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In 1999, relying on results obtained from the Stanford colony, Mignot and his colleagues discovered that narcolepsy in Dobermans and other breeds results from a mutation in Hcrtr2, a gene that encodes a receptor for hypocretin, a neuropeptide produced in two forms in neurons of the dorsolateral hypothalamus and disseminated through the brain stem, spinal cord, and forebrain.
Still, the link from animal model to human behavior is seldom direct. Dobermans and Labradors have slightly different mutations of the Hcrtr2 gene, Mignot and his colleagues found. And humans appear to develop narcolepsy for a completely different reason: They simply don't produce hypocretins at all.
Interestingly, some dogs, such as Bear, also suffer from this form of narcolepsy. “Bear is especially interesting for us to keep for the future,” Mignot says. “Because he lacks hypocretin, like human patients, he could be an ideal model to test a new medication aimed at replacing hypocretins in humans.”
Earlier this year, Stanford University began disbanding its renowned colony of narcoleptic dogs, as functional genetic studies are more easily done in other models, and most of the dogs have been adopted. Lately, Mignot has been thinking that Bear will need a very special retirement home: the scientist's own.
Mignot suspects that, in humans with narcolepsy, hypocretin-producing cells are destroyed by an autoimmune process. To examine that possibility, he is isolating genes and proteins expressed in hypocretin-producing cells and testing them as autoantigens. He is also turning to an entirely new model: zebrafish. They have hypocretin and receptors, and narcoleptic zebrafish demonstrate abnormal sleep patterns similar to those in their human counterparts.
“The decision regarding which animal to use depends on the question you want to address,” says Mignot. “Screening hundreds of thousands of dogs or mice for genetic mutations is just not practical.” But zebrafish, which share a vast number of genes with humans, Mignot notes, can be easily screened in great quantities. Already Mignot has isolated a promoter gene that permits manipulation of zebrafish hypocretin cells in vivo.
That zebrafish doze at all may come as a surprise. But they are not the most esoteric model used to study sleep—that honor likely belongs to the fruit fly. For the last several years, HHMI investigator Amita Sehgal, a neuroscientist at the University of Pennsylvania School of Medicine, has relied on Drosophila melanogaster to help define the body's regulation of sleep.
“Sleep researchers were once of the view that sleep is restricted to mammals and a few birds,” Sehgal says. “But sleep is so important, you die if you don't get it. So why should it be restricted to just a few species? Sleep-like states are present in other organisms that we just haven't looked at yet.”
Simply ascertaining that flies sleep was no small task—it's not as if they close their eyes and snore. But Sehgal notes that fruit flies are immobile for long periods—during that time their threshold for arousal declines, one of the hallmarks of sleep—and these periods fall into regular circadian patterns. Also, like humans, the fly must compensate for any lost rest, which suggests a homeostatic function similar to sleep.
So if fruit flies sleep, what can they tell us of human slumber? In June, Sehgal and her colleagues published two studies demonstrating that a region of the fly brain, called the adult mushroom body, is responsible for regulating the animal's sleep patterns. Also involved in learning and memory consolidation, the adult mushroom body bears some semblance to both the human hippocampus and thalamus.
Humans spend about a third of their lives asleep, but nobody really knows why. Sehgal's finding provides evidence for the theory that sleep is necessary for consolidating memory. “In addition to doing other things, like replenishing energy stores and getting rid of the toxic by-products of metabolism, sleep may permit the nervous system to remodel itself and store information,” she says.