Abstract
BackgroundFamilial amyloidotic polyneuropathy (FAP) is a peripheral neuropathy caused by the extracellular accumulation and deposition of insoluble transthyretin (TTR) aggregates. However the molecular mechanism that underlies TTR toxicity in peripheral nerves is unclear. Previous studies have suggested that amyloidogenic proteins can aggregate into oligomers which disrupt intracellular calcium homeostasis by increasing the permeability of the plasma membrane to extracellular calcium. The aim of the present study was to examine the effect of TTR on calcium influx in dorsal root ganglion neurons.ResultsLevels of intracellular cytosolic calcium were monitored in dorsal root ganglion (DRG) neurons isolated from embryonic rats using the calcium-sensitive fluorescent indicator Fluo4. An amyloidogenic mutant form of TTR, L55P, induced calcium influx into the growth cones of DRG neurons, whereas wild-type TTR had no significant effect. Atomic force microscopy and dynamic light scattering studies confirmed that the L55P TTR contained oligomeric species of TTR. The effect of L55P TTR was decreased by blockers of voltage-gated calcium channels (VGCC), as well as by blockers of Nav1.8 voltage-gated sodium channels and transient receptor potential M8 (TRPM8) channels. siRNA knockdown of TRPM8 channels using three different TRPM8 siRNAs strongly inhibited calcium influx in DRG growth cones.ConclusionsThese data suggest that activation of TRPM8 channels triggers the activation of Nav1.8 channels which leads to calcium influx through VGCC. We suggest that TTR-induced calcium influx into DRG neurons may contribute to the pathophysiology of FAP. Furthermore, we speculate that similar mechanisms may mediate the toxic effects of other amyloidogenic proteins such as the β-amyloid protein of Alzheimer's disease.
Highlights
Familial amyloidotic polyneuropathy (FAP) is a peripheral neuropathy caused by the extracellular accumulation and deposition of insoluble transthyretin (TTR) aggregates
Effect of ion channel blockers As voltage-gated calcium channels (VGCC) were previously found to mediate calcium entry in SH-SY5Y cells [12], and as dorsal root ganglion (DRG) sensory neurons express a variety of VGCCs [29], we examined whether L55P could influence calcium permeability through a similar mechanism at DRG growth cones
Consistent with the possibility that voltagegated sodium channels mediate the TTR-induced opening of VGCC, we found that while TTX had no significant effect on calcium influx, ambroxol and carbamazepine, blockers of TTXR Nav1.8 channels [43,44], inhibited up to 80% of the calcium influx
Summary
Familial amyloidotic polyneuropathy (FAP) is a peripheral neuropathy caused by the extracellular accumulation and deposition of insoluble transthyretin (TTR) aggregates. Previous studies have suggested that amyloidogenic proteins can aggregate into oligomers which disrupt intracellular calcium homeostasis by increasing the permeability of the plasma membrane to extracellular calcium. Protein misfolding is a common feature of many neurodegenerative diseases. In some of these diseases, such as the synucleinopathies and the tauopathies, cytoplasmic proteins aggregate to form intracellular deposits. Familial amyloidotic polyneuropathy (FAP) is a rare autosomal dominant disease characterised by the deposition of transthyretin (TTR) protein in peripheral nerves. The rare L55P mutation produces a more aggressive amyloidosis than the more common V30M mutation, and in-vitro studies show that L55P TTR aggregates more much readily than V30M TTR [12,13,14,15,16]
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