Analysis of cellular release using capillary electrophoresis and matrix assisted laser desorption/ionization-time of flight-mass spectrometry

In order to increase our understanding of the mechanisms of learning and me mory in the central nervous system, it is necessary to know the neurotransm itters; and neuromodulators used in the specific neuronal circuits under st udy. Methods have been developed to identify the peptides released from sin gle neurons and neuronal clusters from the common neuronal model Aplysia ca lifornica. Specifically, solid-phase extraction (SPE), capillary electropho resis (CE) and matrix assisted laser desorption/ionization-time of flight-m ass spectrometry (MALDI-TOF-MS) are combined for profiling neuropeptide rel easates. A variety of combinations of SPE and CE were coupled off-line with MALDI-TOF-MS to reduce the high physiological salts, to concentrate the an alytes, and to reduce the complexity of the mass spectra using separation. With these protocols, peptides and proteins up to 11 000 Da were detected i n releasates, offering a much wider mass range compared to direct MALDI ana lysis of the same releasates. A number of expected and unknown neuropeptide s, including egg-laying hormone (ELH) and the partially processed delta/gam ma -bag cell peptide were observed in the SPE-treated releasates from a sin gle Aplysia-cultured bag cell neuron. However, by adding a CE separation af ter the SPE step preceding off-line MALDI-TOF-MS detection, the most comple te neuropeptide profiles were obtained.