Thin frozen-thawed brain sections were incubated at room temperature for 90 min in the absence of calcium (lane 2, Cont), in the presence of 2.0 mM calcium (lane 3, Ca++), and in the presence of 2.0 mM calcium and10 μM calpain inhibitor III (lane 4, Ca ++ + CI), or calpain inhibitor alone (lane 5, CI). They were then processed for western blots with anti-stargazin antibodies. Membrane fractions were prepared as described under Materials and Methods, and aliquots were treated without (lane 6) or with calpain and calcium (lane 7). Lane 1: molecular weight scale. Right arrow points to the doublet band labeled with the stargazin antibody with an apparent molecular weight of 33–35 kD.
Top: representative image of a western blot under each condition.
Bottom: quantitative analysis of western blots similar to those shown on top. Results were expressed as percent of the values found under control conditions and are means ± S.E.M. of 3 experiments. *p<0.001.