A key feature of potassium channel function is the ability to switch between conducting and non-conducting states by undergoing conformational changes in response to cellular or extracellular signals. Such switching is facilitated by the mechanical coupling of gating domain movements to pore opening and closing. Two-pore domain potassium channels (K(2P)) conduct leak or background potassium-selective currents that are mostly time- and voltage-independent. These channels play a significant role in setting the cell resting membrane potential and, therefore modulate cell responsiveness and excitability. Thus, K(2P) channels are key players in numerous physiological processes and were recently shown to also be involved in human pathologies. It is well established that K(2P) channel conductance, open probability and cell surface expression are significantly modulated by various physical and chemical stimuli. However, in understanding how such signals are translated into conformational changes that open or close the channels gate, there remain more open questions than answers. A growing line of evidence suggests that the outer pore area assumes a critical role in gating K(2P) channels, in a manner reminiscent of C-type inactivation of voltage-gated potassium channels. In some K(2P) channels, this gating mechanism is facilitated in response to external pH levels. Recently, it was suggested that K(2P) channels also possess a lower activation gate that is positively coupled to the outer pore gate. The purpose of this review is to present an up-to-date summary of research describing the conformational changes and gating events that take place at the K(2P) channel ion-conducting pathway during the channel regulation.