Effects of protein phosphorylation on the regulation of capacitative calcium influx in Xenopus oocytes.
The regulation of capacitative Ca2+ influx in Xenopus oocytes was investigated using both the two electrode voltage-clamp (where Ca2+ is monitored through the Ca2+-dependent Cl- current) and patch-clamp techniques. Following stimulation of expressed 5-hydroxytryptamine (5-HT) receptors, capacitative Ca2+ influx deactivated in around 15 min. Following injection of [adenosine 5'-O-(3-Thiotriphosphate)] (ATP [gamma-S]), an ATP analogue that is readily used by protein kinases, capacitative Ca2+ influx activated by 5-HT application either did not deactivate or was prolonged around twofold. However, injection of adenylyl 5'-(beta,gamma-methylene)-diphosphonate (AMP-PCP), another ATP analogue that is not utilised by kinases, did not affect the time-course of Ca2+ influx. When capacitative Ca2+ influx was activated by readmission of Ca2+ to oocytes incubated in thapsigargin/0 Ca2+ solution for several hours, Ca2+ influx occurred and a weakly saturating relationship between external Ca2+ and Ca2+ influx was found. Ca2+ influx in thapsigargin-treated cells was unaffected by ATP [gamma-S]. ATP [gamma-s] and several kinases had no effect on the Ca2+-dependent Cl- current when the latter was activated by elevation of Ca2+ independent of capacitative Ca2+ influx. Protein kinase C slowly and partially inhibited the Cl- current. Outside-out patches taken from thapsigargin-treated cells failed to demonstrated any Ca2+ current or Ca2+-dependent Cl- current on reapplying high Ca2+ to the patch, despite the oocyte showing a large capacitative Ca2+ influx. The results suggest that a kinase, activated on receptor stimulation, prolongs the activation time-course of capacitative Ca2+ influx.
Parekh
A B
AB
Max-Planck-Institut für biophysikalische Chemie, Am Fassberg, D-37077 Göttingen, Germany.
Terlau
H
H
D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002712:Chlorides / Q000502:physiology; D004553:Electric Conductivity; D005260:Female; D006493:Heparin / Q000494:pharmacology; D015759:Ionomycin / Q000494:pharmacology; D009865:Oocytes / Q000187:drug effects / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D010750:Phosphoproteins / Q000096:biosynthesis*; D011985:Receptors, Serotonin / Q000502:physiology; D013997:Time Factors; D014982:Xenopus laevis
D002121:Calcium Channel Blockers; D002712:Chlorides; D010750:Phosphoproteins; D011985:Receptors, Serotonin; C005147:5'-adenylyl (beta,gamma-methylene)diphosphonate; C022571:adenosine 5'-O-(3-thiotriphosphate); D015759:Ionomycin; D000255:Adenosine Triphosphate; D006493:Heparin; C002630:alpha,beta-methyleneadenosine 5'-triphosphate; D002118:Calcium