M‐channels are voltage‐gated K+ channels that regulate the excitability of many neurons. They are composed of Kv7 (KCNQ) family subunits, usually Kv7.2 + Kv7.3. Native M‐channels and expressed Kv7.2 + 7.3 channels are inhibited by stimulating G_q/11‐coupled receptors – prototypically the M1 muscarinic acetylcholine receptor (M1‐mAChR). The channels require membrane phosphatidylinositol‐4,5‐bisphosphate (PIP₂) to open and the effects of mAChR stimulation result primarily from the reduction in membrane PIP₂ levels following G_q/phospholipase C‐catalysed PIP₂ hydrolysis. However, in sympathetic neurons, M‐current inhibition by bradykinin appears to be mediated through the release and action of intracellular Ca²+ by inositol‐1,4,5‐trisphosphate (IP₃), a product of PIP₂ hydrolysis, rather than by PIP₂ depletion. We have therefore compared the effects of bradykinin and oxotremorine‐M (a muscarinic agonist) on membrane PIP₂ in sympathetic neurons using a fluorescently tagged mutated C‐domain of the PIP₂ binding probe, ‘tubby’. In concentrations producing equal M‐current inhibition, bradykinin produced about one‐quarter of the reduction in PIP₂ produced by oxotremorine‐M, but equal reduction when PIP₂ synthesis was blocked with wortmannin. Likewise, wortmannin restored bradykinin‐induced M‐current inhibition when Ca²+ release was prevented with thapsigargin. Thus, inhibition by bradykinin can use product (IP₃/Ca²+)‐dependent or substrate (PIP₂) dependent mechanisms, depending on Ca²+ availability and PIP₂ synthesis rates.