Name: Video: Preparation of OP50 for Caenorhabditis elegans Culture
Uploaded: 2024-02-23T14:45:00
Duration: 1 min 40 s
Description: 605 Views. Preparation of OP50 for Caenorhabditis elegans Culture

preparation of op50 for caenorhabditis elegans culture

High-Throughput Assay to Measure Caenorhabditis elegans Food Intake and Lifespan

Caenorhabditis elegans has been extensively used in aging research. &#160;It has been a powerful model to study the genetics underlying metabolism-mediated longevity induced by dietary restriction, reduced mitochondrial activity, or reduced insulin signaling.1,2,3,4,5,6,7. However, measuring feeding in the context of longevity has been proven difficult for C. elegans due to the small size of the animal3,8,9,10,11,12,13,14,15.
Feeding is a fundamental biological behavior required for survival, and its tight regulation lies at the core of maintaining metabolic health, reproduction, and aging. Feeding behavior is controlled by multiple signaling pathways in both the nervous system and periphery and, thus, can only be studied in vivo16,17,18,19. Feeding represents the first of three pillars: (i) energy intake, (ii) energy storage, and (iii) energy expenditure that govern an organism&#39;s energy homeostasis. Feeding in C. elegans has been chiefly investigated by measuring the pharyngeal pumping, as defined by the rate of contractions of the pharynx. This approach has provided crucial insights into C. elegans&#39; feeding behavior3,4,8,12,13,20,21; however, pharyngeal pumping, especially measured over short intervals of minutes, is not necessarily correlated with food intake.
Food intake is not only determined by the pharyngeal pumping rate but also by parameters such as duration and frequency of feeding bouts and the density of food bacteria. An animal may have a very high pharyngeal pumping, but the length of its feeding bouts may be shorter, offsetting the increased rate. Another confounding factor is the efficiency by which pumping may or may not ingest and grind up bacteria. An extreme example of uncoupling food intake from pharyngeal pumping is the addition of serotonin without any food (bacteria) present. In the presence of exogenous serotonin, animals pump at a high rate, but without bacteria present, the high pumping rate does not lead to food intake2,6,13,22,23.
The bacterial clearance method presented here measures the food intake of worm populations in individual wells as the decline of bacterial concentrations over time (Figure&#160;1). This approach is similar to those used in other species where the disappearance of food represents a measure of food intake10,11,24,25,26,27,28. In the original publication, we validated this method of measuring food intake by feeding C. elegans with 15N isotope-labeled bacteria11. Subsequent measures by mass spectrometry showed that measuring the disappearance of bacteria correlated strongly with the occurrence of isotope labeling, revealing the actual uptake of the bacteria into the animal11. Thus, we are confident that the bacterial clearance assay represents food intake in most circumstances. The assay&#39;s purpose is not to replace the pharyngeal pumping assay but to add to the toolbox of assays to study feeding and metabolism in C. elegans and how it relates to aging and longevity.

Author Spotlight: Exploring the Relationship Between Lipotoxicity and HFpEF 

Quantifying Food Intake in Caenorhabditis elegans by Measuring Bacterial Clearance

Preparation of OP50 for Caenorhabditis elegans Culture

preparation-of-op50-for-caenorhabditis-elegans-culture

Research

JoVE Journal

Behavior

605 Views. Preparation of OP50 for Caenorhabditis elegans Culture

Video: Preparation of OP50 for Caenorhabditis elegans Culture

Feeding is an essential biological process for an organism&#39;s growth, reproduction, and survival. This assay aims to measure the food intake of Caenorhabditis elegans&#160;(C. elegans), an important parameter when studying the genetics of aging or metabolism. In most species, feeding is determined by measuring the difference between the amount of food provided and the amount left after a given time interval. The method presented here uses the same strategy to determine the feeding of C. elegans. It measures the amount of bacteria, the food source of C. elegans, cleared within 72 h. This method uses 96-well microtiter plates and has allowed the screening of hundreds of drugs for their ability to modulate food intake at a speed and depth not possible in other animal models. The strength of this assay is that it allows to measure feeding and lifespan simultaneously and directly measures the disappearance of food and, thus, is based on the same principles used for other organisms, facilitating species-to-species comparison.

This protocol describes an assay to quantify Caenorhabditis elegans feeding rate based on measuring the clearance of bacteria in liquid culture.