Preparation of Neuropeptide-Containing Fractions from Biological Materials

Neuropeptides vary appreciably in terms of then molecular mass, charge, and hydrophobicity so that there is no single optimum method for their extraction from biological materials such as tissues, cultured neurons, plasma, or cerebrospinal fluid (CSF). As all neuropeptides are rapidly degraded by a range of relatively nonspecific peptidases (1 ), the extraction procedure must release the peptides from storage vesicles into an environment in which the enzymes are inactive. Several general methods have been used to inactivate the neuropeptide-degrading enzymes while efficiently releasing the neuropeptides into the extraction medium. These include the use of boiling aqueous solvents at low or neutral pH; organic solvents, or a mixture of aqueous and organic solvents, at low temperature; and aqueous solutions of chaotropic agents such as 6M guanidine hydrochloride containing a cocktail of protease inhibitors. This article will describe protocols using these different approaches.