There is a long-standing debate concerning the localization of the primary insult that results in distal axonal degeneration, or 'dying back' neuropathy. To address this question, we created an in vitro model of vincristine neuropathy in rat dorsal root ganglia (DRG). DRGs were grown in compartmentalized chambers, allowing for isolated exposure of the cell body or the axon to vincristine. Initial dose-finding studies identified a dose of vincristine that showed differential effects on cell death when delivered to either the cell body or the axonal compartment. At this dose of 0.05 microM, exposure of the cell bodies had no effect on the growth of axons, whereas addition of vincristine to the axonal compartment caused axonal shortening without affecting the growth of unexposed 'sister' axons. Toxicity was seen only with exposure of the growing axonal tips. These data support localized axonal toxicity as a cause of distal axonal degeneration due to vincristine.

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The distal to proximal degeneration of axons, or "dying back" is a common pattern of neuropathology in many diseases of the PNS and CNS. A long-standing debate has centered on whether this pattern of neurodegeneration is due to an insult to the cell body or to the axon itself, although it is likely that mechanisms are different for specific disease entities. We have addressed this question in a model system of vincristine-induced axonal degeneration. Here, we created a novel experimental apparatus combining a microfluidic divider with a multielectrode array substrate to allow for independent monitoring of injury-induced electrical activity from dorsal root ganglion (DRG) cell bodies and axons while isolating them into their own culture microenvironments. At specified doses, exposure of the cell body to vincristine caused neither morphological neurodegeneration nor persistent hyperexcitablility. In comparison, exposure of the distal axon to the same dose of vincristine first caused a decrease in the excitability of the axon and then axonal degeneration in a dying back pattern. Additionally, exposure of axons to vincristine caused an initial period of hyperexcitability in the cell bodies, suggesting that a signal is transmitted from the distal axon to the soma during the progression of vincristine-induced axonal degeneration. These data support the proposition that vincristine has a direct neurotoxic effect on the axon.

This paper describes the design, fabrication, and characterization of a microfabricated compartmented culture system (micro-CCS) useful for electrophysiological signaling studies in cultured neurons. The focus of the paper is the process of interfacing the micro-CCS with cultured neurons and to demonstrate the applicability of the system for biochemical-mediated electrophysiological studies. Moreover, we show that we can record action potentials from cultured neurons through the extracellular compartmented application of elevated levels of K(+) ions. Finally, we show that we can isolate the electrophysiological effects of the sodium channel blocker tetrodotoxin in one of the compartments of a two compartment culture while recording electrophysiological data from both compartments.