1Department of Physiology and Biophysics, Zilkha Neurogenetic Institute, University of Southern California, Los Angeles, CA 90089; 2Max-Planck-Institut für Experimentelle Medizin, Abteilung Molekulare Neurobiologie, D-37075 Göttingen, Germany
Complexin (CPX) protein is a 13-15 kd protein of unknown function, that has been suggested to play a role in regulated exocytosis. Hormones and neurotransmitters are stored in vesicles, and the regulated exocytosis of their contents is the basis of neuronal communication and many other cellular functions. At least 3 distinct steps are involved before release: (1) docking of vesicles at the plasma membrane; (2) priming of vesicles to a readily releasable state; and (3) fusion of vesicle membrane and plasma membrane, resulting in the release of the vesicle contents. The SNARE hypothesis is a current working model to explain these steps in molecular terms. It postulates that the vesicle membrane protein synaptobrevin (also known as VAMP) pairs up with the plasma membrane proteins syntaxin and SNAP25 to form a core complex. This interaction may bring the two membranes into close proximity, thereby initiating membrane fusion and release. Complexin protein binds to the assembled SNARE core complex in a one-to-one stoichiometry. Neurons lacking complexin show a dramatically reduced Ca2+-dependent neurotransmitter release, but why release is reduced is still unknown. We review our studies using adrenal chromaffin cells from complexin knock-out mice to study the role of complexin. We combine biophysical approaches to tease out the role of complexin in the three distinct steps outlined above. These approaches include membrane capacitance measurements, which is an approach that tracks membrane addition and removal; amperometry, which is a quantitative electrochemical measurement of the released catecholamines; and total internal reflection fluorescence (TIRF) microscopy, which is an optical sectioning approach that allows selective imaging of vesicles near the plasma membrane.