Hugin-AstA circuitry is a novel central energy sensor that directly regulates sweet sensation in Drosophila and mouse

Reviewer #1 (Public review):

In this manuscript, Qin and colleagues aim to delineate a neural mechanism by which the internal satiety levels modulate the intake of sugar solution. They identified a three-step neuropeptidergic system that downregulates the sensitivity of sweet-sensing gustatory sensory neurons in sated flies. First, neurons that release a neuropeptide Hugin (which is an insect homolog of vertebrate Neuromedin U (NMU)) are in an active state when the concentration of glucose is high. This activation does not require synaptic inputs, suggesting that Hugin-releasing neurons sense hemolymph glucose levels directly. Next, the Hugin neuropeptides activate Allatostatin A (AstA)-releasing neurons via one of Hugin's receptors, PK2-R1. Finally, the released AstA neuropeptide suppresses sugar response in sugar-sensing Gr5a-expressing gustatory sensory neurons through AstA-R1 receptor. Suppression of sugar response in Gr5a-expressing neurons reduces the fly's sugar intake motivation (measured by proboscis extension reflex). They also found that NMU-expressing neurons in the ventromedial hypothalamus (VMH) of mice (which project to the rostral nucleus of the solitary tract (rNST)) are also activated by high concentrations of glucose, independent of synaptic transmission, and that injection of NMU reduces the glucose-induced activity in the downstream of NMU-expressing neurons in rNST. These data suggest that the function of Hugin neuropeptide in the fly is analogous to the function of NMU in the mouse.

Generally, their central conclusions are well-supported by multiple independent approaches. The parallel study in mice adds a unique comparative perspective that makes the paper interesting to a wide range of readers. It is easier said than done: the rigor of this study, which effectively combined pharmacological and genetic approaches to provide multiple lines of behavioral and physiological evidence, deserves recognition and praise.

A perceived weakness is that the behavioral effects of the manipulations of Hugin and AstA systems are modest compared to a dramatic shift of sugar solution-induced PER (the behavioral proxy of sugar sensitivity) induced by hunger, as presented in Figure 1B and E. It is true that the mutation of tyrosine hydroxylase (TH), which synthesizes dopamine, does not completely abolish the hunger-induced PER change, but the remaining effect is small. Moreover, the behavioral effect of the silencing of the Hugin/AstA system (Figure Supplement 13B, C) is difficult to interpret, leaving a possibility that this system may not be necessary for shifting PER in starved flies. These suggest that the Hugin-AstA system accounts for only a minor part of the behavioral adaptation induced by the decreased sugar levels. Their aim to "dissect out a complete neural pathway that directly senses internal energy state and modulates food-related behavioral output in the fly brain" is likely only partially achieved. While this outcome is not a shortcoming of a study per se, the depth of discussion on the mechanism of interactions between the Hugin/AstA system and the other previously characterized molecular circuit mechanisms mediating hunger-induced behavioral modulation is insufficient for readers to appreciate the novelty of this study and future challenges in the field. In this context, authors are encouraged to confront a limitation of the study due to the lack of subtype-level circuit characterization, despite their intriguing finding that only a subtype of Hugin- and AstA-releasing neurons are responsive to the elevated level of bath-applied glucose.

Reviewer #2 (Public review):

Summary:

The question of how caloric and taste information interact and consolidate remains both active and highly relevant to human health and cognition. The authors of this work sought to understand how nutrient sensing of glucose modulates sweet sensation. They found that glucose intake activates hugin signaling to AstA neurons to suppress feeding, which contributes to our mechanistic understanding of nutrient sensation. They did this by leveraging the genetic tools of Drosophila to carry out nuanced experimental manipulations and confirmed the conservation of their main mechanism in a mammalian model. This work builds on previous studies examining sugar taste and caloric sensing, enhancing the resolution of our understanding.

Strengths:

Fully discovering neural circuits that connect body state with perception remains central to understanding homeostasis and behavior. This study expands our understanding of sugar sensing, providing mechanistic evidence for a hugin/AstA circuit that is responsive to sugar intake and suppresses feeding. In addition to effectively leveraging the genetic tools of Drosophila, this study further extends their findings into a mammalian model with the discovery that NMU neural signaling is also responsive to sugar intake.

