2,5-Hexanedione-induced changes in the monomeric neurofilament protein content of rat spinal cord fractions

Richard M. LoPachin, Deke He, Maria L. Reid, Lisa A. Opanashuk

Research output: Contribution to journalArticle

25 Scopus citations

Abstract

Quantitative morphometric analyses have demonstrated that axon atrophy is the primary neuropathic feature in the CNS and PNS of rats intoxicated with 2,5-hexanedione (HD). Axon caliber is maintained by the exchange of mobile neurofilament (NF) subunits with the stationary polymer and, therefore, HD might produce atrophy by disrupting cytoskeletal turnover. To evaluate this possibility, groups of rats were exposed to HD at dosing schedules (175 mg/kg × 101 days or 400 mg/kg × 26 days) that produced moderate levels of neurological deficits and prevalent axon atrophy in spinal cord white matter tracts. Lumbar spinal cord regions from HD-intoxicated rats and their age-matched controls were Triton-extracted and separated by differential fractionation into a low-speed, insoluble pellet (P1) of NF polymer and a high-speed supernatant fraction (S2), which presumably contained mobile monomer. Cytoskeletal protein contents (NF-L, -M, -H, and β-tubulin) in each fraction were determined by immunoblot analysis. Results show that regardless of HD dose-rate, the NF polymer in P1 remained unaffected, although soluble monomer in the S2 fraction was depleted significantly (60-80% reduction). Fractional β-tubulin contents were inconsistently affected and abnormal higher-molecular-weight NF proteins were detected in the P1 fraction only. Studies with antibodies directed against phosphorylated (RT97) and nonphosphorylated (SMI32) epitopes on NF-H and measurements of corresponding isoelectric range suggested that alterations in phosphorylation were not involved. The selective depletion of Triton-soluble protein suggested that HD adduction of NFs interfered with the dynamic interactions of the polymeric and mobile monomeric pools.

Original languageEnglish (US)
Pages (from-to)61-73
Number of pages13
JournalToxicology and Applied Pharmacology
Volume198
Issue number1
DOIs
StatePublished - Jul 1 2004

Keywords

  • 2,5-Hexanedione
  • Axon atrophy
  • Distal axonopathy
  • Neurofilaments

ASJC Scopus subject areas

  • Toxicology
  • Pharmacology

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