Effect of enzymic (collagenase) harvesting on the intracellular Na+/K+ ratio of Swiss/3T3 cells as revealed by X-ray microanalysis.
Swiss/3T3 cell cultures were harvested with 0.05% collagenase and after centrifugation the pellet was prepared by the freeze-fracture/freeze-drying (FFFD) method for bulk-specimen X-ray microanalysis. Time-dependent variations in the intracellular monovalent elemental concentrations (Na+, K+ and Cl-) as well as of the Na+/K+ ratio were followed for 120 min subsequent to harvesting. The quantitative measurements revealed a very considerable increase in the intracellular Na+ and Cl- accompanied by a decrease in the K+ concentration as soon as 5 min after harvesting. The Na+/K+ ratio had increased by this time to about 1.5 on average. These changes indicate a sustained depolarization of the cell membrane. During the first 60 min this depolarization tended to normalize as demonstrated by an exponential decrease in the intracellular Na+ and Cl- and an increase in the K+ content involving a decrease in the Na+/K+ ratio. The total intracellular monovalent ion concentration remained almost constant during this post-harvesting period. These results suggest that harvesting represents a serious depolarizing stimulus to the cells, the consequences of which are restored only after 1-2h. These alterations should be taken into consideration during various experimental designs when using anchorage-dependent cell cultures.
Szállási
Z
Z
Verzár International Laboratory for Experimental Gerontology, Debrecen, Hungary.
Szállási
A
A
Boján
F
F
Zs-Nagy
I
I
D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000502:physiology*; D003584:Cytological Techniques; D004577:Electron Probe Microanalysis; D005347:Fibroblasts / Q000502:physiology; D051379:Mice; D008815:Mice, Inbred Strains; D003012:Microbial Collagenase / Q000378:metabolism; D008855:Microscopy, Electron, Scanning; D011188:Potassium / Q000378:metabolism*; D012964:Sodium / Q000378:metabolism*; D013997:Time Factors
D012964:Sodium; D003012:Microbial Collagenase; D011188:Potassium