Weaknesses:

The effect of Glut1 knockdown on PER in hugin neurons is modest, and does not show a clear difference between fed and starved flies as might be expected if this mechanism acts as a sensor of internal energy state. This could suggest that glucose intake through Glut1 may only be part of the mechanism.

Reviewer #3 (Public review):

Summary:

This study identifies a novel energy-sensing circuit in Drosophila and mice that directly regulates sweet taste perception. In flies, hugin+ neurons function as a glucose sensor, activated through Glut1 transport and ATP-sensitive potassium channels. Once activated, hugin neurons release hugin peptide, which stimulates downstream Allatostatin A (AstA)+ neurons via PK2-R1 receptors. AstA+ neurons then inhibit sweet-sensing Gr5a+ gustatory neurons through AstA peptide and its receptor AstA-R1, reducing sweet sensitivity after feeding. Disrupting this pathway enhances sweet taste and increases food intake, while activating the pathway suppresses feeding.

The mammalian homolog of neuromedin U (NMU) was shown to play an analogous role in mice. NMU knockout mice displayed heightened sweet preference, while NMU administration suppressed it. In addition, VMH NMU+ neurons directly sense glucose and project to rNST Calb2+ neurons, dampening sweet taste responses. The authors suggested a conserved hugin/NMU-AstA pathway that couples energy state to taste perception.

Strengths:

Interesting findings that extend from insects to mammals. Very comprehensive.

Weaknesses:

Coupling energy status to taste sensitivity is not a new story. Many pathways appear to be involved, and therefore, it raises a question as to how this hugin-AstA pathway is unique.

Author response:

Reviewer #1 (Public review):

In this manuscript, Qin and colleagues aim to delineate a neural mechanism by which the internal satiety levels modulate the intake of sugar solution. They identified a three-step neuropeptidergic system that downregulates the sensitivity of sweet-sensing gustatory sensory neurons in sated flies. First, neurons that release a neuropeptide Hugin (which is an insect homolog of vertebrate Neuromedin U (NMU)) are in an active state when the concentration of glucose is high. This activation does not require synaptic inputs, suggesting that Hugin-releasing neurons sense hemolymph glucose levels directly. Next, the Hugin neuropeptides activate Allatostatin A (AstA)-releasing neurons via one of Hugin's receptors, PK2-R1. Finally, the released AstA neuropeptide suppresses sugar response in sugar-sensing Gr5a-expressing gustatory sensory neurons through AstA-R1 receptor. Suppression of sugar response in Gr5a-expressing neurons reduces the fly's sugar intake motivation (measured by proboscis extension reflex). They also found that NMU-expressing neurons in the ventromedial hypothalamus (VMH) of mice (which project to the rostral nucleus of the solitary tract (rNST)) are also activated by high concentrations of glucose, independent of synaptic transmission, and that injection of NMU reduces the glucose-induced activity in the downstream of NMU-expressing neurons in rNST. These data suggest that the function of Hugin neuropeptide in the fly is analogous to the function of NMU in the mouse.

Generally, their central conclusions are well-supported by multiple independent approaches. The parallel study in mice adds a unique comparative perspective that makes the paper interesting to a wide range of readers. It is easier said than done: the rigor of this study, which effectively combined pharmacological and genetic approaches to provide multiple lines of behavioral and physiological evidence, deserves recognition and praise.

A perceived weakness is that the behavioral effects of the manipulations of Hugin and AstA systems are modest compared to a dramatic shift of sugar solution-induced PER (the behavioral proxy of sugar sensitivity) induced by hunger, as presented in Figure 1B and E. It is true that the mutation of tyrosine hydroxylase (TH), which synthesizes dopamine, does not completely abolish the hunger-induced PER change, but the remaining effect is small. Moreover, the behavioral effect of the silencing of the Hugin/AstA system (Figure Supplement 13B, C) is difficult to interpret, leaving a possibility that this system may not be necessary for shifting PER in starved flies. These suggest that the Hugin-AstA system accounts for only a minor part of the behavioral adaptation induced by the decreased sugar levels. Their aim to "dissect out a complete neural pathway that directly senses internal energy state and modulates food-related behavioral output in the fly brain" is likely only partially achieved. While this outcome is not a shortcoming of a study per se, the depth of discussion on the mechanism of interactions between the Hugin/AstA system and the other previously characterized molecular circuit mechanisms mediating hunger-induced behavioral modulation is insufficient for readers to appreciate the novelty of this study and future challenges in the field.

