Four Amino Acids, Four Different Brain Routes to Appetite Control in Layer Chicks

Four common amino acids steer the appetite of young layer-type chicks through strikingly different brain pathways, according to a new study published in Frontiers in Physiology. When researchers injected L-alanine, L-glutamine, L-arginine hydrochloride, or glycine directly into the brain ventricles of five-day-old chicks, three of the four compounds suppressed feeding within two hours, while one actually stimulated it. Crucially, the team found that these behavioral outcomes were mirrored by distinct patterns of gene expression in the hypothalamus, the brain region that acts as a central command center for hunger and satiety signals.

The research, conducted at Shandong Agricultural University in China with collaborators from the University of Lome in Togo, the University of Tehran, and other institutions, addressed a persistent gap in poultry physiology. Although the effects of these amino acids on feed intake had been studied in broiler chicks, their impact on hypothalamic neuropeptides in layers remained unclear. This distinction matters because, according to the authors, long-term selective breeding has produced measurable differences between broiler and layer chickens in how their central nervous systems regulate energy balance, meaning that each strain should be studied individually rather than assuming that findings from one apply to the other.

To deliver the amino acids directly to the brain, the team used intracerebroventricular injection, a technique in which a tiny volume of solution is introduced into the third ventricle of the brain, immediately adjacent to the hypothalamus. This approach bypasses the gastrointestinal tract entirely, allowing researchers to isolate the central neurochemical effects of each amino acid from its peripheral metabolic roles. Each chick received just 10 microliters of solution, with doses of 1, 2, or 3 micrograms depending on the treatment group. The solutions were carefully adjusted to a physiological pH of 7.4 and a calculated osmolarity of 308 to 312 milliosmoles per liter, ensuring that any observed effects could not be attributed to osmotic disturbance or pressure changes within the brain.

A small amount of Evans Blue dye in every solution allowed the researchers to visually confirm, after the experiment, that the injected fluid had actually reached the hypothalamic region; all experimental chicks passed this verification. The subjects were 128 one-day-old male layer chicks of the Hyland Brown strain, obtained from a local incubator. Only males were used to minimize sex-related biological variation during the neonatal period, a stage at which male and female chicks show broadly similar feeding-related physiology. The chicks spent their first two days in a group cage at 33 degrees Celsius and 60 percent humidity, then moved at three days of age into individual cages measuring 42 by 35 by 11 centimeters. Individual housing allowed precise recording of each bird’s feed consumption while eliminating social interactions that could confound intake measurements.

On the day of injection, food was removed at 8 a.m. for a three-hour fast, ensuring that all chicks began the trial under identical conditions. Feed intake was then measured cumulatively at 30 minutes, one hour, and two hours after injection, and expressed relative to body weight to account for size differences among birds. Two hours after injection, each chick was euthanized and the hypothalamus was collected and frozen for RNA extraction. Gene expression of four appetite-related targets was quantified by RT-PCR: neuropeptide Y (NPY) and agouti-related peptide (AgRP), both of which stimulate appetite, and proopiomelanocortin (POMC) and the melanocortin 4 receptor (MC4R), key components of the satiety-promoting melanocortin axis. Each amino acid treatment was compared only with its own control group, because gene expression was measured at a single time point.

The results revealed a remarkable diversity of mechanisms. L-alanine at 1 and 2 micrograms significantly reduced feed intake at the two-hour mark, and gene expression analysis at the 1-microgram dose showed a significant decrease in NPY messenger RNA, one of the most potent appetite-stimulating peptides in birds. POMC and MC4R expression rose only slightly and not significantly, while AgRP remained unchanged. This pattern suggests that L-alanine curbs feeding primarily by dampening the central hunger signal rather than by amplifying satiety. The authors interpret this as evidence that alanine may act as a hypothalamic energy sensor, mimicking a state of nutritional abundance and selectively signaling to NPY neurons to suppress food-seeking behavior. Notably, in broilers, the same amino acid’s intake-reducing effect has been associated with decreased AgRP and increased MC4R expression, so the underlying mechanisms may differ between the strains even when the behavioral outcome is similar.

L-glutamine behaved in the opposite direction. At a dose of 3 micrograms, it significantly increased feed intake at 60 minutes after injection, yet none of the four measured neuropeptide genes showed statistically significant changes, and there were even slight trends in the opposite direction of what a transcriptional mechanism would predict. The researchers propose that glutamine’s feeding-stimulatory effect is likely mediated by rapid synaptic neurotransmission rather than by gene transcription. As a direct metabolic precursor of glutamate, a fast-acting excitatory neurotransmitter with orexigenic activity in the hypothalamus, glutamine could influence feeding through central glutamate and GABA pool dynamics within minutes, well before any change in gene expression could occur. In broiler chicks, by contrast, glutamine’s stimulatory effect has been associated with increased NPY and decreased POMC expression, meaning the two strains can arrive at similar behavioral outcomes through different hypothalamic routes.

