The timing of puberty involves the brain’s regulation of hormones in the body. Could environmental chemicals stimulate the receptors for those signals? In 2024, Shu Yang and colleagues screened a large number of compounds in cells, then investigated candidates in more detail using human-derived neurons and fish.
One candidate was musk ambrette, a synthetic fragrance ingredient. However, it is not representative of modern perfumes as a whole. The International Fragrance Association (IFRA) prohibits its use as a fragrance ingredient on the grounds of photosensitization and neurotoxicity. It must be distinguished from natural ambrette seeds and from other musk odorants.
- Two receptors involved in regulating puberty
- Narrowing down candidates from approximately 10,000 compound entries
- Musk ambrette increased expression of a GnRH-related gene
- In fish, the fluorescent area associated with relevant neurons expanded
- Another pathway also yielded candidates for investigation
- Effects on human puberty remain to be established
- Summary
- References
Two receptors involved in regulating puberty
Receptors are proteins that receive specific signals and translate them into cellular responses. One target in this study was KISS1R, which receives signals from a substance called kisspeptin. It regulates the activity of neurons that produce the hormone GnRH in the brain’s hypothalamus.
The other was GnRHR, which receives the GnRH signal. Reception of this signal in the pituitary gland sets in motion the regulation of other hormones involved in reproduction. By searching for chemicals that stimulate these two receptors, the researchers hoped to find clues to the phenomenon of earlier puberty in girls.
Narrowing down candidates from approximately 10,000 compound entries
The researchers used the Tox21 10K library, a collection of pharmaceuticals and environmental chemicals. It contains approximately 10,000 entries, representing 8,312 unique compounds after duplicates are excluded. This was neither a study of 10,000 people nor a study of 10,000 fragrance ingredients.
Each compound was added at 15 concentrations to cultured cells engineered to produce high levels of the human receptors, with three independent screening runs. The initial readout was a change in intracellular calcium, used to detect a rapid response associated with receptor stimulation.
However, calcium can change for reasons other than an effect on the intended receptor. The researchers therefore compared the results with cells that had not been given additional receptors and checked candidates using another signaling readout. At this confirmation stage, some substances that initially produced a response were not judged to act specifically on the target receptor.
Musk ambrette increased expression of a GnRH-related gene
After these checks, the findings supported musk ambrette as a candidate that stimulates KISS1R. The researchers then examined responses downstream of the receptor in mouse hypothalamic cells and hypothalamic neurons made from human induced pluripotent stem cells.
In the human-derived neurons, adding musk ambrette at 25 micromoles per liter temporarily increased messenger RNA for the gene that produces GnRH. Messenger RNA carries genetic information into the process of protein production. The response was seen at four hours and subsequently returned to baseline.
This result indicates a change in the activity of a gene involved in making a hormone. It was not a measurement of the amount of hormone actually secreted or the timing of puberty in humans. Nor can a concentration applied directly to cells be translated directly into an amount of perfume used or a concentration reaching a target inside the body.
The study also used calculations based on molecular three-dimensional structures to explore where musk ambrette might fit within the receptor. Such modeling informs hypotheses about binding, but is not direct observation proving that the molecule bound at that location.
In fish, the fluorescent area associated with relevant neurons expanded
The researchers next used zebrafish in which GnRH-related neurons could be visualized by fluorescence. They raised the fish in water containing musk ambrette from one to five days after fertilization and measured the fluorescent area in the brain.
The concentrations tested were 0.1, 0.3, 0.5, and 1 microgram per milliliter. No significant difference in fluorescent area was detected at 0.1, but the area was larger than in controls at concentrations of 0.3 and above. Analysis of gene expression in the head also showed increases in GnRH-related RNA and other transcripts at the highest concentration.
This area measures the extent of the fluorescently labeled region. It is not a direct measurement of neuron number, hormone secretion, or the age at which the fish mature. Together with the cell experiments, it supports the possibility of effects on the developing neuroendocrine system, but does not by itself demonstrate earlier puberty.
Developmental delay and abnormal swim-bladder formation were also observed in the fish. The conditions therefore cannot be described as a harmless low exposure that changed only one specific pathway.
Another pathway also yielded candidates for investigation
The paper was not solely a study of musk ambrette. A group of substances with acetylcholine-like activity also showed activity at GnRHR. The researchers examined changes in genes involved in downstream signaling in cells including those derived from the mouse pituitary.
Many substances in this group are used as pharmaceuticals; the finding does not mean they are all everyday fragrance ingredients. The authors emphasized the possibility of activity at reproductive receptor pathways in addition to the compounds’ known effects. They proposed studying structurally similar environmental chemicals that were not included in the library.
Effects on human puberty remain to be established
The authors proposed the hypothesis that musk ambrette and related nitro-musks might contribute to earlier puberty through KISS1R. At the same time, they explicitly stated that their findings in cells and fish were not direct evidence of an effect on puberty.
Separate research is needed to establish which substances people actually encounter, the extent of that exposure, the concentrations reaching targets within the body, and how these relate to developmental timing. Receptor function and regulation may also differ between animals and humans. The authors therefore identified additional human-cell studies and epidemiological research tracking exposure and development as next steps.
The work received support from US NIH research programs and other sources, and the authors declared no conflicts of interest to disclose.
Summary
This study screened many compounds to identify candidates that stimulate receptors involved in regulating puberty, then examined musk ambrette further through changes in cellular gene expression and neuron-associated regions in fish. Its value lies in checking the possibility of activity with different methods, rather than merely listing substances that reacted in a single test.
The result is not a definitive list of ingredients that accelerate human puberty, but clues to compounds and pathways deserving closer investigation. The next stage is to connect nitro-musks, cholinergic substances, and related compounds with studies of actual exposure and development.
References
Yang, S., et al. (2024). Identification of Environmental Compounds That May Trigger Early Female Puberty by Activating Human GnRHR and KISS1R. Endocrinology, 165(10), bqae103. DOI: 10.1210/endocr/bqae103 / Full original article. As a work by US government employees, the original article is in the public domain in the United States.
International Fragrance Association. IFRA Standard: Musk ambrette (CAS 83-66-9). Specifies the prohibition on use as a fragrance ingredient and the reasons for it.
