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Fibroblasts in culture apparently require an intact system of microtubules in order to adopt and maintain a polarized morphology. In contrast, the polarization of a number of epithelial cell types has been shown to be microtubule-independent. Reconciliation of these apparently contradictory data is difficult, however, because the epithelial cells were studied in short-term primary cultures while the fibroblasts were studied in secondary or longer-term cultures. To clarify the situation we have examined the effects of the microtubule-disrupting drugs, colcemid and nocodazole, on the polarization of a single cell type, the chick heart fibroblast (HF), maintained in both primary (1 degree) and secondary (2 degree) cultures. Immunofluorescence observations of both types of culture showed that in control medium the cells contained abundant microtubules, which were absent if the cells were cultured in medium containing either colcemid or nocodazole. The effects of microtubule-disrupting drugs on the polarization of the cells were quantified using two measures of cell shape, elongation and dispersion, both of which increase with increasing polarization. The results show that microtubule-disrupting drugs do not have a significant effect on the polarization of HF spreading in 1 degree culture but significantly reduce the polarization of HF spreading in 2 degree cultures. The effects of microtubule disruption on HF that had been maintained in 1 degree culture for 6 h, 24 h or 48 h were also quantified.(ABSTRACT TRUNCATED AT 250 WORDS)

A new form of chamber for studying chemotaxis, similar in principle to the Zigmond chamber, allows the behaviour of the cells in a linear concentration gradient to be observed directly. The chamber was developed mainly for studying chemotaxis in fibroblasts using interferometric microscopy and the main design criteria were that it should have better optical characteristics, a higher dimensional precision and better long-term stability than the Zigmond chamber. It is made entirely from glass by grinding a blind circular well centrally in the counting platform of a Helber bacteria counting chamber. This procedure leaves an annular ‘bridge’, approximately 1 mm wide, between the new inner circular well and the original outer annular well. This bridge fulfils the same function as the linear bridge of the Zigmond chamber but the precise construction of the counting chamber ensures that a gap of 20 microns between bridge and coverslip can be accurately and repeatedly achieved when the chamber is assembled. It is envisaged that the improved optical clarity, dimensional accuracy and long-term stability of the new chamber will be advantageous in other applications, particularly in studies requiring critical microscopy or a precise knowledge of the gradient and in studies of cells, such as fibroblasts, that move much more slowly than neutrophils.