Forsøksdyr: Neural mechanisms of wakefulness, sleep and general anesthesia in rodents

Godkjenningsdato 23.09.2020

Godkjenningsperiode 23.09.2020-22.09.2024

The proposed experiments will explore the neural mechanisms that are involved in the generation of conscious experience during wakefulness, dreams during rapid-eye-movement (REM) sleep and ketamine anesthesia and unconsciousness in non-REM (NREM) sleep and general anesthesia. Experiments will focus on both cellular and circuit levels. We will use methods and equipment previously developed (FOTS 9781, 11812) to measure EEG or ECoG in rodents, to assess the complexity of cortical activity, which is associated to different conscious states (physiological or pharmacologically induced). We will further couple this method with investigations at specific circuit and cellular elements in the brain using single-cell electrophysiology, optical imaging and optogenetic or selective pharmacological manipulation.

Both rats and mice will undergo general surgical anesthesia and receive analgesia to implant probes and prepare animals for the experiments. Post-operative care and analgesia will be given to alleviate suffering. Electrodes will be placed over the cranium, directly on the dural surface or intra-cranially. Cannulae and optic fibers will be implanted intra-cranially. Chronic cranial windows will be created for imaging, optogenetic stimulation or single cell recording. Brain activity will be measured electrically and optically (single and multi-photon), during natural states (awake, sleeping) or while receiving different anesthetics (propofol, midazolam, ketamine, xylazine, pentobarbital, thiopental, sevoflurane, xenon). Brain activity will be also modulated optically (by optic fibers or wide-field optogenetic illumination) or chemically (by local delivering of drugs or photo-switchable molecules).

Methods to measure conscious states are being developed in multiple laboratories around the world, with the intention of being translatable to the investigation of consciousness in humans, particularly in patients with consciousness disorders following brain trauma or patients undergoing general anesthesia that may occasionally consciously experience pain during surgical procedures. Most of the current methods cannot reliably detect consciousness in humans, causing uncertainty about the proper treatment and hence increased risk of suffering in patients with disorders of consciousness. This uncertainty is mainly due to the lack of detailed knowledge of the neuronal mechanisms that sustain awareness and its transition to unconsciousness. The aims and expected value of the proposed experiments is to deepen our understanding of the neural circuits involved in the generation and interruption of conscious states, to permit a more reliable assessment of consciousness in humans and to establish the necessary knowledge to develop effective interventions in patients affected by disorders of consciousness.

Animals to be used: 160 rats (Sprague Dawley), adults males, minimum 200 grams, and 160 mice (wildtype and transgenic C57bl), adults, male (also female if transgenic), minimum 20 grams to be used over a period of 4 years. Additionally, 800 mice will be required for the breeding of transgenic strains for the lifetime of the experiments. We have used power analysis to estimate the number of animals required for these experiments, and will conduct experiments that adhere to the aims of the three R's in animal research: Replacement, Reduction and Refinement.