Optogenetic urothelial cell stimulation induces bladder contractions and pelvic nerve afferent firing

Abstract

Urothelial cells, which play an essential role in the barrier function, are also thought to play a sensory role in bladder physiology by releasing signaling molecules in response to sensory stimuli that act upon adjacent sensory neurons. However, it is challenging to study this communication due to the overlap in receptor expression and proximity of urothelial cells to sensory neurons. To overcome this challenge, we have developed a mouse model where we can directly stimulate urothelial cells using optogenetics. We have crossed a uroplakin II-cre mouse (UPK2-Cre) with a mouse that expresses the light-activated cation channel, Channelrhodopsin-2 (ChR2), in the presence of cre-expression. Optogenetic stimulation of urothelial cells cultured from UPK2-ChR2 initiates cellular depolarization and release of adenosine triphosphate. Cystometry recordings demonstrate that optical stimulation of urothelial cells increases bladder pressure and pelvic nerve activity. Increases in bladder pressure persisted, albeit to a lesser extent, when the bladder was excised in an in vitro preparation. The P2X receptor antagonist, PPADS, significantly reduced optically evoked bladder contractions in vivo and ex vivo. Further, corresponding nerve activity was also inhibited with PPADS. Our data suggest that urothelial cells can initiate robust bladder contractions via sensory nerve signaling or contractions through local signaling mechanisms. This data supports a foundation of literature demonstrating communication between sensory neurons and urothelial cells. Importantly, with further use of these optogenetic tools, we hope to scrutinize this signaling mechanism, its importance for normal micturition and nociception, and how it may be altered in pathophysiologic conditions. Animal StudiesAll the procedures involving mice and mouse tissue performed in this study were approved by the University of Florida Institutional Animal Care and Use Committee and in strict accordance with the US National Institute of Health (NIH) Guide for the Care and Use of Laboratory Animals. SignificanceIt has been appreciated for almost two decades that urothelial cells play a sensory role in bladder function. However, it has been particularly challenging to study this communication as both sensory neurons, and urothelial cells express the same sensory receptors. Here we demonstrate an optogenetic technique to specifically stimulate urothelial cells and evaluate the effects on bladder physiology. We used this technique to show that specific urothelial stimulation resulted in bladder contractions. This approach will have a long-lasting impact on how we study urothelial-to-sensory neuron communication and the changes that occur under disease conditions.