Axoplasm

Microtubules are universally present in eukaryotes from protozoa to the cells of higher animals and plants (Porter, 1966 Hardham and Gunning, 1978 Lloyd, 1987), but they are absent in mammalian erythrocytes and in prokaryotes. Microtubules participate in a number of cellular functions including the maintenance of cell shape and polarity, mitosis, cytokinesis, the positioning of organelles, intracellular transport to specific domains, axoplasmic transport, and cell locomotion. The diversity of microtubule fimctions suggests that not all microtubules are identical and that different classes of microtubules are present in different cell types or are localized in distinct domains in the same cell type (Ginzburg et al., 1989). 

Another example of parallel arrays of microtubules but with a much looser pattern is provided by axons, showing the involvement of such arrays in axoplasmic transport. This is illustrated in Figure 3. 

The cytoskeleton is involved in anterograde and retrograde axoplasmic transport (Hollenbeck, 1989 Coy and Howard, 1994). 

Lasek, R.J. Brady, S.T. (1985). Attachment of transported vesicles to micrombules in axoplasm is facilitated by AMP-PNP. Nature 316, 645-647. 

Vale, R.D., Schnapp, B.J., Mitchison, T., Steuer, E., Reese, T.S., Sheetz, M.P. (1985b). Different axoplasmic proteins generate movement in opposite directions along microtubules in vitro. Cell 43,623-fr32. 

Microtubules, an integral component of the cellular cy-toskeleton, consist of cytoplasmic tubes 25 nm in diameter and often of extreme length. Microtubules are necessary for the formation and function of the mitotic spindle and thus are present in all eukaryotic cells. They are also involved in the intracellular movement of endocytic and exocytic vesicles and form the major structural components of cilia and flagella. Microtubules are a major component of axons and dendrites, in which they maintain structure and participate in the axoplasmic flow of material along these neuronal processes. 

The short delays between Ca2+ influx and exocytosis have important implications for the mechanism of fusion of synaptic vesicles (see Ch. 9). In this short time, a synaptic vesicle cannot move significant distances and must be already at the release site. From the diffusion constant of Ca2+ in squid axoplasm, one can calculate that Ca2+ could diffuse a distance of only 850 A, somewhat greater than the diameter of a synaptic vesicle. Therefore, in fast synapses, vesicle exocytosis sites must be close to the triggering Ca2+ channels.. Vesicles are exposed to [Ca2+] of a few hundred micromoles near the cytoplasmic mouth of the channels. 

Video microscopy allows study of molecular mechanisms through direct observation of organelle movements while precise control of experimental conditions is maintained. Fast axonal transport continues unabated in isolated axoplasm from giant axons of the squid Loligo pealeii for hours [14]. Video microscopy applied to isolated axoplasm permits a more rigorous dissection of the molecular mechanisms for fast axonal transport... 

Dephosphorylated synapsin inhibits axonal transport of MBOs in isolated axoplasm, while phosphorylated synapsin at similar concentrations has no effect [21]. When a synaptic vesicle passes through a region rich in dephosphorylated synapsin, it may be cross-linked to the available MF matrix by synapsin. Such cross-linked vesicles would be removed from fast axonal transport and are effectively targeted to a synapsin- and MF-rich domain, the presynaptic terminal. 

Brady, S. T., Lasek, R. J. and Allen, R. D. Fast axonal transport in extruded axoplasm from squid giant axon. Cell Mot. 3 (Video Supplement), 1983. 

Lasek, R. J. and Brady, S. T. The Structural Hypothesis of axonal transport Two classes of moving elements. In D. G. Weiss (ed.), Axoplasmic Transport. Berlin Springer-Verlag, 1982, pp. 397-405. 

Schroer, T. A., Brady, S. T. and Kelly, R. Fast axonal transport of foreign vesicles in squid axoplasm. /. Cell Biol. 101 568-572, 1985. 

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