Sleep-Like Neurophysiology Under  Chloral Hydrate and Urethane Anesthesia
Creators: Ward-Flanagan, Rachel
Publisher: name: University of Alberta Library
Year: 2023

<p>Sleep is a vital neurobiological process, yet despite its fundamental significance, delineating the endogenous neural pathways involved has been slow to progress due to a lack of diverse sleep models. Anesthesia, which has direct behavioural parallels to natural sleep, and which is often linked with endogenous sleep-wake systems on both mechanistic and physiological levels, is an obvious choice to model specific components of sleep. In particular, the research anesthetic urethane has long been regarded as an unparalleled model for the archetypical electrophysiological dynamics of natural sleep; specifically, the spontaneous alternations between both activated and deactivated forebrain states, and accompanying physiological measures. <br>
Yet, there have remained a few drawbacks for urethane as a model for sleep. First, it remained undetermined how urethane promoted its sleep-like neurophysiological effects, and second that urethane is limited to acute experimental paradigms due to ethical considerations. Consequently, the overall aim of my thesis was twofold: to determine if any anesthetics not limited to acute experimental paradigms produced sleep-like alternations in neurophysiology at a surgical plane, and to assess if sleep and urethane had similar profiles of neural activity in known sleep-wake nuclei. <br>
In Chapter 2, I used identical recording conditions to compare the spontaneous brain states of five common research anesthetics with urethane anesthesia. At a surgical plane of anesthesia, three of the anesthetics tested produced a coma-like brain state of burst-suppression activity (pentobarbital, isoflurane, and propofol), ketamine-xylazine produced synchronized slow-oscillatory state akin to slow-wave sleep. Critically, any changes to a state of forebrain activation under these anesthetics coincided with a loss of the surgical plane of anesthesia. Conversely, chloral hydrate produced sleep-like spontaneous alternations of brain state, comparable to those observed under urethane. Unlike the other anesthetics, these alternations were not due to changes in anesthetic depth. Thus, I showed that chloral hydrate could provide an alternate model to urethane for sleep-like brain state alternations in non-acute experimental paradigms.<br>
In Chapter 3, using c-Fos immunoreactivity as a proxy for neural activity, I showed that urethane anesthesia produced a similar pattern of neural activation in known sleep-wake nuclei as unpressured natural sleep. Animals engaging in pressured sleep (also referred to as recovery sleep) following sleep-deprivation had slightly different patterns of neural activation, likely due to the rebound of slow-wave sleep following sleep deprivation. Patterns of neural activity across unpressured (no sleep-deprivation) sleep, and both pressured and unpressured urethane were highly comparable. It is important to note, that all sleep and urethane groups had patterns of neural activation that were vastly divergent from the forced wake behavioural group, indicating that differences between sleep and urethane groups were not due to arousal. This study shows that neural activity under urethane is highly stereotyped, and most representative of normal sleep state architecture in rats.<br>
In Chapter 4, I investigated the assertion that chloral hydrate provides insufficient analgesia for surgical manipulations. The majority of these claims were in reference textbooks, and were in direct conflict with the evidence in primary literature. In agreement with the primary literature, my results showed that intravenous chloral hydrate significantly increased tail withdrawal latencies to a noxious thermal stimulus. Furthermore, as some trials did result in a tail flick response, it is clear that this effect is due to analgesia, not immobility. This was also confirmed by the absence of changes in heart or respiration rate to the nociceptive stimulus. With these experiments I showed that chloral hydrate provides sufficient analgesia to be used as a sole anesthetic, and therefore if used as a model of sleep will not require additional analgesics which could influence experimental outcomes.<br>
In Chapter 5, I characterized in detail the strengths and weaknesses of chloral hydrate as a pharmacological model of sleep. Chloral hydrate produces an incredibly similar neurophysiological profile to both sleep and urethane, including individual brain states of forebrain activation and deactivation, the timing of the alternations between these states, and the sensitivity of these alternations to cholinergic manipulations. Furthermore, respiratory rate, respiratory variability and sighs all fluctuate as a function of brain state the same way under sleep, urethane and chloral hydrate. There are some minor differences between sleep, urethane and chloral hydrate, including an overall slowing of hippocampal theta during forebrain activation that occurs in both anesthetics, although this occurs more profoundly under chloral hydrate. Additionally, there were no discernable changes in heart rate across state under chloral hydrate. However, when these caveats are accounted for in an experimental design, chloral hydrate represents an exciting new tool to model sleep-like brain state and breathing without the constraints of an acute paradigm.<br>
    Increasingly, the overlaps and divergences between components of sleep and specific anesthetic agents are offering insight into pharmacological mechanisms and fundamental nuclei involved in the induction and maintenance of unconsciousness. The results of my thesis suggest that urethane and chloral hydrate could be invaluable tools to probe the intersections of sleep and anesthesia.</p>