Pannexins (Panx) are proteins with a similar membrane topology to connexins, the 
integral membrane protein of gap junctions. Panx1 channels are generally of 
major importance in a large number of system and cellular processes and their 
function has been thoroughly characterized. In contrast, little is known about 
channel structure and subcellular distribution. We therefore determine the 
subcellular localization of Panx1 channels in cultured cells and aim at the 
identification of channel morphology in vitro. Using freeze-fracture replica 
immunolabeling on EYFP-Panx1-overexpressing HEK 293 cells, large particles were 
identified in plasma membranes, which were immunogold-labeled using either GFP 
or Panx1 antibodies. There was no labeling or particles in the nuclear membranes 
of these cells, pointing to plasma membrane localization of Panx1-EYFP channels. 
The assembly of particles was irregular, this being in contrast to the regular 
pattern of gap junctions. The fact that no counterparts were identified on 
apposing cells, which would have been indicative of intercellular signaling, 
supported the idea of Panx1 channels within one membrane. Control cells 
(transfected with EYFP only, non-transfected) were devoid of both particles and 
immunogold labeling. Altogether, this study provides the first demonstration of 
Panx1 channel morphology and assembly in intact cells. The identification of 
Panx1 channels as large particles within the plasma membrane provides the 
knowledge required to enable recognition of Panx1 channels in tissues in future 
studies. Thus, these results open up new avenues for the detailed analysis of 
the subcellular localization of Panx1 and of its nearest neighbors such as 
purinergic receptors in vivo.