Protein kinase C alpha modulates the Ca2+ influx phase of the Ca2+ response to 1 alpha,25-dihydroxy-vitamin-D-3 in skeletal muscle cells

Treatment of chick skeletal muscle cells with 1 alpha ,25-dihydroxy-vitamin D-3 [1 alpha ,25(OH)(2)D-3] triggers a rapid and sustained increase in cyt osolic Ca2+ ([Ca2+](i)), which depends on Ca2+ mobilization from inner stor es and extracellular Ca2+ entry. Fluorimetric analysis of changes in [Ca2+] (i) in Fura-2-loaded cells revealed that the hormone significantly stimulat es the Ca2+ influx phase within the concentration range of 10(-12)-10(-6) M , with maximal effects (3.5-fold increase) at 10(-9) M 1 alpha ,25(OH)(2)D- 3. The effects of the sterol on the Ca2+ entry pathway were abolished by th e PKC inhibitors bisindolylmaleimide and calphostin. We have recently shown that, in these cells, 1 alpha ,25(OH)(2)D-3 activates and translocates PKC alpha to the membrane, suggesting that this isozyme accounts for PKC-depen dent 1 alpha ,25(OH)(2)D-3 modulation of Ca2+ entry. The role of PKC alpha was specifically addressed here using antisense technology. When the expres sion of PKC alpha was selectively knocked out by intranuclear microinjectio n of an antisense oligonucleotide against PKC alpha mRNA, the Ca2+ influx c omponent of the response to 1 alpha ,25(OH)(2)D-3 was markedly reduced (-60 %). These results demonstrate that 1 alpha ,25(OH)(2)D-3-induced activation of PKC alpha enhances extracellular Ca2+ entry partially contributing to m aintainance of the sustained phase of the Ca2+ response to the sterol.