplete cartilage ring,and there are cartilaginous bars and spikes behind the rear trachea segment and then nothing backwards.The multilevel septa occur within the lung sac.Lung epithelium consists of a single type of a layer of lung cells.There are rich blood-air barriers;in the composition ratio of the blood-air barrier,the interstitial connective tissue layer accounts for 50%of the blood-air barrier,and the thickness of the epithelial tissue is nearly close to endothelial thickness.The average of blood-air barrier at the thinnest place is 2.08?m,and the span is among 0.68-7.86?m;the thinnest place is 0.68?m.The muscle-controlled bodies portruding into the cavity,accompanying a synaptic knob located on the top are dispersed on the surface of lung sac lumen.2.Evolution Rule of I.Bannanicus Respiratory System in There Different Living Environments-Oviparous,Aquatic and Terrestrial Environments(1)Morphological Evolution of Respiratory System in Three EnvironmentsMorphological Evolution of Skin:at Stage 34,the epidermis only consists of two layers of thin pavement cells,and there are rich blood capillaries under epidermis.At Stage 35,the epidermic cells increase to three layers,and the shapes are mostly flat,with other differentiated shapes;some cells are mature early,such as Leydig cells and glandular cells;the capillary network density under epidermis reaches a peak.When I.Bannanicus lives in the water,the epidermal cells may increase to five layers,and the epidermis continue to be thickened,and the surface layer epithelia become keratinized and the diameter of blood capillaries under epidermis are gradually largened.After landing,the epidermis increased to seven or eight layers,and the epidermal cells polarization are obvious;one and two layers of the outer epidermis are keratinized gradually,and the periodic molting occurs;and the diameter of subepidermal blood vessel is further increased.Morphological evolution of gill:at stage 21,gill begins to appear;at stage 22,the gill filament grows;at stage 33,the size of gill ramus reaches a peak;at stage 34,the gill ramus becomes shrinked and degenerated,and softened after turning pale;at stage 36,the gill ramus tissue falls off.Morphological evolution of lung:at stage 21,lung body is bright,and the meshes surrounded by septa on the internal surface of lung is apparent;the lung sacs are shape[substantially;pulmonary artery and vein in the lung wall have no branches.At stage 22,pulmonary artery and vein in the lung wall begin to branch broadwise,and the tracheal lung and front and middle section of left and right lungs become indistinct and the mesh shape can not be seen clearly;the lung sacs in the tracheal lung and front and middle section of left and right +ungs is substantially shape ? or ?.After entering the water,pulmonary artery and vein in the larva lung branch further and the degree of vascularization is further improved,and the tracheal lung and front and middle section of left and right lungs are more indistinct;lung sacs are radially expanded and raised outward;the tracheal lung and front and middle section of left and right lungs begin to form lung sacs which are much more complex than shape ? lung sacs;lung sacs at the rear of lung is shape U or deep V shape and the cracks(cavity)occur on the bottom;cartilaginous bars occur on the tracheal lung section;lung cells are raised towards the lumen;blood-air barriers are formed;rich exocytosis substances appear in the lung cavity.After landing,the lung sacs are extremely expanded all around along the middle lung axis;the evaginated extrapulmonary lung sacs are more obvious;the incomplete cricoid cartilages are formed in the front section of trachea;blood-air barriers in the lung are rich.(2)Maintenance of Respiration Function of Respiratory System in Three EnvironmentsIn the oviparous environment,the maintenance of respiration is that the breath gradually and orderly passes through the respiratory organs from skin ? skin + gill ? skin.Specifically:before stage 22,the air exchange required by embryonic development is borne by the skin;after stage 22,gill begin to assist in the air exchange;as individual respiration demand increased,the assist role of gill turns into the main role;until stage 33,air exchange borne by gill reaches a peak,while the proportion of the skin achieve a minimum;after stage 33,gill is gradually degenerated and the portion of air exchange borne by it is gradually reduced and may be zero before the hatch(approximately at stage 36);at the same time,the respiration portion borne by skin may be increased accordingly and reaches a peak until hatching.In aquatic environment,the maintenance of respiration is that the breath gradually and orderly passes through the respiratory organs from skin ? skin + lung ? lung.Specifically:over a period of time after the larvae is hatched,respiration is borne by skin fully,and then lung will participate in air exchange in the individual in the mode of air breathing;with the increasing of use frequency of the lung,the portion of air exchange accounted for the respiratory volume by the lung is increasing,while the portion borne by the skin is getting smaller and smaller,and this trend may be continued until an individual landing is complete.Finally,skin may not participate in air exchange,and the lung almost become the only respiratory organ in the daily life of individual animal.Conclusions:The respiratory organs of I.bannanicus consist of skin,gill and lung;in order to meet the respiratory needs of different living environments,different respiratory organs combination is applied in the species.The structure characteristics of organs composition of I.bannanicus respiratory system which can adapt to different living environments and play the role of air exchange is the structure characteristics which represent the phylogeny position of this species.There are many common characteristics between each organ and structure constituting respiratory system of I,bannanicus and that of other caecilian amphibians,and there are personality characteristics which represent the species of I.bannanicus as the basic group members of caecilians;when comparing the structure characteristics of urodeles and Anura respiratory organs,a variety of comparison results show:compared to Anuran,the caecilian and urodela have closer genetic relationship in the phylogeny.In the life history of individuals,the skin of I.bannanicus may develop in the direction which does not help for air exchange;and gill is the aquatic respiratory organ which may appear in case of respiratory needs in a specific period,which is vigorous first and degenerated later;lung may develop in the direction which helps for air exchange.In the different environments of the life history of individuals,the respiration of I.bannanicus is maintained through the gradual and orderly transformation of different combinations of respiratory organs.The morphological evolution of I.bannanicus respiratory system reflects the history of biological evolution and the history of the species phylogeny,and represents the system evolutionary history of respiratory system.The elimination of external factors which hinder the smooth transfer of respiratory function is the approach to improve the survival of the species.