| The Chinese sturgeon is the first level of the protect animals in our country. Theability of osmoregulation when juvenile Chinese sturgeon transform from freshwater toseawater is the key weather it can live in the sea.In recent years, many scholars hadresearched of the physiological changes of the osmoregulation of the Chinesesturgeon,but the research of chloride cells (CCs) in gills of it remains to besupplemented. Hence, the localization, quantification and ultrastructure of chloride cellsin the gills of the juvenile Chinese sturgeon were studied for the object of mechanism inosmoregulation in this study.1. The reasearch of the optimization for the experimental conditions ofimmunofluorescence staining and localization of the CCs in the gills of juvenileChinese sturgeonThis paper analyzed the distribution of CCs in the gills of juvenile Chinese sturgeonthrough the immunofluorescence staining for frozen sections in order to study thelocalization of it. First, this experiment optimized the experimental conditions ofimmunofluorescence staining for better results. The best optimal dilutions of the firstantibody α5and the fluorescent second antibody Alexa488donkey-anti-mousewas1:1500and1:500, respectively, by the checkerboard method. The results confirmedthat the CCs in the freshwater(FW) and seawater(SW, salinity is20) are both distributeon the gill filament especially at the base,with a few amount on gill lamella through theimmunofluorescence staining, which indicates the CCs of the juvenile Chinese sturgeonare mostly found on the gill filaments.2. The reasearch of the quantification of the CCs in the gills and biochemical analysisof juvenile Chinese sturgeon In order to analyze the quantification of CCs in the gills of the juvenile Chinesesturgeon when it transform from freshwater to the salinity20by immunofluorescencestaining,then quantitative it by IPP6.0. The results indicated that the number of CCs ofFW in the gill filaments (4.72±0.34ind./100μm) is larger than those in the gill lamella(0.64±0.19ind./100μm), and the difference was significant (P<0.01). The number ofCCs of SW in the gill filaments (5.08±0.31ind./100μm) is also larger than those in thegill lamella (0.76±0.16ind./100μm), and the difference was significant (P<0.01). It isvisible that the the CCs in the FW and SW are both distribute on the gill filamentespecially at the base,with a few amount on gill lamella which had a further prove forthe distribution of CCs. The mean aera of CCs of FW in the gill filaments and gilllamella (187.28±18.30μm2,159.53±16.72μm2) is both smaller than those in the SW(361.90±27.88μm2,275.61±14.74μm2). But the difference is not significant (P>0.05)and the mean area of CCs in the gill filaments of the two experiments are both largerthan those in the gill lamella. The shape factor of CCs in the gill filaments and gilllamella of FW and SW was0.72±0.33,0.68±0.46,0.67±0.15,0.62±0.20, respectively,and the difference is not significant (P>0.05). It is visible that the shape of CCs in thesetwo experiments is all oval. We also analyzed the changes of the serum osmolality,plasma osmolality and Na+-K+-ATPase (NKA enzyme) activity of the gills in theprocess of osmoregulation of the juvenile Chinese sturgeon. The serum osmolality andthe Na+,Cl-concentrations of the juvenile Chinese sturgeon were all increased,but theK+concentrations was decrease,and the NKA enzyme activity was increased either. It isfurther confirmed the CCs adjust the balance of ions through the increase of NKAenzyme activity during osmoregulation.3. The reasearch of the electron microscopy structure of the CCs in the gills of thejuvenile Chinese sturgeonWe analyzed the changes of surface structure and the internal structure of the CCs inthe gills of the juvenile Chinese sturgeon by scanning electron microscopy (SEM) andtransmission electron microscopy (TEM) separately in order to research it during theprocess of salinity adaption. The results show the surface structures in the CCs of theFW and SW are different. The surface structures in the CCs of the FW were moreslightly, and the apical opening is larger. And it has a number of microvilli on the surface. The surface structures in the CCs of the SW were invaginated to form a apicalcrypt, and the surface opening was smaller. The pictures of the TEM show that: the mostremarkable structure of the CCs of the FW and SW were all had an abundant ofmitochondria and an extensive tubular system in their cytoplasm. As was seen withTEM the apical membrane of CCs in the FW was flat and slightly projecting andequipment with abundant microvilli. However, the apical membrane of CCs in the SWwas invaginated to form a apical crypt, and the number of mitochondria was increased,the tubular system was extensive, the diameter of the tubular was thickness. Thus, thesurface area of CCs in the FW was larger than the SW. In the hypertonic environment,the CCs were interdigitated with neighboring accessory cells, forming a multicellularcomplex. The CCs and accessory cells shared an apical pit in the hypertonicenvironment, linked by shallow leaky junctions. The apical membrane of CCs hadmicroridges and possessed some glycocalyx which can remain the mucous of themucous cells. The mucous cells were linked with the accessory cells, and it had a lot ofmucous granules of their internal. The results of this paper indicated that the surfacestructure and internal structure of the CCs in the gills of the juvenile Chinese sturgeonchanged with the elevated of the salinity then regulated the balance of the osmotic. |
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