Mechanical forces regulate many important biological processes from stem cell differentiation to digestion. Deciphering the enigmatic language of mechanical signaling could help researchers uncover novel biomarkers of disease that go undetected with traditional chemical, optical and electrical sensing modalities. However, existing mechanosensing tools are either too large and invasive to operate in vivo (atomic force microscopy, traction force microscopy) or are limited in their dynamic range and propensity to rapidly photobleach (FRET tethers, which operate below 100 pN). Mechanosensitive upconverting nanoparticles (UCNP) are a promising new class of optical mechanosensors to study micronewton scale biological forces because they are small (<20 nm), relatively non-toxic, photostable and exhibit a strong anti-Stokes shift that allow them to be excited in the near-IR biological window (980 nm). Our group was the first to utilize high pressure diamond anvil cell experiments to demonstrate the ratiometric sensitivity of Erbium red (⁴S_3/2 → ⁴F_9/2) and green (⁴F_9/2 → ⁴I_9/2) emission lines within core@shell cubic phase NaY_0.8Yb_0.18Er_0.02F₄@NaYF₄ subjected to gigapascals of hydrostatic pressure. Using a dual confocal-atomic force microscope (confocal-AFM), we demonstrate here that UCNPs embedded into polystyrene microbeads (microgauges) exhibit a micronewton colorimetric sensitivity to applied compressive force that is linear and robust across multiple compression-relaxation cycles (0.5% change in I_Red:I_Green per µN). Microgauges are fabricated via an emulsion polymerization and sized to mimic the natural bacterial food source of the C. elegans worm; thus, they accumulate readily in the pharyngeal and intestinal lumen upon passive feeding, without adversely affecting worm fecundity or pharyngeal pumping rate. This allows for rapid imaging (>40 Hz) of pharyngeal muscle contraction induced force dynamics in gently immobilized, actively feeding worms. The resulting ratiometric changes induced during the contraction-relaxation cycle are further correlated with the muscle membrane depolarizations that induce them via a simultaneous electropharyngeogram. As C. elegans pharyngeal pumping is an analog for human cardiac pumping functions, we envision this biosensor could enable the first direct assay of force generation for screening potentially lifesaving antihypertensive drugs.

Chris Siefe

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