Calcium uptake and calcium-dependent phosphorylation during development of rat brain neurons in culture.
The emergence of the capacities for calcium uptake and calcium-regulated protein phosphorylation during the development of embryonic brain neurons in tissue culture was examined. In the maturing cells, the enhancement in 45Ca2+-uptake upon stimulation with high K+ increased by 3-4 fold during the second week in vitro, in parallel to an increase in the capacity for high K+-induced Ca2+-dependent release of prelabeled [3H]dopamine. The pattern of incorporation of [32Pi]phosphate into the major phosphoproteins in maturing cells under nonstimulating conditions also changed during cell development: the incorporation of 32Pi into two proteins of apparent molecular weights--55,000 and 43,000 dalton--increased, but decreased in a 45,000 dalton protein. Stimulation of mature cells (after 10-11 days in vitro) resulted in a Ca2+-dependent increase in the amount of 32Pi incorporated into the 43,000 dalton protein and a decrease in the amount incorporated into the 55,000 dalton protein. This calcium-regulated phosphorylation pattern was not observed until 6 days in vitro. Introduction of Ca2+ into the immature cells by means of the Ca2+ ionophore A23187 did not alter the phosphorylation pattern and did not cause neurotransmitter release. The amount of [35S]methionine incorporated into a 43,000 dalton protein which comigrated with the 43,000 dalton phosphoprotein also increased upon cell maturation. The results suggest that this phosphoprotein (which does not comigrate with nonphosphorylated actin on two-dimensional polyacrylamide gels) develops in the cells in parallel to the emerging processes of the stimulation-induced calcium entry and calcium-dependent neurosecretion.
D000818:Animals; D001921:Brain / Q000378:metabolism*; D001923:Brain Chemistry; D002118:Calcium / Q000378:metabolism*; D002463:Cell Membrane Permeability; D002478:Cells, Cultured; D004298:Dopamine / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D005333:Fetus; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D018377:Neurotransmitter Agents / Q000378:metabolism; D010750:Phosphoproteins / Q000302:isolation & purification; D010766:Phosphorylation; D051381:Rats
D009419:Nerve Tissue Proteins; D018377:Neurotransmitter Agents; D010750:Phosphoproteins; D002118:Calcium; D004298:Dopamine