What Happens While We Sleep? The Cellular Intersection in the Sleep-Epilepsy-Cerebellum Relationship

Since ancient Greece, the relationship between epilepsy and sleep has been observed with great interest. Aristotle noted the link between the frequency of epileptic seizures and sleep, and that, occasionally, some people only convulse during sleep. Another interesting observation dating from that time is that sleep deprivation increases the likelihood of epileptic seizures. It was evident at that time that sleep appeared to be a “seizure” similar to epilepsy, as the body disconnects from its conscious state. But what areas of the brain are involved in these seemingly distinct phenomena, and how does this occur at the cellular level?

For many years brain electrical activity has been detected through electroencephalographic recordings. This has allowed researchers to determine that the exacerbated neuronal activity that occurs during epileptic seizures is very similar to what occurs in certain sleep states. Key molecular components for neuronal function are ion channels, proteins that form pores in cell membranes allowing the selective passage of ions and responsible for communication between neurons.

THE EXACERBATED NEURONAL ACTIVITY THAT OCCURS DURING EPILEPTIC SEIZURES IS VERY SIMILAR TO WHAT OCCURS IN CERTAIN SLEEP STATES

TRANSGENIC FISH
In UNAM’s Institute of Neurobiology’s Molecular and Cellular Neurobiology Laboratory we have studied the functional and pharmacological characteristics of ion channels and found that some mutations in the genes that code for these ion channels cause changes in neuronal activity that, when introduced into experimental animals, make them susceptible to epileptic seizures. The model organism we used for our experiments is the zebrafish (Danio rerio). We managed to manipulate the species genome to induce mutations in the ion channel genes and found that these modified fish have seizures from the first days of their development, manifested by uncontrolled body movements followed by “absences.” To understand whether these are changes in neuronal activity and not just changes in movement, we developed an electroencephalographic system for fish, which allowed us to determine that the changes are indeed manifested by alterations in the electrical characteristics of neurons.

The larvae of these fish are transparent during the first days of development, which occurs in just a few days. Thanks to this characteristic, we can observe fluorescent molecules using high spatial and temporal resolution microscopy in genetically modified fish.

At Stanford University, California, United States, important technological advances have been developed in the application of genetically encoded fluorescent calcium detectors and “light sheet” microscopy for zebrafish. These advances have potentiated the study of sleep as a global activity of brain function. The technology, called fluorescence polysomnography (fPSG), allows simultaneous recording of neuronal activity, heart rate, eye movements, and muscle activity in transgenic fish.

Thanks to these experimental approaches, it has been demonstrated that the neuronal characteristics of sleep in humans are similar to those in fish, suggesting that they are functional properties of the brain that emerged millions of years ago and are conserved in most vertebrates.

Given these new findings on sleep physiology, our observations of the functional role of ion channels in controlling neuronal activity in the brain, and the induction of epileptic activity when ion channel genes are mutated, we began a collaboration with researchers at Stanford University to understand whether sleep activity is affected in our mutant fish and to determine which brain areas are compromised.

NEURONAL CHARACTERISTICS OF SLEEP IN HUMANS ARE SIMILAR TO THOSE IN FISH, SUGGESTING THAT THEY ARE FUNCTIONAL PROPERTIES OF THE BRAIN THAT EMERGED MILLIONS OF YEARS AGO

EXPERIMENTAL OBSERVATIONS
The first observations in our experiments have revealed interesting processes. First, we observed that our mutant fish are highly mobile and die more frequently during the night. Although we do not yet know whether this is the result of an epileptic seizure or of another phenomenon, this observation seems to reinforce what has long been known: that seizures very frequently occur during sleep. Second, an unexpected finding was the observation that our mutants have heigh neuronal activity in the cerebellum, an area of ​​the brain that controls and coordinates our movements, but very little is known about its contribution to sleep regulation or whether it plays a role in sleep disorders. Elucidating whether the cerebellum plays a role in controlling the sleep-wake cycle has potential clinical and therapeutic implications for sleep disorders and epilepsy.

The cerebellum is highly organized and contains the largest number of neurons of any brain region. There are millions of inhibitory and excitatory neurons and small local circuits that maintain the proper balance to perform its function. Our next steps will require a very detailed interpretation that must be validated with further light-sheet microscopy observations, in addition to electrophysiological recordings of individual cells or even groups of neurons. The cerebellum has connections with other brain regions involved in the control of cardiovascular function and breath, indicating that its dysfunction could contribute to sleep-related cardiorespiratory problems.

These experiments have established a new working hypothesis for future studies and show that much remains to be learned about the cerebellum’s involvement in sleep and epilepsy.

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