Retrograde degeneration of neurite membrane structural integrity of nerve growth cones following in vitro exposure to mercury

Inhalation of mercury vapor (Hg-0) inhibits binding of GTP to rat brain tub ulin, thereby inhibiting tubulin polymerization into microtubules. A simila r molecular lesion has also been observed in 80% of brains from patients wi th Alzheimer disease (AD) compared to age-matched controls. However the pre cise site and mode of action of Hg ions remain illusive. Therefore, the pre sent study examined whether Hg ions could affect membrane dynamics of neuri te growth cone morphology and behavior. Since tubulin is a highly conserved cytoskeletal protein in both vertebrates and invertebrates, we hypothesize d that growth cones from animal species could be highly susceptible to Hg i ons. To test this possibility, the identified, large Pedal A (PeA) neurons from the central ring ganglia of the snail Lymnaea stagnalis were cultured for 48 h in 2 ml brain conditioned medium (CM). Following neurite outgrowth , metal chloride solution (2 mul) of Hg, Al, Pb, Cd, or Mn (10(-7) M) was p ressure applied directly onto individual growth cones. Time-lapse images wi th inverted microscopy were acquired prior to, during. and after the metal ion exposure. We demonstrate that Hg ions markedly disrupted membrane struc ture and linear growth rates of imaged neurites in 77% of all nerve growth cones. When growth cones were stained with antibodies specific for both tub ulin and actin, it was the tubulin/microtubule structure that disintegrated following Hg exposure. Moreover, some denuded neurites were also observed to form neurofibrillary aggregates. in contrast, growth cone exposure to ot her metal ions did not effect growth cone morphology, nor was their motilit y rate compromised. To determine the growth suppressive effects of Hg ions on neuronal sprouting. cells were cultured either in the presence or absenc e of Hg ions. We found that in the presence of Hg ions, neuronal somata fai led to sprout, whereas other metalic ions did not effect growth patterns of cultured PeA cells. We conclude that this visual evidence and previous bio chemical data strongly implicate Hg as a potential etiological factor in ne urodegeneration. NeuroReport 12:733-737 (C) 2001 Lippincott Williams & Wilk ins.