Biology subjects

Raizen, D. M.

Publications and source records attributed to Raizen, D. M..

8 recordsLinked to original sources

bioRxiv2026

An automated genetic screen identifies modulators of stress-induced sleep in Caenorhabditis elegans

Animals sleep more when they are sick. In C. elegans, stress-induced sleep (SIS) follows cellular injury such as exposure to ultraviolet (UV) light. The genetic regulators of SIS remain incompletely defined. Using a worm picking robot, multi-well WorMotel imaging, and association analysis we performed a semi-automated screen of 941 whole genome sequenced Million Mutation Project (MMP) strains. We quantified behavioral activity and quiescence before and after ultraviolet (UV) radiation. We applied the Sequence Kernel Association Test (SKAT) to this behavioral data to prioritize 6,663 genes and observed significant enrichment of known SIS genetic regulators. Based on these results, we conducted a candidate validation screen for additional genes regulating SIS. We identified three genes (strd-1, egl-8, cla-1), mutations in which reproducibly influence SIS. Further exploration of these genes holds potential for enhancing our understanding of the molecular basis of SIS. These findings establish a pipeline for automated behavioral phenotyping coupled with gene-based association to accelerate studies of C. elegans neurogenetics.

genetics↗

bioRxiv2023

Structural neuroplasticity after sleep loss modifies behavior and requires neurexin and neuroligin

Sleep is conserved across species, and alterations in sleep are associated with a multitude of neurologic conditions. Structural neuroplasticity, or changes in the size, strength, number, and targets of neuronal synaptic connections, can be modified by sleep and sleep disruption. However, the causal relationships between sleep loss and neuroplasticity, behavioral impacts, and genetic perturbations are poorly understood. The C. elegans GABAergic DVB neuron undergoes structural plasticity in adult males that rewires synaptic connections and alters behavior; the rewiring is dependent on the conserved autism-associated genes NRXN1/nrx-1 and NLGN3/nlg-1. Stress exposure during sexual maturation, which overlaps with a developmental sleep period, alters DVB structural plasticity in early adulthood. Here, we use four distinct methods to ask whether sleep deprivation results in altered structural and behavioral plasticity of the DVB neuron. Following sleep deprivation using the genetic mutants (aptf-1 and npr-1), vibration stimulus, or chemo-genetic silencing of a sleep-producing neuron, we find that DVB neurite outgrowth at day 1 of adulthood. Sleep deprivation also leads to an increase in the time to spicule protraction, the behavioral output of DVB function. We find that nrx-1 and nlg-1 both function to regulate sleep loss induced DVB structural and functional changes. Lastly, differences in DVB neurite outgrowth in sleep deprived animals at day 1 are transient, with DVB morphology being indistinguishable from controls by day 3. These results show that sleep loss modifies DVB structural plasticity and its behavioral consequences. Further, we show nrx-1 and nlg-1 mediate the effect of sleep deprivation on behavior at the level of DVB structural plasticity. These insights into sleep, neuroplasticity, behavior, and conserved autism genes have important implications for the role of sleep in neurologic disease. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=128 SRC="FIGDIR/small/545788v1_figu1.gif" ALT="Figure 1"> View larger version (34K): org.highwire.dtl.DTLVardef@2320c4org.highwire.dtl.DTLVardef@3c3b86org.highwire.dtl.DTLVardef@67dc96org.highwire.dtl.DTLVardef@c3033d_HPS_FORMAT_FIGEXP M_FIG Graphical AbstractGABAergic morphologic and behavioral plasticity induced by developmental sleep deprivation depends on the conserved autism genes, nrx-1 and nlg-1 A) Sleep depriving male C. elegans during all larval sleep periods in development (aptf-1 or npr-1) or selectively inhibiting sleep during L4-adult molt (vibration or RIS silencing) results in an increase in DVB outgrowth. B) Mutations in nrx-1 (and nlg-1 in npr-1 animals) prevent sleep deprivation induced morphologic plasticity changes. C) DVB neurite outgrowth results in a functional change in spicule protraction rate as a result of increased GABAergic inhibitory signaling, which also depends on nrx-1 and nlg-1. C_FIG

neuroscience↗

bioRxiv2022

A robotic system for automated genetic manipulation and analysis of Caenorhabditis elegans on agar media

