EXPERIMENTAL ANALYSIS 



Inhibition of the mitotic spindle 



A large number of substances are known to interfere with the formation 

 and function of the mitotic spindle. The final effect on the cell is not 

 always the same ; if the chromosomes can undergo reconstruction even 

 where their normal distribution by the spindle mechanism has been 

 omitted, then either one nucleus will be formed from the whole double 

 set of chromosomes, or the chromosomes which are scattered throughout 

 the cell may form a number of small nuclei (Miszurski and Doljan- 

 5j^j232^ If, however, no stimulus to reconstruction occurs in the absence 

 of the normal movements of anaphase, then the cell may remain in- 

 definitely in a condition of arrested 'metaphase'. The paired chromatids 

 may either clump together (Plate XV (23) ), or remain discrete. Stages 

 of increasing severity of effect on the mitotic spindle can be dis- 

 tinguished. Gaulden and Carlson^ ^^ have shown that a spindle 

 subnormal in size is formed in grasshopper neuroblasts when treated 

 with marginal concentrations of colchicine. In living tissue cultures 

 (HuGHES^"^) when partial metaphase inhibitors such as urethane, 

 cyanide, and fluoride are added to a cell in prophase, the spindle is 

 formed, and usually anaphase ultimately occurs after a much prolonged 

 metaphase, during which the normal random movement of the chromo- 

 somes along the spindle is maintained (Hughes and Swann^^^). In one 

 cell treated with mustard gas, metaphase was followed for 135 minutes 

 after which anaphase suddenly took place (Hughes and Fell^^^). 

 More specific inhibitors of the spindle will prevent its formation in a 

 cell treated during prophase and will inhibit the random movement of 

 the chromosomes in a cell in metaphase. Such agents are colchicine, 

 aminopterin, and among sulphydryl reactants, iodoacetamide, and 

 chloracetophenone. Once this random metaphase movement ceases, 

 the cell will never proceed to anaphase. Chloracetophenone will halt 

 the movement of the chromosomes within 30 seconds if added to a 

 cell when anaphase has already just begun; aminopterin acts with 

 nearly the same rapidity (Figure 68) ; but inhibition during anaphase 

 has not otherwise yet been observed. 



Of the substances which act as spindle inhibitors, some are effective 

 at very great dilutions. The classical example of these is the alkaloid 

 colchicine, which Ludford^^* found to be effective on cells in mitosis 

 in tissue cultures at a concentration of io~^*^ molar. The action of 

 colchicine on dividing cells in the crypts of Lieberkuhn was discovered 

 as early as 1889 (Pernice;^^^ Eigsti et alii^^^). The unrelated compound 

 I, 4 naphthohydroquinone diphosphate has a similar action at a dilution 

 of the same order (Friedmann et alii^^"^). Cells in culture probably are 

 particularly sensitive to metaphase poisons; 2-5 X lO"^ m colchicine is 

 needed for complete inhibition of the spindle in the neuroblasts of the 



191 



