Renstrom, Jacco
(2024)
Prefrontal GABAergic Inhibition and Reversal Learning.
PhD thesis, University of Nottingham.
Abstract
Neural disinhibition, that is reduced GABAergic inhibition, within the prefrontal cortex (PFC) has been suggested to play a key role in the presentation of cognitive impairments in clinical populations, such as schizophrenia. Additionally, several reports have highlighted period of hypofrontality (i.e., reduced activation of the prefrontal cortex or PFC) in patients, particularly during tasks of executive function. Alongside this pathophysiology of schizophrenia, patients have been found to exhibit marked reversal learning deficits, correlating with other symptoms of the condition. Although reversal learning is primarily associated with the orbitofrontal cortex rather than the PFC, earlier evidence suggests that the PFC may still play a role in reversal learning, especially during more demanding reversals. Moreover, we propose that even when the PFC's involvement isn't required, prefrontal neural disinhibition could still hinder reversal performance. This is due to the resulting aberrant firing of prefrontal neurons potentially disrupting processing in projection sites, including the OFC.
To explore a potential relationship between medial prefrontal GABAergic dysfunction and reversal learning, we set out to measure the effect of bi- directional manipulations of prefrontal GABAA activity in rats on two variations of an operant two-lever reversal task. Of particular interest were early task stages where rats were unfamiliar with task demands, potentially resulting in a reliance on prefrontal activity, and later task stages where extensive training resulted in ‘established’ reversal performance.
For this, we first validated a within-subject design of the common operant reversal learning protocol, which could subsequently build the foundations of a serial reversal task (chapter 2). At this stage we also applied a novel Bayesian strategy model which estimated rats’ probabilities of applying certain response strategies at a trial-by-trial resolution. Findings from this experiment underlined a clear distinction between early and late stages of reversal learning in terms of reversal speed and underlying strategy implementation. Subsequently, we applied both paradigms (early and serial) in a pharmacological investigation of reversal performance under the influence of GABAA agonist (muscimol) resulting in prefrontal functional inhibition, or antagonist (picrotoxin) resulting in neural disinhibition (chapter 3). Findings from these experiments indicated a reliance on prefrontal functioning to guide exploration in order to overcome the marked reversal cost during early task stages, but once the task had been established the PFC was no longer required. On the other hand, neural disinhibition disrupted serial, but not early, reversal learning. Further examination revealed a dual impairment in exploration and exploitation underlying this reversal deficit. We hypothesised that the impairment following prefrontal disinhibition to be, at least in part, related to changes in neural firing within the PFC resulting in aberrant neural projection, ultimately disrupting functional processing in prefrontal projection sites more directly implicated in reversal learning.
Finally, we set out to build on findings from chapter 3 by validating a novel chemogenetic approach of prefrontal manipulation, via designer receptors exclusively activated by designer drugs (DREADDs). This approach utilised a transgenic rat line, expressing non-native Cre-recombinase mRNA under the control of the endogenous vesicular GABA transporter (VGAT) promoter. The cell- type specific presence of Cre-recombinase enabled the readout and expression of the inhibitory hM4Di DREADD at GABAergic cells only. Subsequent activation of these receptors via pharmacologically inert actuators, such as clozapine-N-oxide (CNO), would result in the silencing of inhibitory GABA-releasing cells pre- synaptically. However, whilst several similar models have already been established in mice, none to date have been validated in rat lines. Therefore, chapter 4 discusses several histological investigations evaluating feasibility of the rat line, as well as penetrance and cell-type specificity of the DREADD expression within the PFC of a transgenic Long Evans rats. Finally, electrophysiological recordings within the PFC of anaesthetised animals supported functionality of the DREADD, with neural burst firing and local field potential patterns resembling those seen following pharmacologically induced prefrontal disinhibition (Pezze et al., 2014).
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