ADAPTATION OF THE DISECTOR METHOD TO RARE SMALL ORGANELLES IN TEM SECTIONS EXEMPLIFIED BY COUNTING SYNAPTIC BODIES IN THE RAT PINEAL-GLAND

The dissector is the only objective method for quantifying particles o f variable size in a given volume. With this method, cell organelles a re identified on adjacent sections, but only those present in one sect ion are counted. When counting extremely rare structures in transmissi on electron microscope sections (physical dissector), the usual proced ure of counting on electron micrographs is limited for economic reason s (e.g. micrographs highly outnumbering the investigated structures:). Hence, to apply this unbiased stereological method, a modification of the physical dissector concerning 3 aspects has been developed. (1) T he prerequisite of screening large corresponding tissue areas (here si milar to 65000 mu m(2)) was fulfilled by examining tissue areas along the edges of ultrathin sections. (2) The size of the counting frame wa s determined by measuring the lengths of the section margins (minus a guard area) by means of a Morphomat. This value was multiplied by the width of the investigated tissue zone, corresponding to the diameter o f the electron microscope viewing screen. (3) Dissector counting was c arried out simultaneously on both sections (bidirectional dissector) t o improve efficiency. In the present study tiny synaptic bodies (SBs) were quantitated by dissector in a rat pineal gland, yielding similar to 30 SBs/1000 mu m(3). By contrast, single section profile counts of SBs amounted to 90 SBs/20000 mu m(2). Since the presently described ad aptation of the dissector is time-consuming, it is proposed to determi ne a proportion factor allowing to estimate number of structures per v olume based on single section profile counts. This would decrease the evaluation time by more than 50%.