ANAPHYLACTIC DE-GRANULATION OF GUINEA-PIG BASOPHILIC LEUKOCYTES .1. FUSION OF GRANULE MEMBRANES AND CYTOPLASMIC VESICLES - FORMATION AND RESOLUTION OF DE-GRANULATION SACS

Abstract

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