Label-Free Imaging of Lipid Storage Dynamics in <em>Caenorhabditis elegans</em> using Stimulated Raman Scattering Microscopy

Lipid metabolism is a fundamental physiological process necessary for cellular and organism health. Dysregulation of lipid metabolism often elicits ...

Baylor College of Medicine

Advances in modern optical microscopy have provided unparalleled tools to study intracellular structure and function, yet visualizing lipid molecules within a cell remains challenging. Stimulated Raman Scattering (SRS) microscopy is a recently developed imaging modality that addresses this challenge. By selectively imaging the vibration of chemical moieties enriched in lipids, this technique allows for rapid imaging of lipid molecules in vivo without the need for perturbative extrinsic labels. SRS microscopy has been effectively employed in the study of fat metabolism, helping uncover novel regulators of lipid storage. This unit provides a brief introduction to the principle of SRS microscopy, and describes methods for its use in imaging lipids in cells, tissues, and whole organisms.

Label-free biomedical imaging of lipids by stimulated Raman scattering microscopy.

Reproductive plasticity describes the ability of organisms to adjust parameters such as volume, rate, or timing of progeny production in order to maximize successful reproduction under different environmental conditions. Reproductive plasticity in response to environmental variation has been observed in a wide range of animals; however, the mechanisms involved in translating environmental cues into reproductive outcomes remain unknown. Here, we show that olfaction modulates reproductive timing and senescence through neuroendocrine signaling in Caenorhabditis elegans. On their preferred diet, worms demonstrate an increased rate of reproduction and an early onset of reproductive aging. Perception of the preferred diet's odor by AWB olfactory neurons elicits these adjustments by increasing germline proliferation, and optogenetic stimulation of AWB neurons is sufficient to accelerate reproductive timing in the absence of dietary inputs. Furthermore, AWB neurons act through neuropeptide signaling to regulate reproductive rate and senescence. These findings reveal a neuroendocrine nexus linking olfactory sensation and reproduction in response to environmental variation and indicate the significance of olfaction in the regulation of reproductive decline during aging.

Olfaction Modulates Reproductive Plasticity through Neuroendocrine Signaling in Caenorhabditis elegans.

High-throughput screens at microscopic resolution can uncover molecular mechanisms of cellular dynamics, but remain technically challenging in live multicellular organisms. Here we present a genetic screening method using photo-highlighting for candidate selection on microscopes. We apply this method to stimulated Raman scattering (SRS) microscopy and systematically identify 57 Caenorhabditis elegans mutants with altered lipid distribution. Four of these mutants target the components of the Bone Morphogenetic Protein (BMP) signaling pathway, revealing that BMP signaling inactivation causes exhaustion of lipid reserves in somatic tissues. Using SRS-based isotope tracing assay to quantitatively track lipid synthesis and mobilization, we discover that the BMP signaling mutants have increased rates of lipid mobilization. Furthermore, this increase is associated with the induction of mitochondrial β-oxidation and mitochondrial fusion. Together these studies demonstrate a photo-highlighting microscopic strategy for genome-scale screens, leading to the discovery of new roles for BMP signaling in linking mitochondrial homeostasis and lipid metabolism.High-throughput genetic screens in animals could benefit from an easy way to mark positive hits. Here the authors introduce photo-highlighting using a photoconvertible fluorescent protein, and in combination with stimulated Raman scattering (SRS) microscopy, define a role for BMP signaling in lipid metabolism in C. elegans.

High-throughput screens using photo-highlighting discover BMP signaling in mitochondrial lipid oxidation.

Retinoids play critical roles in development, immunity, and lipid metabolism, and their deficiency leads to various human disorders. Yet, tools for sensing retinoids in vivo are lacking, which limits the understanding of retinoid distribution, dynamics and functions in living organisms. Here, using hyperspectral stimulated Raman scattering microscopy, we discover a previously unknown cytoplasmic store of retinoids in Caenorahbditis elegans. Following the temporal dynamics of retinoids, we reveal that their levels are positively correlated with fat storage, and their supplementation slows down fat loss during starvation. We also discover that retinoids promote fat unsaturation in response to high-glucose stress, and improve organism survival. Together, our studies report a new method for tracking the spatiotemporal dynamics of retinoids in living organisms, and suggest the crucial roles of retinoids in maintaining metabolic homeostasis and enhancing organism fitness upon developmental and dietary stresses.

Fingerprint Stimulated Raman Scattering Imaging Reveals Retinoid Coupling Lipid Metabolism and Survival.

Olfactory and metabolic dysfunctions are intertwined phenomena associated with obesity and neurodegenerative diseases; yet how mechanistically olfaction regulates metabolic homeostasis remains unclear. Specificity of olfactory perception integrates diverse environmental odors and olfactory neurons expressing different receptors. Here, we report that specific but not all olfactory neurons actively regulate fat metabolism without affecting eating behaviors in Caenorhabditis elegans, and identified specific odors that reduce fat mobilization via inhibiting these neurons. Optogenetic activation or inhibition of the responsible olfactory neural circuit promotes the loss or gain of fat storage, respectively. Furthermore, we discovered that FLP-1 neuropeptide released from this olfactory neural circuit signals through peripheral NPR-4/neuropeptide receptor, SGK-1/serum- and glucocorticoid-inducible kinase, and specific isoforms of DAF-16/FOXO transcription factor to regulate fat storage. Our work reveals molecular mechanisms underlying olfactory regulation of fat metabolism, and suggests the association between olfactory perception specificity of each individual and his/her susceptibility to the development of obesity.

Olfactory specificity regulates lipid metabolism through neuroendocrine signaling in Caenorhabditis elegans.

Lipids play crucial roles in regulating aging and longevity. In the past few decades, a series of genetic pathways have been discovered to regulate lifespan in model organisms. Interestingly, many of these regulatory pathways are linked to lipid metabolism and lipid signaling. Lipid metabolic enzymes undergo significant changes during aging and are regulated by different longevity pathways. Lipids also actively modulate lifespan and health span as signaling molecules. In this review, we summarize recent insights into the roles of lipid metabolism and lipid signaling in aging and discuss lipid-related interventions in promoting longevity.

Ayse Sena Mutlu

Huffington Center on Aging

Baylor College of Medicine

Label-Free Imaging of Lipid Storage Dynamics in Caenorhabditis elegans using Stimulated Raman Scattering Microscopy

Ayse Sena Mutlu

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Baylor College of Medicine

Label-Free Imaging of Lipid Storage Dynamics in Caenorhabditis elegans using Stimulated Raman Scattering Microscopy

Huffington Center on Aging