The nematode Caenorhabditis elegans is one of the most widely studied organisms in biology due to its small size, rapid life cycle, and manipulable genetics. Research with C. elegans depends on labor-intensive and time-consuming manual procedures, imposing a major bottleneck for many studies, especially those involving large numbers of animals. Here we describe the first general-purpose tool, WormPicker, a robotic system capable of performing complex genetic manipulations and other tasks by imaging, phenotyping, and transferring C. elegans on standard agar media. Our system uses a motorized stage to move an imaging system and a robotic arm over an array of plates. Machine vision tools identify animals and assay developmental stage, morphology, sex, expression of fluorescent reporters, and other phenotypes. Based on the results of these assays the robotic arm selectively transfers individual animals using an electrically self-sterilized wire loop, with the aid of machine vision and electrical capacitance sensing. Automated C. elegans manipulation shows reliability and throughput comparable to standard manual methods. We developed software to enable the system to autonomously carry out complex protocols. To validate the effectiveness and versatility of our methods we used the system to perform a collection of common C. elegans procedures, including genetic crossing, genetic mapping, and genomic integration of a transgene. Our robotic system will accelerate C. elegans research and opens possibilities for performing genetic and pharmacological screens that would be impractical using manual methods. Significance StatementThe nematode Caenorhabditis elegans is a powerful genetic model organism in life sciences due to its compact anatomy, short life cycle, and optical transparency. Current methods for worm genetics rely on laborious, time-consuming, and error-prone manual work. Here, we describe the first general-purpose automated tool capable of genetically manipulating C. elegans. Our robotic system will accelerate a broad variety of C. elegans research and opens possibilities for performing genetic and pharmacological screens that would be impractical using manual methods.

bioengineering↗

bioRxiv2022

G Protein-Coupled Receptor Kinase-2 (GRK-2) regulates exploration through neuropeptide signaling in Caenorhabditis elegans

Animals alter their behavior in manners that depend on environmental conditions as well as their developmental and metabolic states. For example, C. elegans is quiescent during larval molts or during conditions of satiety. By contrast, worms enter an exploration state when removed from food. Sensory perception influences movement quiescence (defined as a lack of body movement), as well as the expression of additional locomotor states in C. elegans that are associated with increased or reduced locomotion activity, such as roaming (exploration behavior) and dwelling (local search). Here we find that movement quiescence is enhanced, and exploration behavior is reduced in G protein-coupled receptor kinase grk-2 mutant animals. grk-2 was previously shown to act in chemosensation, locomotion, and egg-laying behaviors. Using neuron-specific rescuing experiments, we show that GRK-2 acts in multiple ciliated chemosensory neurons to control exploration behavior. grk-2 acts in opposite ways from the cGMP-dependent protein kinase gene egl-4 to control movement quiescence and exploration behavior. Analysis of mutants with defects in ciliated sensory neurons indicates that grk-2 and the cilium-structure mutants act in the same pathway to control exploration behavior. We find that GRK-2 controls exploration behavior in an opposite manner from the neuropeptide receptor NPR-1 and the neuropeptides FLP-1 and FLP-18. Finally, we show that secretion of the FLP-1 neuropeptide is negatively regulated by GRK-2 and that overexpression of FLP-1 reduces exploration behavior. These results define neurons and molecular pathways that modulate movement quiescence and exploration behavior. Author summaryMany modulatory neurotransmitters affect behavior by binding to G protein-coupled receptors (GPCRs) and initiating signals that modify neuronal activity. GPCRs are regulated by G protein-coupled receptor kinases (GRKs). GRKs phosphorylate and promote the inactivation of GPCRs. Here we identify GRK-2 as a regulator of distinct locomotor states in C. elegans. We find that GRK-2 acts in olfactory sensory neurons to promote exploration and suppress movement quiescence. Additionally, we show that GRK-2 acts in opposition to a neuropeptide signaling pathway that acts in interneurons. Thus, this study demonstrates critical roles for GRK-2 in regulating neuromodulatory signaling and locomotor behavior.

genetics↗

bioRxiv2021

Pharyngeal timing and particle transport defects in Caenorhabditis elegans feeding mutants

