Shifts in the Distribution of Mass Densities Is a Signature of Caloric Restriction in Caenorhabditis elegans

Abstract

Although the starvation response of the model multicellular organism Caenorhabditis elegans is a subject of much research, there is no convenient phenotypic readout of caloric restriction that can be applicable to large numbers of worms. This paper describes the distribution of mass densities of populations of C. elegans, from larval stages up to day one of adulthood, using isopycnic centrifugation, and finds that density is a convenient, if complex, phenotypic readout in C. elegans. The density of worms in synchronized populations of wildtype N2 C. elegans grown under standard solid-phase culture conditions was normally distributed, with distributions peaked sharply at a mean of 1.091 g/cm3 for L1, L2 and L3 larvae, 1.087 g/cm3 for L4 larvae, 1.081 g/cm3 for newly molted adults, and 1.074 g/cm3 at 24 hours of adulthood. The density of adult worms under starvation stress fell well outside this range, falling to a mean value of 1.054 g/cm3 after eight hours of starvation. This decrease in density correlated with the consumption of stored glycogen in the food-deprived worms. The density of the worms increased when deprived of food for longer durations, corresponding to a shift in the response of the worms: worms sacrifice their bodies by retaining larvae, which consume the adults from within. Density-based screens with the drug Ivermectin on worms cultured on single plates resulted in a clear bimodal (double-peaked) distribution of densities corresponding to drug exposed and non-exposed worms. Thus, measurements of changes in density could be used to conduct screens on the effects of drugs on several populations of worms cultured on single plates.

Separation, Sizing, and Quantitation of Engineered Nanoparticles in an Organism Model Using Inductively Coupled Plasma Mass Spectrometry and Image Analysis

“…Similar to the first two experiments, free AuNPs were predominately retained in the sample (gradient layers 11 to 13) and salt layers (gradient layers 9 to 11) of the gradient (Figure 3), while the nematodes moved down the gradient until they encountered the level of the sucrose density gradient that inhibited further movement (the average density of the nematodes closely matched the density of the 20 % (w/v) sucrose layer in the sucrose gradient). 38 …”

“…35–37 During centrifugation, high-density ENPs sediment at a faster rate than lower-density ENPs, thus promoting localized separation of the ENPs within different density layers of the gradient. Recently, researchers in the Whitesides group 38 utilized isopycnic density gradient centrifugation with Percoll™ centrifugation medium to quantitatively measure mass density distributions in C. elegans populations. In addition, conventional sucrose density gradient centrifugation protocols are commonly used for separating viable C. elegans from bacteria, dead nematodes, and debris.…”

High-throughput, motility-based sorter for microswimmers such as C. elegans

“…The collection chamber is located upstream of the holding chamber beneath the conduit’s level. Since C. elegans is denser than the carrier water 38 , when the animal arrives at the collection chamber, it sinks to the chamber’s bottom, unable to escape. Although not attempted here, one can envision expanding the L sorter to include a cascade of collection chambers, each doubling as a holding chamber with a judiciously designed separation conduit to sort animals with various motilities.…”

“…Our preliminary observations (see supplement) suggest that such a separation method works, allowing for reduced cost and even greater economy of scale. Such a sorter assumes that the animals have a narrow density (mass per unit volume) distribution as indicated by a recent study 38 .…”

Sucrose density gradient centrifugation for efficient separation of engineered nanoparticles from a model organism, Caenorhabditis elegans

“…[22] Recently, a Percoll medium centrifugation separation allowed the measurement of mass density distributions in C. elegans populations. [23] In fact, sucrose density gradients are widely used in C. elegans maintenance for separating viable nematodes from dead nematodes, eggs, bacteria, and other debris. [3,24,25] However, sucrose density gradient centrifugation (SDGC) separation had not been employed until recently for the separation of C. elegans from NPs after exposure.…”