Aim: The intestinal epithelium of the euryhaline teleost, Anguilla anguilla, is a good model for cell volume regulation research. It is able to actively respond to an osmotic stress by changes in plasma membrane ion transport mechanisms; these responses require the integrity of both F-actin and microtubules, as well as the activity of myosin light chain kinase (MLCK) (Lionetto et al., 2002). Therefore, the aim of the present work was to address the immunolocalization of myosin II in eel intestinal epithelium and to clarify its role in the response to osmotic stress using confocal laser scanning microscopy. Methods: Small segments of middle eel intestine were incubated respectively in isotonic (290 mosm/Kg,), hypotonic (160 mosm/Kg,) and hypertonic (365 mosm/Kg) Ringer solution for 5 and 30 minutes. After incubation, the segments were fixed and frozen at -80°C; then, sections of 10-15 m thickness were cut, placed on poly-L-lysine coated slides and double stained for F-actin and myosin II immunolocalization. They were viewed by a confocal microscope (NIKON TE300- C1). Results: It was clearly demonstrated the presence of myosin II in eel enterocyte, and its localization mainly in the terminal web region just below the actin cortical network. Moreover, myosin II labelling appeared to be increased by hypotonicity in the first 5 min exposure, in parallel with a decrease in the intensity of the apical brush border F-actin labelling; on the other hand, hypertonicity did not seem to exert any significant effect on either myosin redistribution or F-actin labelling. Conclusion: Myosin II is localized in the terminal web area of eel intestinal epithelium in close association with F-actin network, giving rise to an acto-myosin complex that could play a fundamental role in the activation of the osmotic stress response by the involvement of MLCK. However, the specific mechanisms underlining myosin involvement seems to be different in hypotonic or hypertonic stress
Roles of myosin II in the osmotic stress response of eel intestinal epithelium.
GIORDANO, Maria Elena;LIONETTO, Maria Giulia;SCHETTINO, Trifone
2006-01-01
Abstract
Aim: The intestinal epithelium of the euryhaline teleost, Anguilla anguilla, is a good model for cell volume regulation research. It is able to actively respond to an osmotic stress by changes in plasma membrane ion transport mechanisms; these responses require the integrity of both F-actin and microtubules, as well as the activity of myosin light chain kinase (MLCK) (Lionetto et al., 2002). Therefore, the aim of the present work was to address the immunolocalization of myosin II in eel intestinal epithelium and to clarify its role in the response to osmotic stress using confocal laser scanning microscopy. Methods: Small segments of middle eel intestine were incubated respectively in isotonic (290 mosm/Kg,), hypotonic (160 mosm/Kg,) and hypertonic (365 mosm/Kg) Ringer solution for 5 and 30 minutes. After incubation, the segments were fixed and frozen at -80°C; then, sections of 10-15 m thickness were cut, placed on poly-L-lysine coated slides and double stained for F-actin and myosin II immunolocalization. They were viewed by a confocal microscope (NIKON TE300- C1). Results: It was clearly demonstrated the presence of myosin II in eel enterocyte, and its localization mainly in the terminal web region just below the actin cortical network. Moreover, myosin II labelling appeared to be increased by hypotonicity in the first 5 min exposure, in parallel with a decrease in the intensity of the apical brush border F-actin labelling; on the other hand, hypertonicity did not seem to exert any significant effect on either myosin redistribution or F-actin labelling. Conclusion: Myosin II is localized in the terminal web area of eel intestinal epithelium in close association with F-actin network, giving rise to an acto-myosin complex that could play a fundamental role in the activation of the osmotic stress response by the involvement of MLCK. However, the specific mechanisms underlining myosin involvement seems to be different in hypotonic or hypertonic stressI documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
