Expression of calbindin-D-28k in C6 glial cells stabilizes intracellular calcium levels and protects against apoptosis induced by calcium ionophore and amyloid beta-peptide

The calcium binding protein, calbindin-D-28k is normally present in neurons . Recently we reported that brain injury and tumor necrosis factors (TNFs) induce calbindin-D-28k in astrocytes. TNF-treated calbindin expressing astr ocytes were resistant to acidosis and calcium ionophore toxicity, suggestin g that calbindin may have a cytoprotective role in astrocytes in the injure d brain (M.P. Mattson, B. Cheng, S.A. Baldwin, V.L. Smith-Swintosky, J. Kel ler, J. Geddes, Scheff, J.W., Christakos, S., Brain injury and tumor necros is factors induce calbindin-D-28k in astrocytes: evidence for a cytoprotect ive response, J. Neurosci. Res., 42 (1995) 257). In order to obtain direct evidence for a role of calbindin, using the eukaryotic expression vector pR EP4, rat calbindin-D-28k was stably expressed in C6 rat astocytoma glial ce lls. Cytotoxicity in response to calcium ionophore or amyloid beta-peptide (which accumulates in the brain in Alzheimer's disease and has been reporte d to be neurotoxic) was measured by MTT reduction in vector transfected cel ls and in calbindin transfected clones. Stably expressed calbindin resulted in increased cell survival in the presence of calcium ionophore (1-10 mu M ) or amyloid beta-peptide (10-100 mu M) In addition, the calcium ionophore or amyloid beta-peptide mediated rise in intracellular calcium in vector tr ansfected cells was significantly attenuated in calbindin expressing cells. Apoptotic cell death was detected by the Hoechst method in vector transfec ted C6 glial cells treated with calcium ionophore or beta-amyloid (34-36% a poptotic cells/culture). The number of apoptotic nuclei was significantly a ttenuated in similarly treated calbindin-D-28k transfected clones (10-13% a poptotic cells/culture; p < 0.01). Our results support the involvement of c alcium fluxes in apoptosis and suggest that calbindin-D-28k, by buffering c alcium, can suppress death in apoptosis susceptible cells in the central ne rvous system. (C) 1999 Elsevier Science B.V. All rights reserved.