The mechanisms that link visceral mechanosensation to the perception of internal organ status (i.e., interoception) remain elusive. In response to bladder filling, the urothelium releases ATP, which is hypothesized to stimulate voiding function by communicating the degree of bladder fullness to subjacent tissues, including afferent nerve fibers. To determine if PIEZO channels function as mechanosensors in these events, we generated conditional urothelial Piezo1 -, Piezo2 -, and dual Piezo1/2 -knockout (KO) mice. While functional PIEZO1 channels were expressed in all urothelial cell layers, Piezo1 -KO mice had a limited phenotype. Piezo2 expression was limited to a small subset of superficial umbrella cells, yet male Piezo2 -KO mice exhibited incontinence (i.e., leakage) when their voiding behavior was monitored during their active dark phase. Dual Piezo1/2 -KO mice had the most affected phenotype, characterized by decreased urothelial responses to mechanical stimulation, diminished ATP release, bladder hypoactivity in anesthetized Piezo1/2 -KO females but not males, and urinary incontinence in both male and female Piezo1/2 -KO mice during their dark phase but not inactive light one. Our studies reveal that the urothelium functions in a sex- and circadian rhythm–dependent manner to link urothelial PIEZO1/2 channel–driven mechanotransduction to normal voiding function and behavior, and in the absence of these signals, bladder dysfunction ensues.

Ca²⁺-activated BK channels in renal intercalated cells (ICs) mediate luminal flow–induced K⁺ secretion (FIKS), but how ICs sense increased flow remains uncertain. We examined whether PIEZO1, a mechanosensitive Ca²⁺-permeable channel expressed in the basolateral membranes of ICs, is required for FIKS. In isolated cortical collecting ducts (CCDs), the mechanosensitive cation-selective channel inhibitor GsMTx4 dampened flow-induced increases in intracellular Ca²⁺ concentration ([Ca²⁺]_i), whereas the PIEZO1 activator Yoda1 increased [Ca²⁺]_i and BK channel activity. CCDs from mice fed a high-K⁺ (HK) diet exhibited a greater Yoda1-dependent increase in [Ca²⁺]_i than CCDs from mice fed a control K⁺ diet. ICs in CCDs isolated from mice with a targeted gene deletion of Piezo1 in ICs (IC-Piezo1-KO) exhibited a blunted [Ca²⁺]_i response to Yoda1 or increased flow, with an associated loss of FIKS in CCDs. Male IC-Piezo1-KO mice selectively exhibited an increased blood [K⁺] in response to an oral K⁺ bolus and blunted urinary K⁺ excretion following a volume challenge. Whole-cell expression of BKα subunit was reduced in ICs of IC-Piezo1-KO mice fed an HK diet. We conclude that PIEZO1 mediates flow-induced basolateral Ca²⁺ entry into ICs, is upregulated in the CCD in response to an HK diet, and is necessary for FIKS.

Sensitization of primary afferents is essential for the development of pain, but the molecular events involved in this process and its reversal are poorly defined. Recent studies revealed that acid-sensing ion channels (ASICs) control the excitability of nociceptors in the urinary bladder. Using genetic and pharmacological tools we show that ASICs are functionally coupled with voltage-gated Ca²⁺ channels to mediate Ca²⁺ transients evoked by acidification in sensory neurons. Genetic deletion of Asic3 of these sensory neurons does not alter the mechanical response of bladder afferents to distension in naïve mice. Both control and sensory neuron conditional Asic3 knockout (Asic3-KO) mice with chemical cystitis induced by cyclophosphamide (CYP) administration exhibit frequent low volume voiding events. However, these changes are transient and revert over time. Of major significance, in Asic3-KO mice, CYP treatment results in the sensitization of a subset of bladder afferents and pelvic allodynia that persist beyond the resolution of the inflammatory process. Thus, ASICs function is necessary to prevent long-term sensitization of visceral nociceptors.

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