Primary cultures of cerebellar rat granule neurons were grown for 18-22 days in vitro in the absence of antibiotics. When the cultures were placed in a low calcium (no EGTA) balanced salt solution at room temperature, rapid cell death occurred usually within 30 min of placing cells in the buffer. Changes in the cells were evident within 10 min and included an apparent cellular granulation with a partial loss of cell body birefringence at 10 x magnification which was complete by 30 min. This rapid death was prevented by (1) replacing chloride in the buffer with acetate; (2) increasing the osmolarity of the buffer by 30% with sucrose; (3) the addition of the selective excitatory amino acid (EAA) antagonist, 2-amino-7-phosphonoheptanoic acid (APH, 200 microM) but not by the selective kainate-quisqualate antagonist, glutamylaminomethylsulfonic acid (GAMS, 400 microM); or (4) the addition of one of the following calcium channel antagonists, verapamil (400 microM) diltiazem (150 microM) or lanthanum (5 microM). Placing cells in low calcium buffer resulted in a 3.7- and 3.2-fold increase in the non-selective secretion of aspartate and glutamate (as well as other amino acids) over baseline secretion (same buffer except containing 2.5 mM calcium). This increase was partially prevented by verapamil, but not by APH or chloride deletion. Verapamil only partially prevented the efflux of glutamate in buffer containing 1 mM EGTA. These results indicate that placing cells in low calcium buffer results in neurotoxicity secondary to both the influx of chloride and water in conjunction with the efflux of amino acids, some of which stimulate an excitatory amino acid receptor.(ABSTRACT TRUNCATED AT 250 WORDS)