Use of optical tweezers to probe epithelial mechanosensation

Cellular mechanosensation mechanisms have been implicated in a variety of disease states. Specifically in renal tubules, the primary cilium and associated mechanosensitive ion channels are hypothesized to play a role in water and salt homeostasis, with relevant disease states including polycystic kidney disease and hypertension. Previous experiments investigating ciliary-mediated cellular mechanosensation have used either fluid flow chambers or micropipetting to elicit a biological response. The interpretation of these experiments in terms of the "ciliary hypothesis" has been difficult due the spatially distributed nature of the mechanical disturbance-several competing hypotheses regarding possible roles of primary cilium, glycocalyx, microvilli, cell junctions, and actin cytoskeleton exist. I report initial data using optical tweezers to manipulate individual primary cilia in an attempt to elicit a mechanotransduction response-specifically, the release of intracellular calcium. The advantage of using laser tweezers over previous work is that the applied disturbance is highly localized. I find that stimulation of a primary cilium elicits a response, while stimulation of the apical surface membrane does not. These results lend support to the hypothesis that the primary cilium mediates transduction of mechanical strain into a biochemical response in renal epithelia.