An increase in intracellular concentration of Ca²⁺ ([Ca²⁺]_i) initiates contraction in all types of smooth muscle cells including uterine myocytes. On a tissue level, temporal and spatial summation of [Ca²⁺]_i transients in individual cells triggers a contraction pattern specific to a particular type of smooth muscle. The mechanisms responsible for the initiation and spread of tissue-level Ca²⁺ signals differ in different smooth muscles, involving in some cases, a specialised type of cell called interstitial cells of Cajal (ICCs). The transition of myometrium from relative quiescence throughout pregnancy to powerful contractions during labour is a very complex and precisely timed process. An increase in intercellular coupling through gap-junctions and paracrine mechanisms is thought to be important for synchronising myocyte activity at the end of pregnancy, yet the mechanism of myometrial autorhythmicity remains unclear. Recent work has established that ICC-like cells are present in both rodent and human myometrium, although they are unlikely to be the pacemakers, at least in the rat [1]. In the present study, we used laser scanning confocal microscopy to investigate the initiation and propagation of Ca²⁺ signals between uterine myocytes in their natural environment (i.e. within thin slices of intact myometrium). Myometrial biopsies were taken from the patients undergoing Caesarean section. Samples from three different patients were used in this study. Ten to fifteen slices of myometrium (200-300 μm thick) were cut using a vibroslicer (Integraslice 7550 PSDS, Campden Instruments, UK) in ice-cold physiological saline. The Ca²⁺ signalling events were recorded from Fluo-4 loaded slices using a Zeiss LSM 510 META confocal microscope. Confocal imaging of tissue slices revealed bundles of smooth muscle cells separated by interstitial spaces. We observed synchronous and large (3.25 ± 0.35 F/F₀, n=32) rises in [Ca²⁺]_i elicited by action potentials in bundles of smooth muscle cells within the field of view (10x and 20x objective lenses). This was followed by non-propagating asynchronous Ca²⁺ transients of smaller amplitude (1.05-1.75 F/F₀, n=102), presumably due to spontaneous Ca²⁺ release from the sarcoplasmic reticulum. Immediately preceding high-amplitude [Ca²⁺]_i transients in the smooth muscle cell bundle, there were spikes of [Ca²⁺]_i originating in the interstitial space outside the bundle, and propagating towards the muscle bundle. Immunohistochemistry of fixed myometrial slices revealed vimentin-positive ICC-like cells within the interstitium and surrounding the smooth muscle cell bundles. In conclusion, our data suggest that in human myometrium, vimentin positive ICC-like cells may play a role as a pacemaker.