Calcium/calmodulin-dependent protein kinase activity in primary astrocyte cultures

Calcium, calmodulin-dependent protein kinase (Ca/CaM kinase) is an important component of calcium signalling mechanisms in the brain, but little is known about the properties of this protein phosphorylation system in astrocytes. Addition of calcium and calmodulin to supernatant or membrane fractions obtained from rat astrocytes in primary culture increased phosphate incorporation into an exogenously added substrate, casein, and into endogenous protein substrates; this increase was greater than that observed with either calcium alone or calmodulin alone. The calcium, calmodulin-stimulated increase was inhibited by trifluoperazine, and this inhibition could be overcome by the addition of excess calmodulin. The major substrates for Ca/CaM kinase activity were proteins with molecular weights of 59 and 53 kDa, which were similar, but not identical, to the subunits of Ca/CaM kinase type II from brain. The specific activity of Ca/CaM kinase and the phosphorylation of 59 kDa were increased in astrocyte cultures treated and maintained in dibutyryl cyclic adenosine monophosphate (dBcAMP). These results indicate that astrocytes contain Ca/CaM kinase activity and suggest an interaction between the cAMP and calcium/calmodulin messenger systems in these cells.