Endocytosis at Nerve Terminals: Timing Is Everything

range of capacitance measurements. However, in two The nervous system has evolvedto make use of a variety different types of presynaptic terminals, it has been posof mechanisms that allow information to flow and be sible to measure the simultaneous fusion of vesicles processed among a large collection of individual cells. from a large number of release sites acting in parallel. Perhaps one of the most elegant forms of this communiIn both hair cells of the frog sacculus (Parsons et al., cation occurs at chemical synapses, where the exo1994) and giant terminals of goldfish retinal bipolar cells cytosis of synaptic vesicles is regulated to provide rapid (von Gersdorff and Matthews, 1994a), synaptic vesicles (< ms) secretion of a few thousand molecules of neurofuse with the plasma membrane at multiple specialized transmitter on demand. The orchestration of this cellular release sites (z20 dense body synpases in hair cells, process is initiated by a rise in intracellular Ca21 (Zucker, z60 ribbon synapses in bipolar cells) within a single 1996, this issue of Neuron) and is thought to be critically terminal. The synchronous fusion increases the surface dependent upon the correct functioning of specific promembrane area in the entire terminal and, therefore, teins on both the synaptic vesicle and plasma memprovides a measurable change in capacitance. branes (Scheller, 1995; Südhof, 1995). Equally intriguing The development of vital probes of membrane recyis the subsequent retrieval of synaptic vesicle compocling, in particular the amphipathic fluorescent probe nents, which is accomplished by a remarkably efficient FM1-43 (Betz and Bewick, 1992), has permitted mearecycling system that operates using local endocytic surements of many steps of synaptic vesicle recycling machinery in the presynaptic terminal (DeCamilli and at more conventional fast synapses, where capacitance Takei, 1996). measurements have so far proven impractical. These At synapses where the action of the secreted neuroprobes are used as tracers of membrane traffic that transmitter is terminated promptly (within milliseconds, can be applied or washed away at specific times with such as for the fast acting neurotransmitters acetylchorespect to the exocytotic fusion of the synaptic vesicle line, glutamate, GABA, and glycine), synaptic transmiswith the plasma membrane. The approach is similar to sion can in principle occur at high frequency. Unless the original tracer studies used with electron microsthe recycling of synaptic vesicles occurs with speeds copy, which were central to identifying synaptic vesicle similar to, or faster than, exocytosis, the maintenance recycling (Heuser, 1989). With the new vital fluorescent of synaptic transmission during repetitive stimulation probes, measurements in living tissue have successfully will be limited by the size of the releasable pool and the been applied to the study of synaptic vesicle recycling relative balance of the rates of recycling and release. at the frog neuromuscular junction (NMJ) and synapses Thus, obtaining good estimates of all of these parameformed between hippocampal neurons from rat or ters is crucial to understanding the efficacy of fast synmouse grown in culture. apses over a full range of physiological stimuli. In this The Timing of Endocytosis minireview, I shall focus on recent measurements of The earliest estimates of the kinetics of endocytosis the speed of the first step in the recycling process, came from ultrastructural observations of the uptake of endocytosis. extracellular tracers (horseradish peroxidase, HRP) at The job of endocytosis is to recover synaptic vesicle varied times after a burst of exocytosis (reviewed by components after the synaptic vesicle has fused with Heuser, 1989). These studies indicated that endocytosis the plasma membrane. The efficiency of this recovery proceeded rather slowly, over thecourse of many tens of process is considerable; integral membrane proteins of seconds. Measurable uptake of HRP was only obtained synaptic vesiclesare only transiently accessible to labelusing rather intense, nonphysiological stimulation. This ing with specific antibodies following nerve stimulation was thought to provide a possible explanation for the (Valtorta et al., 1988; Kraszewski et al., 1995). This indidiscrepancy between the time scale of endocytosis cates that although these proteins become part of the measured with this method and more recent observaplasma membrane, their residence in that membrane is tions of much faster, but stimulus intensity-dependent, short lived. The recovered components are then reused membrane retrieval. Capacitance measurements in retilocally in the assembly of new vesicles. This distributed nal bipolar cells (von Gersdorff and Matthews, 1994a)