Glowing green brains in Petri dishes may sound like science fiction, but in Yoshitaka Oka's lab at the University of Tokyo, Japan, they are more like fine art. In a study recently published in Endocrinology, his lab used transgenics – where specific bits of DNA are added to an organism in order to study the function of a gene – to make the brains of medaka fish fluoresce green wherever a protein called kisspeptin is made. Kisspeptin is produced by neurons in discrete regions of the brain and these neurons release the protein to communicate with other neurons in the brain that, in turn, regulate many aspects of reproduction, including sexual maturation. In other words, kisspeptin begins the conversation between the brain and the gonads. While kisspeptin is known to be essential for puberty in mammals, its role in regulating the reproductive cycles of seasonal breeders such as medaka, is not known, so Oka and his colleagues used their transgenic medaka to figure this out.
Knowing that medaka that are raised under 14 h light/10 h dark conditions become sexually mature whereas fish grown in 10 h light/14 h dark conditions do not, the scientists raised their transgenic medaka under the two different lighting conditions to produce a group of sexually mature fish and another group that was not sexually mature. Next, they carefully removed the brains and placed them in dishes. The team could easily see neurons expressing kisspeptin glowing green by looking at the brains under a fluorescent microscope. Then, they recorded the kisspeptin neurons in action using an electrophysiology technique called whole-cell patch-clamping and compared the neuron activity in the two groups of fish. Oka's team found a clear difference between the kisspeptin neurons of mature and immature fish. In mature fish that were ready to breed, kisspeptin neurons glowed brighter, meaning more kisspeptin was being made, and showed higher firing frequencies compared with kisspeptin neurons in immature fish. Since a high firing frequency, or activity level, is required for a neuron to release its peptides and communicate with other neurons in the brain, the team concluded that more kisspeptin is made and released in sexually mature fish.
In a second set of experiments, Oka's team wanted to understand how these kisspeptin neurons are regulated during a breeding cycle. They raised transgenic female medaka to sexual maturity and then surgically removed their ovaries – to effectively end the breeding cycle – and allowed the fish to recover. Since ovaries are the major source of the female sex hormone, estrogen, Oka's team supplemented half of the recovering fish with estrogen while the other half received none. After 3 days of recovery, the team removed the brains and analyzed the kisspeptin neurons as before. They found that the kisspeptin neurons of mature females that had their ovaries removed resembled those of immature fish – they glowed faintly and were mostly inactive, so little kisspeptin was made and released by these neurons. In comparison, giving female fish estrogen after their ovaries were removed returned the kisspeptin neurons to an expression and activity level reminiscent of sexually mature fish.
So, Oka and his crew showed that the production and release of kisspeptin in the medaka brain coincides with sexual maturation and the activity of kisspeptin neurons is sensitive to changes in estrogen. Their results support a conserved role for kisspeptin in regulating sexual maturation among vertebrates with different types of reproductive cycles. Now pucker up and go smerch a perch!
