African cichlids, a group of thousands of freshwater fish species, present a wide range of distinctive mating and parental care behaviors. However, the mechanisms that drive these behaviors are poorly understood. Cheng-Yu Li, assistant professor of biological sciences and a member of the UM FIRST faculty cohort, and colleagues have discovered a new clue in this mystery and published their results in Current Biology. A wide range of vertebrate species exhibit versions of these behaviors, so Li and colleagues hope their research will contribute to a broader understanding of the neural mechanisms that are behind them.
Using an African cichlid species, Astatotilapia burtoni, as the model organism, the authors identified that a group or neurons called ciliated olfactory sensory neurons (OSNs) are the key type of cell responsible for detecting female reproductive pheromones. They further pinpointed a specific pheromone receptor, Or113a, that regulates males’ reproductive behavior.
In male cichlids, sensory neurons expressing the Or113a receptor detect cues derived from a pheromone called prostaglandin F2ɑ, or PGF2ɑ. PGF2ɑ functions both as a female hormone that regulates female mating behavior and as a pheromone that attracts males when females are reproductively receptive. Knocking out their Or113a receptors limited males’ preference for water taken from near a sexually receptive female. Silencing the males’ relevant sensory neurons by knocking out a protein called Cnga2b eliminated the preference entirely.
PGF2ɑ on its own is attractive to males in some fish species, such as zebrafish and goldfish, however, but male A. burtoni cichlids are insensitive to PGF2ɑ. This suggests that PGF2ɑ must be converted into a different chemical in reproductive females or that more than one pheromone is involved in the male attraction response. That makes evolutionary sense, Li explains, because many species of cichlids and other fish living in the African Great Lakes may use PGF2ɑ for pheromone signaling. If it was the only signal, there would be a high probability of interbreeding between cichlid species, or a male could mistakenly approach females of predator species. Using more than one signal would solve this problem. Li and colleagues are actively working on finding additional pheromone clues to further unravel this complex signaling system.
A puzzle worth brooding over
Discovering a neurological mechanism driving males’ attraction to females was exciting for Li, but an unusual observation during their trials added to the study’s impact. Many cichlids “mouth brood,” or carry their eggs in their mouths for several weeks until the larvae can swim on their own. In most species, including the one used in Li’s study, the female is the exclusive mouth brooder. However, Li’s study found that males with mutations in the genes of cnga2b or or113a would take eggs into their mouths as a female would. But only 30 percent of the males who picked up eggs would mouth brood them until hatching; the other 70 percent immediately swallowed the eggs.
Li and colleagues had expected the males with mutated receptors to alter their preference for water containing female pheromones, but the mouth brooding was a surprise. “That was kind of outside of our expectations,” Li says. “It’s something that we had never heard before, and we didn’t even predict that would happen.”
Left: A female A. burtoni holds eggs in her mouth. Right: A male A. burtoni, the cichlid species Li studies, carries eggs in his mouth and ignores nearby food pellets. (Photos courtesy of Li)
To explain the male mouth brooding, “Our first guess is that the PGF2ɑ-derived pheromone cues play dual roles,” Li says. “The first role is to attract males during mating. But it may also inhibit male mouth-brooding behavior. So once you knock out this receptor, this circuit turns on in the males, and they become mouth brooders as well.” In other words, mouth brooding may be the default behavior in both species, but some mechanism driven by pheromones normally turns it off in males.
Supporting this hypothesis, Li and his colleagues found that in blackchin tilapia, a different cichlid species where males normally mouth brood the eggs until hatching, the males may have a naturally occurring mutation in the same Or113a gene. That supports the idea that the phenomenon of male mouth-brooding, and the mechanism that regulates it, “ is a regular mechanism in the field, not just some weird example we saw in the lab,” says Li.
The next question is what additional cues may be required to induce full-term mouth brooding. Li and colleagues are investigating.
Connections across species
Cichlids have a complex social status system, demonstrate significant parental care, and present a diverse set of distinctive mating behaviors, traits they have in common with a wide range of species. Even some of the mechanisms are shared across large evolutionary distances. For example, the hormone prolactin drives parental care in fishes, mice, and even humans.
“We are using this simple model in cichlids, which are easy to breed and easy to study in the lab, and exploring mechanisms that may have broader biological relevance. It’s a wonderful model where we can bridge ecology and evolution concepts with neuroscience,” Li says. “By gaining deeper insights into what regulates parenting behavior, we can identify key neurons and genes in the brain involved in this process, with the goal of uncovering mechanisms that may be shared across various species.”
