PLASMODESMATAL FREQUENCY IN RELATION TO SHORT-DISTANCE TRANSPORT AND PHLOEM LOADING IN LEAVES OF BARLEY (HORDEUM-VULGARE) - PHLOEM IS NOT LOADED DIRECTLY FROM THE SYMPLAST

We investigated the phloem loading pathway in barley, by determining p lasmodes-matal frequencies at the electron microscope level for both i ntermediate and small blade bundles of mature barley leaves. Lucifer y ellow was injected intercellularly into bundle sheath, vascular parenc hyma, and thin-walled sieve tubes. Passage of this symplastically tran sported dye was monitored with an epifluorescence microscope under blu e light. Low plasmodesmatal frequencies endarch to the bundle sheath c ells are relatively low for most interfaces terminating at the thin- a nd thick-walled sieve tubes within this C-3 species. Lack of connectio ns between Vascular parenchyma and sieve tubes, and low frequencies (0 .5% plasmodesmata per mu m cell wall interface) of connections between vascular parenchyma and companion cells, as well as the very low freq uency of pore-plasmodesmatal connections between companion cells and s ieve tubes in small bundles (0.2% plasmodesmata per mu m cell wall int erface), suggest that the companion cell-sieve tube complex is symplas tically isolated from other Vascular parenchyma cells in small bundles . The degree of cellular connectivity and the potential isolation of t he companion cell-sieve tube complex was determined electrophysiologic ally, using an electrometer coupled to microcapillary electrodes. The less negative cell potential (average -52 mV) from mesophyll to the va scular parenchyma cells contrasted sharply with the more negative pote ntial (-122.5 mV) recorded for the companion cell-thin-walled sieve tu be complex. Although intercellular injection of lucifer yellow clearly demonstrated rapid (0.75 mu m s(-1)) longitudinal and radial transpor t in the bundle sheath-vascular parenchyma complex, as well as from th e bundle sheath through transverse veins to adjacent longitudinal vein s, we were neither able to detect nor present unequivocal evidence in support of the symplastic connectivity of the sieve tubes to the vascu lar parenchyma. Injection of the companion cell-sieve tube complex, di d not demonstrate backward connectivity to the bundle sheath. We concl ude that the low plasmodesmatal frequencies, coupled with a two-domain electropotential zonation configuration, and the negative transport e xperiments using lucifer yellow, precludes symplastic phloem loading i n barley leaves.