Membrane modifications in chick osteoclasts revealed by freeze‐fracture replicas

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Abstract
Remarkable differences among various membranes of bone cells became evident by examination of freeze-fracture replicas. In osteoclasts, three types of intramembranous particles (IMPs) were identified based on their size and shape: two sizes of isolated globular particles (8 and 12 nm in diameter) and rod-shaped, linear aggregates (8 × 30 nm in dimension). Furthermore, the density and distribution pattern of these IMPs enabled us to distinguish three different domains of membranes of osteoclasts including reffled border, clear zone, and basolateral regions, as were also observed in thin sections. The highest density of IMPs was 3,500-4,000/μm2 in the ruffled border membrane, and these IMPs included linear aggregates among the usual globular particles. Linear aggregated particles were also observed in the membrane of cytoplasmic vesicles in the vicinity of the ruffled border region, but not in this membrane in other bone cells. In attached osteoclasts, the distribution pattersn and densities of IMPs in each ruffled-finger and -plate were extremely variable, from closely to the loosely packed membrane particles. Focal aggregates of membrane particles were also frequently encountered. An important outcome of the present study was the finding that the presence of linear aggregated particles proved to be an additional criterion for distinguishing membrane domains in freeze-replicas of osteoclasts. The surface of the clear zone membrane was not smooth in profile, but revealed a number of eminences that were almost free of particles. Basolateral membranes exhibited a particle density of 2,400/μm2. Globular particles were homogeneously scattered in random fashion on their exposed fracture faces. In some cases, aggregates of IMPs on the basolateral membranes were encountered. In comparison with the ruffled fingers, microprojections from the basolateral surface showed a lesser density of IMPs and were devoid of rod-shaped or linear aggregated particles. Differences between osteoblasts and osteocytes were apparent in the density and the size of IMPs. The membranes of osteoblasts and osteocytes contained the same types of globular particles as seen in osteoclasts. Various sizes of gap junctions were located only on basolateral membranes of the osteoblasts. In contrast, no cellular junctions were observed between osteoclasts and any other type of cells.