Techniques in Neuropeptide Processing, Trafficking, and Secretion

Neuropeptides function as neurotransmitters and neuromodulators. They are synthesized as larger precursors at the rough endoplasmic reticulum (RER), trafficked to the trans-Golgi network (TGN), and sorted into granules of the regulated secretory pathway (RSP) for secretion in an activity-dependent manner. Polymorphisms found in human neuropeptide genes can lead to defects in trafficking and processing of the neuropeptide precursors, resulting in disease. Examples of mutations of human neuropeptide genes that have led to biosynthesis of precursors that were misrouted and only partially processed include insulin (1), and cocaine and amphetamine-regulated transcript (CART) peptide (2), giving rise to diabetes and obesity, respectively. A human valine to methionine mutation in the prodomain of brain-derived neurotrophic factor (BDNF) causes its inefficient sorting to the RSP and diminished activity-dependent secretion of BDNF from hippocampal neurons, resulting in memory deficits in these humans (3). With the sequencing of the human genome, increasing numbers of polymorphisms in neuropeptide genes will be identified. Studies on the trafficking, processing, and activity-dependent secretion of the mutant neuropeptide precursors will be useful in elucidating the molecular and cellular basis of diseases associated with the mutations. There are currently many paradigms and tools to study neuropeptide precursor trafficking, processing, and secretion, and these will be described in this chapter. These procedures are also applicable to studying the processing of other proteins such as neurotrophins.First Editio