To characterize further the Ca²⁺ signalling mechanisms of myenteric neurones, we studied the effect of thapsigargin, a blocker of the Ca²⁺‐store ATPase, and the mechanisms involved in restoring the intracellular Ca²⁺ concentration ([Ca²⁺]_i) after activation. Thapsigargin (5 × 10⁻⁶ mol L⁻¹) induced an oscillatory [Ca²⁺]_i response in 86.6% of the neurones (n=276), which was blocked by the removal of extracellular Ca²⁺ and by ω‐conotoxin MVIIA (5 × 10⁻⁷ mol L⁻¹). The IP₃‐blocker, 2‐aminoethyl‐diphenyl‐borate (75 × 10⁻⁶ mol L⁻¹), blocked or reduced the responses in 74.5% of the neurones. The oscillatory responses induced by the depletion of Ca²⁺ stores suggest that myenteric neurones might recruite N‐type Ca²⁺ channels as a refill mechanism. Thapsigargin pretreatment increased the amplitude, the upstroke and duration of the K⁺‐induced [Ca²⁺]_i responses. Mitochondrial blockers (rotenone and antimycin/oligomycin) also prolonged the responses, but without affecting the amplitude. Furthermore, it was found that for high [Ca²⁺]_i, the thapsigargin‐sensitive Ca²⁺ uptake was crucial, while mitochondrial blockade affected the Ca²⁺ uptake over a wide range of concentrations. The Ca²⁺‐sequestering components might also have been compensating for each other, as most drugs only delayed and not inhibited Ca²⁺ removal.