The nematode C. elegans uses rhythmic muscle contractions (pumps) of the pharynx, a tubular feeding organ, to filter, transport, and crush food particles. A number of feeding mutants have been identified, including those with slow pharyngeal pumping rate, weak muscle contraction, defective muscle relaxation, and defective grinding of bacteria. Many aspects of these pharyngeal behavioral defects and how they affect pharyngeal function are not well understood. For example, the behavioral deficits underlying inefficient particle transport in slippery mutants have been unclear. Here we use high speed video microscopy to describe pharyngeal pumping behaviors and particle transport in wild-type animals and in feeding mutants. Different slippery mutants exhibit distinct defects including weak isthmus contraction, failure to trap particles in the anterior isthmus, and abnormal timing of contraction and relaxation in pharyngeal compartments. Our results show that multiple deficits in pharyngeal timing or contraction can cause defects in particle transport.

neuroscience↗

bioRxiv2020

Dehydrated Caenorhabditis elegans stocks are resistant to multiple freeze-thaw cycles

Ultracold preservation is widely used for storage of genetic stocks of Caenorhabditis elegans. Current cryopreservation protocols are vulnerable to refrigeration failures, which can result in the loss of stock viability due to damage during re-freezing. Here we present a method for preserving worms in a dehydrated and frozen form that retains viability after multiple freeze/thaw cycles. After dehydration in the presence of trehalose or glycerol, C. elegans stocks can be frozen and thawed multiple times while maintaining viability. While both dauer and non-dauer larvae survive desiccation and freezing, the dauer defective mutant daf-16 does not survive desiccation. Our technique is useful for storing stocks in a manner robust to freezer failures, and potentially for shipping strains between laboratories.

genetics↗

bioRxiv2019

A quiescent state following mild sensory arousal in Caenorhabditis elegans is potentiated by stress

An animals behavioral and physiological response to stressors includes changes to its responses to stimuli. How such changes occur is not well understood. Here we describe a Caenorhabditis elegans quiescent behavior, post-response quiescence (PRQ), which is modulated by the C. elegans response to cellular stressors. Following an aversive mechanical or blue light stimulus, worms respond first by briefly moving, and then become more quiescent for a period lasting tens of seconds. PRQ occurs at low frequency in unstressed animals, but is more frequent in animals that have experienced cellular stress due to ultraviolet light exposure as well as in animals following overexpression of epidermal growth factor (EGF). PRQ requires the function of the carboxypeptidase EGL-21 and the calcium-activated protein for secretion (CAPS) UNC-31, suggesting it has a neuropeptidergic mechanism. Although PRQ requires the sleep-promoting neurons RIS and ALA, it is not accompanied by decreased arousability, and does not appear to be homeostatically regulated, suggesting that it is not a sleep state. PRQ represents a simple, tractable model for studying how neuromodulatory states like stress alter behavioral responses to stimuli.

neuroscience↗

bioRxiv2019

A salt-induced kinase (SIK) is required for the metabolic regulation of sleep

Many lines of evidence point to links between sleep regulation and energy homeostasis, but mechanisms underlying these connections are unknown. During C. elegans sleep, energetic stores are allocated to non-neural tasks with a resultant drop in the overall fat stores and energy charge. Mutants lacking KIN-29, the C. elegans homolog of a mammalian Salt-Inducible Kinase (SIK) that signals sleep pressure, have low ATP levels despite high fat stores, indicating a defective response to cellular energy deficits. Liberating energy stores corrects adiposity and sleep defects of kin-29 mutants. kin-29 sleep and energy homeostasis roles map to a small number of sensory neurons that act upstream of fat regulation as well as of central sleep-controlling neurons, suggesting hierarchical somatic/neural interactions regulating sleep and energy homeostasis. Genetic interaction between kin-29 and the histone deacetylase hda-4 coupled with subcellular localization studies indicate that KIN-29 acts in the nucleus to regulate sleep. We propose that KIN-29/SIK acts in nuclei of sensory neuroendocrine cells to transduce low cellular energy charge into the mobilization of energy stores, which in turn promotes sleep.\n\nHighlightsO_LISleep is associated with fat mobilization and low ATP levels\nC_LIO_LIMetabolic regulation of sleep requires the salt-induced kinase (SIK) homolog KIN-29\nC_LIO_LIKIN-29 acts in sensory neurons upstream of sleep-promoting neurons\nC_LIO_LINuclear localization of KIN-29 is required for the metabolic regulation of sleep\nC_LIO_LIA type 2 histone deacetylase acts down stream of KIN-29 in the regulation of sleep.\nC_LIO_LILiberation of energy from fat-storage cells promotes sleep.\nC_LIO_LIBeta-oxidation promotes sleep.\nC_LI

neuroscience↗