Joint Lab Publication: Novel Means to Quantify Physiological Sleepiness

For years, sleep researchers have characterized physiological sleepiness observed by EEG through one measure: slow wave activity. Slow wave activity is fairly ResearchBlogging.orgstraightforward to identify and represents EEG power at slower frequencies that essentially look like a roadside view of the Rocky Mountain Range. The amount of slow wave activity is directly proportional to the length of time that an animal is awake meaning that slow wave activity dominates the EEG after long periods of wake or sleep deprivation. While  slow wave activity has helped determine how a host of environmental, physiological, and genetic factors influence the ability to recover from sleep loss, it is limiting because it does not accurately quantify how an animal feels during sleep loss. And so, our lab teamed up with another electrophysiology lab that studies epilepsy to provide a means to quantify physiological sleepiness as it occurs, not after.

This was achieved through a period-amplitude analysis which looks at an individual EEG within a specific time frame, say 10 seconds, rather than all EEG waveforms within a specific time frame.   With this period-amplitude analysis, the number of slow wave peaks were counted across sleep loss with the mice being deprived of sleep for up to 24 hours. Early into the sleep deprivation, there were slow wave peaks here and there.

Early into Sleep Deprivation

But as time passed, particularly after the animal had been deprived of sleep for 6 hours, slow wave peaks began to dominate the EEG and persists until the end of 24 hours of sleep deprivation.
Late into Sleep Deprivation
We also found that the number of slow wave peaks was negatively correlated to the time it took the animal to fall asleep, in particular NREM sleep, which makes up 85-90% of total sleep and is the first type of sleep that an animal has unless the animal is narcoleptic. This relationship between slow wave peaks and time to NREM sleep corroborates the previously identified negative relationship between slow wave activity and time to NREM sleep.
Number of Slow Wave Peaks With Continued Sleep Deprivation
Finally, we found a time-dependent difference in the quantity of slow wave peaks based on whether the animal got sufficient sleep or was recovering from sleep loss. With sufficient sleep, slow wave peaks were more common during the light or rest/sleeping period of a nocturnal rodent. There were also more slow wave peaks during the middle of the night when most nocturnal rodents nap. This distinctive rhythm in slow wave peaks was absent in a mouse recovering from sleep loss. This is largely because there were more slow wave peaks during the night than that present with sufficient sleep.
Loss of Rhythm of Peaks After Sleep Deprivation

This study provides a means to better characterize changes in sleep and wake with inadvertent or voluntary sleep deprivation.

Ehlen JC, Jefferson F, Brager AJ, Benveniste M, & Paul KN (2013). Period-Amplitude Analysis Reveals Wake-Dependent Changes in the Electroencephalogram during Sleep Deprivation. Sleep, 36 (11), 1723-35 PMID: 24179307

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Tosini Lab Publication: Melatonin and Vision

Very recently, the Tosini lab published a paper in Science Signaling! Many congratulations! The paper investigated the impact of melatonin receptor cascades on eye function at physiological and molecular levels. Using a combination of transgenic mice and mutant cell lines coupled with electrophysiological recordings, pharmacology, histology, and wet lab work, it was found that the MT1 and MT2 receptors–of which melatonin binds to– form a heterodimer in the photoreceptors and this heterodimer mediates the action of melatonin on these cells.

ResearchBlogging.orgIn the first series of experiments, electrophysiological output from the retina was measured. The extent of retina activity was quantified by measuring two discrete, waveforms known as a and b waves. In mice lacking the MT2 receptor, the difference in electrophysiological output between the mid-night, when mice are active, and mid-day, when mice are typically asleep, was gone. The usual time-dependent difference could also not be restored by melatonin injections. This is indicates the importance of the MT2 receptor. A similar result was also found in mice lacking the MT1 receptor.

Next, an examination of structure and location of MT1/MT2 receptors was undertaken. It was found that MT1/MT2 receptors heteromize–expressed together–and are found primarily in the photoreceptors. What’s cool about this experiment is that an antibody for MT2 is not commercially available. To get around this, a fluorescent tag for MT receptors was inserted in the mice so that a typical immunofluorescent stain could be done.

In an additional series of experiments, the researchers also examined the impact of a mutant form of the MT2 receptor on formation of the MT1/MT2 heterodimer and photoreceptors  function in the presence an absence of melatonin. There again, eye function was disrupted. Finally, the researchers investigated the intracellular signaling cascades that are activated by the MT1/MT2 heterodimer in the photoreceptors.

In all, the study strengthens the argument that melatonin has a significant impact on photorecepto function. In the world of biomedical research, the melatonin cascade may be an ideal target for the treatment of vision problems.

Baba K, Benleulmi-Chaachoua A, Journé AS, Kamal M, Guillaume JL, Dussaud S, Gbahou F, Yettou K, Liu C, Contreras-Alcantara S, Jockers R, & Tosini G (2013). Heteromeric MT1/MT2 Melatonin Receptors Modulate Photoreceptor Function. Science signaling, 6 (296) PMID: 24106342