Omega images at the active zone may be endocytotic rather than exocytotic: implications for the vesicle hypothesis of transmitter release - PubMed (original) (raw)

Omega images at the active zone may be endocytotic rather than exocytotic: implications for the vesicle hypothesis of transmitter release

J H Koenig et al. Proc Natl Acad Sci U S A. 1998.

Abstract

A Ca2+-dependent synaptic vesicle-recycling pathway emanating from the plasma membrane adjacent to the dense body at the active zone has been demonstrated by blocking pinch-off of recycling membrane by using the Drosophila mutant, shibire. Exposure of wild-type Drosophila synapses to low Ca2+/high Mg2+ saline is shown here to block this active zone recycling pathway at the stage in which invaginations of the plasma membrane develop adjacent to the dense body. These observations, in combination with our previous demonstration that exposure to high Ca2+ causes "docked" vesicles to accumulate in the identical location where active zone endocytosis occurs, suggest the possibility that a vesicle-recycling pathway emanating from the active zone may exist that is stimulated by exposure to elevated Ca2+, thereby causing an increase in vesicle recycling, and is suppressed by exposure to low Ca2+ saline, thereby blocking newly forming vesicles at the invagination stage. The presence of a Ca2+-dependent endocytotic pathway at the active zone opens up the following possibilities: (i) electron microscopic omega-shaped images (and their equivalent, freeze fracture dimples) observed at the active zone adjacent to the dense body could represent endocytotic images (newly forming vesicles) rather than exocytotic images; (ii) vesicles observed attached to the plasma membrane adjacent to the dense body could represent newly formed vesicles rather than vesicles "docked" for release of transmitter.

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Figures

Figure 1

Figure 1

Typical coxal terminal. (A) Cross section of terminal in normal saline containing mitochondria (m), synaptic vesicles, and two dense bodies at active zones (arrows). (B) Active zone in normal saline demonstrating dense body, made up of the dense body base (large arrow) and filamentous cap (small arrows). Note that the postsynaptic muscle fiber (p) extends a finger-like projection to the active zone. (Inset) Dense body base sectioned horizontally, parallel to plasma membrane. (C) Active zone in normal saline demonstrating attachment of dense body plate to the presynaptic membrane (arrow). Note the double lobed appearance of dense body base (D_–_F, arrowhead also). (D) Active zone in normal saline sectioned slightly tangentially to the plasma membrane to demonstrate the attachment of a subpopulation of vesicles to the dense body by thin fibrils. (E and F) Active zones exposed to 18 mM Ca2+ saline for 10 min, demonstrating many vesicles “docked” at the plasma membrane under the dense body plate (large arrows). Note the attachment of the vesicles to the plate by thin fibrils in E (small arrows). (G) Active zone of shi retinula cell terminal exposed to 29°C to induce vesicle depletion, followed by exposure to 26°C for 1 min. Note the membrane emanating from the plasma membrane adjacent to the dense body base (large arrows). The dense body plate (small arrows) appears to have been displaced away from the plasma membrane by the invaginating membrane. [Bars: A, 1 μm (×30,000); C, 100 nm (×75,000); B and D, 100 nm (×90,000); and E_–_G, 100 nm (×135,000).]

Figure 2

Figure 2

Typical coxal terminal after 30 min exposure to 1 mM Ca2+/35 mM Mg2+ saline. (A) Terminal with three abnormal active zones (arrows) with invaginating plasma membrane under dense body plate. (B_–_D) Active zones in A shown at higher magnification. Note omega-shaped images (large arrows) under dense body plate (small arrows) Dense body base–arrowhead. [Bars: A, 0.5 μm (×68,000); and B_–_D, 100 nm (×135,000).]

Figure 3

Figure 3

Further examples of active zones of coxal terminals exposed to 1 mM Ca2+/20 mM Mg2+ saline for 10 min. The pre- and postsynaptic membranes at the invagination have been traced and are presented as Insets in each figure. All invaginations presented are from active zone membrane aligned to the specialized postsynaptic membrane. (A_–_D) Note long, thin necks of invaginations (arrows). The dense body is out of the plane of sectioning in A_–_C. In D, dense body (arrowhead) is only lightly stained. (E_–_H) Note large size of invaginations (large arrows). In E_–_G, dense body is out of plane of sectioning. In H, the dense body is sectioned slightly tangentially, demonstrating attachment of vesicles to it by thin fibrils (small arrows). (I_–_L) Note apparent displacement of invagination by dense body plate (large arrows). In I, J, and L, dense body base (arrowhead) and plate (small arrows) are lightly stained. [Bars: A, D, F and J, 100 nm (×135,000); B, C, E, G, and H, 100 nm (×90,000); and I, K, and L, 100 nm (×75,000).]

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