L-arginine hydrochloride produced the most comprehensive transcriptional response. At just 1 microgram, it significantly reduced feed intake at two hours while simultaneously increasing POMC and MC4R expression and decreasing NPY expression, with AgRP showing a slight, non-significant decline. This dual action, activating the anorexigenic melanocortin pathway while suppressing the orexigenic NPY network, represents a coordinated shift of the hypothalamic balance toward satiety. The authors hypothesize, though they did not measure it directly, that arginine’s effect may involve strong local production of nitric oxide, since arginine is the sole substrate of neuronal nitric oxide synthase in chicks. Nitric oxide could act as a retrograde messenger capable of both activating POMC and MC4R neurons and inhibiting NPY neurons. Consistent with earlier reports, central arginine reduced intake in layers but had shown no effect in broilers, further underscoring the strain-specific nature of central amino acid signaling. The authors also note that the dose-response relationship was limited because only a few concentrations were tested.

Glycine, tested at 3 micrograms, was the most consistent suppressor of feeding, significantly reducing intake at every measured time point. Its gene expression profile was distinctive: POMC was unaffected, MC4R was significantly increased, and both NPY and AgRP trended downward without reaching statistical significance, with the NPY reduction larger than that of AgRP. The authors conclude that glycine’s effect operates primarily through sensitization of melanocortin receptor signaling rather than through changes in neuropeptide levels. In poultry, central glycine interacts with N-methyl-D-aspartate receptors and the adrenergic system, and the observed MC4R upregulation may reflect synaptic convergence whereby glycine potentiates satiety circuits independently of POMC transcription. This finding aligns with previous work showing that glycine reduces feed intake in both broilers and laying hens.

The study also carries an important methodological caveat regarding dose interpretation. Because the four amino acids differ substantially in molecular mass, from 75.07 grams per mole for glycine to 210.66 grams per mole for arginine hydrochloride, identical mass doses delivered very different molar loads. At the 3-microgram dose, glycine provided 39.96 nanomoles while arginine hydrochloride provided only 14.24 nanomoles, a nearly threefold difference. The fact that higher-molecular-weight amino acids produced significant effects despite much smaller molar loads suggests greater sensitivity or receptor affinity of avian neuronal networks, such as the nitrergic and melanocortin systems, for these molecular structures, whereas glycine’s effect may depend partly on its higher molar density activating ionotropic glycine receptors and NMDA receptors more broadly. The authors therefore caution that their results should be read as an assessment of behavioral efficacy under mass-based dosing, not as a comparison of intrinsic stoichiometric affinity for a single receptor.

All findings in the study are strictly acute, measured within two hours of injection and at a single time point for gene expression, so long-term effects remain unexplored. Only messenger RNA levels were quantified, leaving the upstream signaling events and protein-level changes unresolved, and the authors acknowledge that peripheral responses downstream of the central administration cannot be ruled out despite the bidirectional communication between brain and body. Nevertheless, the authors argue that the work provides proof of concept for nutritional strategies in poultry production. If central amino acid signaling can be translated to oral delivery, glutamine-rich pre-starter or in-ovo supplements might stimulate early post-hatch feeding, while arginine and glycine could help manage intake under stressful conditions. Future studies, the team notes, must evaluate dietary routes of administration and long-term effects on growth performance before these central mechanisms can inform commercial feeding practice.

Subject of Research: Central effects of four amino acids on feed intake and hypothalamic neuropeptide expression in layer-type chicks

Article Title: Differential effects of L-alanine, L-glutamine, L-arginine HCl, and glycine on feed intake and hypothalamic appetite-related responses in layer-type chicks

Article References: Ghashghaei, E., Mijiyawa, A., Riaz, M. F., Wang, M., Li, K., Wang, D., Okai, M. A., Maa Maa Temhoul, C., Nadiedjoa, Y., Rahmani, B., Obianwuna, U. E., Zendehdel, M., & Lin, H. (2026). Differential effects of L-alanine, L-glutamine, L-arginine HCl, and glycine on feed intake and hypothalamic appetite-related responses in layer-type chicks. Frontiers in Physiology, 17, 1983120. https://doi.org/10.3389/fphys.2026.1983120

Image Credits: AI Generated

DOI: 10.3389/fphys.2026.1983120

Keywords: amino acids, feed intake, hypothalamus, intracerebroventricular injection, layer chicks, neuropeptide Y, POMC, MC4R, L-glutamine, L-arginine, glycine, avian physiology