Abstract
ObjectiveDietary proteins are sensed by hypothalamic neurons and strongly influence multiple aspects of metabolic health, including appetite, weight gain, and adiposity. However, little is known about the mechanisms by which hypothalamic neural circuits controlling behavior and metabolism sense protein availability. The aim of this study is to characterize how neurons from the mediobasal hypothalamus respond to a signal of protein availability: the amino acid l-leucine. MethodsWe used primary cultures of post-weaning murine mediobasal hypothalamic neurons, hypothalamic neurons derived from human induced pluripotent stem cells, and calcium imaging to characterize rapid neuronal responses to physiological changes in extracellular l-Leucine concentration. ResultsA neurochemically diverse subset of both mouse and human hypothalamic neurons responded rapidly to l-leucine. Consistent with l-leucine's anorexigenic role, we found that 25% of mouse MBH POMC neurons were activated by l-leucine. 10% of MBH NPY neurons were inhibited by l-leucine, and leucine rapidly reduced AGRP secretion, providing a mechanism for the rapid leucine-induced inhibition of foraging behavior in rodents. Surprisingly, none of the candidate mechanisms previously implicated in hypothalamic leucine sensing (KATP channels, mTORC1 signaling, amino-acid decarboxylation) were involved in the acute activity changes produced by l-leucine. Instead, our data indicate that leucine-induced neuronal activation involves a plasma membrane Ca2+ channel, whereas leucine-induced neuronal inhibition is mediated by inhibition of a store-operated Ca2+ current. ConclusionsA subset of neurons in the mediobasal hypothalamus rapidly respond to physiological changes in extracellular leucine concentration. Leucine can produce both increases and decreases in neuronal Ca2+ concentrations in a neurochemically-diverse group of neurons, including some POMC and NPY/AGRP neurons. Our data reveal that leucine can signal through novel mechanisms to rapidly affect neuronal activity.
Highlights
Dietary proteins strongly influence metabolic health via their effect on appetite, weight gain, and adiposity [1e3]
We assessed the responsiveness of these cultures to glucose and leptin, two metabolic signals sensed by mediobasal hypothalamic neurons
We found that a transition from 2.5 mM to 5 mM glucose triggered an increase in intracellular Ca2þ concentration [Ca2þ]i in 25% of neurons in culture, and a decrease in [Ca2þ]i in 4.75% of neurons (Figure 1DeF), consistent with previous observations [30,31]. 10 nM Leptin increased [Ca2þ]i in 15% of neurons, and decreased [Ca2þ]i in 6.7% of neurons, consistent with previous reports (Figure 1GeI) [32]
Summary
Dietary proteins strongly influence metabolic health via their effect on appetite, weight gain, and adiposity [1e3]. In humans and rodents, circulating leucine levels rapidly increase following the ingestion of a protein dense meal [8e10], and consumption of a protein-diluted diet chronically reduces serum leucine levels [11]. It is well documented in the literature that leucine signals protein abundance to regulate key anabolic functions including muscle protein synthesis [12] and beta-cell insulin release [13] at least in part via mTORC1 signaling, a pathway for which leucine is the main activator [14]. Protein sources with higher anabolic value (assessed by their ability to promote growth or activate TOR signaling) or rich in branched-chain amino acids produce
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