Anaphylactic degranulation of guinea pig basophilic leukocytes. I. Fusion of granule membranes and cytoplasmic vesicles formation and resolution of degranulation sacs

Abstract

Anaphylactic degranulation of guinea pig basophilic leukocytes, induced in vitro either with Concanavalin A or sheep serum (antigen), was resolved by transmission electron microscopy into two phases: (1) fusion of cytoplasmic granule membranes to form degranulation sacs communicating with the extracellular space by narrow pores and (2) resolution of degranulation sacs with concomitant granule matrix extrusion. Fusion of granule membranes occurred in the absence of obvious alterations of cytoplasmic filaments or microtubules but was preceded by a rapid increase in the number of 50- to 70-nm. cytoplasmic vesicles, a process evident 1 minute after exposure to lectin. By 5 minutes and at later intervals up to 20 minutes, as individual granule membranes fused to form degranulation sacs, vesicle frequency plunged to values one-half or less of control levels. Cytoplasmic vesicles were apparently incorporated into degranulation sacs and may have had a role in joining together the membranes of adjacent granules. Histamine release, detected at 5 minutes and maximal at 20 minutes, occurred at times when communications between degranulations sacs and the extracellular space were so narrow as to retain most recognizable granule matrix material. Resolution of degranulation sacs proceeded over a period of a day in culture and, in Concanavalin A-induced anaphylaxis, was sometimes incomplete even after 36 hours. During this phase, the frequency of cytoplasmic vesicles returned to normal or supernormal values, and the thin cytoplasmic processes forming the walls of degranulation sacs developed prominent, longitudinally disposed cytoplasmic filaments and ultimately retracted into the main cell body, depositing the membrane-free cytoplasmic granule matrix material outside the perimeter of the cell. Guinea pig basophil anaphylactic degranulation thus differs morphologically and kinetically from mast cell and basophil degranulation in other species in which granule membrane fusion and granule matrix extrusion occur nearly stimultaneously and are complete within minutes. The guinea pig basophil provides a useful model for dissociating these two intrinsic components of the degranulation process.