Computational Analysis of the Effects of Antineoplastic Agents on Axonal Transport

Abstract

Abstract. Axonal transport plays a crucial role in neuronal morphogenesis, survival, and function. Despite its importance, however, the molecular mechanisms of axonal transport remain mostly unknown because a simple and quantitative assay system for axonal transport has been lacking. In order to better characterize the molecular mechanisms involved in axonal transport, we here developed a computer-assisted monitoring system. Using lipophilic fluorochrome chloromethylbenzamido dialkylcarbocyanine (CM-DiI) as a labeling dye, we have successfully labeled membranous organelles in cultured chick dorsal root ganglia neurons. We confirmed that sodium azide, an ATPase inhibitor, and nocodazole, a microtubule-destabilizing agent, markedly suppressed anterograde and retrograde axonal transport of CM-DiI-labeled particles. We further tested the effects of several anti-neoplastic drugs on axonal transport. Paclitaxel, vincristine, cisplatin, and oxaliplatin, all of which are known to be neurotoxic and to cause neurological symptoms, suppressed anterograde and retrograde axonal transport. Another series of anti-neoplastic drugs, including methotrexate and 5-fluorouracil, did not affect the axonal transport. This is the first report of an automated monitoring system for axonal transport. This system will be useful for toxicity assays, characterizing axonal transport, or screening drugs that may modify neuronal functions.

Computational Analysis of Axonal Transport: A Novel Assessment of Neurotoxicity, Neuronal Development and Functions

“…Recently, similar attempts have been reported independently to detect and track vesicles along neuronal axons [4,29,30]. The time-sequence images of moving vesicles, which are observed by fluorescence microscopy with probes, such as fluorescence recovery after FRAP, GFP, and CM-DiI, are used for analyzing axonal transport as inputs.…”

“…The time-sequence images of moving vesicles, which are observed by fluorescence microscopy with probes, such as fluorescence recovery after FRAP, GFP, and CM-DiI, are used for analyzing axonal transport as inputs. The methods are classified into two approaches: the particle tracking in time-sequence image frames [4,29,30] and analysis using a kymograph image [29–31]. We will briefly introduce these methods in this section.…”

“…As discussed in the following section, in order to differentiate between those vesicles with different velocities and directions in a narrow passageway, strategies have been explored, such as use of Bayesian estimation [30], particle movement diagram [4] and kymograph image [29,31]. …”

“…This transport machinery depends on two families of motor proteins to transport molecules along microtubules, kinesin and dynein [1,3]. Defects in axonal transport reportedly can lead to neurodegenerative pathology and related disorders, such as Schizophrenia, Alzheimer’s (AD), Parkinson’s, other motor neuron diseases and drug-induced neuropathies [4–6], highlighting the importance of understanding the mechanisms of axonal transport.…”

“…Therefore, the development of a simple and objective assay system to evaluate axonal transport has long been awaited. We have recently developed a computer-assisted monitoring system for axonal transport [4]. In this review, we will just briefly summarize the physiology and pathophysiology of axonal transport, and discuss about the validity and general versatility of this method, and our tasks for the future.…”