Age- and stress-associated C. elegans granulins impair lysosomal function and induce a compensatory HLH-30/TFEB transcriptional response.

The progressive failure of protein homeostasis is a hallmark of aging and a common feature in neurodegenerative disease. As the enzymes executing the final stages of autophagy, lysosomal proteases are key contributors to the maintenance of protein homeostasis with age. We previously reported that expression of granulin peptides, the cleavage products of the neurodegenerative disease protein progranulin, enhance the accumulation and toxicity of TAR DNA binding protein 43 (TDP-43) in Caenorhabditis elegans (C. elegans). In this study we show that C. elegans granulins are produced in an age- and stress-dependent manner. Granulins localize to the endolysosomal compartment where they impair lysosomal protease expression and activity. Consequently, protein homeostasis is disrupted, promoting the nuclear translocation of the lysosomal transcription factor HLH-30/TFEB, and prompting cells to activate a compensatory transcriptional program. The three C. elegans granulin peptides exhibited distinct but overlapping functional effects in our assays, which may be due to amino acid composition that results in distinct electrostatic and hydrophobicity profiles. Our results support a model in which granulin production modulates a critical transition between the normal, physiological regulation of protease activity and the impairment of lysosomal function that can occur with age and disease

The tryptophan metabolite kynurenic acid is a neuromodulator mediating learning, brain aging, and neurodegeneration

The ability to change behaviors based on past experience is one of the most important and complex functions of the brain. Elucidating how activity of molecules, cells, and circuits all ultimately control learning and memory is difficult, and the simplicity of the nematode C. elegans offers an opportunity to connect these lower-level changes to full-scale changes in behavior. This allows for a depth of understanding of the brain’s biology that is largely impossible in other model organisms. Here we show that the tryptophan metabolite kynurenic acid (KYNA) functions as an inhibitory neuromodulator, communicating information about peripheral physiology to the nervous system to alter behavior. We have found that dietary restriction and many pathways involved in the body’s responses to dietary restriction – long known to affect brain function – enhance learning in C. elegans because they reduce production of KYNA, releasing KYNA-induced inhibition of neuronal activity in a specific pair of interneurons required for learning. In contrast, KYNA production increases with age, accounting for a significant portion of the decline in learning that occurs as animals get older. Furthermore, a C. elegans model of neurodegeneration may also increase KYNA production to impair learning. These studies demonstrate that metabolism can regulate brain function, having profound effects on cellular activity, learning, and memory. They establish that KYNA production is a crucial physiological indicator of peripheral status to the nervous system, offering insight into how the brain integrates information to direct behavior

The beneficial effects of dietary restriction on learning are distinct from its effects on longevity and mediated by depletion of a neuroinhibitory metabolite.

In species ranging from humans to Caenorhabditis elegans, dietary restriction (DR) grants numerous benefits, including enhanced learning. The precise mechanisms by which DR engenders benefits on processes related to learning remain poorly understood. As a result, it is unclear whether the learning benefits of DR are due to myriad improvements in mechanisms that collectively confer improved cellular health and extension of organismal lifespan or due to specific neural mechanisms. Using an associative learning paradigm in C. elegans, we investigated the effects of DR as well as manipulations of insulin, mechanistic target of rapamycin (mTOR), AMP-activated protein kinase (AMPK), and autophagy pathways-processes implicated in longevity-on learning. Despite their effects on a vast number of molecular effectors, we found that the beneficial effects on learning elicited by each of these manipulations are fully dependent on depletion of kynurenic acid (KYNA), a neuroinhibitory metabolite. KYNA depletion then leads, in an N-methyl D-aspartate receptor (NMDAR)-dependent manner, to activation of a specific pair of interneurons with a critical role in learning. Thus, fluctuations in KYNA levels emerge as a previously unidentified molecular mechanism linking longevity and metabolic pathways to neural mechanisms of learning. Importantly, KYNA levels did not alter lifespan in any of the conditions tested. As such, the beneficial effects of DR on learning can be attributed to changes in a nutritionally sensitive metabolite with neuromodulatory activity rather than indirect or secondary consequences of improved health and extended longevity

Kynurenic acid accumulation underlies learning and memory impairment associated with aging.

A general feature of animal aging is decline in learning and memory. Here we show that in Caenorhabditis elegans, a significant portion of this decline is due to accumulation of kynurenic acid (KYNA), an endogenous antagonist of neural N-methyl-D-aspartate receptors (NMDARs). We show that activation of a specific pair of interneurons either through genetic means or by depletion of KYNA significantly improves learning capacity in aged animals even when the intervention is applied in aging animals. KYNA depletion also improves memory. We show that insulin signaling is one factor in KYNA accumulation

Kynurenic acid accumulation underlies learning and memory impairment associated with aging.

A general feature of animal aging is decline in learning and memory. Here we show that in Caenorhabditis elegans, a significant portion of this decline is due to accumulation of kynurenic acid (KYNA), an endogenous antagonist of neural N-methyl-D-aspartate receptors (NMDARs). We show that activation of a specific pair of interneurons either through genetic means or by depletion of KYNA significantly improves learning capacity in aged animals even when the intervention is applied in aging animals. KYNA depletion also improves memory. We show that insulin signaling is one factor in KYNA accumulation

The steroid hormone ADIOL promotes learning by reducing neural kynurenic acid levels.

