Permeability of reconstituted sarcoplasmic reticulum vesicles. Reconstitution of the K+, Na+ channel.
Permeability properties of reconstituted rabbit skeletal muscle sarcoplasmic reticulum vesicles were characterized by measuring efflux rates of [3H]inulin, [3H]choline+, 86Rb+, and 22Na+, as well as membrane potential changes using the voltage-sensitive probe, 3,3'-dipentyl-2,2'-oxacarbocyanine. Native vesicles were dissociated with deoxycholate and were reconstituted by dialysis. Energized Ca2+ accumulation was partially restored. About 1/2 of the reconstituted vesicles were found to be 'leaky', i.e., permeable to choline+ of Tris+ but not to inulin. The remaining reconstituted vesicles were 'sealed', i.e., impermeable to choline+, Tris+ and inulin. Sealed reconstituted vesicles could be further subdivided according to their K+, Na+ permeability. About 1/2, previously designated Type I, were readily permeable to K+ and Na+, indicating the presence of the K+, Na+ channel of sarcoplasmic reticulum. The remaining sealed vesicles (Type II) formed a permeability barrier to K+ and Na+, suggesting that they lacked the K+, Na+ channel. These studies show that the K+, Na+ channel of sarcoplasmic reticulum can be solubilized with detergent and reconstituted with retention of activity. Furthermore, our results suggest that part or all of the decreased Ca2+-loading efficiency of reconstituted vesicles may be due to the presence of a significant fraction of leaky vesicles.
D000818:Animals; D002118:Calcium / Q000378:metabolism; D002794:Choline / Q000378:metabolism; D003840:Deoxycholic Acid; D007444:Inulin / Q000378:metabolism; D007473:Ion Channels / Q000378:metabolism*; D008564:Membrane Potentials; D010539:Permeability; D011188:Potassium / Q000378:metabolism*; D011817:Rabbits; D012413:Rubidium / Q000378:metabolism; D012519:Sarcoplasmic Reticulum / Q000378:metabolism*; D012964:Sodium / Q000378:metabolism*
D007473:Ion Channels; D003840:Deoxycholic Acid; D007444:Inulin; D012964:Sodium; D012413:Rubidium; D002794:Choline; D011188:Potassium; D002118:Calcium