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Methods for Observing Protozoan Contractile Vacuoles

  • Author: Michael Witty 1
  • VIEW AFFILIATIONS HIDE AFFILIATIONS
    Affiliations: 1: Math and Science Department, Florida SouthWestern State College, Fort Myers, FL, 33919
  • Citation: Michael Witty. 2009. Methods for observing protozoan contractile vacuoles.
  • Publication Date : August 2009
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Introduction



Protozoans that live in freshwater have greater water potential outside than inside their cells.  This means water may enter the cell by osmosis and create internal pressures too great for survival.  In naked cells or cells with flexible cell walls, this problem may be solved by the action of the contractile vacuole, which pumps excess water out of cells (2, 4).  Contractile vacuoles are sometimes difficult to observe because their action can be hard to see against a background of other organelles, there  are multiple planes of focus, and their action can make them periodically invisible.  This movie shows several methods for making contractile vacuoles clearly visible.
 
Some protozoans are inherently easier to observe and photograph.  They are larger cells with slower motions; one may be seen in this movie.  However, having an eye for spotting contractile vacuoles is still an advantage, i.e., knowing what one is looking for gives one a better chance of seeing it.  Having a video camera attachment adds to this advantage because the action of contractile vacuoles may be viewed as slow motion.  The protozoan mentioned above is presented at 0.5x natural speed to make the action of the contractile vacuole more prominent.  Media which increase viscosity significantly but do not fatally alter water potential are a third useful method for observing microbial actions that are otherwise too rapid to observe.  These media include methyl cellulose (1) and polyethylene oxide (3), but the preparations Protoslo (Carolina Biological Supply Company, Burlington, NC) or Detain (Ward's Natural Science, Rochester, NY) may be useful and convenient options.  The action of flagella, cilia, and contractile vacuoles can be observed clearly using these materials.  The best results are seen when these three methods are combined, i.e., when an organism with a prominent contractile vacuole is immobilized in a viscose mounting medium and video images are recorded.  Because the protozoan is generally immobile, images can be presented in fast motion making the action of the contractile vacuole very prominent, as shown in the last frames of this video.



Methods

 
Water from submerged mud in a temporary puddle was drawn into a Pasteur pipette then transported to the laboratory in an Eppendorf tube.  One drop of water and one drop of 1% methyl cellulose were mixed on a glass slide and then a cover slip was applied.  This material was examined under conventional bright-field microscopy.  Protozoa were located, then their motion was recorded using a Pupil Cam attachment (Ken-A-Vision, Kansas City, MO) and replayed using fast motion.



Discussion



Contractile vacuoles are common features in many freshwater protozoa .  However, they are difficult to identify because protozoa are almost always in rapid motion.  After seeing contractile vacuoles in this movie, students will be experienced enough to identify this organelle in their own samples of protozoa.



References

1.  Brokaw, C. J.  1963.  Movement of the flagella of Polytoma uvella.  J. Exp. Biol. 40:149–156.

2.  Patterson, D. L.  1980.  Contractile vacuoles and associated structures: their organization and function.  Biological Reviews of the Cambridge Philosophical Society 55:1–46.

3.  Spoon, D. M., C. O. Feise, and R. S. Youn.  1977. Poly(ethylene oxide), a new slowing agent for protozoa.  J. Eukaryot. Microbiol. 24 :471–474.

4. Taylor, W. D., and R. W. Sanders.  2001.  Protozoa, p. 43–89.  In J. H. Thorp and A. P. Covich (ed.), Ecology and classification of North American freshwater invertebrates, 2nd ed.  Academic Press, New York, NY.

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