Nature research paper: Wake-like skin patterning and neural activity during octopus sleep
) were made by placing an 8K camera fitted with a Canon Macro Lens EF 180 mm lens on a gimbal, and filming top-down on a single animal using a 45° mirror . Three white bar lights were used to light the tank. All recordings were made using the following camera settings: 1/200, F32, ISO 3,200. Because AS events are periodic, AS recording was started a few minutes before the expected AS time, and ended after AS pattern dynamics ceased.
To put lighting levels in context, daylight ranges from 10 to 25 klx when sunny, 1,000 lx when overcast. Nighttime light levels range from roughly 0.3 lx during a full moon to 0.002 lx when the moon is not visible. Animal sleep time, AS duration and interval seemed normal under various experimental lighting conditions . Darker skin patterns did not appear under 3 klx lighting while awake, but were observed under 1 klx lighting.
For sleep homeostasis, octopuses were recorded continuously for 48 h before the start of sleep deprivation. The following day, animals were kept awake from 07:00 to 17:00 by gently brushing their skin with a paintbrush every 2–3 min. Movement, elevated posture and eye opening were considered indicators of wakefulness. Animals were left to behave freely after 17:00. Sleep deprivation was then repeated, using the above procedure, for a second day.
For measuring circadian rhythm, octopuses were acclimated to a 12-h light/dark cycle for 2 weeks. After 48 h of continuous filming, they were subject to either 72 h of continuous daylight or 72 h of continuous darkness. Feeding was halted during this period to prevent cues from live crabs. Animals were then switched back to a 12-h light/dark cycle.
For measuring temperature dependence, the temperature of the water circulating in the behavioural filming system was cooled to 19, 22 and 24 °C using a cooler attached to the water circulation system. Each temperature was maintained for 2 days. Higher temperatures were achieved by turning off the cooler. Tank water temperature was measured using a temperature sensor .
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