Food availability and nutritional status modulate olfactory acuity and behavior in most organisms 
. In C. elegans
, food stimulates ASH-mediated aversive responses and the present studies highlight the complexity of the serotonergic signal defining food availability (). 5-HT released from the serotonergic NSMs and ADFs appears to have antagonistic effects on octanol-stimulated aversive behavior, with NSM 5-HT stimulating aversive responses and ADF 5-HT inhibiting food-stimulation. For example, increasing serotonergic signaling from the NSMs by knocking down NSM mod-5
or overexpressing tph-1
stimulates basal aversive responses to levels observed on food or exogenous 5-HT. Conversely, increasing serotonergic signaling from the ADFs by knocking down ADF mod-5
or overexpressing tph-1
abolishes the stimulation of aversive responses by food or exogenous 5-HT. These genetic manipulations have been interpreted as having acute effects on serotonergic signaling, but it is worth noting that altered serotonergic signaling can potentially also have effects on neuronal development in C. elegans 
. These results highlight the complexity of serotonergic modulation and the potential limitations of exogenous ligands in characterizing serotonergic signaling.
Model for the food dependent modulation of the ASH-mediated aversive behavior.
Interactions between NSM and ADF signaling are complex, with both neuron-specific and cooperative interactions described previously. The branched NSMs make synapses onto the pharyngeal basement membrane and muscle, but also secrete 5-HT and neuropeptides directly into the pseudocoelomic fluid from vesicle-filled varicosities 
. Many mammalian monoaminergic neurons are also “asynaptic” 
. In contrast, the ADFs are innervated by the ASHs directly and synapse onto interneurons modulating key locomotory transitions 
. The NSMs are required for the enhanced slowing response and both the NSMs and ADFs are required for dispersal during starvation 
. However, these studies often have relied on NSM ablation, so whether NSM 5-HT, glutamate and/or neuropeptides were involved is unclear. In contrast, the food-sensitization of hyperoxia avoidance and aversive learning were rescued in tph-1
animals by tph-1
expression in the ADFs, but not the NSMs, supporting a role for ADF 5-HT in food-sensitization 
. Whether the ADFs sense food directly or respond to signals from other neurons in response to food availability/quality is unclear. However, ADFp::tph-1
expression is elevated in daf-7
encodes a TGF-β homologue and the ASI expression of DAF-7 decreases during stress/starvation, suggesting that starvation stimulates ADF 5-HT and food stimulates NSM 5-HT 
. Serotonergic signaling from ADFs requires MOD-1 for avoidance to pathogenic bacteria 
. Bacteria should stimulate NSM 5-HT and activate feeding, but pathogenic bacteria may also stimulate ADF 5-HT to antagonize NSM signaling, with products from the pathogens either activating the ADFs directly or indirectly through other sensory neurons. In the present studies, SER-1, not MOD-1, was essential for the ADF 5-HT inhibition of food-stimulation and it is unclear if SER-1 was required for aversive learning to pathogenic bacteria, highlighting the potential complexity of ADF serotonergic signaling.
Food and 5-HT stimulate ASH-mediated aversive responses through at least three different 5-HT receptors, operating within the ASH-mediated circuit 
. 5-HT from the NSMs, ADFs and potentially other serotonergic neurons differentially interacts with subsets of these 5-HT receptors and serotonergic signaling appears to involve precisely modulated changes in local 5-HT levels, mediated primarily by extra-synaptic 5-HT receptors (). In fact, C. elegans
5-HT receptors are expressed on many neurons that are not directly innervated by serotonergic neurons, suggesting that most serotonergic signaling is humoral and extra-synaptic 
. Our data suggest that stimulation of aversive responses by NSM 5-HT requires SER-5 and SER-1, and confirm the role of the RIAs in NSM 5-HT stimulation 
. In contrast, MOD-1 is not required, although it is essential for the sensitization of aversive responses by food or exogenous 5-HT, suggesting that additional sources of 5-HT may contribute to food-sensitization or that the levels of 5-HT released from the NSMs in these experiments overcome the need for MOD-1. The inhibition of food-stimulation by ADF 5-HT requires only SER-1, not in the RIAs but in the RICs. Presumably the Gαq
-coupled SER-1 stimulates OA release from the RICs that, in turn, inhibits ASH signaling through the Gαo
-coupled OCTR-1, as we have demonstrated previously 
. Together, these results demonstrate that SER-1 is required for both ADF inhibition and NSM stimulation, highlighting the complexity of serotonergic modulation.
Nutritional status modulates multiple aspects of the ASH-mediated aversive response, including reversal duration and trajectory after reversal is complete. For example, off food, animals initiate reversal more slowly, back up further and are more likely to turn from their initial trajectory, often initiating an omega turn. These post-initiation responses are independent of the intensity of the initiating stimulus (30 vs 100% 1-octanol), suggesting that the decision to continue forward or change direction after contact with an aversive stimulus is dictated largely by nutritional state. The 5-HT receptors modulating the initiation of reversal also modulate post-initiation behaviors, suggesting that nutritional status is defined by an extrasynaptic “serotonergic circuit.” The presence of food is perceived by a subset of sensory neurons and modulates many sensory-mediated behaviors. For example, the AWCs, ASKs, and ASIs are involved in locomotory changes associated with removal from food 
. The AWCs are required for turning and reversals 
, the ASKs and AWCs for long reversals and omega turns, and the ASIs for dispersal during starvation 
. Nutritional status also modulates odor conditioning in the AWCs and salt conditioning in the ASEs 
. Interestingly, the ablation of sensory input has no apparent effect on reversal frequency on food, but dramatically alters reversal/turning off food, i.e., off food animals with compromised sensory input behave as if they are on food 
. Since, tph-1
null animals on food behave as if they are off food, the “serotonergic circuit” identified in the present study may also mediate other food-dependent locomotory transitions.
The present study has demonstrated that food availability is translated by complex changes in both monoaminergic and peptidergic signaling to modulate aversive responses mediated by the ASHs and highlights the advantages of this model system for dissecting nutritional modulation, as these same excitatory and inhibitory food signals most certainly also modulate other nutritionally-dependent behaviors, including attraction, feeding, locomotion, and egg-laying. Given the advantages of the C. elegans model, it should now be possible to fully dissect these nutritionally-sensitive signaling pathways in the modulation of individual neurons or circuits.