We thank the reviewer for the thoughtful comment. We agree that the behavioral effects of manipulating the Hugin–AstA system alone were considerably weaker than the pronounced PER shifts induced by starvation. We will revise our Discussion to address it by positioning our findings within the broader context of energy regulation.

More specifically, we will discuss that feeding behavior is controlled by two distinct, yet synergistic, types of mechanisms:

(1) Hunger-driven 'accelerators': as the reviewer notes, pathways involving dopamine and NPF are powerful drivers of sweet sensitivity. These systems are strongly activated by hunger to promote food-seeking and consumption.

(2) Satiety-driven 'brakes': our study identifies the counterpart to those systems above, aka. a satiety-driven 'brake'. The Hugin–AstA pathway acts as a direct sensor of high internal energy (glucose), which is specifically engaged during satiety to actively suppress sweet sensation and prevent overconsumption.

This framework explains the seemingly discrepancy in effect size. The dramatic PER shift seen upon starvation is a combined result of engaging the 'accelerators' (hunger pathways like TH/NPF) while simultaneously releasing the 'brake' (our Hugin–AstA pathway being inactive).

Our manipulations, which specifically target only the 'brake' system, are therefore expected to have a more modest effect than this combined physiological state. Thus, rather than being a "minor part," the Hugin–AstA pathway is a mechanistically defined, satiety-specific circuit that is essential for the precise "braking" required for energy homeostasis. We will update our Discussion to emphasize how these 'accelerator' and 'brake' circuits must work in concert to ensure precise energy regulation.

In this context, authors are encouraged to confront a limitation of the study due to the lack of subtype-level circuit characterization, despite their intriguing finding that only a subtype of Hugin- and AstA-releasing neurons are responsive to the elevated level of bath-applied glucose.

We thank the reviewer for highlighting the critical issue of subtype-level specialization within the Hugin and AstA populations.

We fully agree that the Hugin system is known for its functional heterogeneity (pleiotropy), with different Hugin neuron subclusters implicated in regulating a variety of behaviors, including feeding, aversion, and locomotion (we will cite relevant literature here). Our finding that only a specific subcluster of Hugin neurons is responsive to glucose elevation provides a crucial first step in functionally dissecting this complexity.

We will add a dedicated paragraph to elaborate on this functional partitioning. We propose that this subtype-level specialization allows the Hugin system to precisely link specific physiological states (like high circulating glucose) to appropriate behavioral outputs (like the suppression of sweet taste), demonstrating an elegant solution to coordinating multiple survival behaviors. Future work using high-resolution tools such as split-GAL4 and single-cell sequencing will be invaluable in fully mapping the specific functional roles corresponding to each Hugin and AstA subcluster.

Reviewer #2 (Public review):

Summary:

The question of how caloric and taste information interact and consolidate remains both active and highly relevant to human health and cognition. The authors of this work sought to understand how nutrient sensing of glucose modulates sweet sensation. They found that glucose intake activates hugin signaling to AstA neurons to suppress feeding, which contributes to our mechanistic understanding of nutrient sensation. They did this by leveraging the genetic tools of Drosophila to carry out nuanced experimental manipulations and confirmed the conservation of their main mechanism in a mammalian model. This work builds on previous studies examining sugar taste and caloric sensing, enhancing the resolution of our understanding.

Strengths:

Fully discovering neural circuits that connect body state with perception remains central to understanding homeostasis and behavior. This study expands our understanding of sugar sensing, providing mechanistic evidence for a hugin/AstA circuit that is responsive to sugar intake and suppresses feeding. In addition to effectively leveraging the genetic tools of Drosophila, this study further extends their findings into a mammalian model with the discovery that NMU neural signaling is also responsive to sugar intake.

