Characterization of ryanodine-sensitive Ca2+ release from microsomal vesicles of rat parotid acinar cells: regulation by cyclic ADP-ribose.
We have measured ryanodine (caffeine)-sensitive 45Ca2+ release from isolated microsomal vesicles of endoplasmic reticulum prepared from rat parotid acinar cells. After a steady state of ATP-dependent 45Ca2+ uptake, the addition of caffeine (40 mm), ryanodine (10 approximately 500 microm) or an NAD+ metabolite, cyclic ADP-ribose (cADPR, 4 microm) released about 10% of the 45Ca2+ that had been taken up. The 45Ca2+ release was not inhibited by heparin, an antagonist of IP3 receptor. The effects of caffeine, ryanodine and cADPR on 45Ca2+ release were also tested in the presence of thapsigargin (TG), an inhibitor of microsomal Ca2+-ATPase. When caffeine (10 approximately 40 mm), ryanodine (10 microm) or cADPR (1 approximately 10 microm) was added in the medium with 100 nm TG, a significant 45Ca2+ release was seen, while higher concentrations of ryanodine (>100 microm) did not cause any 45Ca2+ release in the presence of TG. The initial rate of caffeine (40 mm)-induced 45Ca2+ release was increased by a pretreatment with 10 microm ryanodine, whereas the caffeine-induced 45Ca2+ release was strongly inhibited by the presence of a higher concentration (500 microm) of ryanodine. cADPR-induced 45Ca2+ release was also inhibited by 500 microm ryanodine. Caffeine (40 mm)- or cADPR (4 microm)-induced 45Ca2+ release was abolished by a presence of ruthenium red (50 approximately 100 microm). The presence of a low concentration (0.5 microm) of cADPR shifted the dose-response curve of caffeine-induced 45Ca2+ release to the left. These results indicate the presence of a ryanodine sensitive Ca2+ release mechanism in the endoplasmic reticulum of rat parotid acinar cells that is distinct from the IP3-sensitive Ca2+ channel and is activated by caffeine, cADPR and a low concentration (10 microm) of ryanodine, but is inhibited by higher concentrations (>100 microm) of ryanodine and ruthenium red. The properties of the ryanodine-sensitive mechanism are similar to that of the ryanodine receptor as described in muscle cells.
Ozawa
T
T
Department of Physiology, Tohoku University School of Medicine, Seiryocho 2, 980-77, Sendai, Japan.
Nishiyama
A
A
D000246:Adenosine Diphosphate Ribose / Q000031:analogs & derivatives* / Q000502:physiology; D000818:Animals; D002110:Caffeine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000187:drug effects / Q000378:metabolism*; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors; D002478:Cells, Cultured; D036563:Cyclic ADP-Ribose; D004305:Dose-Response Relationship, Drug; D004721:Endoplasmic Reticulum / Q000187:drug effects / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006493:Heparin / Q000494:pharmacology; D053496:Inositol 1,4,5-Trisphosphate Receptors; D008297:Male; D008861:Microsomes / Q000187:drug effects / Q000378:metabolism*; D009124:Muscle Proteins / Q000187:drug effects / Q000378:metabolism*; D010306:Parotid Gland / Q000166:cytology / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar; D018160:Receptors, Cytoplasmic and Nuclear / Q000037:antagonists & inhibitors; D012430:Ruthenium Red / Q000494:pharmacology; D012433:Ryanodine / Q000494:pharmacology; D019837:Ryanodine Receptor Calcium Release Channel; D019284:Thapsigargin / Q000494:pharmacology
D015220:Calcium Channels; D004791:Enzyme Inhibitors; D053496:Inositol 1,4,5-Trisphosphate Receptors; D009124:Muscle Proteins; D018160:Receptors, Cytoplasmic and Nuclear; D019837:Ryanodine Receptor Calcium Release Channel; D012430:Ruthenium Red; D036563:Cyclic ADP-Ribose; D012433:Ryanodine; D000246:Adenosine Diphosphate Ribose; D002110:Caffeine; D019284:Thapsigargin; D006493:Heparin; D000252:Calcium-Transporting ATPases; D002118:Calcium