The hypoxic response extends lifespan through a bioaminergic and peptidergic neural circuit

Reviewer #1 (Public review):

Summary:

In this study by Kitto et al., the authors set out to identify specific signaling components regulating the hypoxic response from the neurons to the periphery and which components are required for lifespan extension. Their previous work had shown that expression of a stabilized HIF-1 mutant in the nervous system extends lifespan through the serotonin receptor SER-7 and leads to the induction of fmo-2 in the intestine. In the current study, they mapped the precise neural circuits required for this response, as well as the signaling mediators. Their work reveals that neurotransmitters GABA and tyramine, and the neuropeptide NLP-17, act downstream of neuronal HIF-1 to convey a "hypoxic signal" to peripheral tissues. Through cell-type-specific expression studies, targeted knockouts, and comprehensive lifespan analysis, the authors provide robust evidence to support their conclusions. The insights gained from the study are both moving the field forward as they advance our understanding of neuro-peripheral hypoxic signaling, but they also lay the groundwork for potential therapeutic strategies aimed at the modulation of such signaling pathways.

Strengths:

(1) This study provides new evidence further delineating signaling components required for hypoxic signaling-mediated longevity, from the nervous system to the periphery. Using a rigorous approach where they express stabilized HIF-1 mutant selectively in ADF, NSM, and HSN serotonergic neurons, followed by cell-type-specific tph-1 knockouts to pinpoint ADF-dependent serotonin signaling as essential for both lifespan extension and intestinal fmo-2 induction.

This was followed by generating 11 transgenic lines that drive SER-7 expression under distinct neuron-specific promoters, to systematically tease out in which of 27 candidate neurons SER-7 functions to mediate hypoxia-induced longevity. This ultimately highlighted the RIS interneuron as the required signaling hub.

(2) As the intestine lacks direct neuronal innervation, the authors employ neuron-specific RNAi (TU3311 strain) and dense core vesicle analyses to identify that the neuropeptide NLP-17 is required to transmit the hypoxic signal from RIS to induce fmo-2 in the intestine.

(3) Overall, the paper is very well written. The experiments were carried out carefully and thoroughly, and the conclusions drawn are also well supported by the results they are showing.

Weaknesses:

Overall, I don't see many weaknesses. One point relates to their read-outs, which rely heavily on lifespan measurements and fmo-2 induction without evaluating other physiological processes that serotonin or NLP-17 might affect. For translational relevance, it would be valuable to assess or mention potential adverse effects, such as changes in reproduction, pharyngeal pumping, or proteostasis capacity (proteostasis capacity specifically in the tissue showing fmo-2 upregulation).

While lifespan assays and fmo-2 expression do provide strong evidence, incorporating additional markers of stress resistance could strengthen the link between hypoxic signaling and organismal health as well.

Reviewer #2 (Public review):

Summary:

The authors aimed to identify the specific neurons, neurotransmitters, and neuropeptides that mediate the longevity effects of the hypoxic response in C. elegans. By genetically dissecting the pathway downstream of HIF-1, they define a neural circuit involving ADF serotonergic neurons, the SER-7 receptor in the RIS interneuron, tyraminergic signaling from RIM, and neuropeptide NLP-17, ultimately linking neuronal hypoxic sensing to pro-longevity signaling in the intestine.

Strengths:

The study employs a diverse genetic toolkit, including neuron-specific transgenes, tissue-specific knockouts and rescues, RNAi knockdowns, allowing the authors to pinpoint causality, sufficiency, and necessity with high resolution. The comprehensive mapping of cell-nonautonomous signaling adds depth to our understanding of how HIF and serotonin signaling interface with aging pathways. The conclusions are supported by consistent survival assays and fmo-2 gene expression analyses.

Weaknesses:

A key limitation is the lack of clear evidence showing epistasis of so many identified molecular/neuronal components downstream of HIF-1 and serotonin. Thus, the mechanisms of how a diverse set of molecules/neurons coordinate and mediate neuronal HIF-1 effects on intestinal fmo-2 and longevity remain murky. Some rescue strategies may inadvertently cause non-physiological expression. Additionally, environmental hypoxia was not tested in parallel, so the claim on "hypoxia respone" throughout the manuscript is not justified by genetic manipulation alone, and the translational relevance of the genetic manipulations remains somewhat uncertain.

Reviewer #3 (Public review):

Summary:

This study found that ADF serotonergic neurons have a significant role in extending lifespan mediated by HIF-1, as well as serotonin receptor SER-7 in the GABAergic RIS interneurons. The author focuses on the sufficiency and necessity of components from the central nervous system and how they contribute to aging upon hypoxia.

Previous work from the lab has identified that the stabilization of HIF-1 in neurons is sufficient to extend lifespan through the serotonin receptor, SER-7, which subsequently activates fmo-2 in the intestine and leads to lifespan extension. Building on this, the author sought to determine which serotonergic neurons are involved and found that serotonin signaling in ADF neurons is required for lifespan extension mediated by HIF-1.

The author next tested which subset of neurons requires Ser-7 expression to rescue hypoxic response. They found that ser-7 expression in multiple neurons is sufficient to induce fmo-2, with the top candidate being the RIS neuron. Ablation of the RIS neuron did not extend lifespan, suggesting that ser-7 expression in the RIS neuron is required for lifespan extension, positioning it as a key component in the longevity signaling pathway.

The author also investigated neurotransmitters and found that GABA and tyramine are important components in this circuit. They showed that the tyramine receptor called tyra-3 is required for vhl-1-mediated longevity. Given that tyra-3 is expressed in oxygen- and carbon dioxide-sensing neurons, the author demonstrated that these sensing neurons work downstream of serotonin signaling. Lastly, the author screened neuropeptide/receptor binding pairs and identified NLP-17 as playing a role in hypoxia-mediated longevity.

Originality and Significance:

This research is significant in that it uncovers components that are sufficient and necessary for lifespan extension via the hypoxic response. It provides comprehensive data supporting longevity induced by HIF-1-mediated hypoxic response, in conjunction with fmo-2, a longevity gene, as demonstrated in previous work from the lab. Moreover, it provides a number of new transgenic worm tools for C. elegans and aging communities.

Data and Methodology:

(1) The experiments were thoroughly conducted, especially the generations of strains using different neuron-type promoters and crossing into mutant strains to demonstrate sufficiency and necessity.

(2) Some figure legends from the text do not match what the data show. (Figure 6E, F, G).

(3) The lifespan graph legends are confusing and could use some revamping for better clarification.

Conclusions:

This study provides insights into how hypoxic response regulates aging in a cell non-autonomous manner, outlining a potential circuit involving neurons, neurotransmitters, and neuropeptides.