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In vivo application of upconverting force sensors to elucidate neuromuscular pump action in C. elegans

Wednesday, November 22, 2017 - 12:00
Place: 
Donostia International Physics Center
Who: 
PhD. Student Alice Lay, Stanford University, Department of materials science and engineering
Source Name: 
DIPC

The
feeding behavior of C. elegans is a strong indicator of
health; changes indicate environmental toxins, scarce or abundant food
resources, aging, and neurodegenerative disease. In particular, the pharyngeal
pump action is a rhythmic contraction and relaxation of muscles that allows the
worm to pull in and concentrate bacteria, crush and chew them in
the grinder, and then pass them through the intestinal tract [1]. Because the
pump action is regulated by motor neurons (MC, M3, M4) [2-5], it serves as a
model system for neuromuscular pumps like the heart. Here, we investigate the
magnitude of forces exerted by muscles in the pharynx, combining extracellular
electrophysiological recordings, or electrophargyngograms (EPGs), with optical
force measurements from upconverting nanoparticles (UCNPs).  Sub-25
nm Mn2+-doped NaYF4:Er,Yb UCNPs provide a photostable and
consistent color response to stress [6]. The nano- to micro-Newton sensitivity
of these nanoparticles relies on the energetic coupling between the crystal
field sensitive d-metal and upconverting lanthanides, which under
stress, yields a positive or negative change in the red to green Er3+ emission
ratio for cubic- and hexagonal-phase NaYF4, respectively.
Further, we investigate new geometries (e.g. core-shell) for more efficient and
force-sensitive nanoparticles. 

We
demonstrate the first in vivo capabilities of these
nanosensors to image and quantify forces exerted along the pharynx. First, we
incubate the worms with water-soluble UCNPs (5 mg/mL) overnight for feeding,
which yields no significant chronic cytotoxicity effects on their
fertility. Then, we load the worms in a microfluidic device and collect upconversion
spectra at key anatomical features.  Based on ratiometric differences
in emission peaks, we find that forces exerted in the grinder (~10 uN) are
nearly an order of magnitude higher than those exerted at the
pharyngeal-intestinal valve (~1 uN). Furthermore, we compare these optical
force measurements to muscle contraction and relaxation events, characterized
by voltage spikes in the EPGs. We determine pump action parameters (e.g.,
duration, frequency, amplitude) and muscular forces in wild-type and
neurotransmitter-treated (5 mM serotonin) worms. From these results,
we work towards mapping neuromuscular pump dynamics and providing the first
in-vivo determination of the forces required for healthy function in C.
elegans
.

[1] Fang-Yen, C., L. Avery, and S.
Aravinthan. PNAS (2009)

[2] Raizen, D.M. and L. Avery. Neuron (1994)

[3] Niacaris, T. and L. Avery. Journal
of Experimental Biology
 (2003)

[4] Trojanowski, N.F., D.M. Raizen,
and C. Fang-Yen. Scientific reports (2016)

[5] Lee, K.S. et al. Nature
Communications 
(2017)

[6] Lay, A. et al. Nano
Letters 
(2017)

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