Opaque deposits on gap junction membranes after glutaraldehyde-calcium fixation

When cloned hybrid cells (A/Bm-5) were grown to confluence and fixed in glutaraldehyde-calcium, electron-opaque deposits were observed on the cytoplasmic faces of plasma membrane. Deposits were most abundant at gap junctions. Deposits were often precisely paired, cell-to-cell, across the gap junctional membranes, and these paired deposits were frequently equivalent in size. This relationship was most often observed on long profiles of gap junctions, in contrast to the asymmetric distribution of larger deposits commonly found on short junctional profiles. Deposits were present with or without heavy metal staining but did not appear when calcium was omitted from the fixative. Fixation at room temperature yielded more and larger deposits than fixation at 0 degrees C. The significance of these observations is discussed with regard to the possible binding of calcium at fixed membrane sites or the precipitation of calcium by anions produced by enzymes located at the gap junction.