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Caffeine was used to study the intracellular Ca²⁺ pools of bovine chromaffin cells. Its effects on cytosolic Ca²⁺ concentration ([Ca²⁺]_i) were examined using fura‐2. Caffeine caused a transient increase in [Ca²⁺]_i in the presence or absence of extracellular Ca²⁺. In the former case, the caffeineinduced [Ca²⁺]_i increase was higher and stayed above the basal value for several minutes. In the latter case, the [Ca²⁺]_i rise was lower and fell to the basal level within 1 min. These results suggest that caffeine increases [Ca²⁺]_i by causing both Ca²⁺ influx and Ca²⁺ release from intracellular pools. In the absence of extracellular Ca²⁺, ionomycin but not caffeine caused a further increase in [Ca²⁺]_i in cells that had been treated with caffeine. Apparently there are at least two intracellular Ca²⁺ pools, only one of which is sensitive to caffeine. The caffeine‐induced [Ca²⁺]_i rise became smaller when the cells were pretreated with …