Oscillating calcium signals in smooth muscle cells underlie the persistent basal tone of internal anal sphincter

A persistent basal tone in the internal anal sphincter (IAS) is essential for keeping the anal canal closed and fecal continence; its inhibition via the rectoanal inhibitory reflex (RAIR) is required for successful defecation. However, cellular signals underlying the IAS basal tone remain enigmatic. Here we report the origin and molecular mechanisms of calcium signals that control the IAS basal tone, using a combination approach including a novel IAS slice preparation that retains cell arrangement and architecture as in vivo, 2-photon imaging, and cell-specific gene-modified mice. We found that IAS smooth muscle cells generate two forms of contractions (i.e., phasic and sustained contraction) and Ca²⁺ signals (i.e., synchronized Ca²⁺ oscillations [SCaOs] and asynchronized Ca²⁺ oscillations [ACaOs]) that last for hours. RyRs, TMEM16A, L-type Ca²⁺ channels, and gap junctions are required for SCaOs, which account for phasic contraction and 75% of sustained contraction. Nevertheless, only RyRs are required for ACaOs, which contribute 25% of sustained contraction. Nitric oxide, the primary neurotransmitter mediating the RAIR, blocks both types of Ca²⁺ signals, leading to IAS's full relaxation. Our results show that the oscillating nature of Ca²⁺ signals generates and maintains the basal tone without causing cytotoxicity to IAS. Our study provides insight into fecal continence and normal defecation.