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Recent studies using atomic force microscopy demonstrate for the first time the …


Biology Articles » Biophysics » Fusion Pore in Live Cells » Introduction

Introduction
- Fusion Pore in Live Cells

Introduction 

The fusion of membrane-bound secretory vesicles at the cell plasma membrane and consequent expulsion of vesicular contents is a fundamental cellular process regulating basic physiological functions such as neurotransmission, enzyme secretion, and hormone release. Secretory vesicles dock and fuse at specific plasma membrane locations following secretory stimuli. Earlier electrophysiological studies on mast cells suggested the existence of "fusion pores" at the cell plasma membrane, which become continuous with the secretory vesicle membrane following stimulation of secretion (13). By using atomic force microscopy (AFM), the existence of the fusion pore was confirmed, and its structure and dynamics in both exocrine (16, 19) and neuroendocrine cells (7, 9) were determined at near-nanometer resolution and in real time.

Why had this new cellular structure (the fusion pore) eluded visualization in live cells for so long? The answer lies simply in the resolution limit of the light microscope, which is dependent on the wavelength of the light used, and hence the resolving power would be at best 300–400 nm. The recently discovered fusion pore in live cells is cone shaped, measuring 100–150 nm at its wide end and 15–30 nm in relative depth. As a result, it had evaded visual detection. With the development of AFM (4) and its improved capabilities to image biological samples at near-nanometer resolution, cellular structures such as the fusion pore and its dynamics could be examined at nanometer resolution and in real time (1, 2, 18). In AFM, a probe tip microfabricated from silicon or silicon nitride and mounted on a cantilever spring is used to scan the surface of the sample at a constant force (1). Either the probe or the sample can be precisely moved in a raster pattern by using an xyz piezo tube to scan the surface of the sample (5). The deflection of the cantilever measured optically is used to generate an isoforce relief map of the sample (2). AFM therefore allows imaging at nanometer resolution and in real time of live cells, subcellular structures, or single molecules submerged in physiological buffer solutions. Structure and dynamics of the fusion pore at nanometer resolution is just the first of many structures waiting to be identified in the living cell. This finding has opened the window to a new understanding of the workings of a living cell. In this review, the structure and dynamics of the fusion pore in live cells, as determined by using the AFM, is presented.



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