In this study, we explored the causal relationship between cell type-specific neuron activation and feeding behavior for AGRP and POMC neurons in the hypothalamic arcuate nucleus. Our results show that simple stimulus patterns in AGRP neurons are sufficient to rapidly induce the complex behavioral sequences required to seek and consume food. AGRP neuron stimulation was not simply a trigger for feeding; instead, we report multiple lines of evidence indicating that the level of AGRP neuron activity is integral to the magnitude, dynamics, and duration of evoked feeding. We find greater food consumption and a decreasing latency to eat with increasing numbers of photoexcitable neurons. Also, food intake is reduced as the stimulus frequency is decreased, consistent with a graded behavioral response to different neuron activities. Finally, continuation of evoked feeding requires ongoing stimulation of AGRP neurons, which suggests that activity in downstream circuits is tied to AGRP neuron signaling during the behavior. Such measurements were not previously accessible without genetically encoded tools for cell type-specific, temporally precise manipulation of neuron activity. Together, these experiments show a close relationship of AGRP neuron signaling with downstream circuits that influence the diverse sensory, motor, and motivational components that underlie this behavior.
Selective stimulation of POMC neurons gave the opposite behavioral outcome, reducing food intake and body weight over 24 hours of continuous photostimulation, an effect which required signaling through melanocortin receptors. However, in contrast to pharmacological studies with melanocortin receptor agonists26,27
, POMC neuron stimulation for two hours straddling dark period onset did not reduce food intake. This difference likely stems from the constraints associated with activation of intrinsic neurons as compared to intracranial injection of a neuromodulator. Specifically, the axon projections of a neuron and the natural expression level of an endogenous neuromodulator limit its distribution and the amount available to interact with downstream receptors. Another aspect of selective neuron activation is that it allows the contribution of other co-released substances to be measured. For POMC neurons, no effect on ad libitum
feeding behavior was observed with melanocortin receptors blocked in Ay
mice, indicating that co-released modulators and neurotransmitters were not sufficient to inhibit food intake under our experimental conditions.
Because the melanocortin pathway regulates feeding behavior, AGRP suppression of POMC-derived melanocortin signaling has been widely considered as a pathway through which these neurons could rapidly increase feeding behavior11
. However, although these cell populations interact, we find that inhibition of melanocortin receptors is not necessary for an acute AGRP neuron-evoked feeding response. While these photostimulation experiments do not exclude a role for this pathway to regulate food intake over time scales longer than we have investigated here, they demonstrate the capability of AGRP neurons to independently engage other downstream circuits that coordinate feeding. Future experiments could focus on the signaling pathways necessary for evoked feeding, which are likely mediated by other co-released components of AGRP neurons such as the neuromodulator NPY and the fast neurotransmitter γ-aminobutyric acid14
A striking property of AGRP neurons was the capacity for simple photostimulus patterns to drive feeding behavior selectively without training or prior exposure to AGRP neuron stimulation. Previous experiments have shown that simple activity patterns played into cortical neurons could be used to train a behavioral task or activate a learned behavior28,29
. In other studies, stimulation of molecularly-defined populations conditioned a place preference30
, reduced latency to waking31
, and elevated respiration32
. However, the capability of a small molecularly-defined neuron population to generate a goal-directed behavior without training has not been previously described. One explanation for this result is that AGRP neurons have a dedicated role for controlling feeding behavior and that these exogenously applied activity patterns can substitute for patterns observed during normal activation of these neurons, for example in a state of energy deficit. While natural activity patterns in behaving animals are not known for AGRP neurons, optogenetic techniques allowed us to probe the capacity of these neurons and their downstream signaling pathways to influence feeding behavior, which showed that a range of activity patterns were sufficient to activate a proportional feeding response.
Thus, these neurons, which transduce circulating signals of metabolic state such as leptin, ghrelin, and glucose33
into electrical activity, act as interoceptive sensory neurons which can control feeding behavior and may contribute to mediating the internal representation of hunger. Analogous to externally oriented sensory systems where anatomical and functional properties of sensory neurons have facilitated elucidation of neural pathways responsible for sensation and perception, the axon projections of AGRP neurons34
may guide the identification of the motivational, hedonic, and autonomic circuits that underlie control of feeding behavior. Because AGRP neurons also provide a genetically-accessible entry point into these circuits, we propose that new genetically encoded tools for rapid manipulation of neuronal and synaptic function could be applied to dissect these discrete pathways under normal conditions as well as in pathological states of over- and under-eating.