Although a considerable amount of circumstantial evidence exists which indicates the involvement of Ca²&43 in the regulation of phytochrome-mediated responses (Bossen et al., 1988; Viner et al., 1988; Roux, 1986), the exact nature of this involvement has remained unclear. Etiolated wheat leaf protoplasts swell in response to red light (Bossen et al., 1988) by a mechanism which is phytochrome-mediated and is related to leaf growth and unrolling (Zhou et al., 1990). In this study fluorescent Ca²+ indicators have been used in conjunction with Laser Scanning Confocal Microscopy to measure changes in [Ca²+ ]_i following red light irradiation of these protoplasts. In most cases (n = 18 of 23), an increase in [Ca²+ ]_i of approximately 400-700nM was observed, followed by a decrease to below resting level. This response preceded the physiological response of protoplast swelling, although the timing of the [Ca²+ ]_i response varied between protoplasts. Transient increases in [Ca²+ ]_i initiated by photoactivation of caged Ca²+ or caged InsP₃ mimicked the swelling response, and photoactivation of Diazo-2, a caged Ca²+ scavenger, within these protoplasts prevented the swelling response. Data is also presented here which suggests that cAMP, released from caged cAMP, can also induce an increase in protoplast volume, possibly by causing an increase in [Ca²+ ]_i. These data support the hypothesis that phytochrome signals are transduced through Ca²+ and suggest a possible role for cAMP in signal transduction within plant cells.