Fine structure of synaptic vesicles in two types of nerve terminals in crayfish stretch receptor organs: influence of fixation methods

Abstract

Crayfish stretch receptor organs were used. The standard procedure consisted of primary fixation with a 1.6% glutaraldehyde solution containing either buffer which was 60% hyposmotic or buffer which was isosmotic with the physiological solution (440 milliosmols), washing with isosmotic buffer, and postfixation with an isosmotic 1% osmium tetroxide solution. Under these conditions, we encountered two types of nerve terminals; small-vesicle terminals (SVTs) containing small elongated vesicles (300 approximately 330 A) and large-vesicle terminals (LVTs) containing larger round vesicles (430 A). Their location and physiological evidence suggest that SVTs and LVTs are inhibitory and excitatory, respectively. A hyperosmotic primary fixation solution due to increased glutaraldehyde concentration gave results similar to the standard procedures, while a hyperosmotic primary fixation solution due to increased buffer concentration caused shrinkage of the nerve terminal. A hyperosmotic buffer wash produced elongation of vesicles in SVTs and LVTs, while washing with hypoosmotic buffer rendered vesicles in both types round. Direct fixation with isosmotic osmium tetroxide yielded less elongated vesicles in SVTs and irregularly round vesicles in LVTs. However, under all conditions, vesicles in SVTs were smaller than those in LVTs. These results suggest that the consistent morphological difference in vesicles between SVTs and LVTs is size rather than shape, and it is important to standardize the osmolarity of primary fixation and washing solutions when discussing the differences of vesicle shape in various kinds of synapses.