RESUMEN
A method is described for measuring the volume of individual specimens of Amoeba proteus which utilizes an easily constructed compressor to flatten the specimen to a known thickness. The microscopic image of the flattened specimen is captured on tape, digitized and analysed with the NIH Image software. The results from one specimen are given to illustrate the sources and magnitude of errors affecting these volume measurements.
Asunto(s)
Amoeba/citología , Tamaño de la Célula , Procesamiento de Imagen Asistido por Computador , Programas Informáticos , Animales , National Institutes of Health (U.S.) , Estados UnidosRESUMEN
The position of a mono-oriented chromosome changes as it oscillates to and from the pole to which it is attached. Such oscillatory behavior reveals that the net force on a mono-oriented chromosome is constantly changing. Fluctuations may occur in both the polewardly directed force acting at the kinetochore and the opposing outwardly directed force associated with the aster. We have examined the ejection properties of the aster--as well as the oscillatory behavior and positioning of mono-oriented chromosomes--in relation to astral microtubule turnover. We treated cells containing monopolar spindles with drugs that affect microtubule turnover, either by promoting the depletion of dynamically unstable astral microtubules (nocodazole and colcemid) or by augmenting their numbers and stability (taxol). Both types of drugs stopped the oscillatory behavior of mono-oriented chromosomes within seconds. The final position of the chromosomes depended on how microtubule turnover was affected. In the case of nocodazole and colcemid, non-kinetochore astral microtubules were depleted first and the kinetochore-to-pole distance shortened. In these cells chromosome fragments generated by laser microsurgery were no longer expelled from the center of the aster. By contrast, with taxol the number of non-kinetochore microtubules increased and the astral ejection force became stronger as shown by the finding that the chromosomes moved away from the pole to the periphery of the monaster. Moreover, arms severed from chromosomes at the periphery of the taxol monaster failed to move further away from the aster's center. From these observations we conclude that the oscillatory movements and changing position of a mono-oriented chromosome relative to the pole are mediated by changes in the number of astral microtubules. The dynamic instability of astral microtubules that leads to a rapid turnover may contribute to the astral ejection force by allowing the continual growth of microtubules out from the aster. Growing astral microtubules may exert a pushing force that their rigidity maintains until their depolymerization.