One of the critical problems in the development of fibre tracking techniques is that there are no other available methods to assess the course of nerve fibres in vivo, or reference standards to which data can be compared. Indeed, knowledge of white matter fibre anatomy derives from postmortem studies, where even in the best conditions only the main fibre bundles can be followed and the resolution is insufficient to constitute a reference for validation .
Ablation studies of animal models make it possible to document the course of axons using specific tracers. Though representing the reference standard for connectivity studies, these data cannot however be extended to humans because tracer methods follow single axons at the cell level, and axons may cross different fibre bundles along their course. Owing to the practical impossibility of obtaining adequate statistics for 1011 neurons, these methods cannot be used to validate effectively data obtained using tractographic techniques.
Despite the limitations outlined above, a good agreement between fibre tracking and anatomy has however been demonstrated [24,44].
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