Weaknesses:

The effect of Glut1 knockdown on PER in hugin neurons is modest, and does not show a clear difference between fed and starved flies as might be expected if this mechanism acts as a sensor of internal energy state. This could suggest that glucose intake through Glut1 may only be part of the mechanism.

We thank the reviewer for this insightful comment and agree that the modest behavioral effect of Glut1 knockdown is a critical finding that warrants further clarification. This observation strongly supports the idea that internal energy state is monitored by a sophisticated and robust network, not a single, fragile component. We believe the effect size is modest for two main reasons, which we will further address in revised Discussion.

Firstly, the effect size is likely attenuated by technical and molecular redundancy. Specifically, the RNAi-mediated knockdown of Glut1 may be incomplete, leaving residual transporter function. Furthermore, Glut1 is likely only one part of the Hugin neuron's intrinsic sensing mechanism; other components, such as alternative glucose transporters or downstream K_ATP channel signaling, may provide molecular redundancy, meaning that the full energy-sensing function is not easily abolished by a single manipulation.

Secondly, and more importantly, the final feeding decision is an integrated output of competing circuits. While hunger-sensing pathways like the dopamine and NPF circuits act as powerful "accelerators" to drive sweet consumption, the Hugin–AstA pathway serves as a satiety-specific "brake". The modest effect of partially inhibiting just one component of this 'brake' system is the hallmark of a precisely regulated, multi-layered homeostatic system. We will further clarify in the Discussion that the Hugin pathway represents one essential inhibitory circuit within this cooperative network that works together with the hunger-promoting systems to ensure precise control over energy intake.

Reviewer #3 (Public review):

Summary:

This study identifies a novel energy-sensing circuit in Drosophila and mice that directly regulates sweet taste perception. In flies, hugin+ neurons function as a glucose sensor, activated through Glut1 transport and ATP-sensitive potassium channels. Once activated, hugin neurons release hugin peptide, which stimulates downstream Allatostatin A (AstA)+ neurons via PK2-R1 receptors. AstA+ neurons then inhibit sweet-sensing Gr5a+ gustatory neurons through AstA peptide and its receptor AstA-R1, reducing sweet sensitivity after feeding. Disrupting this pathway enhances sweet taste and increases food intake, while activating the pathway suppresses feeding.

The mammalian homolog of neuromedin U (NMU) was shown to play an analogous role in mice. NMU knockout mice displayed heightened sweet preference, while NMU administration suppressed it. In addition, VMH NMU+ neurons directly sense glucose and project to rNST Calb2+ neurons, dampening sweet taste responses. The authors suggested a conserved hugin/NMU-AstA pathway that couples energy state to taste perception.

Strengths

Interesting findings that extend from insects to mammals. Very comprehensive.

Weaknesses:

Coupling energy status to taste sensitivity is not a new story. Many pathways appear to be involved, and therefore, it raises a question as to how this hugin-AstA pathway is unique.

The reviewer is correct that several energy-sensing pathways are known. However, we now clarify that these previously established mechanisms, such as the dopaminergic and NPF pathways, primarily function as hunger-driven "accelerators." They are activated by low energy states to promote sweet sensitivity and drive consumption.

The crucial, missing piece of the puzzle—which our study provides—is the satiety-specific "brake" mechanism. We identify the Hugin–AstA circuit as one of the “brakes”: a dedicated, central sensor that responds directly to high circulating glucose (satiety) to suppress sweet sensation and prevent overconsumption.

Thus, our work is unique because it defines the essential counterpart to the hunger pathways. In the revised Discussion, we will further explain how these 'accelerator' (hunger) and 'brake' (satiety) systems work in concert to allow for the precise, bidirectional regulation of energy intake. Furthermore, by demonstrating that this Hugin/NMU 'brake' circuit is evolutionarily conserved in mice, our findings reveal a fundamental energy-sensing strategy and suggest that this pathway could represent a promising new therapeutic target for managing conditions of excessive food intake.