Reductions in brain kynurenic acid levels, a neuroinhibitory metabolite, improve cognitive function in diverse organisms. Thus, modulation of kynurenic acid levels is thought to have therapeutic potential in a range of brain disorders. Here we report that the steroid 5-androstene 3β, 17β-diol (ADIOL) reduces kynurenic acid levels and promotes associative learning in Caenorhabditis elegans We identify the molecular mechanisms through which ADIOL links peripheral metabolic pathways to neural mechanisms of learning capacity. Moreover, we show that in aged animals, which normally experience rapid cognitive decline, ADIOL improves learning capacity. The molecular mechanisms that underlie the biosynthesis of ADIOL as well as those through which it promotes kynurenic acid reduction are conserved in mammals. Thus, rather than a minor intermediate in the production of sex steroids, ADIOL is an endogenous hormone that potently regulates learning capacity by causing reductions in neural kynurenic acid levels

The beneficial effects of dietary restriction on learning are distinct from its effects on longevity and mediated by depletion of a neuroinhibitory metabolite

<div><p>In species ranging from humans to <i>Caenorhabditis elegans</i>, dietary restriction (DR) grants numerous benefits, including enhanced learning. The precise mechanisms by which DR engenders benefits on processes related to learning remain poorly understood. As a result, it is unclear whether the learning benefits of DR are due to myriad improvements in mechanisms that collectively confer improved cellular health and extension of organismal lifespan or due to specific neural mechanisms. Using an associative learning paradigm in <i>C</i>. <i>elegans</i>, we investigated the effects of DR as well as manipulations of insulin, mechanistic target of rapamycin (mTOR), AMP-activated protein kinase (AMPK), and autophagy pathways—processes implicated in longevity—on learning. Despite their effects on a vast number of molecular effectors, we found that the beneficial effects on learning elicited by each of these manipulations are fully dependent on depletion of kynurenic acid (KYNA), a neuroinhibitory metabolite. KYNA depletion then leads, in an N-methyl D-aspartate receptor (NMDAR)-dependent manner, to activation of a specific pair of interneurons with a critical role in learning. Thus, fluctuations in KYNA levels emerge as a previously unidentified molecular mechanism linking longevity and metabolic pathways to neural mechanisms of learning. Importantly, KYNA levels did not alter lifespan in any of the conditions tested. As such, the beneficial effects of DR on learning can be attributed to changes in a nutritionally sensitive metabolite with neuromodulatory activity rather than indirect or secondary consequences of improved health and extended longevity.</p></div

Genetic and pharmacological manipulations that mimic dietary restriction (DR) enhance learning by depleting kynurenic acid (KYNA).

<p>(A) RNAi interference (RNAi)-mediated reductions in the insulin receptor (<i>daf-2</i>), the mechanistic target of rapamycin (mTOR) kinase (<i>let-363</i>), Raptor (<i>daf-15</i>), Rictor (<i>rict-1</i>), and a negative regulator of autophagy (<i>mx1-3</i>), as well as animals treated with an activator of AMP-activated protein kinase (AMPK) (phenformin), have enhanced learning capacity even when fed ad libitum. <i>n</i> = 3–6, *<i>p</i> < 0.05, ***<i>p</i> < 0.001 by 2-way ANOVA (Bonferroni). (B) The elevated learning capacities of genetic and pharmacological mimetics of DR are dependent on N-methyl D-aspartate receptor (NMDAR) signaling. <i>n</i> = 3, *<i>p</i> < 0.05, **<i>p</i> < 0.01, ***<u><i>p</i></u> < 0.001 by 2-way ANOVA (Bonferroni). (C) Learning index values for additional mutants in various neural nutrient sensing pathways: <i>eat-2</i> mutants have a pharyngeal pumping defect, <i>tph-1</i> mutants do not produce serotonin, <i>flp-18</i> mutants lack a neuropeptide Y-like peptide, <i>tdc-1</i> mutants do not produce tyramine or octopamine, <i>tbh-1</i> mutants do not produce octopamine, and <i>dbl-1</i> mutants lack a transforming growth factor β (TGFβ) ligand. <i>n</i> = 3–6, *<i>p</i> < 0.05, ***<i>p</i> < 0.001 by 1-way ANOVA (Tukey). (D) Average total intensity of RIM GCaMP fluorescence over the entire 250-second imaging window in animals exposed to genetic and pharmacological DR mimetics. <i>n</i> = 6–10, *<i>p</i> < 0.05, **<i>p</i> < 0.01, ***<i>p</i> < 0.001 by 1-way ANOVA (Tukey). (E) Learning index values for mutants with high KYNA exposed to genetic and pharmacological DR mimetics. <i>n</i> = 3, *<i>p</i> < 0.05, **<i>p</i> < 0.01, ***<i>p</i> < 0.001 by 2-way ANOVA (Bonferroni). (F) Learning index values for wild-type and <i>nkat-1</i> animals given DR mimetics. To ensure that effects of DR mimetics in the context of KYNA depletion could be observed, animals were conditioned for only 15 minutes. <i>n</i> = 3, ***<i>p</i> < 0.001 by 2-way ANOVA (Bonferroni). (G) High-performance liquid chromatography (HPLC) measurements of steady-state KYNA levels for animals exposed to genetic and pharmacological DR mimetics. <i>n</i> = 5–18, *<i>p</i> < 0.05, **<i>p</i> < 0.01, ***<i>p</i> < 0.001 by 1-way ANOVA (Tukey). (H) HPLC measurements of steady-state KYNA levels for wild-type and <i>daf-2(e1370)</i> mutant animals. <i>n</i> = 2, *<i>p</i> < 0.05 by 2-tailed Student <i>t</i> test. Animals in panels (B), (C), (E), and (F) were ad libitum fed and conditioned. All data are represented as mean ± SEM. Underlying data can be found in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.2002032#pbio.2002032.s009" target="_blank">S1 Data</a>. n.s., not significant.</p