Transient membrane hyperpolarizations due to spontaneous fluctuations of the cytosolic Ca2+ in osteoblast-like cells.
The recently developed nystatin modification of the patch clamp technique allows stable whole-cell recordings without affecting the intracellular Ca2+ buffering capacity and thereby may provide a means to indirectly monitor spontaneous changes in the intracellular Ca2+ concentrations. To test this hypothesis, we applied the nystatin method to the well-characterized ROS 17/2.8 osteoblast-like cell system, where rises of the intracellular Ca2+ are known to cause transient hyperpolarizations via activation of Ca2+ -dependent K+ channels. Additionally to minor fluctuations (10-20 mV) around a mean potential of -42.1 +/- 4.2 mV, we observed spontaneously occurring, transient hyperpolarizations to membrane potentials as negative as -80 mV. These transient hyperpolarizations were not eliminated by Ca2+ entry blockers but abolished by intracellular infusion of 10 mM EGTA. Thapsigargin, a specific inhibitor of the endoplasmic reticulum Ca(2+)-ATPase, hyperpolarized the cells close to the K+ reversal potential. Moreover, voltage-clamp studies revealed an intermittendly activating Ca2+-dependent K+ conductance. These results strongly suggest that the nystatin method is particularly suitable to study Ca(2+)-dependent channels and thereby spontaneous changes in the intracellular Ca2+.
Scherübl
H
H
Pharmakologisches Institut der Freien Universität Berlin, Federal Republic of Germany.
Hescheler
J
J
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors; D003600:Cytosol / Q000378:metabolism*; D004594:Electrophysiology; D005110:Extracellular Space / Q000378:metabolism; D008564:Membrane Potentials; D009994:Osmolar Concentration; D010006:Osteoblasts / Q000378:metabolism* / Q000502:physiology; D011188:Potassium / Q000378:metabolism; D013729:Terpenes / Q000494:pharmacology; D019284:Thapsigargin
D013729:Terpenes; D019284:Thapsigargin; D000252:Calcium-Transporting ATPases; D011188:Potassium; D002118:Calcium