Exp Physiol 95.11 pp 1049–1060 Ca2+ events in living cells 1055 was provided for an inhibitory function. Interventions that inhibited sparks resulted in constriction of pressurized arteries, suggesting that sparks normally exert a dilatory inﬂuence (Nelson et al. 1995). It was proposed that this reﬂects negative feedback via spark activation of BK channels, resulting in outward, hyperpolarizing current (Fig. 7). Activation of voltage-operated Ca2+ channels is reduced in these circumstances, limiting Ca2+ inﬂux. Localized Ca2+ release in the form of Ca2+ sparks could, therefore, favour a reduction in global [Ca2+]i, relaxing smooth muscle. Considerable evidence supports this model in many different smooth muscles, particularly in vascular and urinary myocytes (Perez et al. 2001; Heppner et al. 2003; Burdyga & Wray, 2005; McGahon et al. 2007). This is conceptually quite different from their proposed role in cardiac muscle, where sparks are used to construct global Ca2+ signals. Evidence is now accumulating, however, to suggest that Ca2+ sparks may also play an excitatory role in some smooth muscles. We have used confocal linescan imaging to investigate Ca2+ signalling in the smooth muscle of retinal arterioles, which regulate blood ﬂow to the inner retina. These myocytes generate both brief localized Ca2+ sparks and more prolonged Ca2+ oscillations (Fig. 8). Rather than reducing global [Ca2+]i, sparks often appear to summate, generating oscillations, which sometimes result in obvious contraction of the relevant myocyte (Curtis et al. 2004). In more recent experiments using faster two-dimensional Ca2+ imaging, Ca2+ sparks again appear to initiate cell-wide Ca2+ oscillations and contraction, Figure 6. Indirect and direct evidence for Ca2+ sparks in smooth muscle A, electrophysiological recordings from voltage-clamped smooth muscle reveals spontaneous outward transient currents (STOCs) carried by K+. Activation of RyRs with caffeine to release the sarcoplasmic reticulum Ca2+ store resulted in an initial burst of STOC activity and then quiescence. Inclusion of EGTA in the recording pipette to buffer changes in [Ca2+]i inhibited STOCs. This indirect but highly persuasive evidence suggests that there must be spontaneous Ca2+ release via RyRs, resulting in the activation of Ca2+-activated BK channels. Reproduced with permission from Benham & Bolton (1986). B, confocal linescan image of an arterial smooth muscle cell loaded with the indicator ﬂuo-3. Normalized ﬂuorescence (F/F0) is used as a measure of [Ca2+]i. Two spontaneous Ca2+ sparks have been recorded. Reproduced with permission from Nelson et al. (1995). C 2010 The Author. Journal compilation C 2010 The Physiological Society Figure 7. Sparks activate Ca2+-sensitive membrane currents in smooth muscle A, activation of Ca2+-activated K+ channels by Ca2+ sparks results in spontaneous transient outward currents (STOCs), hyperpolarizing the plasma membrane (increased membrane potential; V m). This reduces activation of voltage-operated Ca2+ channels (VOCCs), decreasing Ca2+ inﬂux and global [Ca2+]. This can act as a negative feedback mechanism, whereby localized Ca2+ release from the sarcoplasmic reticulum (SR) lowers mean cytosolic [Ca2+], restricting further store loading. B, spark-dependent activation of Ca2+-activated Cl− channels causes spontaneous transient inward currents (STICs), with membrane depolarization and Ca2+ channel activation. This results in positive feedback, with Ca2+ release from the SR promoting further Ca2+ inﬂux.