THE ACTIN CYTOSKELETON IN UNFIXED POLLEN TUBES FOLLOWING MICROWAVE-ACCELERATED DMSO-PERMEABILISATION AND TRITC-PHALLOIDIN STAINING

The disposition of the actin cytoskeleton in pollen tubes of Narcissus pseudonarcissus has been investigated using microwave-accelerated DMSO-permeabilisation and TRITC-phalloidin (Tr-Ph) staining. Extending tubes were transferred from growth medium into a calcium-free medium containing 1 mu-g ml-1 Tr-Ph and 5% DMSO. After 10 s irradiation in a 500 W microwave oven, when the temperature in the sample was estimated at 52-degrees-C, some two-thirds of the tubes retained essentially normal apical zonation; in the remainder the cytoplasm was coarsely granular, and the zonation had been lost. Optimal Tr-Ph staining of the actin cytoskeleton was obtained about 1 h after irradiation. In the most favourable cases, the transition from a longitudinally oriented system of fine fibrils in the sub-apical region to a mass of shorter fibrils in the centre of the apex could be traced, with a peripheral population of more extended fibrils continuing further along the forming wall towards the growing point. This organisation can be reconciled with that revealed in recent fine-structural studies of the microfilament system of the pollen tube apex using freeze-substitution. The relationship of the system with the pattern of movement observed in the apical region of the living tube and with the probable mechanism of tip growth is briefly discussed.