The researchers bred mice with orexin neurons that could be selectively and temporally destroyed by a toxin. This specific cell death was controlled by the presence or absence of a chemical added to the animals’ food. Mice lacking orexin neurons were found to be more sensitive to nociception than control mice, as demonstrated by pain-related behaviors such as withdrawing their paws from a hot plate or licking paws injected with a chemical irritant. Conversely, the artificial activation of orexin neurons significantly decreased such pain-related behaviors, suggesting its analgesic effect.

Selective removal of orexin neurons also reduced neurons expressing another chemical messenger, dynorphin, within part of the hypothalamus. “This indicates that the analgesic effect of activating orexin neurons could involve the combined action of orexin and dynorphin, or other messengers,” first author Ayumu Inutsuka says.

Development of a novel measurement system enabled the researchers to record real-time orexin neuronal activity. “We selectively engineered orexin neurons to express a calcium indicator that could be illuminated by an LED light source, and the resultant fluorescence was detected,” corresponding author Akihiro Yamanaka explains. “Strong mechanical stimuli applied to the mouse tail in the form of pinching with forceps brought an increase in the indicator signal. No such response was elicited when weaker stimuli were used, nor when the mice were anesthetized.”

Temperature-dependent increases of heat stimuli also induced a similar response, which again could not be detected under anesthesia, suggesting that anesthetics may function by inhibiting orexin neurons.