Figure 3.

Resting [Ca²⁺]_c (A) and ATP-induced effects on [Ca²⁺]_c (B and C), in CHO cells transiently expressing cytAEQ alone (control), or together with PrP^c, or Dpl. (A) Resting [Ca²⁺]_c, monitored by the Ca²⁺-indicator fura-2, is expressed as the ratio of the fluorescence emitted by fura-2 after cell excitation at 340 and 380 nm (see Materials and Methods). No statistically significant difference was found in the three cell types. (B) [Ca²⁺]_c transients, after ATP (100 μM) addition at the indicated time point, were monitored with cytAEQ reconstituted with wild-type coelenterazine (5 μM; see Materials and Methods). Although a similar peak was found in controls (light gray trace) and Dpl-containing cells (dark gray trace), cells with PrP^c (black trace) gave rise to a transient of smaller magnitude. (C) Using the above described cytAEQ, a protocol was applied so as to evaluate the contribution to the ATP-induced [Ca²⁺]_c movements shown in B, first of the InsP₃-induced Ca²⁺ mobilization (first peak, ATP added in the presence of 100 μM EGTA, i.e., with no external Ca²⁺) and then of the Ca²⁺ influx from the external medium (second peak, in the presence of 1 mM CaCl₂). On Ca²⁺ release from the ER (first peak), a transient of smaller magnitude was again observed in cells with PrP^c, whereas no difference in the second peak was evident in the three cell types. Presented data are typical of at least seven independent experiments, which yielded equivalent results.